You are here

Caron, Marc G.

Overview:

Studies of the mechanisms of action and regulation of hormones and neurotransmitters at the cellular and molecular levels constitute the main goals our of research activities. G protein-coupled receptors (GPCR) mediate the actions of signaling molecules from unicellular organisms to man. We have used adrenergic and dopamine receptors to characterize the structure/function and regulation mechanisms of these prototypes of G protein-coupled receptors. Another approach has been to characterize the nature of neurotransmitter transporters for dopamine, serotonin and other neurotransmitters in an attempt to understand their function in normal and pathological neurotransmission. Another goal of our laboratory is to define the genes and pathways involved in the reinforcing properties of drugs of abuse using both forward and reverse genetic approaches. Our laboratory uses biochemical, molecular biology and gene targeting approaches to examine these questions.

Positions:

James B. Duke Professor of Cell Biology

Cell Biology
School of Medicine

Professor of Cell Biology

Cell Biology
School of Medicine

Vice Chair for Science and Research in the Department of Cell Biology

Cell Biology
School of Medicine

Professor in Neurobiology

Neurobiology
School of Medicine

Professor in Medicine

Medicine, Cardiology
School of Medicine

Faculty Network Member of the Duke Institute for Brain Sciences

Duke Institute for Brain Sciences
Institutes and Provost's Academic Units

Member of the Duke Cancer Institute

Duke Cancer Institute
School of Medicine

Affiliate of the Regeneration Next Initiative

Regeneration Next Initiative
School of Medicine

Education:

Ph.D. 1973

Ph.D. — University of Miami

News:

Grants:

Mechanistic studies of Mas receptor activation and its role in aortic aneurysm formation

Administered By
Medicine, Cardiology
AwardedBy
National Institutes of Health
Role
Collaborator
Start Date
July 15, 2017
End Date
June 30, 2022

Duke Training Grant in Nephrology

Administered By
Medicine, Nephrology
AwardedBy
National Institutes of Health
Role
Preceptor
Start Date
September 20, 1995
End Date
June 30, 2022

Exploiting Dopamine Receptor Functional Selectivity as an Approach to Treat Parkinson's Symptoms

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
May 15, 2017
End Date
February 28, 2022

Translational Research in Surgical Oncology

Administered By
Surgery, Surgical Sciences
AwardedBy
National Institutes of Health
Role
Mentor
Start Date
January 01, 2002
End Date
August 31, 2021

Multidisciplinary Heart and Vascular Diseases

Administered By
Medicine, Cardiology
AwardedBy
National Institutes of Health
Role
Mentor
Start Date
July 01, 1975
End Date
March 31, 2021

New Methods for the Synthesis and Study of Bioactive Nitrogen-Containing Molecules

Administered By
Chemistry
AwardedBy
National Institutes of Health
Role
Collaborator
Start Date
January 05, 2017
End Date
December 31, 2020

Akt/GSK-3 Signaling Cascade and the Actions of Dopamine

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
May 17, 2005
End Date
December 31, 2020

Establishing the molecular and cellular mechanisms of Lgr5 signaling for controlling cancer stem cell behavior

Administered By
Surgery, Surgical Sciences
AwardedBy
National Institutes of Health
Role
Collaborator
Start Date
September 01, 2017
End Date
August 31, 2020

Control of Excitatory Synapse Formation by Huntingtin

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Co-Mentor
Start Date
May 01, 2017
End Date
April 30, 2020

Biased Agonism at Neurotensin Receptor 1 for the Treatment of Methamphetamine Addiction

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
May 01, 2017
End Date
April 30, 2020

Dissecting the Distinct Arrestin Signaling Capabilities of the Short and Long Dopamine D2 Receptors as a Paradigm for Psychostimulant Addiction Treatment

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
July 01, 2016
End Date
June 30, 2019

Drug Abuse: Discovering Ligands for Pertinent GPCRs

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
July 01, 2010
End Date
April 30, 2019

Basic predoctoral training in neuroscience

Administered By
Neurobiology
AwardedBy
National Institutes of Health
Role
Training Faculty
Start Date
July 01, 1992
End Date
June 30, 2018

Cellular and Molecular Signatures of Functionally-Selective D2R Ligands

Administered By
Cell Biology
AwardedBy
University of North Carolina - Chapel Hill
Role
Principal Investigator
Start Date
August 10, 2012
End Date
April 30, 2018

A Cancer Rainbow Mouse for Simultaneous Assessment of Multiple Oncogenes

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
February 09, 2015
End Date
January 31, 2018

The 5-HT theory of depression tested in a naturalistic model of 5-HT deficiency

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
December 15, 2006
End Date
August 31, 2017

NARSAD Prize - Marc Caron / Nikhil Urs

Administered By
Cell Biology
AwardedBy
Brain and Behavior Research Foundation
Role
Principal Investigator
Start Date
December 01, 2013
End Date
November 30, 2016

Screening and identification of beta-arrestin biased dopamine receptor ligands for PD symptom therapy.

Administered By
Cell Biology
AwardedBy
Michael J. Fox Foundation for Parkinson's Research
Role
Co Investigator
Start Date
March 01, 2016
End Date
October 31, 2016

Chemical Biology Approaches to Combat Parkinson's Disease and Dyskinesia

Administered By
Chemistry
AwardedBy
National Institutes of Health
Role
Collaborator
Start Date
August 01, 2013
End Date
July 31, 2016

Ultra-resolution imaging of brain circuitry and its development in mental health

Administered By
Duke-UNC Center for Brain Imaging and Analysis
AwardedBy
National Institutes of Health
Role
Collaborator
Start Date
May 03, 2013
End Date
June 30, 2016

Role of domaine D2 receptor biased signaling in antipsychotic responsive behaviours in genetically engineered mouse models

Administered By
Cell Biology
AwardedBy
Universite Laval
Role
Principal Investigator
Start Date
February 11, 2015
End Date
March 31, 2016

Beta-arrestin Regulation of Ghrelin Signaling in Modulating Addictive Behavior

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Co Investigator
Start Date
April 01, 2014
End Date
March 31, 2016

Cancer Biology Training Grant

Administered By
Pharmacology & Cancer Biology
AwardedBy
National Cancer Institute
Role
Mentor
Start Date
July 01, 1993
End Date
March 31, 2016

Validation of a novel mouse model of dyskinesia in Parkinson's disease

Administered By
Cell Biology
AwardedBy
Canadian Institutes of Health Research
Role
Principal Investigator
Start Date
April 01, 2015
End Date
March 01, 2016

Training in Fundamental &Translational Neuroscience

Administered By
Neurobiology
AwardedBy
National Institutes of Health
Role
Training Faculty
Start Date
July 01, 2005
End Date
February 29, 2016

Consequences of naturalistic serotonin deficiency for suicide-related social endophenotypes in mice

Administered By
Cell Biology
AwardedBy
American Foundation for Suicide Prevention
Role
Co Investigator
Start Date
October 01, 2013
End Date
October 01, 2015

Deciphering Drug Abuse Induced Mechanisms of Cellular Plasticity

Administered By
Cell Biology
AwardedBy
Ruth K. Broad Biomedical Research Foundation
Role
Principal Investigator
Start Date
July 01, 2014
End Date
August 31, 2015

Clinical Oncology Research Career Development Program

Administered By
Surgery, Surgical Sciences
AwardedBy
National Institutes of Health
Role
Mentor
Start Date
September 29, 2009
End Date
July 31, 2015

Novel high-throughput screening for modifiers of TorsinA pathology

Administered By
Neurology, Movement Disorders
AwardedBy
National Institutes of Health
Role
Collaborator
Start Date
April 01, 2013
End Date
March 31, 2015

A Cancer Rainbow Mouse for the Simultaneous Assessment of Multiple Oncogenes

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
September 12, 2012
End Date
February 28, 2015

Whole brain multimodal microscopy of an apoptosis reporter mouse

Administered By
Psychiatry & Behavioral Sciences, Translational Neuroscience
AwardedBy
National Institutes of Health
Role
Collaborator
Start Date
September 01, 2012
End Date
August 31, 2014

Negative regulation of NF-kB by optineurin in primary open angle glaucoma

Administered By
Ophthalmology
AwardedBy
National Institutes of Health
Role
Co-Mentor
Start Date
September 30, 2011
End Date
July 31, 2014

Discovering Ligands for a5-a3-b4 Nicotinic Acetylcholine Receptors.

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
July 01, 2010
End Date
June 30, 2014

Non-invasive Chemical Genetic Control of Neuronal Activity

Administered By
Neurobiology
AwardedBy
National Institutes of Health
Role
Collaborator
Start Date
September 30, 2009
End Date
May 31, 2014

The Impact of Serotonin Deficiency

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
March 01, 2011
End Date
February 28, 2014

Agilent Direct Drive 9.4T MRS/MRI Console

Administered By
Radiology
AwardedBy
National Institutes of Health
Role
Major User
Start Date
May 15, 2012
End Date
November 14, 2013

Beta-catenin modulates dopamine dependent signal transduction and behavior.

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
August 08, 2011
End Date
September 30, 2013

Role of beta-arrestin-2 signaling complexes in the actions of psychostimulants

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
July 01, 2010
End Date
June 30, 2013

Small molecule screen to enhance epithelial repair

Administered By
Medicine, Pulmonary, Allergy, and Critical Care Medicine
AwardedBy
National Institutes of Health
Role
Collaborator
Start Date
July 01, 2011
End Date
December 31, 2012

Small Molecule Agonsists for the Neurotensin 1 Receptor

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Collaborator
Start Date
December 04, 2009
End Date
October 31, 2012

IsoCyte Laser Scanning Plate Cytometer for High-throughput, High-content Assays

Administered By
Molecular Genetics and Microbiology
AwardedBy
National Institutes of Health
Role
Major User
Start Date
April 01, 2011
End Date
March 31, 2012

Behavioral and Synaptic Consequences of MeCP2 Phosphorylation

Administered By
Neurobiology
AwardedBy
National Institutes of Health
Role
Co-Sponsor
Start Date
December 01, 2010
End Date
December 31, 2011

Effects of Chronic Antidepressants on Mice with A Mutation in Tph2

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
May 01, 2008
End Date
May 31, 2011

Pharmacometabolomics Research Network

Administered By
Psychiatry & Behavioral Sciences, Translational Neuroscience
AwardedBy
National Institutes of Health
Role
Co Investigator
Start Date
March 15, 2010
End Date
February 28, 2011

Toxicant evaluation in mice with selective dopamine transporter overexpression

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Co-Mentor
Start Date
September 01, 2008
End Date
August 31, 2010

Drug Abuse: Discovering Ligands for Pertinent GPCRs

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
September 30, 2006
End Date
June 30, 2010

An ENU Mutagenesis Screen for Genes of Addiction

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Mentor
Start Date
April 01, 2006
End Date
July 01, 2009

Dopamine Receptor Regulation by GRKs in Drug Abuse

Administered By
Medicine, Gastroenterology
AwardedBy
National Institutes of Health
Role
Consultant
Start Date
August 15, 2004
End Date
March 31, 2009

Dopamine Receptors: Characterization And Regulation

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
December 01, 1983
End Date
May 31, 2008

Elucidating the actions of acetylcholine in the brain

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
April 15, 2005
End Date
March 31, 2008

Molecular studies of osmotic neural sensing

Administered By
Neurology, Headache and Pain
AwardedBy
National Institute of Mental Health
Role
Co-Sponsor
Start Date
January 10, 2002
End Date
December 31, 2006

Arrestins in G Protein-Coupled Receptor Trafficking

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
August 01, 2003
End Date
August 31, 2006

Genetic Analysis of Dopaminergic Reward Mechanisms

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
May 15, 2000
End Date
April 30, 2006

Molecular fingerprinting of GPCR ligands in Cancer

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Co Investigator
Start Date
March 01, 2004
End Date
February 28, 2006

GRK/Beta-arrestin in Sonic hedgehog signaling.

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
September 01, 2004
End Date
August 31, 2005

Proteomic/Genetic Approaches to Monoamine Transporters

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Mentor
Start Date
September 01, 2003
End Date
June 30, 2004

Mechanisms of G Protein Coupled Receptor Regulation

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Collaborator
Start Date
July 01, 1999
End Date
May 31, 2004

Dopamine & Glutamate Pathways in Models of Schizophrenia

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
July 01, 2000
End Date
January 31, 2004

Cell/Molecular Regulation of Human Dopamine Transporter

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
March 01, 2001
End Date
September 08, 2003

Presynaptic Dopamine Mechanisms After Chronic Cocaine

Administered By
Psychiatry & Behavioral Sciences, Addictions
AwardedBy
National Institutes of Health
Role
Co Investigator
Start Date
February 01, 1997
End Date
November 30, 2002

Novel secretin receptor interacting proteins (Small Grant Program)

Administered By
Medicine, Gastroenterology
AwardedBy
National Institutes of Health
Role
Mentor
Start Date
January 01, 2001
End Date
September 30, 2002

Morphine Sensitivity in GRK and Arrestin Knockout Mice

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
April 01, 2000
End Date
March 31, 2002

Presynaptic Dopamine Mechanisms After Chronic Cocaine

Administered By
Psychiatry & Behavioral Sciences, Translational Neuroscience
AwardedBy
National Institutes of Health
Role
Co-Principal Investigator
Start Date
February 01, 1997
End Date
November 30, 1999

Crc/Pe For The Study Of Depression

Administered By
Psychiatry & Behavioral Sciences, Geriatric Behavioral Health
AwardedBy
National Institutes of Health
Role
Co-Principal Investigator
Start Date
September 30, 1984
End Date
July 31, 1999

Comprehensive Cancer Center Core Support Grant

Administered By
Medicine, Medical Oncology
AwardedBy
National Institutes of Health
Role
Co-Principal Investigator
Start Date
September 01, 1976
End Date
December 31, 1998

Comprehensive Cancer Center Core Support Grant

Administered By
Medicine, Medical Oncology
AwardedBy
National Institutes of Health
Role
Co-Principal Investigator
Start Date
September 01, 1976
End Date
December 31, 1998

Dopamine Receptors: Characterization And Regulation

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
December 01, 1994
End Date
November 30, 1998

Dopamine Receptors: Characterization And Regulation

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
December 15, 1991
End Date
November 30, 1998

Dopamine Receptors - Characterization And Regulation

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
December 01, 1983
End Date
November 30, 1998

Pesticides And Dopaminergic Function

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Co-Principal Investigator
Start Date
September 30, 1997
End Date
September 30, 1998

Molecular Properties Of Cardiac Adrenergic Receptors

Administered By
Medicine, Cardiology
AwardedBy
National Institutes of Health
Role
Co-Principal Investigator
Start Date
September 01, 1976
End Date
August 31, 1996

An Investigation Of The Specificity Of Novel Ligans For Me

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
April 26, 1993
End Date
April 25, 1994

Dopamine Receptors: Characterization & Endocrine Regula.

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
December 01, 1990
End Date
November 30, 1991

Dopamine Receptors: Characterization & Endocrine Regulati.

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
December 01, 1989
End Date
November 30, 1991

Dopamine Receptors: Characterization & Endocrine Regulatio

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
December 01, 1988
End Date
November 30, 1991

Dopamine Receptors: Characterization And Endocrine Regulat

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
December 01, 1987
End Date
November 30, 1991

Dopamine Receptors: Characterization/Endocrine Regulation

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
December 01, 1985
End Date
November 01, 1986
Show More

Awards:

Julius Axelrod Award. American Society for Pharmacology and Experimental Therapeutics.

Type
National
Awarded By
American Society for Pharmacology and Experimental Therapeutics
Date
January 01, 2005

Investigator/Alumni Investigator. Howard Hughes Medical Institute.

Type
National
Awarded By
Howard Hughes Medical Institute
Date
January 01, 1992

Publications:

Ghrelin receptor antagonism of hyperlocomotion in cocaine-sensitized mice requires βarrestin-2.

The "brain-gut" peptide ghrelin, which mediates food-seeking behaviors, is recognized as a very strong endogenous modulator of dopamine (DA) signaling. Ghrelin binds the G protein-coupled receptor GHSR1a, and administration of ghrelin increases the rewarding properties of psychostimulants while ghrelin receptor antagonists decrease them. In addition, the GHSR1a signals through βarrestin-2 to regulate actin/stress fiber rearrangement, suggesting βarrestin-2 participation in the regulation of actin-mediated synaptic plasticity for addictive substances like cocaine. The effects of ghrelin receptor ligands on reward strongly suggest that modulation of ghrelin signaling could provide an effective strategy to ameliorate undesirable behaviors arising from addiction. To investigate this possibility, we tested the effects of ghrelin receptor antagonism in a cocaine behavioral sensitization paradigm using DA neuron-specific βarrestin-2 KO mice. Our results show that these mice sensitize to cocaine as well as wild-type littermates. The βarrestin-2 KO mice, however, no longer respond to the locomotor attenuating effects of the GHSR1a antagonist YIL781. The data presented here suggest that the separate stages of addictive behavior differ in their requirements for βarrestin-2 and show that pharmacological inhibition of βarrestin-2 function through GHSR1a antagonism is not equivalent to the loss of βarrestin-2 function achieved by genetic ablation. These data support targeting GHSR1a signaling in addiction therapy but indicate that using signaling biased compounds that modulate βarrestin-2 activity differentially from G protein activity may be required.

Authors
Toth, K; Slosky, LM; Pack, TF; Urs, NM; Boone, P; Mao, L; Abraham, D; Caron, MG; Barak, LS
MLA Citation
Toth, K, Slosky, LM, Pack, TF, Urs, NM, Boone, P, Mao, L, Abraham, D, Caron, MG, and Barak, LS. "Ghrelin receptor antagonism of hyperlocomotion in cocaine-sensitized mice requires βarrestin-2." Synapse (New York, N.Y.) (September 23, 2017).
PMID
28941296
Source
epmc
Published In
Synapse
Publish Date
2017
DOI
10.1002/syn.22012

Hepatic β-arrestin 2 is essential for maintaining euglycemia.

An increase in hepatic glucose production (HGP) represents a key feature of type 2 diabetes. This deficiency in metabolic control of glucose production critically depends on enhanced signaling through hepatic glucagon receptors (GCGRs). Here, we have demonstrated that selective inactivation of the GPCR-associated protein β-arrestin 2 in hepatocytes of adult mice results in greatly increased hepatic GCGR signaling, leading to striking deficits in glucose homeostasis. However, hepatocyte-specific β-arrestin 2 deficiency did not affect hepatic insulin sensitivity or β-adrenergic signaling. Adult mice lacking β-arrestin 1 selectively in hepatocytes did not show any changes in glucose homeostasis. Importantly, hepatocyte-specific overexpression of β-arrestin 2 greatly reduced hepatic GCGR signaling and protected mice against the metabolic deficits caused by the consumption of a high-fat diet. Our data support the concept that strategies aimed at enhancing hepatic β-arrestin 2 activity could prove useful for suppressing HGP for therapeutic purposes.

Authors
Zhu, L; Rossi, M; Cui, Y; Lee, RJ; Sakamoto, W; Perry, NA; Urs, NM; Caron, MG; Gurevich, VV; Godlewski, G; Kunos, G; Chen, M; Chen, W; Wess, J
MLA Citation
Zhu, L, Rossi, M, Cui, Y, Lee, RJ, Sakamoto, W, Perry, NA, Urs, NM, Caron, MG, Gurevich, VV, Godlewski, G, Kunos, G, Chen, M, Chen, W, and Wess, J. "Hepatic β-arrestin 2 is essential for maintaining euglycemia." Journal of Clinical Investigation 127.8 (August 2017): 2941-2945.
PMID
28650340
Source
epmc
Published In
Journal of Clinical Investigation
Volume
127
Issue
8
Publish Date
2017
Start Page
2941
End Page
2945
DOI
10.1172/jci92913

Design, synthesis and biological evaluation of GPR55 agonists.

GPR55, a G protein-coupled receptor, is an attractive target to alleviate inflammatory and neuropathic pain and treat osteoporosis and cancer. Identifying a potent and selective ligand will aid to further establish the specific physiological roles and pharmacology of the receptor. Towards this goal, a targeted library of 22 compounds was synthesized in a modular fashion to obtain structure-activity relationship information. The general route consisted of coupling a variety of p-aminophenyl sulfonamides to isothiocyanates to form acylthioureas. For the synthesis of a known naphthyl ethyl alcohol motif, route modification led to a shorter and more efficient process. The 22 analogues were analyzed for their ability to serve as agonists at GPR55 and valuable information for both ends of the molecule was ascertained.

Authors
Fakhouri, L; Cook, CD; Al-Huniti, MH; Console-Bram, LM; Hurst, DP; Spano, MBS; Nasrallah, DJ; Caron, MG; Barak, LS; Reggio, PH; Abood, ME; Croatt, MP
MLA Citation
Fakhouri, L, Cook, CD, Al-Huniti, MH, Console-Bram, LM, Hurst, DP, Spano, MBS, Nasrallah, DJ, Caron, MG, Barak, LS, Reggio, PH, Abood, ME, and Croatt, MP. "Design, synthesis and biological evaluation of GPR55 agonists." Bioorganic & medicinal chemistry 25.16 (August 2017): 4355-4367.
PMID
28673732
Source
epmc
Published In
Bioorganic & Medicinal Chemistry
Volume
25
Issue
16
Publish Date
2017
Start Page
4355
End Page
4367
DOI
10.1016/j.bmc.2017.06.016

Protamine is an antagonist of apelin receptor, and its activity is reversed by heparin.

Apelin signaling plays an important role during embryo development and regulates angiogenesis, cardiovascular activity, and energy metabolism in adulthood. Overexpression and hyperactivity of this signaling pathway is observed in various pathologic states, such as cardiovascular diseases and cancer, which highlights the importance of inhibiting apelin receptor (APJ); therefore, we developed a cell-based screening assay that uses fluorescence microscopy to identify APJ antagonists. This approach led us to identify the U.S. Food and Drug Administration-approved compound protamine-already used clinically after cardiac surgery-as an agent to bind to heparin and thereby reverse its anticlotting activity. Protamine displays a 390-nM affinity for APJ and behaves as a full antagonist with regard to G protein and β-arrestin-dependent intracellular signaling. Ex vivo and in vivo, protamine abolishes well-known apelin effects, such as angiogenesis, glucose tolerance, and vasodilatation. Remarkably, protamine antagonist activity is fully reversed by heparin treatment both in vitro and in vivo Thus, our results demonstrate a new pharmacologic property of protamine-blockade of APJ-that could explain some adverse effects observed in protamine-treated patients. Moreover, our data reveal that the established antiangiogenic activity of protamine would rely on APJ antagonism.-Le Gonidec, S., Chaves-Almagro, C., Bai, Y., Kang, H. J., Smith, A., Wanecq, E., Huang, X.-P., Prats, H., Knibiehler, B., Roth, B. L., Barak, L. S., Caron, M. G., Valet, P., Audigier, Y., Masri, B. Protamine is an antagonist of apelin receptor, and its activity is reversed by heparin.

Authors
Le Gonidec, S; Chaves-Almagro, C; Bai, Y; Kang, HJ; Smith, A; Wanecq, E; Huang, X-P; Prats, H; Knibiehler, B; Roth, BL; Barak, LS; Caron, MG; Valet, P; Audigier, Y; Masri, B
MLA Citation
Le Gonidec, S, Chaves-Almagro, C, Bai, Y, Kang, HJ, Smith, A, Wanecq, E, Huang, X-P, Prats, H, Knibiehler, B, Roth, BL, Barak, LS, Caron, MG, Valet, P, Audigier, Y, and Masri, B. "Protamine is an antagonist of apelin receptor, and its activity is reversed by heparin." FASEB journal : official publication of the Federation of American Societies for Experimental Biology 31.6 (June 2017): 2507-2519.
PMID
28242772
Source
epmc
Published In
The FASEB journal : official publication of the Federation of American Societies for Experimental Biology
Volume
31
Issue
6
Publish Date
2017
Start Page
2507
End Page
2519
DOI
10.1096/fj.201601074r

Inhibiting clathrin-mediated endocytosis of the leucine-rich G protein-coupled receptor-5 diminishes cell fitness.

The leucine-rich G protein-coupled receptor-5 (LGR5) is expressed in adult tissue stem cells of many epithelia, and its overexpression is negatively correlated with cancer prognosis. LGR5 potentiates WNT/β-catenin signaling through its unique constitutive internalization property that clears negative regulators of the WNT-receptor complex from the membrane. However, both the mechanism and physiological relevance of LGR5 internalization are unclear. Therefore, a natural product library was screened to discover LGR5 internalization inhibitors and gain mechanistic insight into LGR5 internalization. The plant lignan justicidin B blocked the constitutive internalization of LGR5. Justicidin B is structurally similar to more potent vacuolar-type H+-ATPase inhibitors, which all inhibited LGR5 internalization by blocking clathrin-mediated endocytosis. We then tested the physiological relevance of LGR5 internalization blockade in vivo A LGR5-rainbow (LBOW) mouse line was engineered to express three different LGR5 isoforms along with unique fluorescent protein lineage reporters in the same mouse. In this manner, the effects of each isoform on cell fate can be simultaneously assessed through simple fluorescent imaging for each lineage reporter. LBOW mice express three different forms of LGR5, a wild-type form that constitutively internalizes and two mutant forms whose internalization properties have been compromised by genetic perturbations within the carboxyl-terminal tail. LBOW was activated in the intestinal epithelium, and a year-long lineage-tracing course revealed that genetic blockade of LGR5 internalization diminished cell fitness. Together these data provide proof-of-concept genetic evidence that blocking the clathrin-mediated endocytosis of LGR5 could be used to pharmacologically control cell behavior.

Authors
Snyder, JC; Rochelle, LK; Ray, C; Pack, TF; Bock, CB; Lubkov, V; Lyerly, HK; Waggoner, AS; Barak, LS; Caron, MG
MLA Citation
Snyder, JC, Rochelle, LK, Ray, C, Pack, TF, Bock, CB, Lubkov, V, Lyerly, HK, Waggoner, AS, Barak, LS, and Caron, MG. "Inhibiting clathrin-mediated endocytosis of the leucine-rich G protein-coupled receptor-5 diminishes cell fitness." The Journal of biological chemistry 292.17 (April 2017): 7208-7222.
PMID
28275053
Source
epmc
Published In
The Journal of biological chemistry
Volume
292
Issue
17
Publish Date
2017
Start Page
7208
End Page
7222
DOI
10.1074/jbc.m116.756635

Distinct cortical and striatal actions of a β-arrestin-biased dopamine D2 receptor ligand reveal unique antipsychotic-like properties.

The current dopamine (DA) hypothesis of schizophrenia postulates striatal hyperdopaminergia and cortical hypodopaminergia. Although partial agonists at DA D2 receptors (D2Rs), like aripiprazole, were developed to simultaneously target both phenomena, they do not effectively improve cortical dysfunction. In this study, we investigate the potential for newly developed β-arrestin2 (βarr2)-biased D2R partial agonists to simultaneously target hyper- and hypodopaminergia. Using neuron-specific βarr2-KO mice, we show that the antipsychotic-like effects of a βarr2-biased D2R ligand are driven through both striatal antagonism and cortical agonism of D2R-βarr2 signaling. Furthermore, βarr2-biased D2R agonism enhances firing of cortical fast-spiking interneurons. This enhanced cortical agonism of the biased ligand can be attributed to a lack of G-protein signaling and elevated expression of βarr2 and G protein-coupled receptor (GPCR) kinase 2 in the cortex versus the striatum. Therefore, we propose that βarr2-biased D2R ligands that exert region-selective actions could provide a path to develop more effective antipsychotic therapies.

Authors
Urs, NM; Gee, SM; Pack, TF; McCorvy, JD; Evron, T; Snyder, JC; Yang, X; Rodriguiz, RM; Borrelli, E; Wetsel, WC; Jin, J; Roth, BL; O'Donnell, P; Caron, MG
MLA Citation
Urs, NM, Gee, SM, Pack, TF, McCorvy, JD, Evron, T, Snyder, JC, Yang, X, Rodriguiz, RM, Borrelli, E, Wetsel, WC, Jin, J, Roth, BL, O'Donnell, P, and Caron, MG. "Distinct cortical and striatal actions of a β-arrestin-biased dopamine D2 receptor ligand reveal unique antipsychotic-like properties." Proceedings of the National Academy of Sciences of the United States of America 113.50 (December 2016): E8178-E8186.
PMID
27911814
Source
epmc
Published In
Proceedings of the National Academy of Sciences of USA
Volume
113
Issue
50
Publish Date
2016
Start Page
E8178
End Page
E8186
DOI
10.1073/pnas.1614347113

Adjunctive 5-Hydroxytryptophan Slow-Release for Treatment-Resistant Depression: Clinical and Preclinical Rationale.

Serotonin transporter (SERT) inhibitors treat depression by elevating brain extracellular 5-hydroxytryptamine (5-HTExt). However, only one-third of patients respond adequately. Treatment-resistant depression (TRD) is a major unmet need. Interestingly, elevating 5-HTExt beyond what is achieved by a SERT inhibitor appears to treat TRD. Adjunctive administration of 5-hydroxytryptophan (5-HTP) safely elevates 5-HTExt beyond the SERT inhibitor effect in humans; however, 5-HTP cannot be a clinically viable drug because of its poor pharmacokinetics. A slow-release (SR) delivery mode would be predicted to overcome the pharmacokinetic limitations of 5-HTP, substantially enhancing the pharmacological action and transforming 5-HTP into a clinically viable drug. Animal studies bear out this prediction. Thus, adjunct 5-HTP SR could be an important new treatment for TRD. Here, we review the clinical and preclinical evidence for this treatment.

Authors
Jacobsen, JPR; Krystal, AD; Krishnan, KRR; Caron, MG
MLA Citation
Jacobsen, JPR, Krystal, AD, Krishnan, KRR, and Caron, MG. "Adjunctive 5-Hydroxytryptophan Slow-Release for Treatment-Resistant Depression: Clinical and Preclinical Rationale." Trends in pharmacological sciences 37.11 (November 2016): 933-944. (Review)
PMID
27692695
Source
epmc
Published In
Trends in Pharmacological Sciences
Volume
37
Issue
11
Publish Date
2016
Start Page
933
End Page
944
DOI
10.1016/j.tips.2016.09.001

SSRI Augmentation by 5-Hydroxytryptophan Slow Release: Mouse Pharmacodynamic Proof of Concept.

Drugs, notably SSRIs, that elevate brain extracellular 5-HT (5-HTExt) are antidepressants. Unfortunately, most patients fail to remit. Multipronged clinical evidence suggests that elevating 5-HTExt beyond the SSRI effect enhances antidepressant efficacy, but previous such drug strategies had prohibitive limitations. In humans, adjunct treatment with the 5-HT precursor 5-hydroxytryptophan (5-HTP) elevates 5-HTExt beyond the SSRI effect. Small pilot trials suggest that adjunct 5-HTP can confer antidepressant response in treatment-resistant depression (TRD). However, sustained, stable 5-HTExt elevation is required for antidepressant effect; therefore, the rapid absorption and elimination of standard 5-HTP immediate release (IR) likely curtail 5-HTP IR's antidepressant potential. Slow-release (SR) drug delivery can crucially improve efficacy and safety of rapidly absorbed and eliminated compounds. Here we tested in mice the hypothesis that SR delivery will substantially improve 5-HTP's drug properties, by minimizing adverse effects and securing sustained 5-HTExt elevation beyond the SSRI effect. We modeled 5-HTP SR with minipumps, 5-HTP IR with injections, and chronic SSRI with dietary fluoxetine. We tested adjunct 5-HTP SR in wild-type mice and in mice with low brain 5-HT owing to expression of a mutant form of the brain 5-HT synthesis enzyme, tryptophan hydroxylase 2. In both lines of mice, adjunct 5-HTP SR synergized with SSRI to elevate 5-HTExt beyond the SSRI effect. We observed no adverse effect. Adjunct 5-HTP IR could not produce this therapy-like profile, producing transient 5-HTExt spikes and marked adverse effects. Integrated with a body of clinical data, our mouse data suggest that an adjunct 5-HTP SR drug could safely and effectively elevate 5-HTExt beyond the SSRI effect and represent a novel treatment for TRD.

Authors
Jacobsen, JP; Rudder, ML; Roberts, W; Royer, EL; Robinson, TJ; Oh, A; Spasojevic, I; Sachs, BD; Caron, MG
MLA Citation
Jacobsen, JP, Rudder, ML, Roberts, W, Royer, EL, Robinson, TJ, Oh, A, Spasojevic, I, Sachs, BD, and Caron, MG. "SSRI Augmentation by 5-Hydroxytryptophan Slow Release: Mouse Pharmacodynamic Proof of Concept." Neuropsychopharmacology : official publication of the American College of Neuropsychopharmacology 41.9 (August 2016): 2324-2334.
PMID
26932820
Source
epmc
Published In
Neuropsychopharmacology (Nature)
Volume
41
Issue
9
Publish Date
2016
Start Page
2324
End Page
2334
DOI
10.1038/npp.2016.35

ML314: A Biased Neurotensin Receptor Ligand for Methamphetamine Abuse.

Pharmacological treatment for methamphetamine addiction will provide important societal benefits. Neurotensin receptor NTR1 and dopamine receptor distributions coincide in brain areas regulating methamphetamine-associated reward, and neurotensin peptides produce behaviors opposing psychostimulants. Therefore, undesirable methamphetamine-associated activities should be treatable with druggable NTR1 agonists, but no such FDA-approved therapeutics exist. We address this limitation with proof-of-concept data for ML314, a small-molecule, brain penetrant, β-arrestin biased, NTR1 agonist. ML314 attenuates amphetamine-like hyperlocomotion in dopamine transporter knockout mice, and in C57BL/6J mice it attenuates methamphetamine-induced hyperlocomotion, potentiates the psychostimulant inhibitory effects of a ghrelin antagonist, and reduces methamphetamine-associated conditioned place preference. In rats, ML314 blocks methamphetamine self-administration. ML314 acts as an allosteric enhancer of endogenous neurotensin, unmasking stoichiometric numbers of hidden NTR1 binding sites in transfected-cell membranes or mouse striatal membranes, while additionally supporting NTR1 endocytosis in cells in the absence of NT peptide. These results indicate ML314 is a viable, preclinical lead for methamphetamine abuse treatment and support an allosteric model of G protein-coupled receptor signaling.

Authors
Barak, LS; Bai, Y; Peterson, S; Evron, T; Urs, NM; Peddibhotla, S; Hedrick, MP; Hershberger, P; Maloney, PR; Chung, TDY; Rodriguiz, RM; Wetsel, WC; Thomas, JB; Hanson, GR; Pinkerton, AB; Caron, MG
MLA Citation
Barak, LS, Bai, Y, Peterson, S, Evron, T, Urs, NM, Peddibhotla, S, Hedrick, MP, Hershberger, P, Maloney, PR, Chung, TDY, Rodriguiz, RM, Wetsel, WC, Thomas, JB, Hanson, GR, Pinkerton, AB, and Caron, MG. "ML314: A Biased Neurotensin Receptor Ligand for Methamphetamine Abuse." ACS chemical biology 11.7 (July 2016): 1880-1890.
PMID
27119457
Source
epmc
Published In
ACS Chemical Biology
Volume
11
Issue
7
Publish Date
2016
Start Page
1880
End Page
1890
DOI
10.1021/acschembio.6b00291

Structural biology: Antidepressants at work.

Authors
Caron, MG; Gether, U
MLA Citation
Caron, MG, and Gether, U. "Structural biology: Antidepressants at work." Nature 532.7599 (April 6, 2016): 320-321.
PMID
27049942
Source
epmc
Published In
Nature
Volume
532
Issue
7599
Publish Date
2016
Start Page
320
End Page
321
DOI
10.1038/nature17883

Design, synthesis, and analysis of antagonists of GPR55: Piperidine-substituted 1,3,4-oxadiazol-2-ones.

A series of 1,3,4-oxadiazol-2-ones was synthesized and tested for activity as antagonists at GPR55 in cellular beta-arrestin redistribution assays. The synthesis was designed to be modular in nature so that a sufficient number of analogues could be rapidly accessed to explore initial structure-activity relationships. The design of analogues was guided by the docking of potential compounds into a model of the inactive form of GPR55. The results of the assays were used to learn more about the binding pocket of GPR55. With this oxadiazolone scaffold, it was determined that modification of the aryl group adjacent to the oxadiazolone ring was often detrimental and that the distal cyclopropane was beneficial for activity. These results will guide further exploration of this receptor.

Authors
Meza-Aviña, ME; Lingerfelt, MA; Console-Bram, LM; Gamage, TF; Sharir, H; Gettys, KE; Hurst, DP; Kotsikorou, E; Shore, DM; Caron, MG; Rao, N; Barak, LS; Abood, ME; Reggio, PH; Croatt, MP
MLA Citation
Meza-Aviña, ME, Lingerfelt, MA, Console-Bram, LM, Gamage, TF, Sharir, H, Gettys, KE, Hurst, DP, Kotsikorou, E, Shore, DM, Caron, MG, Rao, N, Barak, LS, Abood, ME, Reggio, PH, and Croatt, MP. "Design, synthesis, and analysis of antagonists of GPR55: Piperidine-substituted 1,3,4-oxadiazol-2-ones." Bioorganic & medicinal chemistry letters 26.7 (April 2016): 1827-1830.
PMID
26916440
Source
epmc
Published In
Bioorganic & Medicinal Chemistry Letters
Volume
26
Issue
7
Publish Date
2016
Start Page
1827
End Page
1830
DOI
10.1016/j.bmcl.2016.02.030

Effects of β-Arrestin-Biased Dopamine D2 Receptor Ligands on Schizophrenia-Like Behavior in Hypoglutamatergic Mice.

Current antipsychotic drugs (APDs) show efficacy with positive symptoms, but are limited in treating negative or cognitive features of schizophrenia. Whereas all currently FDA-approved medications target primarily the dopamine D2 receptor (D2R) to inhibit G(i/o)-mediated adenylyl cyclase, a recent study has shown that many APDs affect not only G(i/o)- but they can also influence β-arrestin- (βArr)-mediated signaling. The ability of ligands to differentially affect signaling through these pathways is termed functional selectivity. We have developed ligands that are devoid of D2R-mediated G(i/o) protein signaling, but are simultaneously partial agonists for D2R/βArr interactions. The purpose of this study was to test the effectiveness of UNC9975 or UNC9994 on schizophrenia-like behaviors in phencyclidine-treated or NR1-knockdown hypoglutamatergic mice. We have found the UNC compounds reduce hyperlocomotion in the open field, restore PPI, improve novel object recognition memory, partially normalize social behavior, decrease conditioned avoidance responding, and elicit a much lower level of catalepsy than haloperidol. These preclinical results suggest that exploitation of functional selectivity may provide unique opportunities to develop drugs with fewer side effects, greater therapeutic selectivity, and enhanced efficacy for treating schizophrenia and related conditions than medications that are currently available.

Authors
Park, SM; Chen, M; Schmerberg, CM; Dulman, RS; Rodriguiz, RM; Caron, MG; Jin, J; Wetsel, WC
MLA Citation
Park, SM, Chen, M, Schmerberg, CM, Dulman, RS, Rodriguiz, RM, Caron, MG, Jin, J, and Wetsel, WC. "Effects of β-Arrestin-Biased Dopamine D2 Receptor Ligands on Schizophrenia-Like Behavior in Hypoglutamatergic Mice." Neuropsychopharmacology 41.3 (February 2016): 704-715.
PMID
26129680
Source
epmc
Published In
Neuropsychopharmacology (Nature)
Volume
41
Issue
3
Publish Date
2016
Start Page
704
End Page
715
DOI
10.1038/npp.2015.196

A rapid and affordable screening platform for membrane protein trafficking.

Membrane proteins regulate a diversity of physiological processes and are the most successful class of targets in drug discovery. However, the number of targets adequately explored in chemical space and the limited resources available for screening are significant problems shared by drug-discovery centers and small laboratories. Therefore, a low-cost and universally applicable screen for membrane protein trafficking was developed.This high-throughput screen (HTS), termed IRFAP-HTS, utilizes the recently described MarsCy1-fluorogen activating protein and the near-infrared and membrane impermeant fluorogen SCi1. The cell surface expression of MarsCy1 epitope-tagged receptors can be visualized by simple addition of SCi1. User-friendly, rapid, and quantitative detection occurs on a standard infrared western-blotting scanner. The reliability and robustness of IRFAP-HTS was validated by confirming human vasopressin-2 receptor and dopamine receptor-2 trafficking in response to agonist or antagonist. The IRFAP-HTS screen was deployed against the leucine-rich G protein-coupled receptor-5 (Lgr5). Lgr5 is expressed in stem cells, modulates Wnt/ß-catenin signaling, and is therefore a promising drug target. However, small molecule modulators have yet to be reported. The constitutive internalization of Lgr5 appears to be one primary mode through which its function is regulated. Therefore, IRFAP-HTS was utilized to screen 11,258 FDA-approved and drug-like small molecules for those that antagonize Lgr5 internalization. Glucocorticoids were found to potently increase Lgr5 expression at the plasma membrane.The IRFAP-HTS platform provides a versatile solution for screening more targets with fewer resources. Using only a standard western-blotting scanner, we were able to screen 5,000 compounds per hour in a robust and quantitative assay. Multi-purposing standardly available laboratory equipment eliminates the need for idiosyncratic and more expensive high-content imaging systems. The modular and user-friendly IRFAP-HTS is a significant departure from current screening platforms. Small laboratories will have unprecedented access to a robust and reliable screening platform and will no longer be limited by the esoteric nature of assay development, data acquisition, and post-screening analysis. The discovery of glucocorticoids as modulators for Lgr5 trafficking confirms that IRFAP-HTS can accelerate drug-discovery and drug-repurposing for even the most obscure targets.

Authors
Snyder, JC; Pack, TF; Rochelle, LK; Chakraborty, SK; Zhang, M; Eaton, AW; Bai, Y; Ernst, LA; Barak, LS; Waggoner, AS; Caron, MG
MLA Citation
Snyder, JC, Pack, TF, Rochelle, LK, Chakraborty, SK, Zhang, M, Eaton, AW, Bai, Y, Ernst, LA, Barak, LS, Waggoner, AS, and Caron, MG. "A rapid and affordable screening platform for membrane protein trafficking." BMC biology 13 (December 17, 2015): 107-.
PMID
26678094
Source
epmc
Published In
BMC Biology
Volume
13
Publish Date
2015
Start Page
107
DOI
10.1186/s12915-015-0216-3

Essential role of D1R in the regulation of mTOR complex1 signaling induced by cocaine.

The mammalian target of rapamycin (mTOR) is a serine/threonine kinase that is involved in neuronal adaptions that underlie cocaine-induced sensitization and reward. mTOR exists in two functionally distinct multi-component complexes known as mTORC1 and mTORC2. In this study, we show that increased mTORC1 activity induced by cocaine is mediated by the dopamine D1 receptor (D1R). Specifically, cocaine treatment increased the phosphorylation on residues Thr2446 and Ser2481 but not on Ser2448 in the nucleus accumbens (NAc) and that this increase in phosphorylated mTOR levels was also apparent when complexed with its binding partner Raptor. Furthermore, the increase in phosphorylated mTOR levels, as well as phosphorylated 4E-BP1 and S6K, downstream targets of mTORC1 were blocked with SCH23390 treatment. Similar results were also observed in the dopamine-transporter knockout mice as the increase in phosphorylated mTOR Thr2446 and Ser2481 was blocked by SCH23390 but not with raclopride. To further validate D1R role in mTORC1 signaling, decrease in phosphorylated mTOR levels were observed in D1R knockout mice, whereas administration of SKF81297 elevated phosphorylated mTOR in the NAc. Lastly deletion of mTOR or Raptor in D1R expressing neurons reduced cocaine-induced locomotor activity. Together, our data supports a mechanism whereby mTORC1 signaling is activated by cocaine administration through the stimulation of D1R.

Authors
Sutton, LP; Caron, MG
MLA Citation
Sutton, LP, and Caron, MG. "Essential role of D1R in the regulation of mTOR complex1 signaling induced by cocaine." Neuropharmacology 99 (December 2015): 610-619.
PMID
26314207
Source
epmc
Published In
Neuropharmacology
Volume
99
Publish Date
2015
Start Page
610
End Page
619
DOI
10.1016/j.neuropharm.2015.08.024

Daytime spikes in dopaminergic activity drive rapid mood-cycling in mice.

Disruptions in circadian rhythms and dopaminergic activity are involved in the pathophysiology of bipolar disorder, though their interaction remains unclear. Moreover, a lack of animal models that display spontaneous cycling between mood states has hindered our mechanistic understanding of mood switching. Here, we find that mice with a mutation in the circadian Clock gene (ClockΔ19) exhibit rapid mood-cycling, with a profound manic-like phenotype emerging during the day following a period of euthymia at night. Mood-cycling coincides with abnormal daytime spikes in ventral tegmental area (VTA) dopaminergic activity, tyrosine hydroxylase (TH) levels and dopamine synthesis. To determine the significance of daytime increases in VTA dopamine activity to manic behaviors, we developed a novel optogenetic stimulation paradigm that produces a sustained increase in dopamine neuronal activity and find that this induces a manic-like behavioral state. Time-dependent dampening of TH activity during the day reverses manic-related behaviors in ClockΔ19 mice. Finally, we show that CLOCK acts as a negative regulator of TH transcription, revealing a novel molecular mechanism underlying cyclic changes in mood-related behavior. Taken together, these studies have identified a mechanistic connection between circadian gene disruption and the precipitation of manic episodes in bipolar disorder.

Authors
Sidor, MM; Spencer, SM; Dzirasa, K; Parekh, PK; Tye, KM; Warden, MR; Arey, RN; Enwright, JF; Jacobsen, JPR; Kumar, S; Remillard, EM; Caron, MG; Deisseroth, K; McClung, CA
MLA Citation
Sidor, MM, Spencer, SM, Dzirasa, K, Parekh, PK, Tye, KM, Warden, MR, Arey, RN, Enwright, JF, Jacobsen, JPR, Kumar, S, Remillard, EM, Caron, MG, Deisseroth, K, and McClung, CA. "Daytime spikes in dopaminergic activity drive rapid mood-cycling in mice." Molecular psychiatry 20.11 (November 2015): 1406-1419.
PMID
25560763
Source
epmc
Published In
Molecular Psychiatry
Volume
20
Issue
11
Publish Date
2015
Start Page
1406
End Page
1419
DOI
10.1038/mp.2014.167

Daytime spikes in dopaminergic activity drive rapid mood-cycling in mice.

Authors
Sidor, MM; Spencer, SM; Dzirasa, K; Parekh, PK; Tye, KM; Warden, MR; Arey, RN; Enwright, JF; Jacobsen, JPR; Kumar, S; Remillard, EM; Caron, MG; Deisseroth, K; McClung, CA
MLA Citation
Sidor, MM, Spencer, SM, Dzirasa, K, Parekh, PK, Tye, KM, Warden, MR, Arey, RN, Enwright, JF, Jacobsen, JPR, Kumar, S, Remillard, EM, Caron, MG, Deisseroth, K, and McClung, CA. "Daytime spikes in dopaminergic activity drive rapid mood-cycling in mice." Molecular psychiatry 20.11 (November 2015): 1479-1480.
PMID
25687774
Source
epmc
Published In
Molecular Psychiatry
Volume
20
Issue
11
Publish Date
2015
Start Page
1479
End Page
1480
DOI
10.1038/mp.2015.8

Elucidation of G-protein and β-arrestin functional selectivity at the dopamine D2 receptor.

The neuromodulator dopamine signals through the dopamine D2 receptor (D2R) to modulate central nervous system functions through diverse signal transduction pathways. D2R is a prominent target for drug treatments in disorders where dopamine function is aberrant, such as schizophrenia. D2R signals through distinct G-protein and β-arrestin pathways, and drugs that are functionally selective for these pathways could have improved therapeutic potential. How D2R signals through the two pathways is still not well defined, and efforts to elucidate these pathways have been hampered by the lack of adequate tools for assessing the contribution of each pathway independently. To address this, Evolutionary Trace was used to produce D2R mutants with strongly biased signal transduction for either the G-protein or β-arrestin interactions. These mutants were used to resolve the role of G proteins and β-arrestins in D2R signaling assays. The results show that D2R interactions with the two downstream effectors are dissociable and that G-protein signaling accounts for D2R canonical MAP kinase signaling cascade activation, whereas β-arrestin only activates elements of this cascade under certain conditions. Nevertheless, when expressed in mice in GABAergic medium spiny neurons of the striatum, the β-arrestin-biased D2R caused a significant potentiation of amphetamine-induced locomotion, whereas the G protein-biased D2R had minimal effects. The mutant receptors generated here provide a molecular tool set that should enable a better definition of the individual roles of G-protein and β-arrestin signaling pathways in D2R pharmacology, neurobiology, and associated pathologies.

Authors
Peterson, SM; Pack, TF; Wilkins, AD; Urs, NM; Urban, DJ; Bass, CE; Lichtarge, O; Caron, MG
MLA Citation
Peterson, SM, Pack, TF, Wilkins, AD, Urs, NM, Urban, DJ, Bass, CE, Lichtarge, O, and Caron, MG. "Elucidation of G-protein and β-arrestin functional selectivity at the dopamine D2 receptor." Proceedings of the National Academy of Sciences of the United States of America 112.22 (June 2015): 7097-7102.
PMID
25964346
Source
epmc
Published In
Proceedings of the National Academy of Sciences of USA
Volume
112
Issue
22
Publish Date
2015
Start Page
7097
End Page
7102
DOI
10.1073/pnas.1502742112

Targeting β-arrestin2 in the treatment of L-DOPA-induced dyskinesia in Parkinson's disease.

Parkinson's disease (PD) is characterized by severe locomotor deficits and is commonly treated with the dopamine (DA) precursor l-3,4-dihydroxyphenylalanine (L-DOPA), but its prolonged use causes dyskinesias referred to as L-DOPA-induced dyskinesias (LIDs). Recent studies in animal models of PD have suggested that dyskinesias are associated with the overactivation of G protein-mediated signaling through DA receptors. β-Arrestins desensitize G protein signaling at DA receptors (D1R and D2R) in addition to activating their own G protein-independent signaling events, which have been shown to mediate locomotion. Therefore, targeting β-arrestins in PD L-DOPA therapy might prove to be a desirable approach. Here we show in a bilateral DA-depletion mouse model of Parkinson's symptoms that genetic deletion of β-arrestin2 significantly limits the beneficial locomotor effects while markedly enhancing the dyskinesia-like effects of acute or chronic L-DOPA treatment. Viral rescue or overexpression of β-arrestin2 in knockout or control mice either reverses or protects against LIDs and its key biochemical markers. In other more conventional animal models of DA neuron loss and PD, such as 6-hydroxydopamine-treated mice or rats and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated nonhuman primates, β-arrestin2 overexpression significantly reduced dyskinesias while maintaining the therapeutic effect of L-DOPA. Considerable efforts are being spent in the pharmaceutical industry to identify therapeutic approaches to block LIDs in patients with PD. Our results point to a potential therapeutic approach, whereby development of either a genetic or pharmacological intervention to enhance β-arrestin2- or limit G protein-dependent D1/D2R signaling could represent a more mechanistically informed strategy.

Authors
Urs, NM; Bido, S; Peterson, SM; Daigle, TL; Bass, CE; Gainetdinov, RR; Bezard, E; Caron, MG
MLA Citation
Urs, NM, Bido, S, Peterson, SM, Daigle, TL, Bass, CE, Gainetdinov, RR, Bezard, E, and Caron, MG. "Targeting β-arrestin2 in the treatment of L-DOPA-induced dyskinesia in Parkinson's disease." Proceedings of the National Academy of Sciences of the United States of America 112.19 (May 2015): E2517-E2526.
PMID
25918399
Source
epmc
Published In
Proceedings of the National Academy of Sciences of USA
Volume
112
Issue
19
Publish Date
2015
Start Page
E2517
End Page
E2526
DOI
10.1073/pnas.1502740112

Serotonin deficiency alters susceptibility to the long-term consequences of adverse early life experience.

Brain 5-HT deficiency has long been implicated in psychiatric disease, but the effects of 5-HT deficiency on stress susceptibility remain largely unknown. Early life stress (ELS) has been suggested to contribute to adult psychopathology, but efforts to study the long-term consequences of ELS have been limited by a lack of appropriate preclinical models. Here, we evaluated the effects of 5-HT deficiency on several long-term cellular, molecular, and behavioral responses of mice to a new model of ELS that combines early-life maternal separation (MS) of pups and postpartum learned helplessness (LH) training in dams. Our data demonstrate that this paradigm (LH/MS) induces depressive-like behavior and impairs pup retrieval in dams. In addition, we show that brain 5-HT deficiency exacerbates anxiety-like behavior induced by LH/MS and blunts the effects of LH/MS on acoustic startle responses in adult offspring. Although the mechanisms underlying these effects remain unclear, following LH/MS, 5-HT-deficient animals had significantly less mRNA expression of the mineralocorticoid receptor in the amygdala than wild-type animals. In addition, 5-HT-deficient mice exhibited reduced mRNA levels of the 5-HT2a receptor and p11 in the hippocampus regardless of stress. LH/MS decreased the number of doublecortin+ immature neurons in the hippocampus in both wild-type (WT) and 5-HT-deficient animals. Our data emphasize the importance of complex interactions between genetic factors and early life experience in mediating long-term changes in emotional behavior. These findings may have important implications for our understanding of the combinatorial roles of 5-HT deficiency, ELS, and postpartum depression in the development of neuropsychiatric disorders.

Authors
Sachs, BD; Rodriguiz, RM; Tran, HL; Iyer, A; Wetsel, WC; Caron, MG
MLA Citation
Sachs, BD, Rodriguiz, RM, Tran, HL, Iyer, A, Wetsel, WC, and Caron, MG. "Serotonin deficiency alters susceptibility to the long-term consequences of adverse early life experience." Psychoneuroendocrinology 53 (March 2015): 69-81.
PMID
25602134
Source
epmc
Published In
Psychoneuroendocrinology
Volume
53
Publish Date
2015
Start Page
69
End Page
81
DOI
10.1016/j.psyneuen.2014.12.019

Lgr4 and Lgr5 drive the formation of long actin-rich cytoneme-like membrane protrusions.

Embryonic development and adult tissue homeostasis require precise information exchange between cells and their microenvironment to coordinate cell behavior. A specialized class of ultra-long actin-rich filopodia, termed cytonemes, provides one mechanism for this spatiotemporal regulation of extracellular cues. We provide here a mechanism whereby the stem-cell marker Lgr5, and its family member Lgr4, promote the formation of cytonemes. Lgr4- and Lgr5-induced cytonemes exceed lengths of 80 µm, are generated through stabilization of nascent filopodia from an underlying lamellipodial-like network and functionally provide a pipeline for the transit of signaling effectors. As proof-of-principle, we demonstrate that Lgr5-induced cytonemes act as conduits for cell signaling by demonstrating that the actin motor and filopodial cargo carrier protein myosin X (Myo10) and the G-protein-coupled receptor (GPCR) signaling effector β-arrestin-2 (Arrb2) transit into cytonemes. This work delineates a biological function for Lgr4 and Lgr5 and provides the rationale to fully investigate Lgr4 and Lgr5 function and cytonemes in mammalian stem cell and cancer stem cell behavior.

Authors
Snyder, JC; Rochelle, LK; Marion, S; Lyerly, HK; Barak, LS; Caron, MG
MLA Citation
Snyder, JC, Rochelle, LK, Marion, S, Lyerly, HK, Barak, LS, and Caron, MG. "Lgr4 and Lgr5 drive the formation of long actin-rich cytoneme-like membrane protrusions." Journal of cell science 128.6 (March 2015): 1230-1240.
PMID
25653388
Source
epmc
Published In
Journal of cell science
Volume
128
Issue
6
Publish Date
2015
Start Page
1230
End Page
1240
DOI
10.1242/jcs.166322

Brain 5-HT deficiency increases stress vulnerability and impairs antidepressant responses following psychosocial stress.

Brain serotonin (5-HT) deficiency and exposure to psychosocial stress have both been implicated in the etiology of depression and anxiety disorders, but whether 5-HT deficiency influences susceptibility to depression- and anxiety-like phenotypes induced by psychosocial stress has not been formally established. Most clinically effective antidepressants increase the extracellular levels of 5-HT, and thus it has been hypothesized that antidepressant responses result from the reversal of endogenous 5-HT deficiency, but this hypothesis remains highly controversial. Here we evaluated the impact of brain 5-HT deficiency on stress susceptibility and antidepressant-like responses using tryptophan hydroxylase 2 knockin (Tph2KI) mice, which display 60-80% reductions in brain 5-HT. Our results demonstrate that 5-HT deficiency leads to increased susceptibility to social defeat stress (SDS), a model of psychosocial stress, and prevents the fluoxetine (FLX)-induced reversal of SDS-induced social avoidance, suggesting that 5-HT deficiency may impair antidepressant responses. In light of recent clinical and preclinical studies highlighting the potential of inhibiting the lateral habenula (LHb) to achieve antidepressant and antidepressant-like responses, we also examined whether LHb inhibition could achieve antidepressant-like responses in FLX-insensitive Tph2KI mice subjected to SDS. Our data reveal that using designer receptors exclusively activated by designer drugs (DREADDs) to inhibit LHb activity leads to reduced SDS-induced social avoidance behavior in both WT and Tph2KI mice. This observation provides additional preclinical evidence that inhibiting the LHb might represent a promising alternative therapeutic approach under conditions in which selective 5-HT reuptake inhibitors are ineffective.

Authors
Sachs, BD; Ni, JR; Caron, MG
MLA Citation
Sachs, BD, Ni, JR, and Caron, MG. "Brain 5-HT deficiency increases stress vulnerability and impairs antidepressant responses following psychosocial stress." Proceedings of the National Academy of Sciences of the United States of America 112.8 (February 9, 2015): 2557-2562.
PMID
25675490
Source
epmc
Published In
Proceedings of the National Academy of Sciences of USA
Volume
112
Issue
8
Publish Date
2015
Start Page
2557
End Page
2562
DOI
10.1073/pnas.1416866112

Increased expression of the dopamine transporter leads to loss of dopamine neurons, oxidative stress and l-DOPA reversible motor deficits.

The dopamine transporter is a key protein responsible for regulating dopamine homeostasis. Its function is to transport dopamine from the extracellular space into the presynaptic neuron. Studies have suggested that accumulation of dopamine in the cytosol can trigger oxidative stress and neurotoxicity. Previously, ectopic expression of the dopamine transporter was shown to cause damage in non-dopaminergic neurons due to their inability to handle cytosolic dopamine. However, it is unknown whether increasing dopamine transporter activity will be detrimental to dopamine neurons that are inherently capable of storing and degrading dopamine. To address this issue, we characterized transgenic mice that over-express the dopamine transporter selectively in dopamine neurons. We report that dopamine transporter over-expressing (DAT-tg) mice display spontaneous loss of midbrain dopamine neurons that is accompanied by increases in oxidative stress markers, 5-S-cysteinyl-dopamine and 5-S-cysteinyl-DOPAC. In addition, metabolite-to-dopamine ratios are increased and VMAT2 protein expression is decreased in the striatum of these animals. Furthermore, DAT-tg mice also show fine motor deficits on challenging beam traversal that are reversed with l-DOPA treatment. Collectively, our findings demonstrate that even in neurons that routinely handle dopamine, increased uptake of this neurotransmitter through the dopamine transporter results in oxidative damage, neuronal loss and l-DOPA reversible motor deficits. In addition, DAT over-expressing animals are highly sensitive to MPTP-induced neurotoxicity. The effects of increased dopamine uptake in these transgenic mice could shed light on the unique vulnerability of dopamine neurons in Parkinson's disease.

Authors
Masoud, ST; Vecchio, LM; Bergeron, Y; Hossain, MM; Nguyen, LT; Bermejo, MK; Kile, B; Sotnikova, TD; Siesser, WB; Gainetdinov, RR; Wightman, RM; Caron, MG; Richardson, JR; Miller, GW; Ramsey, AJ; Cyr, M; Salahpour, A
MLA Citation
Masoud, ST, Vecchio, LM, Bergeron, Y, Hossain, MM, Nguyen, LT, Bermejo, MK, Kile, B, Sotnikova, TD, Siesser, WB, Gainetdinov, RR, Wightman, RM, Caron, MG, Richardson, JR, Miller, GW, Ramsey, AJ, Cyr, M, and Salahpour, A. "Increased expression of the dopamine transporter leads to loss of dopamine neurons, oxidative stress and l-DOPA reversible motor deficits." Neurobiology of disease 74 (February 2015): 66-75.
PMID
25447236
Source
epmc
Published In
Neurobiology of Disease
Volume
74
Publish Date
2015
Start Page
66
End Page
75
DOI
10.1016/j.nbd.2014.10.016

Cariprazine exerts antimanic properties and interferes with dopamine D 2 receptor β -arrestin interactions

Authors
Gao, Y; Peterson, S; Masri, B; Hougland, MT; Adham, N; Gyertyán, I; Kiss, B; Caron, MG; El-Mallakh, RS
MLA Citation
Gao, Y, Peterson, S, Masri, B, Hougland, MT, Adham, N, Gyertyán, I, Kiss, B, Caron, MG, and El-Mallakh, RS. "Cariprazine exerts antimanic properties and interferes with dopamine D 2 receptor β -arrestin interactions." Pharmacology Research & Perspectives 3.1 (February 2015): e00073-e00073.
Source
crossref
Published In
Pharmacology Research and Perspectives
Volume
3
Issue
1
Publish Date
2015
Start Page
e00073
End Page
e00073
DOI
10.1002/prp2.73

Phenotypic regulation of the sphingosine 1-phosphate receptor miles apart by G protein-coupled receptor kinase 2.

The evolutionarily conserved DRY motif at the end of the third helix of rhodopsin-like, class-A G protein-coupled receptors (GPCRs) is a major regulator of receptor stability, signaling activity, and β-arrestin-mediated internalization. Substitution of the DRY arginine with histidine in the human vasopressin receptor results in a loss-of-function phenotype associated with diabetes insipidus. The analogous R150H substitution of the DRY motif in zebrafish sphingosine-1 phosphate receptor 2 (S1p2) produces a mutation, miles apart m(93) (mil(m93)), that not only disrupts signaling but also impairs heart field migration. We hypothesized that constitutive S1p2 desensitization is the underlying cause of this strong zebrafish developmental defect. We observed in cell assays that the wild-type S1p2 receptor is at the cell surface whereas in distinct contrast the S1p2 R150H receptor is found in intracellular vesicles, blocking G protein but not arrestin signaling activity. Surface S1p2 R150H expression could be restored by inhibition of G protein-coupled receptor kinase 2 (GRK2). Moreover, we observed that β-arrestin 2 and GRK2 colocalize with S1p2 in developing zebrafish embryos and depletion of GRK2 in the S1p2 R150H miles apart zebrafish partially rescued cardia bifida. The ability of reduced GRK2 activity to reverse a developmental phenotype associated with constitutive desensitization supports efforts to genetically or pharmacologically target this kinase in diseases involving biased GPCR signaling.

Authors
Burczyk, M; Burkhalter, MD; Blätte, T; Matysik, S; Caron, MG; Barak, LS; Philipp, M
MLA Citation
Burczyk, M, Burkhalter, MD, Blätte, T, Matysik, S, Caron, MG, Barak, LS, and Philipp, M. "Phenotypic regulation of the sphingosine 1-phosphate receptor miles apart by G protein-coupled receptor kinase 2." Biochemistry 54.3 (January 15, 2015): 765-775.
PMID
25555130
Source
epmc
Published In
Biochemistry
Volume
54
Issue
3
Publish Date
2015
Start Page
765
End Page
775
DOI
10.1021/bi501061h

Receptor, Ligand and Transducer Contributions to Dopamine D2 Receptor Functional Selectivity.

Functional selectivity (or biased agonism) is a property exhibited by some G protein-coupled receptor (GPCR) ligands, which results in the modulation of a subset of a receptor's signaling capabilities and more precise control over complex biological processes. The dopamine D2 receptor (D2R) exhibits pleiotropic responses to the biogenic amine dopamine (DA) to mediate complex central nervous system functions through activation of G proteins and β-arrestins. D2R is a prominent therapeutic target for psychological and neurological disorders in which DA biology is dysregulated and targeting D2R with functionally selective drugs could provide a means by which pharmacotherapies could be developed. However, factors that determine GPCR functional selectivity in vivo may be multiple with receptors, ligands and transducers contributing to the process. We have recently described a mutagenesis approach to engineer biased D2R mutants in which G protein-dependent ([Gprot]D2R) and β-arrestin-dependent signaling ([βarr]D2R) were successfully separated (Peterson, et al. PNAS, 2015). Here, permutations of these mutants were used to identify critical determinants of the D2R signaling complex that impart signaling bias in response to the natural or synthetic ligands. Critical residues identified in generating [Gprot]D2R and [βarr]D2R conferred control of partial agonism at G protein and/or β-arrestin activity. Another set of mutations that result in G protein bias was identified that demonstrated that full agonists can impart unique activation patterns, and provided further credence to the concept of ligand texture. Finally, the contributions and interplay between different transducers indicated that G proteins are not aberrantly activated, and that receptor kinase and β-arrestin activities are inextricably linked. These data provide a thorough elucidation of the feasibility and malleability of D2R functional selectivity and point to means by which novel in vivo therapies could be modeled.

Authors
Peterson, SM; Pack, TF; Caron, MG
MLA Citation
Peterson, SM, Pack, TF, and Caron, MG. "Receptor, Ligand and Transducer Contributions to Dopamine D2 Receptor Functional Selectivity." PloS one 10.10 (January 2015): e0141637-.
PMID
26516769
Source
epmc
Published In
PloS one
Volume
10
Issue
10
Publish Date
2015
Start Page
e0141637
DOI
10.1371/journal.pone.0141637

The interaction of escitalopram and R-citalopram at the human serotonin transporter investigated in the mouse.

Escitalopram appears to be a superior antidepressant to racemic citalopram. It has been hypothesized that binding of R-citalopram to the serotonin transporter (SERT) antagonizes escitalopram binding to and inhibition of the SERT, there by curtailing the elevation of extracellular 5-hydroxytryptamine (5-HTExt), and hence anti-depressant efficacy. Further, it has been suggested that a putative allosteric binding site is important for binding of escitalopram to the primary, orthosteric, site, and for R-citalopram's inhibition here of.Primary: Investigate at the human (h)SERT, at clinical relevant doses, whether R-citalopram antagonizes escitalopram-induced 5-HTExt elevation. Secondary: Investigate whether abolishing the putative allosteric site affects escitalopram-induced 5-HTExt elevation and/or modulates the effect of R-citalopram.Recombinant generation of hSERT transgenic mice; in vivo microdialysis; SERT binding; pharmacokinetics; 5-HT sensitive behaviors (tail suspension, marble burying).We generated mice expressing either the wild-type human SERT (hSERT(WT)) or hSERT carrying amino acid substitutions (A505V, L506F, I507L, S574T and I575T) collectively abolishing the putative allosteric site (hSERT(ALI/VFL+SI/TT)). One mg/kg escitalopram yielded clinical relevant plasma levels and brain levels consistent with therapeutic SERT occupancy. The hSERT mice showed normal basal 5-HTExt levels. Escitalopram-induced 5-HTExt elevation was not decreased by R-citalopram co-treatment and was unaffected by loss of the allosteric site. The behavioral effects of the clinically relevant escitalopram dose were small and tended to be enhanced by R-citalopram co-administration.We find no evidence that R-citalopram directly antagonizes escitalopram or that the putative allosteric site is important for hSERT inhibition by escitalopram.

Authors
Jacobsen, JPR; Plenge, P; Sachs, BD; Pehrson, AL; Cajina, M; Du, Y; Roberts, W; Rudder, ML; Dalvi, P; Robinson, TJ; O'Neill, SP; Khoo, KS; Morillo, CS; Zhang, X; Caron, MG
MLA Citation
Jacobsen, JPR, Plenge, P, Sachs, BD, Pehrson, AL, Cajina, M, Du, Y, Roberts, W, Rudder, ML, Dalvi, P, Robinson, TJ, O'Neill, SP, Khoo, KS, Morillo, CS, Zhang, X, and Caron, MG. "The interaction of escitalopram and R-citalopram at the human serotonin transporter investigated in the mouse." Psychopharmacology 231.23 (December 2014): 4527-4540.
PMID
24810106
Source
epmc
Published In
Psychopharmacology
Volume
231
Issue
23
Publish Date
2014
Start Page
4527
End Page
4540
DOI
10.1007/s00213-014-3595-1

G Protein and β-arrestin signaling bias at the ghrelin receptor.

The G protein-coupled ghrelin receptor GHSR1a is a potential pharmacological target for treating obesity and addiction because of the critical role ghrelin plays in energy homeostasis and dopamine-dependent reward. GHSR1a enhances growth hormone release, appetite, and dopamine signaling through G(q/11), G(i/o), and G(12/13) as well as β-arrestin-based scaffolds. However, the contribution of individual G protein and β-arrestin pathways to the diverse physiological responses mediated by ghrelin remains unknown. To characterize whether a signaling bias occurs for GHSR1a, we investigated ghrelin signaling in a number of cell-based assays, including Ca(2+) mobilization, serum response factor response element, stress fiber formation, ERK1/2 phosphorylation, and β-arrestin translocation, utilizing intracellular second loop and C-tail mutants of GHSR1a. We observed that GHSR1a and β-arrestin rapidly form metastable plasma membrane complexes following exposure to an agonist, but replacement of the GHSR1a C-tail by the tail of the vasopressin 2 receptor greatly stabilizes them, producing complexes observable on the plasma membrane and also in endocytic vesicles. Mutations of the contiguous conserved amino acids Pro-148 and Leu-149 in the GHSR1a intracellular second loop generate receptors with a strong bias to G protein and β-arrestin, respectively, supporting a role for conformation-dependent signaling bias in the wild-type receptor. Our results demonstrate more balance in GHSR1a-mediated ERK signaling from G proteins and β-arrestin but uncover an important role for β-arrestin in RhoA activation and stress fiber formation. These findings suggest an avenue for modulating drug abuse-associated changes in synaptic plasticity via GHSR1a and indicate the development of GHSR1a-biased ligands as a promising strategy for selectively targeting downstream signaling events.

Authors
Evron, T; Peterson, SM; Urs, NM; Bai, Y; Rochelle, LK; Caron, MG; Barak, LS
MLA Citation
Evron, T, Peterson, SM, Urs, NM, Bai, Y, Rochelle, LK, Caron, MG, and Barak, LS. "G Protein and β-arrestin signaling bias at the ghrelin receptor." The Journal of biological chemistry 289.48 (November 2014): 33442-33455.
PMID
25261469
Source
epmc
Published In
The Journal of biological chemistry
Volume
289
Issue
48
Publish Date
2014
Start Page
33442
End Page
33455
DOI
10.1074/jbc.m114.581397

Chronic fluoxetine increases extra-hippocampal neurogenesis in adult mice.

Chronic treatment with antidepressants has been shown to enhance neurogenesis in the adult mammalian brain. Although this effect was initially reported to be restricted to the hippocampus, recent work has suggested that fluoxetine, a selective serotonin reuptake inhibitor, also promotes neurogenesis in the cortex. However, whether antidepressants target neural progenitor cells in other brain regions has not been examined.Here, we used BrdU labeling and immunohistochemistry with a transgenic mouse line in which nestin+ neural progenitor cells can be inducibly labeled with the fluorescent protein, Tomato, following tamoxifen administration. We investigated the effects of chronic fluoxetine on cell proliferation and nestin+ progenitor cells in periventricular areas in the medial hypothalamus and medial habenula, two brain areas involved in stress and anxiety responses.Our data provide the first in vivo evidence that fluoxetine promotes cell proliferation and neurogenesis and increases the mRNA levels of BDNF in the hypothalamus and habenula.By identifying novel cellular targets of fluoxetine, our results may provide new insight into the mechanisms underlying antidepressant responses.

Authors
Sachs, BD; Caron, MG
MLA Citation
Sachs, BD, and Caron, MG. "Chronic fluoxetine increases extra-hippocampal neurogenesis in adult mice." The international journal of neuropsychopharmacology 18.4 (October 31, 2014).
PMID
25583694
Source
epmc
Published In
The international journal of neuropsychopharmacology / official scientific journal of the Collegium Internationale Neuropsychopharmacologicum (CINP)
Volume
18
Issue
4
Publish Date
2014
DOI
10.1093/ijnp/pyu029

Selective deletion of GRK2 alters psychostimulant-induced behaviors and dopamine neurotransmission.

GRK2 is a G protein-coupled receptor kinase (GRK) that is broadly expressed and is known to regulate diverse types of receptors. GRK2 null animals exhibit embryonic lethality due to a severe developmental heart defect, which has precluded the study of this kinase in the adult brain. To elucidate the specific role of GRK2 in the brain dopamine (DA) system, we used a conditional gene knockout approach to selectively delete GRK2 in DA D1 receptor (D1R)-, DA D2 receptor (D2R)-, adenosine 2A receptor (A2AR)-, or DA transporter (DAT)-expressing neurons. Here we show that select GRK2-deficient mice display hyperactivity, hyposensitivity, or hypersensitivity to the psychomotor effects of cocaine, altered striatal signaling, and DA release and uptake. Mice with GRK2 deficiency in D2R-expressing neurons also exhibited increased D2 autoreceptor activity. These findings reveal a cell-type-specific role for GRK2 in the regulation of normal motor behavior, sensitivity to psychostimulants, dopamine neurotransmission, and D2 autoreceptor function.

Authors
Daigle, TL; Ferris, MJ; Gainetdinov, RR; Sotnikova, TD; Urs, NM; Jones, SR; Caron, MG
MLA Citation
Daigle, TL, Ferris, MJ, Gainetdinov, RR, Sotnikova, TD, Urs, NM, Jones, SR, and Caron, MG. "Selective deletion of GRK2 alters psychostimulant-induced behaviors and dopamine neurotransmission." Neuropsychopharmacology : official publication of the American College of Neuropsychopharmacology 39.10 (September 2014): 2450-2462.
PMID
24776686
Source
epmc
Published In
Neuropsychopharmacology (Nature)
Volume
39
Issue
10
Publish Date
2014
Start Page
2450
End Page
2462
DOI
10.1038/npp.2014.97

Overlapping and opposing functions of G protein-coupled receptor kinase 2 (GRK2) and GRK5 during heart development.

G protein-coupled receptor kinases 2 (GRK2) and 5 (GRK5) are fundamental regulators of cardiac performance in adults but are less well characterized for their function in the hearts of embryos. GRK2 and -5 belong to different subfamilies and function as competitors in the control of certain receptors and signaling pathways. In this study, we used zebrafish to investigate whether the fish homologs of GRK2 and -5, Grk2/3 and Grk5, also have unique, complementary, or competitive roles during heart development. We found that they differentially regulate the heart rate of early embryos and equally facilitate heart function in older embryos and that both are required to develop proper cardiac morphology. A loss of Grk2/3 results in dilated atria and hypoplastic ventricles, and the hearts of embryos depleted in Grk5 present with a generalized atrophy. This Grk5 morphant phenotype was associated with an overall decrease of early cardiac progenitors as well as a reduction in the area occupied by myocardial progenitor cells. In the case of Grk2/3, the progenitor decrease was confined to a subset of precursor cells with a committed ventricular fate. We attempted to rescue the GRK loss-of-function heart phenotypes by downstream activation of Hedgehog signaling. The Grk2/3 loss-of-function embryos were rescued by this approach, but Grk5 embryos failed to respond. In summary, we found that GRK2 and GRK5 control cardiac function as well as morphogenesis during development although with different morphological outcomes.

Authors
Philipp, M; Berger, IM; Just, S; Caron, MG
MLA Citation
Philipp, M, Berger, IM, Just, S, and Caron, MG. "Overlapping and opposing functions of G protein-coupled receptor kinase 2 (GRK2) and GRK5 during heart development." The Journal of biological chemistry 289.38 (September 2014): 26119-26130.
PMID
25104355
Source
epmc
Published In
The Journal of biological chemistry
Volume
289
Issue
38
Publish Date
2014
Start Page
26119
End Page
26130
DOI
10.1074/jbc.m114.551952

Structural basis for Smoothened receptor modulation and chemoresistance to anticancer drugs.

The Smoothened receptor (SMO) mediates signal transduction in the hedgehog pathway, which is implicated in normal development and carcinogenesis. SMO antagonists can suppress the growth of some tumours; however, mutations at SMO have been found to abolish their antitumour effects, a phenomenon known as chemoresistance. Here we report three crystal structures of human SMO bound to the antagonists SANT1 and Anta XV, and the agonist, SAG1.5, at 2.6-2.8 Å resolution. The long and narrow cavity in the transmembrane domain of SMO harbours multiple ligand binding sites, where SANT1 binds at a deeper site as compared with other ligands. Distinct interactions at D473(6.54f) elucidated the structural basis for the differential effects of chemoresistance mutations on SMO antagonists. The agonist SAG1.5 induces a conformational rearrangement of the binding pocket residues, which could contribute to SMO activation. Collectively, these studies reveal the structural basis for the modulation of SMO by small molecules.

Authors
Wang, C; Wu, H; Evron, T; Vardy, E; Han, GW; Huang, X-P; Hufeisen, SJ; Mangano, TJ; Urban, DJ; Katritch, V; Cherezov, V; Caron, MG; Roth, BL; Stevens, RC
MLA Citation
Wang, C, Wu, H, Evron, T, Vardy, E, Han, GW, Huang, X-P, Hufeisen, SJ, Mangano, TJ, Urban, DJ, Katritch, V, Cherezov, V, Caron, MG, Roth, BL, and Stevens, RC. "Structural basis for Smoothened receptor modulation and chemoresistance to anticancer drugs." Nature communications 5 (July 10, 2014): 4355-.
PMID
25008467
Source
epmc
Published In
Nature Communications
Volume
5
Publish Date
2014
Start Page
4355
DOI
10.1038/ncomms5355

Integrated approaches to understanding antipsychotic drug action at GPCRs.

The G-protein coupled receptor (GPCR) family of genes represents one of the largest druggable families of genes in the human genome. This is evident by the fact that approximately 30% of currently marketed drugs target GPCRs. However, many of these drugs are limited in their clinical potential as they are associated with debilitating side effects-a consequence of our incomplete understanding of their pharmacology and the signaling pathways regulated by GPCRs. Because of the limited range of tools available to resolve these issues, integrated approaches are required to fully understand the pharmacological action of drugs and the biochemical repertoire regulated by GPCRs. In this review we will focus on the action of antipsychotic drugs on certain monoamine GPCRs in the central nervous system (CNS) and the approaches being developed to elucidate their distinct pharmacology.

Authors
Urs, NM; Nicholls, PJ; Caron, MG
MLA Citation
Urs, NM, Nicholls, PJ, and Caron, MG. "Integrated approaches to understanding antipsychotic drug action at GPCRs." Current opinion in cell biology 27 (April 2014): 56-62. (Review)
PMID
24680431
Source
epmc
Published In
Current Opinion in Cell Biology
Volume
27
Publish Date
2014
Start Page
56
End Page
62
DOI
10.1016/j.ceb.2013.11.002

Dopamine D2 receptor relies upon PPM/PP2C protein phosphatases to dephosphorylate huntingtin protein.

Striatal dopamine D2 receptor (D2R) relies upon G protein- and β-arrestin-dependent signaling pathways to convey its action on motor control and behavior. Considering that D2R activation inhibits Akt in the striatum and that huntingtin physiological functions are affected by Akt phosphorylation, we sought to investigate whether D2R-mediated signaling could regulate huntingtin phosphorylation. We demonstrate that D2R activation decreases huntingtin phosphorylation on its Akt site. This dephosphorylation event depends upon the Gαi-dependent engagement of specific members of the protein phosphatase metallo-dependent (PPM/PP2C) family and is independent of β-arrestin 2. These observations identify the PPM/PP2C family as a mediator of G protein-coupled receptor signaling and thereby suggest a novel mechanism of dopaminergic signaling.

Authors
Marion, S; Urs, NM; Peterson, SM; Sotnikova, TD; Beaulieu, J-M; Gainetdinov, RR; Caron, MG
MLA Citation
Marion, S, Urs, NM, Peterson, SM, Sotnikova, TD, Beaulieu, J-M, Gainetdinov, RR, and Caron, MG. "Dopamine D2 receptor relies upon PPM/PP2C protein phosphatases to dephosphorylate huntingtin protein." The Journal of Biological Chemistry 289.17 (April 2014): 11715-11724.
PMID
24619418
Source
epmc
Published In
The Journal of biological chemistry
Volume
289
Issue
17
Publish Date
2014
Start Page
11715
End Page
11724
DOI
10.1074/jbc.m113.544312

Sex differences in response to chronic mild stress and congenital serotonin deficiency.

Women exhibit a nearly twofold increased risk of developing depression and anxiety disorders when compared to men, a fact that has been hypothesized to result in part from increased stress susceptibility. Here, we used the tryptophan hydroxylase-2 R439H knock-in mouse (Tph2KI) and the chronic unpredictable mild stress (CMS) model to examine sex differences in response to congenital 5-HT deficiency and chronic stress. Our results demonstrate that female mice, but not 5-HT-deficient animals, exhibit significantly increased susceptibility to CMS-induced despair-like behavior in the forced swim test. In addition, female 5-HT-deficient mice exhibit anhedonia-like behavior in the sucrose preference test, whereas male 5-HT-deficient animals do not, suggesting that females exhibit increased sensitivity to at least some of the effects of congenital 5-HT deficiency. Although CMS did not reduce cell proliferation in the hippocampus, low levels of brain 5-HT were associated with increased hippocampal cell proliferation, an effect that was predominantly observed in females. Overall, these results highlight the importance of interactions between psychiatric disease risk factors such as sex, chronic stress and congenital 5-HT deficiency in the development of aberrant emotional behavior.

Authors
Sachs, BD; Ni, JR; Caron, MG
MLA Citation
Sachs, BD, Ni, JR, and Caron, MG. "Sex differences in response to chronic mild stress and congenital serotonin deficiency." Psychoneuroendocrinology 40 (February 2014): 123-129.
PMID
24485484
Source
epmc
Published In
Psychoneuroendocrinology
Volume
40
Publish Date
2014
Start Page
123
End Page
129
DOI
10.1016/j.psyneuen.2013.11.008

Congenital brain serotonin deficiency leads to reduced ethanol sensitivity and increased ethanol consumption in mice.

Serotonergic dysfunction has been hypothesized to play an important role in the pathophysiology of alcoholism. However, whether congenital serotonin (5-HT) deficiency leads to increased alcohol consumption or affects ethanol-related behaviors has not been established. Here, we use a transgenic mouse line that expresses a hypofunctional variant of the 5-HT synthesis enzyme, tryptophan hydroxylase 2, to examine the impact of 5-HT deficiency on responses to alcohol. We demonstrate that these 5-HT-deficient transgenic animals (Tph2KI mice) recover their righting reflex more rapidly than wild-type controls following a high dose of ethanol and exhibit blunted locomotor retardation in response to repeated ethanol administration. In addition, compared to WT controls, 5-HT-deficient animals consume significantly more ethanol and exhibit increased preference for ethanol in two-bottle choice tests. Our data also suggest that 5-HT plays a critical role in mediating the effects of ethanol on Akt/GSK3β signaling in the nucleus accumbens. Overall, our results corroborate previous theories regarding the importance of brain 5-HT levels in mediating responsiveness to alcohol and demonstrate, for the first time, that congenital 5-HT deficiency leads to increased ethanol consumption and decreased sensitivity to the sedative-like effects of ethanol, perhaps in part through modulating Akt/GSK3β signaling.

Authors
Sachs, BD; Salahi, AA; Caron, MG
MLA Citation
Sachs, BD, Salahi, AA, and Caron, MG. "Congenital brain serotonin deficiency leads to reduced ethanol sensitivity and increased ethanol consumption in mice." Neuropharmacology 77 (February 2014): 177-184.
PMID
24067926
Source
pubmed
Published In
Neuropharmacology
Volume
77
Publish Date
2014
Start Page
177
End Page
184
DOI
10.1016/j.neuropharm.2013.09.010

Imidazole-derived agonists for the neurotensin 1 receptor

A scaffold-hop program seeking full agonists of the neurotensin-1 (NTR1) receptor identified the probe molecule ML301 (1) and associated analogs, including its naphthyl analog (14) which exhibited similar properties. Compound 1 showed full agonist behavior (79-93%) with an EC 50 of 2.0-4.1 μM against NTR1. Compound 1 also showed good activity in a Ca mobilization FLIPR assay (93% efficacy at 298 nM), consistent with it functioning via the G q coupled pathway, and good selectivity relative to NTR2 and GPR35. In further profiling, 1 showed low potential for promiscuity and good overall pharmacological data. This report describes the discovery, synthesis, and SAR of 1 and associated analogs. Initial in vitro pharmacologic characterization is also presented. © 2013 Elsevier Ltd. All rights reserved.

Authors
Hershberger, PM; Hedrick, MP; Peddibhotla, S; Mangravita-Novo, A; Gosalia, P; Li, Y; Gray, W; Vicchiarelli, M; Smith, LH; Chung, TDY; Thomas, JB; Caron, MG; Pinkerton, AB; Barak, LS; Roth, GP
MLA Citation
Hershberger, PM, Hedrick, MP, Peddibhotla, S, Mangravita-Novo, A, Gosalia, P, Li, Y, Gray, W, Vicchiarelli, M, Smith, LH, Chung, TDY, Thomas, JB, Caron, MG, Pinkerton, AB, Barak, LS, and Roth, GP. "Imidazole-derived agonists for the neurotensin 1 receptor." Bioorganic and Medicinal Chemistry Letters 24.1 (January 1, 2014): 262-267.
Source
scopus
Published In
Bioorganic & Medicinal Chemistry Letters
Volume
24
Issue
1
Publish Date
2014
Start Page
262
End Page
267
DOI
10.1016/j.bmcl.2013.11.026

Selective deletion of GRK2 alters psychostimulant-induced behaviors and dopamine neurotransmission

GRK2 is a G protein-coupled receptor kinase (GRK) that is broadly expressed and is known to regulate diverse types of receptors. GRK2 null animals exhibit embryonic lethality due to a severe developmental heart defect, which has precluded the study of this kinase in the adult brain. To elucidate the specific role of GRK2 in the brain dopamine (DA) system, we used a conditional gene knockout approach to selectively delete GRK2 in DA D1 receptor (D1R)-, DA D2 receptor (D2R)-, adenosine 2A receptor (A2AR)-, or DA transporter (DAT)-expressing neurons. Here we show that select GRK2-deficient mice display hyperactivity, hyposensitivity, or hypersensitivity to the psychomotor effects of cocaine, altered striatal signaling, and DA release and uptake. Mice with GRK2 deficiency in D2R-expressing neurons also exhibited increased D2 autoreceptor activity. These findings reveal a cell-type-specific role for GRK2 in the regulation of normal motor behavior, sensitivity to psychostimulants, dopamine neurotransmission, and D2 autoreceptor function. © 2014 American College of Neuropsychopharmacology.

Authors
Daigle, TL; Ferris, MJ; Gainetdinov, RR; Sotnikova, TD; Urs, NM; Jones, SR; Caron, MG
MLA Citation
Daigle, TL, Ferris, MJ, Gainetdinov, RR, Sotnikova, TD, Urs, NM, Jones, SR, and Caron, MG. "Selective deletion of GRK2 alters psychostimulant-induced behaviors and dopamine neurotransmission." Neuropsychopharmacology 39.10 (January 1, 2014): 2450-2462.
Source
scopus
Published In
Neuropsychopharmacology (Nature)
Volume
39
Issue
10
Publish Date
2014
Start Page
2450
End Page
2462
DOI
10.1038/npp.2014.97

Correction: The Stem Cell-Expressed Receptor Lgr5 Possesses Canonical and Functionally Active Molecular Determinants Critical to β-arrestin-2 Recruitment.

[This corrects the article on p. e84476 in vol. 8.].

Authors
Snyder, JC; Rochelle, LK; Barak, LS; Caron, MG
MLA Citation
Snyder, JC, Rochelle, LK, Barak, LS, and Caron, MG. "Correction: The Stem Cell-Expressed Receptor Lgr5 Possesses Canonical and Functionally Active Molecular Determinants Critical to β-arrestin-2 Recruitment." PloS one 9.1 (2014).
PMID
24427241
Source
epmc
Published In
PloS one
Volume
9
Issue
1
Publish Date
2014
DOI
10.1371/annotation/7f83735d-53d4-42f3-b3b2-b05dd5db059c

The interaction of escitalopram and R-citalopram at the human serotonin transporter investigated in the mouse

© 2014 Springer-Verlag Berlin Heidelberg.Rationale: Escitalopram appears to be a superior antidepressant to racemic citalopram. It has been hypothesized that binding of R-citalopram to the serotonin transporter (SERT) antagonizes escitalopram binding to and inhibition of the SERT, thereby curtailing the elevation of extracellular 5-hydroxytryptamine (5-HT<inf>Ext</inf>), and hence antidepressant efficacy. Further, it has been suggested that a putative allosteric binding site is important for binding of escitalopram to the primary, orthosteric, site, and for R-citalopram's inhibition hereof.Objectives: Primary: Investigate at the human (h)SERT, at clinical relevant doses, whether R-citalopram antagonizes escitalopram-induced 5-HT<inf>Ext</inf> elevation. Secondary: Investigate whether abolishing the putative allosteric site affects escitalopram-induced 5-HT<inf>Ext</inf> elevation and/or modulates the effect of R-citalopram.Methods: Recombinant generation of hSERT transgenic mice; in vivo microdialysis; SERT binding; pharmacokinetics; 5-HT sensitive behaviors (tail suspension, marble burying).Results: We generated mice expressing either the wild-type human SERT (hSERT<sup>WT</sup>) or hSERT carrying amino acid substitutions (A505V, L506F, I507L, S574T and I575T) collectively abolishing the putative allosteric site (hSERT<sup>ALI/VFL+SI/TT</sup>). One mg/kg escitalopram yielded clinical relevant plasma levels and brain levels consistent with therapeutic SERT occupancy. The hSERT mice showed normal basal 5-HT<inf>Ext</inf> levels. Escitalopram-induced 5-HT<inf>Ext</inf> elevation was not decreased by R-citalopram co-treatment and was unaffected by loss of the allosteric site<inf>.</inf> The behavioral effects of the clinically relevant escitalopram dose were small and tended to be enhanced by R-citalopram co-administration.Conclusions: We find no evidence that R-citalopram directly antagonizes escitalopram or that the putative allosteric site is important for hSERT inhibition by escitalopram.

Authors
Jacobsen, JPR; Plenge, P; Sachs, BD; Pehrson, AL; Cajina, M; Du, Y; Roberts, W; Rudder, ML; Dalvi, P; Robinson, TJ; O'Neill, SP; Khoo, KS; Morillo, CS; Zhang, X; Caron, MG
MLA Citation
Jacobsen, JPR, Plenge, P, Sachs, BD, Pehrson, AL, Cajina, M, Du, Y, Roberts, W, Rudder, ML, Dalvi, P, Robinson, TJ, O'Neill, SP, Khoo, KS, Morillo, CS, Zhang, X, and Caron, MG. "The interaction of escitalopram and R-citalopram at the human serotonin transporter investigated in the mouse." Psychopharmacology 231.23 (2014): 4527-4540.
Source
scival
Published In
Psychopharmacology
Volume
231
Issue
23
Publish Date
2014
Start Page
4527
End Page
4540
DOI
10.1007/s00213-014-3595-1

Identification of the GPR55 Antagonist Binding Site Using a Novel Set of High-Potency GPR55 Selective Ligands

Authors
Kotsikorou, E; Sharir, H; Shore, DM; Hurst, DP; Lynch, DL; Madrigal, KE; Heynen-Genel, S; Milan, LB; Chung, TDY; Seltzman, HH; Bai, Y; Caron, MG; Barak, LS; Croatt, MP; Abood, ME; Reggio, PH
MLA Citation
Kotsikorou, E, Sharir, H, Shore, DM, Hurst, DP, Lynch, DL, Madrigal, KE, Heynen-Genel, S, Milan, LB, Chung, TDY, Seltzman, HH, Bai, Y, Caron, MG, Barak, LS, Croatt, MP, Abood, ME, and Reggio, PH. "Identification of the GPR55 Antagonist Binding Site Using a Novel Set of High-Potency GPR55 Selective Ligands." Biochemistry 52.52 (December 31, 2013): 9456-9469.
Source
crossref
Published In
Biochemistry
Volume
52
Issue
52
Publish Date
2013
Start Page
9456
End Page
9469
DOI
10.1021/bi4008885

β-arrestin Regulation of Ghrelin Signaling in modulating Addictive Behavior

Authors
Evron, T; Urs, NM; Sutton, L; Bai, Y; Caron, MG; Barak, LS
MLA Citation
Evron, T, Urs, NM, Sutton, L, Bai, Y, Caron, MG, and Barak, LS. "β-arrestin Regulation of Ghrelin Signaling in modulating Addictive Behavior." (December 1, 2013): 184-. (Chapter)
Source
scopus
Publish Date
2013
Start Page
184
DOI
10.1016/B978-0-12-800044-1.00167-7

Integrated Approaches to Understand the Actions of GPCRs: The b-Arrestin-Dependent D2R Mediated Signaling Through Akt/GSK3

Authors
Caron, MG; Urs, NM; Snyder, JC; Peterson, SM
MLA Citation
Caron, MG, Urs, NM, Snyder, JC, and Peterson, SM. "Integrated Approaches to Understand the Actions of GPCRs: The b-Arrestin-Dependent D2R Mediated Signaling Through Akt/GSK3." (December 1, 2013): 99-100. (Chapter)
Source
scopus
Publish Date
2013
Start Page
99
End Page
100
DOI
10.1016/B978-0-12-800044-1.00086-6

Targeting Beta-Arrestin Dependent Signaling in the Treatment of Parkinson's Disease

Authors
Urs, NM; Daigle, TL; Ting, J; Caron, MG
MLA Citation
Urs, NM, Daigle, TL, Ting, J, and Caron, MG. "Targeting Beta-Arrestin Dependent Signaling in the Treatment of Parkinson's Disease." (December 1, 2013): 103-104. (Chapter)
Source
scopus
Publish Date
2013
Start Page
103
End Page
104
DOI
10.1016/B978-0-12-800044-1.00090-8

D2 Functionally Selective Ligands: Novel Therapeutics?

Authors
Sassano, MF; Allen, JA; Jin, J; Setola, V; Wetsel, WC; Caron, MG; Roth, BL
MLA Citation
Sassano, MF, Allen, JA, Jin, J, Setola, V, Wetsel, WC, Caron, MG, and Roth, BL. "D2 Functionally Selective Ligands: Novel Therapeutics?." (December 1, 2013): 105-. (Chapter)
Source
scopus
Publish Date
2013
Start Page
105
DOI
10.1016/B978-0-12-800044-1.00091-X

Methylphenidate amplifies the potency and reinforcing effects of amphetamines by increasing dopamine transporter expression

Methylphenidate (MPH) is commonly diverted for recreational use, but the neurobiological consequences of exposure to MPH at high, abused doses are not well defined. Here we show that MPH self-administration in rats increases dopamine transporter (DAT) levels and enhances the potency of MPH and amphetamine on dopamine responses and drug-seeking behaviours, without altering cocaine effects. Genetic overexpression of the DAT in mice mimics these effects, confirming that MPH self-administration-induced increases in DAT levels are sufficient to induce the changes. Further, this work outlines a basic mechanism by which increases in DAT levels, regardless of how they occur, are capable of increasing the rewarding and reinforcing effects of select psychostimulant drugs, and suggests that individuals with elevated DAT levels, such as ADHD sufferers, may be more susceptible to the addictive effects of amphetamine-like drugs.© 2013 Macmillan Publishers Limited. All rights reserved.

Authors
Calipari, ES; Ferris, MJ; Salahpour, A; Caron, MG; Jones, SR
MLA Citation
Calipari, ES, Ferris, MJ, Salahpour, A, Caron, MG, and Jones, SR. "Methylphenidate amplifies the potency and reinforcing effects of amphetamines by increasing dopamine transporter expression." Nature Communications 4 (November 5, 2013).
PMID
24193139
Source
scopus
Published In
Nature Communications
Volume
4
Publish Date
2013
DOI
10.1038/ncomms3720

Novel roles for β-arrestins in the regulation of pharmacological sequestration to predict agonist-induced desensitization of dopamine D 3 receptors

Background and Purpose In addition to typical GPCR kinase (GRK)-/β-arrestin-dependent internalization, dopamine D 3 receptor employed an additional GRK-independent sequestration pathway. In this study, we investigated the molecular mechanism of this novel sequestration pathway. Experimental Approach Radioligand binding, flow cytometry and cell surface biotinylation assay were used to characterize trafficking properties of D 2 and D 3 receptors. Serine/threonine and N-linked glycosylation mutants of the D 3 receptor were utilized to locate receptor regions involved in pharmacological sequestration and desensitization. Various point mutants of the D 2 and D 3 receptors, whose sequestration and desensitization properties were altered, were combined with knockdown cells of GRKs or β-arrestins to functionally correlate pharmacological sequestration and desensitization. Key Results The D 3 receptor, but not the D 2 receptor, showed characteristic trafficking behaviour in which receptors were shifted towards the more hydrophobic domains within the plasma membrane without translocation into other intracellular compartments. Among various amino acid residues tested, S145/S146, C147 and N12/19 were involved in pharmacological sequestration and receptor desensitization. Both pharmacological sequestration and desensitization of D 3 receptor required β-arrestins, and functional relationship was observed between two processes when it was tested for D 3 receptor variants and agonists. Conclusions and Implications Pharmacological sequestration of D 3 receptor accompanies movement of cell surface receptors into a more hydrophobic fraction within the plasma membrane and renders D 3 receptor inaccessible to hydrophilic ligands. Pharmacological sequestration is correlated with desensitization of the D 3 receptor in a Gβγ- and β-arrestin-dependent manner. This study provides new insights into molecular mechanism governing GPCR trafficking and desensitization. © 2013 The British Pharmacological Society.

Authors
Min, C; Zheng, M; Zhang, X; Caron, MG; Kim, KM
MLA Citation
Min, C, Zheng, M, Zhang, X, Caron, MG, and Kim, KM. "Novel roles for β-arrestins in the regulation of pharmacological sequestration to predict agonist-induced desensitization of dopamine D 3 receptors." British Journal of Pharmacology 170.5 (November 1, 2013): 1112-1129.
PMID
23992580
Source
scopus
Published In
British Journal of Pharmacology
Volume
170
Issue
5
Publish Date
2013
Start Page
1112
End Page
1129
DOI
10.1111/bph.12357

N-aryl piperazine metabotropic glutamate receptor 5 positive allosteric modulators possess efficacy in preclinical models of NMDA hypofunction and cognitive enhancement.

Impaired transmission through glutamatergic circuits has been postulated to play a role in the underlying pathophysiology of schizophrenia. Furthermore, inhibition of the N-methyl-d-aspartate (NMDA) subtype of ionotropic glutamate receptors (NMDAR) induces a syndrome that recapitulates many of the symptoms observed in patients with schizophrenia. Selective activation of metabotropic glutamate receptor subtype 5 (mGlu5) may provide a novel therapeutic approach for treatment of symptoms associated with schizophrenia through facilitation of transmission through central glutamatergic circuits. Here, we describe the characterization of two novel N-aryl piperazine mGlu5 positive allosteric modulators (PAMs): 2-(4-(2-(benzyloxy)acetyl)piperazin-1-yl)benzonitrile (VU0364289) and 1-(4-(2,4-difluorophenyl)piperazin-1-yl)-2-((4-fluorobenzyl)oxy)ethanone (DPFE). VU0364289 and DPFE induced robust leftward shifts in the glutamate concentration-response curves for Ca(2+) mobilization and extracellular signal-regulated kinases 1 and 2 phosphorylation. Both PAMs displayed micromolar affinity for the common mGlu5 allosteric binding site and high selectivity for mGlu5. VU0364289 and DPFE possessed suitable pharmacokinetic properties for dosing in vivo and produced robust dose-related effects in reversing amphetamine-induced hyperlocomotion, a preclinical model predictive of antipsychotic-like activity. In addition, DPFE enhanced acquisition of contextual fear conditioning in rats and reversed behavioral deficits in a mouse model of NMDAR hypofunction. In contrast, DPFE had no effect on reversing apomorphine-induced disruptions of prepulse inhibition of the acoustic startle reflex. These mGlu5 PAMs also increased monoamine levels in the prefrontal cortex, enhanced performance in a hippocampal-mediated memory task, and elicited changes in electroencephalogram dynamics commensurate with procognitive effects. Collectively, these data support and extend the role for the development of novel mGlu5 PAMs for the treatment of psychosis and cognitive deficits observed in individuals with schizophrenia.

Authors
Gregory, KJ; Herman, EJ; Ramsey, AJ; Hammond, AS; Byun, NE; Stauffer, SR; Manka, JT; Jadhav, S; Bridges, TM; Weaver, CD; Niswender, CM; Steckler, T; Drinkenburg, WH; Ahnaou, A; Lavreysen, H; Macdonald, GJ; Bartolomé, JM; Mackie, C; Hrupka, BJ; Caron, MG; Daigle, TL; Lindsley, CW; Conn, PJ; Jones, CK
MLA Citation
Gregory, KJ, Herman, EJ, Ramsey, AJ, Hammond, AS, Byun, NE, Stauffer, SR, Manka, JT, Jadhav, S, Bridges, TM, Weaver, CD, Niswender, CM, Steckler, T, Drinkenburg, WH, Ahnaou, A, Lavreysen, H, Macdonald, GJ, Bartolomé, JM, Mackie, C, Hrupka, BJ, Caron, MG, Daigle, TL, Lindsley, CW, Conn, PJ, and Jones, CK. "N-aryl piperazine metabotropic glutamate receptor 5 positive allosteric modulators possess efficacy in preclinical models of NMDA hypofunction and cognitive enhancement." J Pharmacol Exp Ther 347.2 (November 2013): 438-457.
PMID
23965381
Source
pubmed
Published In
The Journal of pharmacology and experimental therapeutics
Volume
347
Issue
2
Publish Date
2013
Start Page
438
End Page
457
DOI
10.1124/jpet.113.206623

Cerebral 5-HT2A receptor binding, but not mGluR2, is increased in tryptophan hydroxylase 2 decrease-of-function mice

Transgenic mice with a knock-in (KI) of a tryptophan hydroxylase 2 (Tph2) R439H mutation, analogous to the Tph2 R441H single-nucleotide polymorphism originally identified in a late life depression cohort, have markedly reduced levels of 5-hydroxytryptamine (5-HT). These Tph2KI mice are therefore interesting as a putative translational model of low endogenous 5-HT function that allows for assessment of adaptive changes in different anatomical regions. Here, we determined 5-HT 2A receptor binding in several brain regions using in vitro receptor autoradiography and two different radioligands. When using the 5-HT 2A receptor selective antagonist radioligand 3 H-MDL100907, we found higher binding in the prefrontal cortex (10%, P=0.009), the striatum (26%, P=0.005), and the substantia nigra (21%, P=0.027). The increase was confirmed in the same regions with the 5-HT 2A/C receptor agonist, 3 H-CIMBI-36 (2-(4-Bromo-2,5-dimethoxyphenyl)-N-(2-methoxybenzyl)ethanamine). 5-HT 2A receptors establish heteromeric receptor complexes with metabotropic glutamate 2 receptors (mGluR2), but binding levels of the mGluR2/3 ligand 3 H-LY341495 were unaltered in brain areas with increased 5-HT 2A receptor levels. These data show that in distinct anatomical regions, 5-HT 2A receptor binding sites are up-regulated in 5-HT deficient mice, and this increase is not associated with changes in mGluR2 binding. © 2013 Elsevier Ireland Ltd.

Authors
Jørgensen, CV; Jacobsen, JP; Caron, MG; Klein, AB; Knudsen, GM; Mikkelsen, JD
MLA Citation
Jørgensen, CV, Jacobsen, JP, Caron, MG, Klein, AB, Knudsen, GM, and Mikkelsen, JD. "Cerebral 5-HT2A receptor binding, but not mGluR2, is increased in tryptophan hydroxylase 2 decrease-of-function mice." Neuroscience Letters 555 (October 25, 2013): 118-122.
PMID
24055299
Source
scopus
Published In
Neuroscience Letters
Volume
555
Publish Date
2013
Start Page
118
End Page
122
DOI
10.1016/j.neulet.2013.08.073

The effects of brain serotonin deficiency on behavioural disinhibition and anxiety-like behaviour following mild early life stress.

Aberrant serotonin (5-HT) signalling and exposure to early life stress have both been suggested to play a role in anxiety- and impulsivity-related behaviours. However, whether congenital 5-HT deficiency × early life stress interactions influence the development of anxiety- or impulsivity-like behaviour has not been established. Here, we examined the effects of early life maternal separation (MS) stress on anxiety-like behaviour and behavioural disinhibition, a type of impulsivity-like behaviour, in wild-type (WT) and tryptophan hydroxylase 2 (Tph2) knock-in (Tph2KI) mice, which exhibit ~60-80% reductions in the levels of brain 5-HT due to a R439H mutation in Tph2. We also investigated the effects of 5-HT deficiency and early life stress on adult hippocampal neurogenesis, plasma corticosterone levels and several signal transduction pathways in the amygdala. We demonstrate that MS slightly increases anxiety-like behaviour in WT mice and induces behavioural disinhibition in Tph2KI animals. We also demonstrate that MS leads to a slight decrease in cell proliferation within the hippocampus and potentiates corticosterone responses to acute stress, but these effects are not affected by brain 5-HT deficiency. However, we show that 5-HT deficiency leads to significant alterations in SGK-1 and GSK3β signalling and NMDA receptor expression in the amygdala in response to MS. Together, these findings support a potential role for 5-HT-dependent signalling in the amygdala in regulating the long-term effects of early life stress on anxiety-like behaviour and behavioural disinhibition.

Authors
Sachs, BD; Rodriguiz, RM; Siesser, WB; Kenan, A; Royer, EL; Jacobsen, JPR; Wetsel, WC; Caron, MG
MLA Citation
Sachs, BD, Rodriguiz, RM, Siesser, WB, Kenan, A, Royer, EL, Jacobsen, JPR, Wetsel, WC, and Caron, MG. "The effects of brain serotonin deficiency on behavioural disinhibition and anxiety-like behaviour following mild early life stress." Int J Neuropsychopharmacol 16.9 (October 2013): 2081-2094.
PMID
23672796
Source
pubmed
Published In
The international journal of neuropsychopharmacology / official scientific journal of the Collegium Internationale Neuropsychopharmacologicum (CINP)
Volume
16
Issue
9
Publish Date
2013
Start Page
2081
End Page
2094
DOI
10.1017/S1461145713000321

Discovery of ML314, a brain penetrant nonpeptidic β-arrestin biased agonist of the neurotensin NTR1 receptor

The neurotensin 1 receptor (NTR1) is an important therapeutic target for a range of disease states including addiction. A high-throughput screening campaign, followed by medicinal chemistry optimization, led to the discovery of a nonpeptidic β-arrestin biased agonist for NTR1. The lead compound, 2-cyclopropyl-6,7-dimethoxy-4-(4-(2-methoxyphenyl)-piperazin-1-yl)quinazoline, 32 (ML314), exhibits full agonist behavior against NTR1 (EC 50 = 2.0 μM) in the primary assay and selectivity against NTR2. The effect of 32 is blocked by the NTR1 antagonist SR142948A in a dose-dependent manner. Unlike peptide-based NTR1 agonists, compound 32 has no significant response in a Ca 2+ mobilization assay and is thus a biased agonist that activates the β-arrestin pathway rather than the traditional G q coupled pathway. This bias has distinct biochemical and functional consequences that may lead to physiological advantages. Compound 32 displays good brain penetration in rodents, and studies examining its in vivo properties are underway. © 2013 American Chemical Society.

Authors
Peddibhotla, S; Hedrick, MP; Hershberger, P; Maloney, PR; Li, Y; Milewski, M; Gosalia, P; Gray, W; Mehta, A; Sugarman, E; Hood, B; Suyama, E; Nguyen, K; Heynen-Genel, S; Vasile, S; Salaniwal, S; Stonich, D; Su, Y; Mangravita-Novo, A; Vicchiarelli, M; Roth, GP; Smith, LH; Chung, TDY; Hanson, GR; Thomas, JB; Caron, MG; Barak, LS; Pinkerton, AB
MLA Citation
Peddibhotla, S, Hedrick, MP, Hershberger, P, Maloney, PR, Li, Y, Milewski, M, Gosalia, P, Gray, W, Mehta, A, Sugarman, E, Hood, B, Suyama, E, Nguyen, K, Heynen-Genel, S, Vasile, S, Salaniwal, S, Stonich, D, Su, Y, Mangravita-Novo, A, Vicchiarelli, M, Roth, GP, Smith, LH, Chung, TDY, Hanson, GR, Thomas, JB, Caron, MG, Barak, LS, and Pinkerton, AB. "Discovery of ML314, a brain penetrant nonpeptidic β-arrestin biased agonist of the neurotensin NTR1 receptor." ACS Medicinal Chemistry Letters 4.9 (September 12, 2013): 846-851.
PMID
24611085
Source
scopus
Published In
ACS Medicinal Chemistry Letters
Volume
4
Issue
9
Publish Date
2013
Start Page
846
End Page
851
DOI
10.1021/ml400176n

Grk5l controls heart development by limiting mTOR signaling during symmetry breaking.

The correct asymmetric placement of inner organs is termed situs solitus and is determined early during development. Failure in symmetry breaking results in conditions ranging from randomized organ arrangement to a complete mirror image, often accompanied by severe congenital heart defects (CHDs). We found that the zebrafish homolog of mammalian G protein-coupled receptor kinase 5 (GRK5) employs noncanonical, receptor-independent functions to secure symmetry breaking. Knockdown of GRK5's closest homolog in zebrafish embryos, Grk5l, is sufficient to randomize cardiac looping and left-right asymmetry. Mechanistically, we found that loss of GRK5 increases mammalian target of rapamycin complex 1 (mTORC1) activity. This causes elongation of motile cilia in the organ of laterality, a consequence that is known to be sufficient to trigger aberrant organ arrangement. By fine-tuning mTORC1, GRK5 thus serves an unanticipated function during early development, besides its well-characterized role in the adult heart. These findings could implicate GRK5 as a susceptibility allele for certain cases of CHD.

Authors
Burkhalter, MD; Fralish, GB; Premont, RT; Caron, MG; Philipp, M
MLA Citation
Burkhalter, MD, Fralish, GB, Premont, RT, Caron, MG, and Philipp, M. "Grk5l controls heart development by limiting mTOR signaling during symmetry breaking." Cell Rep 4.4 (August 29, 2013): 625-632.
PMID
23972986
Source
pubmed
Published In
Cell Reports
Volume
4
Issue
4
Publish Date
2013
Start Page
625
End Page
632
DOI
10.1016/j.celrep.2013.07.036

Triphenylmethane dye activation of beta-arrestin.

β-Arrestins regulate G protein-coupled receptor signaling as competitive inhibitors and protein adaptors. Low molecular weight biased ligands that bind receptors and discriminate between the G protein dependent arm and β-arrestin, clathrin-associated arm of receptor signaling are considered therapeutically valuable as a result of this distinctive pharmacological behavior. Other than receptor agonists, compounds that activate β-arrestins are not available. We show that within minutes of exposure to the cationic triphenylmethane dyes malachite green and brilliant green, tissue culture cells recruit β-arrestins to clathrin scaffolds in a receptor-activation independent manner. In the presence of these compounds, G protein signaling is inhibited, ERK and GSK3β signaling are preserved, and the recruitment of the beta2-adaptin, AP2 adaptor complex to clathrin as well as transferrin internalization is reduced. Moreover, malachite green binds β-arrestin2-GFP coated immunotrap beads relative to GFP only coated beads. Triphenylmethane dyes are FDA approved for topical use on newborns as components of triple-dye preparations and are not approved but used effectively as aqueous antibiotics in fish husbandry. As possible carcinogens, their chronic ingestion in food preparations, particularly through farmed fish, is discouraged in the U.S. and Europe. Our results indicate triphenylmethane dyes as a result of novel pharmacology may have additional roles as β-arrestin/clathrin pathway signaling modulators in both pharmacology research and clinical therapy.

Authors
Barak, LS; Bai, Y; Snyder, JC; Wang, J; Chen, W; Caron, MG
MLA Citation
Barak, LS, Bai, Y, Snyder, JC, Wang, J, Chen, W, and Caron, MG. "Triphenylmethane dye activation of beta-arrestin." Biochemistry 52.32 (August 13, 2013): 5403-5414.
PMID
23865508
Source
pubmed
Published In
Biochemistry
Volume
52
Issue
32
Publish Date
2013
Start Page
5403
End Page
5414
DOI
10.1021/bi400217r

The effects of congenital brain serotonin deficiency on responses to chronic fluoxetine.

The importance of reversing brain serotonin (5-HT) deficiency and promoting hippocampal neurogenesis in the mechanisms of action for antidepressants remain highly controversial. Here we examined the behavioral, neurochemical and neurogenic effects of chronic fluoxetine (FLX) in a mouse model of congenital 5-HT deficiency, the tryptophan hydroxylase 2 (R439H) knock-in (Tph2KI) mouse. Our results demonstrate that congenital 5-HT deficiency prevents a subset of the signature molecular, cellular and behavioral effects of FLX, despite the fact that FLX restores the 5-HT levels of Tph2KI mice to essentially the levels observed in wild-type mice at baseline. These results suggest that inducing supra-physiological levels of 5-HT, not merely reversing 5-HT deficiency, is required for many of the antidepressant-like effects of FLX. We also demonstrate that co-administration of the 5-HT precursor, 5-hydroxytryptophan (5-HTP), along with FLX rescues the novelty suppressed feeding (NSF) anxiolytic-like effect of FLX in Tph2KI mice, despite still failing to induce neurogenesis. Thus, our results indicate that brain 5-HT deficiency reduces the efficacy of FLX and that supplementation with 5-HTP can restore some antidepressant-like responses in the context of 5-HT deficiency. Our findings also suggest that feeding latency reductions in the NSF induced by chronic 5-HT elevation are not mediated by drug-induced increments in neurogenesis in 5-HT-deficient animals. Overall, these findings shed new light on the impact of 5-HT deficiency on responses to FLX and may have important implications for treatment selection in depression and anxiety disorders.

Authors
Sachs, BD; Jacobsen, JPR; Thomas, TL; Siesser, WB; Roberts, WL; Caron, MG
MLA Citation
Sachs, BD, Jacobsen, JPR, Thomas, TL, Siesser, WB, Roberts, WL, and Caron, MG. "The effects of congenital brain serotonin deficiency on responses to chronic fluoxetine. (Published online)" Transl Psychiatry 3 (August 13, 2013): e291-.
PMID
23942622
Source
pubmed
Published In
Translational Psychiatry
Volume
3
Publish Date
2013
Start Page
e291
DOI
10.1038/tp.2013.65

Novel adamantyl cannabinoids as CB1 receptor probes.

In previous studies, compound 1 (AM411), a 3-(1-adamantyl) analogue of the phytocannabinoid (-)-Δ(8)-tetrahydrocannabinol (Δ(8)-THC), was shown to have improved affinity and selectivity for the CB1 receptor. In this work, we further explored the role of the 1-adamantyl group at the C-3 position in a series of tricyclic cannabinoid analogues modified at the 9-northern aliphatic hydroxyl (NAH) position. Of these, 9-hydroxymethyl hexahydrocannabinol 11 (AM4054) exhibited high CB1 affinity and full agonist profile. In the cAMP assay, the 9-hydroxymethyl cannabinol analogue 24 (AM4089) had a partial agonist profile, with high affinity and moderate selectivity for rCB1 over hCB2. In vivo results in rat models of hypothermia and analgesia were congruent with in vitro data. Our in vivo data indicate that 3-(1-adamantyl) substitution, within NAH cannabinergics, imparts improved pharmacological profiles when compared to the corresponding, traditionally used 3-dimethylheptyl analogues and identifies 11 and 24 as potentially useful in vivo CB1 cannabinergic probes.

Authors
Thakur, GA; Bajaj, S; Paronis, C; Peng, Y; Bowman, AL; Barak, LS; Caron, MG; Parrish, D; Deschamps, JR; Makriyannis, A
MLA Citation
Thakur, GA, Bajaj, S, Paronis, C, Peng, Y, Bowman, AL, Barak, LS, Caron, MG, Parrish, D, Deschamps, JR, and Makriyannis, A. "Novel adamantyl cannabinoids as CB1 receptor probes." J Med Chem 56.10 (May 23, 2013): 3904-3921.
PMID
23621789
Source
pubmed
Published In
Journal of Medicinal Chemistry
Volume
56
Issue
10
Publish Date
2013
Start Page
3904
End Page
3921
DOI
10.1021/jm4000775

Constitutive internalization of the leucine-rich G protein-coupled receptor-5 (LGR5) to the trans-Golgi network.

LGR5 is a Wnt pathway associated G protein-coupled receptor (GPCR) that serves as a molecular determinant of stem cells in numerous tissues including the intestine, stomach, hair follicle, eye, and mammary gland. Despite its importance as a marker for this critical niche, little is known about LGR5 signaling nor the biochemical mechanisms and receptor determinants that regulate LGR5 membrane expression and intracellular trafficking. Most importantly, in cells LGR5 is predominantly intracellular, yet the mechanisms underlying this behavior have not been determined. In this work we elucidate a precise trafficking program for LGR5 and identify the motif at its C terminus that is responsible for the observed constitutive internalization. We show that this process is dependent upon dynamin GTPase activity and find that wild-type full-length LGR5 rapidly internalizes into EEA1- and Rab5-positive endosomes. However, LGR5 fails to rapidly recycle to the plasmid membrane through Rab4-positive vesicles, as is common for other GPCRs. Rather, internalized LGR5 transits through Rab7- and Rab9-positive vesicles, co-localizes in vesicles with Vps26, a retromer complex component that regulates retrograde trafficking to the trans-Golgi network (TGN) and reaches a steady-state distribution in the TGN within 2 h. Using mutagenesis, particularly of putative phosphorylation sites, we show that the amino acid pair, serine 861 and 864, is the principal C-tail determinant that mediates LGR5 constitutive internalization. The constitutive internalization of LGR5 to the TGN suggests the existence of novel biochemical roles for its Wnt pathway related, but ill defined signaling program.

Authors
Snyder, JC; Rochelle, LK; Lyerly, HK; Caron, MG; Barak, LS
MLA Citation
Snyder, JC, Rochelle, LK, Lyerly, HK, Caron, MG, and Barak, LS. "Constitutive internalization of the leucine-rich G protein-coupled receptor-5 (LGR5) to the trans-Golgi network." J Biol Chem 288.15 (April 12, 2013): 10286-10297.
PMID
23439653
Source
pubmed
Published In
The Journal of biological chemistry
Volume
288
Issue
15
Publish Date
2013
Start Page
10286
End Page
10297
DOI
10.1074/jbc.M112.447540

A Gαs DREADD mouse for selective modulation of cAMP production in striatopallidal neurons.

Here, we describe a newly generated transgenic mouse in which the Gs DREADD (rM3Ds), an engineered G protein-coupled receptor, is selectively expressed in striatopallidal medium spiny neurons (MSNs). We first show that in vitro, rM3Ds can couple to Gαolf and induce cAMP accumulation in cultured neurons and HEK-T cells. The rM3Ds was then selectively and stably expressed in striatopallidal neurons by creating a transgenic mouse in which an adenosine2A (adora2a) receptor-containing bacterial artificial chromosome was employed to drive rM3Ds expression. In the adora2A-rM3Ds mouse, activation of rM3Ds by clozapine-N-oxide (CNO) induces DARPP-32 phosphorylation, consistent with the known consequence of activation of endogenous striatal Gαs-coupled GPCRs. We then tested whether CNO administration would produce behavioral responses associated with striatopallidal Gs signaling and in this regard CNO dose-dependently decreases spontaneous locomotor activity and inhibits novelty induced locomotor activity. Last, we show that CNO prevented behavioral sensitization to amphetamine and increased AMPAR/NMDAR ratios in transgene-expressing neurons of the nucleus accumbens shell. These studies demonstrate the utility of adora2a-rM3Ds transgenic mice for the selective and noninvasive modulation of Gαs signaling in specific neuronal populations in vivo.This unique tool provides a new resource for elucidating the roles of striatopallidal MSN Gαs signaling in other neurobehavioral contexts.

Authors
Farrell, MS; Pei, Y; Wan, Y; Yadav, PN; Daigle, TL; Urban, DJ; Lee, H-M; Sciaky, N; Simmons, A; Nonneman, RJ; Huang, X-P; Hufeisen, SJ; Guettier, J-M; Moy, SS; Wess, J; Caron, MG; Calakos, N; Roth, BL
MLA Citation
Farrell, MS, Pei, Y, Wan, Y, Yadav, PN, Daigle, TL, Urban, DJ, Lee, H-M, Sciaky, N, Simmons, A, Nonneman, RJ, Huang, X-P, Hufeisen, SJ, Guettier, J-M, Moy, SS, Wess, J, Caron, MG, Calakos, N, and Roth, BL. "A Gαs DREADD mouse for selective modulation of cAMP production in striatopallidal neurons." Neuropsychopharmacology 38.5 (April 2013): 854-862.
PMID
23303063
Source
pubmed
Published In
Neuropsychopharmacology
Volume
38
Issue
5
Publish Date
2013
Start Page
854
End Page
862
DOI
10.1038/npp.2012.251

Cortical-amygdalar circuit dysfunction in a genetic mouse model of serotonin deficiency.

Although the majority of first-line antidepressants increase brain serotonin and rare polymorphisms in tryptophan hydroxlase-2 (Tph2), the rate-limiting enzyme in the brain serotonin synthesis pathway, have been identified in cohorts of subjects with major depressive disorder, the circuit level alterations that results from serotonergic hypofunction remain poorly understood. Here we use chronic multicircuit neurophysiological recordings to characterize functional interactions across cortical and limbic circuits in mice engineered to express a human loss-of-function depression allele Tph2-(R441H) [Tph2 knockin (Tph2KI)]. Our results show that Tph2KI mice exhibit increased intra-network synchrony within medial prefrontal cortex (mPFC) and basal amygdala (AMY) and increased inter-network synchrony between these two brain networks. Moreover, we demonstrate that chronic treatment with fluoxetine reverses several of the circuit alterations observed within Tph2KI mice. Together, our findings establish a functional link between functional hyposerotonergia and altered mPFC-AMY network dynamics.

Authors
Dzirasa, K; Kumar, S; Sachs, BD; Caron, MG; Nicolelis, MAL
MLA Citation
Dzirasa, K, Kumar, S, Sachs, BD, Caron, MG, and Nicolelis, MAL. "Cortical-amygdalar circuit dysfunction in a genetic mouse model of serotonin deficiency." J Neurosci 33.10 (March 6, 2013): 4505-4513.
PMID
23467366
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
33
Issue
10
Publish Date
2013
Start Page
4505
End Page
4513
DOI
10.1523/JNEUROSCI.4891-12.2013

Chronic SSRI treatment exacerbates serotonin deficiency in humanized Tph2 mutant mice.

Selective serotonin reuptake inhibitors (SSRIs) are a major class of antidepressants that act by blocking inward transport of serotonin (5-HT) into presynaptic neurons mediated by the serotonin transporter (SERT). Both reuptake and ongoing synthesis are essential in supporting intraneuronal serotonin concentrations in serotonergic neurons. A rare mutation in tryptophan hydroxylase 2 (Tph2), the rate limiting enzyme for 5-HT synthesis, was identified in several patients with major depression, and knock-in mice expressing the analogous mutation (R439H Tph2 KI) show 80% reduction in 5-HT synthesis and tissue levels. Chronic treatment with SSRIs (fluoxetine and paroxetine) resulted in a dramatic further depletion of 5-HT tissue levels in R439H Tph2 KI mice (down to 1-3% of wild type levels) while having little effects in wild-type controls. Treatment with the 5-HT precursor 5-hydroxytryptophan (5-HTP) restored 5-HT tissue content in mutant mice, and cotreatment with 5-HTP and fluoxetine essentially prevented the depleting effect of a chronic SSRI. These data demonstrate that chronic SSRI treatment could further exacerbate the 5-HT deficiency in Tph2 mutation carriers, and this can be prevented by 5-HTP supplementation.

Authors
Siesser, WB; Sachs, BD; Ramsey, AJ; Sotnikova, TD; Beaulieu, J-M; Zhang, X; Caron, MG; Gainetdinov, RR
MLA Citation
Siesser, WB, Sachs, BD, Ramsey, AJ, Sotnikova, TD, Beaulieu, J-M, Zhang, X, Caron, MG, and Gainetdinov, RR. "Chronic SSRI treatment exacerbates serotonin deficiency in humanized Tph2 mutant mice." ACS Chem Neurosci 4.1 (January 16, 2013): 84-88.
PMID
23336047
Source
pubmed
Published In
ACS Chemical Neuroscience
Volume
4
Issue
1
Publish Date
2013
Start Page
84
End Page
88
DOI
10.1021/cn300127h

Pharmacological blockade of a β(2)AR-β-arrestin-1 signaling cascade prevents the accumulation of DNA damage in a behavioral stress model.

Chronic stress is known to have a profound negative impact on human health and has been suggested to influence a number of disease states. However, the mechanisms underlying the deleterious effects of stress remain largely unknown. Stress is known to promote the release of epinephrine, a catecholamine stress hormone that binds to β(2)-adrenergic receptors (β(2)ARs) with high affinity. Our previous work has demonstrated that chronic stimulation of a β(2)AR-β-arrestin-1-mediated signaling pathway by infusion of isoproterenol suppresses p53 levels and impairs genomic integrity. In this pathway, β-arrestin-1, which is activated via β(2)ARs, facilitates the AKT-mediated activation of Mdm2 and functions as a molecular scaffold to promote the binding and degradation of p53 by the E3-ubiquitin ligase, Mdm2. Here, we show that chronic restraint stress in mice recapitulates the effects of isoproterenol infusion to reduce p53 levels and results in the accumulation of DNA damage in the frontal cortex of the brain, two effects that are abrogated by the β-blocker, propranolol and by genetic deletion of β-arrestin-1. These data suggest that the β(2)AR-β-arrestin-1 signaling pathway may represent an attractive therapeutic target to prevent some of the negative consequences of stress in the treatment of stress-related disorders.

Authors
Hara, MR; Sachs, BD; Caron, MG; Lefkowitz, RJ
MLA Citation
Hara, MR, Sachs, BD, Caron, MG, and Lefkowitz, RJ. "Pharmacological blockade of a β(2)AR-β-arrestin-1 signaling cascade prevents the accumulation of DNA damage in a behavioral stress model." Cell Cycle 12.2 (January 15, 2013): 219-224.
PMID
23287463
Source
pubmed
Published In
Cell Cycle
Volume
12
Issue
2
Publish Date
2013
Start Page
219
End Page
224
DOI
10.4161/cc.23368

The stem cell-expressed receptor Lgr5 possesses canonical and functionally active molecular determinants critical to β-arrestin-2 recruitment.

Lgr5 is a membrane protein related to G protein-coupled receptors (GPCR)s whose expression identifies stem cells in multiple tissues and is strongly correlated with cancer. Despite the recent identification of endogenous ligands for Lgr5, its mode of signaling remains enigmatic. The ability to couple to G proteins and βarrestins are classical molecular behaviors of GPCRs that have yet to be observed for Lgr5. Therefore, the goal of this study was to determine if Lgr5 can engage a classical GPCR behavior and elucidate the molecular determinants of this process. Structural analysis of Lgr5 revealed several motifs consistent with its ability to recruit βarr2. Among them, a "SSS" serine cluster located at amino acid position 873-875 within the C-terminal tail (C-tail), is in a region consistent with other GPCRs that bind βarr2 with high-affinity. To test its functionality, a ligand-independent βarr2 translocation assay was implemented. We show that Lgr5 recruits βarr2 and that the "SSS" amino acids (873-875) are absolutely critical to this process. We also demonstrate that for full efficacy, this cluster requires other Lgr5 C-tail serines that were previously shown to be important for constitutive and βarr2 independent internalization of Lgr5. These data are proof of principle that a classical GPCR behavior can be manifested by Lgr5. The existence of alternative ligands or missing effectors of Lgr5 that scaffold this classical GPCR behavior and the downstream signaling pathways engaged should be considered. Characterizing Lgr5 signaling will be invaluable for assessing its role in tissue maintenance, repair, and disease.

Authors
Snyder, JC; Rochelle, LK; Barak, LS; Caron, MG
MLA Citation
Snyder, JC, Rochelle, LK, Barak, LS, and Caron, MG. "The stem cell-expressed receptor Lgr5 possesses canonical and functionally active molecular determinants critical to β-arrestin-2 recruitment. (Published online)" PLoS One 8.12 (2013): e84476-.
PMID
24386388
Source
pubmed
Published In
PloS one
Volume
8
Issue
12
Publish Date
2013
Start Page
e84476
DOI
10.1371/journal.pone.0084476

The Role of Arrestins in Development

Authors
Philipp, M; Evron, T; Caron, MG
MLA Citation
Philipp, M, Evron, T, and Caron, MG. "The Role of Arrestins in Development." MOLECULAR BIOLOGY OF ARRESTINS 118 (2013): 225-242.
PMID
23764056
Source
wos-lite
Published In
Progress in Molecular Biology and Translational Science
Volume
118
Publish Date
2013
Start Page
225
End Page
242
DOI
10.1016/B978-0-12-394440-5.00009-7

Trace amine-associated receptor 1 partial agonism reveals novel paradigm for neuropsychiatric therapeutics.

BACKGROUND: Trace amines, compounds structurally related to classical biogenic amines, represent endogenous ligands of the trace amine-associated receptor 1 (TAAR1). Because trace amines also influence the activity of other targets, selective ligands are needed for the elucidation of TAAR1 function. Here we report on the identification and characterization of the first selective and potent TAAR1 partial agonist. METHODS: The TAAR1 partial agonist RO5203648 was evaluated for its binding affinity and functional activity at rodent and primate TAAR1 receptors stably expressed in HEK293 cells, for its physicochemical and pharmacokinetic properties, for its effects on the firing frequency of monoaminergic neurons ex vivo, and for its properties in vivo with genetic and pharmacological models of central nervous system disorders. RESULTS: RO5203648 showed high affinity and potency at TAAR1, high selectivity versus other targets, and favorable pharmacokinetic properties. In mouse brain slices, RO5203648 increased the firing frequency of dopaminergic and serotonergic neurons in the ventral tegmental area and the dorsal raphe nucleus, respectively. In various behavioral paradigms in rodents and monkeys, RO5203648 demonstrated clear antipsychotic- and antidepressant-like activities as well as potential anxiolytic-like properties. Furthermore, it attenuated drug-taking behavior and was highly effective in promoting attention, cognitive performance, and wakefulness. CONCLUSIONS: With the first potent and selective TAAR1 partial agonist, RO5203648, we show that TAAR1 is implicated in a broad range of relevant physiological, behavioral, and cognitive neuropsychiatric dimensions. Collectively, these data uncover important neuromodulatory roles for TAAR1 and suggest that agonists at this receptor might have therapeutic potential in one or more neuropsychiatric domains.

Authors
Revel, FG; Moreau, J-L; Gainetdinov, RR; Ferragud, A; Velázquez-Sánchez, C; Sotnikova, TD; Morairty, SR; Harmeier, A; Groebke Zbinden, K; Norcross, RD; Bradaia, A; Kilduff, TS; Biemans, B; Pouzet, B; Caron, MG; Canales, JJ; Wallace, TL; Wettstein, JG; Hoener, MC
MLA Citation
Revel, FG, Moreau, J-L, Gainetdinov, RR, Ferragud, A, Velázquez-Sánchez, C, Sotnikova, TD, Morairty, SR, Harmeier, A, Groebke Zbinden, K, Norcross, RD, Bradaia, A, Kilduff, TS, Biemans, B, Pouzet, B, Caron, MG, Canales, JJ, Wallace, TL, Wettstein, JG, and Hoener, MC. "Trace amine-associated receptor 1 partial agonism reveals novel paradigm for neuropsychiatric therapeutics." Biol Psychiatry 72.11 (December 1, 2012): 934-942.
PMID
22705041
Source
pubmed
Published In
Biological Psychiatry
Volume
72
Issue
11
Publish Date
2012
Start Page
934
End Page
942
DOI
10.1016/j.biopsych.2012.05.014

Dopamine Receptors

This chapter focuses on the role of dopamine receptors in neurotransmission, their structures, and their functions. Dopamine (DA) is one of the major monoamine neurotransmitters in the mammalian brain. It is generated by hydroxylation and decarboxylation of the amino acid tyrosine, and further metabolized into epinephrine and norepinephrine. Dopamine receptors belong to the superfamily of GPCRs and are all considered to be in the Rhodopsin-like Class A family of 7-transmembrane receptors based on sequence homology and function. For all the five DARs the N-terminus and extracellular loops are glycosylated and cysteine residues between the loops form disulfide bonds. Higher order GPCR signaling structures have long been hypothesized to exist, however there remains some controversy over the biological significance of such structures. The significance of arrestin dependent signaling for GPCRs may be manifold. First, arrestin dependent signaling is usually more persistent than G protein signaling. Second, it has been shown that for certain GPCRs a given compound can act as an agonist at one signaling pathway and an antagonist at the other or vice versa. © 2012 Elsevier Inc. All rights reserved.

Authors
Peterson, SM; Urs, N; Caron, MG
MLA Citation
Peterson, SM, Urs, N, and Caron, MG. "Dopamine Receptors." (December 1, 2012): 67-70. (Chapter)
Source
scopus
Publish Date
2012
Start Page
67
End Page
70
DOI
10.1016/B978-0-12-386525-0.00013-5

Deletion of GSK3β in D2R-expressing neurons reveals distinct roles for β-arrestin signaling in antipsychotic and lithium action.

Several studies in rodent models have shown that glycogen synthase kinase 3 β (GSK3β) plays an important role in the actions of antispychotics and mood stabilizers. Recently it was demonstrated that GSK3β through a β-arrestin2/protein kinase B (PKB or Akt)/protein phosphatase 2A (PP2A) signaling complex regulates dopamine (DA)- and lithium-sensitive behaviors and is required to mediate endophenotypes of mania and depression in rodents. We have previously shown that atypical antipsychotics antagonize DA D2 receptor (D2R)/β-arrestin2 interactions more efficaciously than G-protein-dependent signaling, whereas typical antipsychotics inhibit both pathways with similar efficacy. To elucidate the site of action of GSK3β in regulating DA- or lithium-sensitive behaviors, we generated conditional knockouts of GSK3β, where GSK3β was deleted in either DA D1- or D2-receptor-expressing neurons. We analyzed these mice for behaviors commonly used to test antipsychotic efficacy or behaviors that are sensitive to lithium treatment. Mice with deletion of GSK3β in D2 (D2GSK3β(-/-)) but not D1 (D1GSK3β(-/-)) neurons mimic antipsychotic action. However, haloperidol (HAL)-induced catalepsy was unchanged in either D2GSK3β(-/-) or D1GSK3β(-/-) mice compared with control mice. Interestingly, genetic stabilization of β-catenin, a downstream target of GSK3β, in D2 neurons did not affect any of the behaviors tested. Moreover, D2GSK3β(-/-) or D1GSK3β(-/-) mice showed similar responses to controls in the tail suspension test (TST) and dark-light emergence test, behaviors which were previously shown to be β-arrestin2- and GSK3β-dependent and sensitive to lithium treatment. Taken together these studies suggest that selective deletion of GSK3β but not stabilization of β-catenin in D2 neurons mimics antipsychotic action without affecting signaling pathways involved in catalepsy or certain mood-related behaviors.

Authors
Urs, NM; Snyder, JC; Jacobsen, JPR; Peterson, SM; Caron, MG
MLA Citation
Urs, NM, Snyder, JC, Jacobsen, JPR, Peterson, SM, and Caron, MG. "Deletion of GSK3β in D2R-expressing neurons reveals distinct roles for β-arrestin signaling in antipsychotic and lithium action." Proceedings of the National Academy of Sciences of the United States of America 109.50 (December 2012): 20732-20737.
PMID
23188793
Source
epmc
Published In
Proceedings of the National Academy of Sciences of USA
Volume
109
Issue
50
Publish Date
2012
Start Page
20732
End Page
20737
DOI
10.1073/pnas.1215489109

The 5-HT deficiency theory of depression: perspectives from a naturalistic 5-HT deficiency model, the tryptophan hydroxylase 2Arg439His knockin mouse.

A decreased level of brain 5-hydroxytryptamine (5-HT) has been theorized to be a core pathogenic factor in depression for half a century. The theory arose from clinical observations that drugs enhancing extracellular levels of 5-HT (5-HT(Ext)) have antidepressant effects in many patients. However, whether such drugs indeed correct a primary deficit remains unresolved. Still, a number of anomalies in putative biomarkers of central 5-HT function have been repeatedly reported in depression patients over the past 40 years, collectively indicating that 5-HT deficiency could be present in depression, particularly in severely ill and/or suicidal patients. This body of literature on putative 5-HT biomarker anomalies and depression has recently been corroborated by data demonstrating that such anomalies indeed occur consequent to severely reduced 5-HT(Ext) levels in a mouse model of naturalistic 5-HT deficiency, the tryptophan hydroxylase 2 His(439) knockin (Tph2KI) mouse. In this review, we will critically assess the evidence for 5-HT deficiency in depression and the possible role of polymorphisms in the Tph2 gene as a causal factor in 5-HT deficiency, the latter investigated from a clinical as well as preclinical angle.

Authors
Jacobsen, JPR; Medvedev, IO; Caron, MG
MLA Citation
Jacobsen, JPR, Medvedev, IO, and Caron, MG. "The 5-HT deficiency theory of depression: perspectives from a naturalistic 5-HT deficiency model, the tryptophan hydroxylase 2Arg439His knockin mouse." Philos Trans R Soc Lond B Biol Sci 367.1601 (September 5, 2012): 2444-2459. (Review)
PMID
22826344
Source
pubmed
Published In
Philosophical Transactions B
Volume
367
Issue
1601
Publish Date
2012
Start Page
2444
End Page
2459
DOI
10.1098/rstb.2012.0109

Elimination of GRK2 from cholinergic neurons reduces behavioral sensitivity to muscarinic receptor activation.

Although G-protein-coupled receptor kinase 2 (GRK2) is the most widely studied member of a family of kinases that has been shown to exert powerful influences on a variety of G-protein-coupled receptors, its role in the brain remains largely unknown. Here we report the localization of GRK2 in the mouse brain and generate novel conditional knock-out (KO) mice to assess the physiological importance of this kinase in cholinergic neurons. Mice with the selective deletion of GRK2 in this cell population (ChAT(IRES-cre)Grk2(f/f) KO mice) exhibit reduced behavioral responsiveness to challenge with oxotremorine-M (Oxo-M), a nonselective muscarinic acetylcholine receptor agonist. Specifically, Oxo-M-induced hypothermia, hypolocomotion, and salivation were markedly reduced in these animals, while analgesic responses were unaltered. In contrast, we found that GRK2 deficiency in cholinergic neurons does not alter cocaine-induced psychomotor activation, behavioral sensitization, or conditioned place preference. These results demonstrate that the elimination of GRK2 in cholinergic neurons reduces sensitivity to select muscarinic-mediated behaviors, while dopaminergic effects remain intact and further suggests that GRK2 may selectively impair muscarinic acetylcholine receptor-mediated function in vivo.

Authors
Daigle, TL; Caron, MG
MLA Citation
Daigle, TL, and Caron, MG. "Elimination of GRK2 from cholinergic neurons reduces behavioral sensitivity to muscarinic receptor activation." J Neurosci 32.33 (August 15, 2012): 11461-11466.
PMID
22895728
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
32
Issue
33
Publish Date
2012
Start Page
11461
End Page
11466
DOI
10.1523/JNEUROSCI.2234-12.2012

Deficient serotonin neurotransmission and depression-like serotonin biomarker alterations in tryptophan hydroxylase 2 (Tph2) loss-of-function mice.

Probably the foremost hypothesis of depression is the 5-hydroxytryptamine (5-HT, serotonin) deficiency hypothesis. Accordingly, anomalies in putative 5-HT biomarkers have repeatedly been reported in depression patients. However, whether such anomalies in fact reflect deficient central 5-HT neurotransmission remains unresolved. We employed a naturalistic model of 5-HT deficiency, the tryptophan hydroxylase 2 (Tph2) R439H knockin mouse, to address this question. We report that Tph2 knockin mice have reduced basal and stimulated levels of extracellular 5-HT (5-HT(Ext)). Interestingly, cerebrospinal fluid (CSF) 5-hydroxyindoleacetic acid (5-HIAA) and fenfluramine-induced plasma prolactin levels are markedly diminished in the Tph2 knockin mice. These data seemingly confirm that low CSF 5-HIAA and fenfluramine-induced plasma prolactin reflects chronic, endogenous central nervous system (CNS) 5-HT deficiency. Moreover, 5-HT(1A) receptor agonist-induced hypothermia is blunted and frontal cortex 5-HT(2A) receptors are increased in the Tph2 knockin mice. These data likewise parallel core findings in depression, but are usually attributed to anomalies in the respective receptors rather than resulting from CNS 5-HT deficiency. Further, 5-HT(2A) receptor function is enhanced in the Tph2 knockin mice. In contrast, 5-HT(1A) receptor levels and G-protein coupling is normal in Tph2 knockin mice, indicating that the blunted hypothermic response relates directly to the low 5-HT(Ext). Thus, we show that not only low CSF 5-HIAA and a blunted fenfluramine-induced prolactin response, but also blunted 5-HT(1A) agonist-induced hypothermia and increased 5-HT(2A) receptor levels are bona fide biomarkers of chronic, endogenous 5-HT deficiency. Potentially, some of these biomarkers could identify patients likely to have 5-HT deficiency. This could have clinical research utility or even guide pharmacotherapy.

Authors
Jacobsen, JPR; Siesser, WB; Sachs, BD; Peterson, S; Cools, MJ; Setola, V; Folgering, JHA; Flik, G; Caron, MG
MLA Citation
Jacobsen, JPR, Siesser, WB, Sachs, BD, Peterson, S, Cools, MJ, Setola, V, Folgering, JHA, Flik, G, and Caron, MG. "Deficient serotonin neurotransmission and depression-like serotonin biomarker alterations in tryptophan hydroxylase 2 (Tph2) loss-of-function mice." Mol Psychiatry 17.7 (July 2012): 694-704.
PMID
21537332
Source
pubmed
Published In
Molecular Psychiatry
Volume
17
Issue
7
Publish Date
2012
Start Page
694
End Page
704
DOI
10.1038/mp.2011.50

Discovery of Small Molecule Kappa Opioid Receptor Agonist and Antagonist Chemotypes through a HTS and Hit Refinement Strategy.

Herein we present the outcome of a high throughput screening (HTS) campaign-based strategy for the rapid identification and optimization of selective and general chemotypes for both kappa (κ) opioid receptor (KOR) activation and inhibition. In this program, we have developed potent antagonists (IC(50) < 120 nM) or agonists of high binding affinity (K(i) < 3 nM). In contrast to many important KOR ligands, the compounds presented here are highly modular, readily synthesized and, in most cases, achiral. The four new chemotypes hold promise for further development into chemical tools for studying the KOR or as potential therapeutic lead candidates.

Authors
Frankowski, KJ; Hedrick, MP; Gosalia, P; Li, K; Shi, S; Whipple, D; Ghosh, P; Prisinzano, TE; Schoenen, FJ; Su, Y; Vasile, S; Sergienko, E; Gray, W; Hariharan, S; Milan, L; Heynen-Genel, S; Mangravita-Novo, A; Vicchiarelli, M; Smith, LH; Streicher, JM; Caron, MG; Barak, LS; Bohn, LM; Chung, TDY; Aubé, J
MLA Citation
Frankowski, KJ, Hedrick, MP, Gosalia, P, Li, K, Shi, S, Whipple, D, Ghosh, P, Prisinzano, TE, Schoenen, FJ, Su, Y, Vasile, S, Sergienko, E, Gray, W, Hariharan, S, Milan, L, Heynen-Genel, S, Mangravita-Novo, A, Vicchiarelli, M, Smith, LH, Streicher, JM, Caron, MG, Barak, LS, Bohn, LM, Chung, TDY, and Aubé, J. "Discovery of Small Molecule Kappa Opioid Receptor Agonist and Antagonist Chemotypes through a HTS and Hit Refinement Strategy." ACS Chem Neurosci 3.3 (March 21, 2012): 221-236.
PMID
22737280
Source
pubmed
Published In
ACS Chemical Neuroscience
Volume
3
Issue
3
Publish Date
2012
Start Page
221
End Page
236
DOI
10.1021/cn200128x

Gene-dose dependent effects of methamphetamine on interval timing in dopamine-transporter knockout mice.

The dopamine transporter (DAT) is the major regulator of the spatial and temporal resolution of dopaminergic neurotransmission in the brain. Hyperdopaminergic mice with DAT gene deletions were evaluated for their ability to perform duration discriminations in the seconds-to-minutes range. DAT -/- mice were unable to demonstrate temporal control of behavior in either fixed-interval or peak-interval timing procedures, whereas DAT +/- mice were similar to DAT +/+ mice under normal conditions. Low to moderate-dose methamphetamine (MAP) challenges indicated that DAT +/- mice were less sensitive to the clock-speed enhancing effects of MAP compared with DAT +/+ mice. In contrast, DAT +/- mice were more vulnerable than DAT +/+ mice to the disruptive effects of MAP at high doses as revealed by the elevation of response rate in the right hand tail of the Gaussian-shaped timing functions. Moreover, this treatment made DAT +/- mice functionally equivalent to DAT -/- mice in terms of the loss of temporal control. Taken together, these results demonstrate the importance of dopaminergic control of interval timing in cortico-striatal circuits and the potential link of timing dysfunctions to schizophrenia and drug abuse.

Authors
Meck, WH; Cheng, R-K; MacDonald, CJ; Gainetdinov, RR; Caron, MG; Cevik, MÖ
MLA Citation
Meck, WH, Cheng, R-K, MacDonald, CJ, Gainetdinov, RR, Caron, MG, and Cevik, MÖ. "Gene-dose dependent effects of methamphetamine on interval timing in dopamine-transporter knockout mice." Neuropharmacology 62.3 (March 2012): 1221-1229.
PMID
21296093
Source
pubmed
Published In
Neuropharmacology
Volume
62
Issue
3
Publish Date
2012
Start Page
1221
End Page
1229
DOI
10.1016/j.neuropharm.2011.01.042

GRK2: multiple roles beyond G protein-coupled receptor desensitization.

G protein-coupled receptor kinases (GRKs) regulate numerous G protein-coupled receptors (GPCRs) by phosphorylating the intracellular domain of the active receptor, resulting in receptor desensitization and internalization. GRKs also regulate GPCR trafficking in a phosphorylation-independent manner via direct protein-protein interactions. Emerging evidence suggests that GRK2, the most widely studied member of this family of kinases, modulates multiple cellular responses in various physiological contexts by either phosphorylating non-receptor substrates or interacting directly with signaling molecules. In this review, we discuss traditional and newly discovered roles of GRK2 in receptor internalization and signaling as well as its impact on non-receptor substrates. We also discuss novel exciting roles of GRK2 in the regulation of dopamine receptor signaling and in the activation and trafficking of the atypical GPCR, Smoothened (Smo).

Authors
Evron, T; Daigle, TL; Caron, MG
MLA Citation
Evron, T, Daigle, TL, and Caron, MG. "GRK2: multiple roles beyond G protein-coupled receptor desensitization." Trends Pharmacol Sci 33.3 (March 2012): 154-164. (Review)
PMID
22277298
Source
pubmed
Published In
Trends in Pharmacological Sciences
Volume
33
Issue
3
Publish Date
2012
Start Page
154
End Page
164
DOI
10.1016/j.tips.2011.12.003

The role of GRK6 in animal models of Parkinson's disease and L-DOPA treatment.

G protein-coupled Receptor Kinase 6 (GRK6) belongs to a family of kinases that phosphorylate GPCRs. GRK6 levels were found to be altered in Parkinson's Disease (PD) and D(2) dopamine receptors are supersensitive in mice lacking GRK6 (GRK6-KO mice). To understand how GRK6 modulates the behavioral manifestations of dopamine deficiency and responses to L-DOPA, we used three approaches to model PD in GRK6-KO mice: 1) the cataleptic response to haloperidol; 2) introducing GRK6 mutation to an acute model of absolute dopamine deficiency, DDD mice; 3) hemiparkinsonian 6-OHDA model. Furthermore, dopamine-related striatal signaling was analyzed by assessing the phosphorylation of AKT/GSK3β and ERK1/2. GRK6 deficiency reduced cataleptic behavior, potentiated the acute effect of L-DOPA in DDD mice, reduced rotational behavior in hemi-parkinsonian mice, and reduced abnormal involuntary movements induced by chronic L-DOPA. These data indicate that approaches to regulate GRK6 activity could be useful in modulating both therapeutic and side-effects of L-DOPA.

Authors
Managò, F; Espinoza, S; Salahpour, A; Sotnikova, TD; Caron, MG; Premont, RT; Gainetdinov, RR
MLA Citation
Managò, F, Espinoza, S, Salahpour, A, Sotnikova, TD, Caron, MG, Premont, RT, and Gainetdinov, RR. "The role of GRK6 in animal models of Parkinson's disease and L-DOPA treatment." Sci Rep 2 (2012): 301-.
Website
http://hdl.handle.net/10161/10773
PMID
22393477
Source
pubmed
Published In
Scientific Reports
Volume
2
Publish Date
2012
Start Page
301
DOI
10.1038/srep00301

Pregnenolone rescues schizophrenia-like behavior in dopamine transporter knockout mice.

Pregnenolone belongs to a class of endogenous neurosteroids in the central nervous system (CNS), which has been suggested to enhance cognitive functions through GABA(A) receptor signaling by its metabolites. It has been shown that the level of pregnenolone is altered in certain brain areas of schizophrenic patients, and clozapine enhances pregnenolone in the CNS in rats, suggesting that pregnenolone could be used to treat certain symptoms of schizophrenia. In addition, early phase proof-of-concept clinical trials have indicated that pregnenolone is effective in reducing the negative symptoms and cognitive deficits of schizophrenia patients. Here, we evaluate the actions of pregnenolone on a mouse model for schizophrenia, the dopamine transporter knockout mouse (DAT KO). DAT KO mice mirror certain symptoms evident in patients with schizophrenia, such as the psychomotor agitation, stereotypy, deficits of prepulse inhibition and cognitive impairments. Following acute treatment, pregnenolone was found to reduce the hyperlocomotion, stereotypic bouts and pre-pulse inhibition (PPI) deficits in DAT KO mice in a dose-dependent manner. At 60 mg/kg of pregnenolone, there were no significant differences in locomotor activities and stereotypy between wild-type and DAT KO mice. Similarly, acute treatment of 60 mg/kg of pregnenolone fully rescued PPI deficits of DAT KO mice. Following chronic treatment with pregnenolone at 60 mg/kg, the cognitive deficits of DAT KO mice were rescued in the paradigms of novel object recognition test and social transmission of food preference test. Pregnenolone thus holds promise as a therapeutic candidate in schizophrenia.

Authors
Wong, P; Chang, CCR; Marx, CE; Caron, MG; Wetsel, WC; Zhang, X
MLA Citation
Wong, P, Chang, CCR, Marx, CE, Caron, MG, Wetsel, WC, and Zhang, X. "Pregnenolone rescues schizophrenia-like behavior in dopamine transporter knockout mice." PLoS One 7.12 (2012): e51455-.
PMID
23240026
Source
pubmed
Published In
PloS one
Volume
7
Issue
12
Publish Date
2012
Start Page
e51455
DOI
10.1371/journal.pone.0051455

Optimizing the temporal resolution of fast-scan cyclic voltammetry

(Figure Presented) Electrochemical detection with carbon-fiber microelectrodes has become an established method to monitor directly the release of dopamine from neurons and its uptake by the dopamine transporter. With constant potential amperometry (CPA), the measured current provides a real time view of the rapid concentration changes, but the method lacks chemical identification of the monitored species and markedly increases the difficulty of signal calibration. Monitoring with fast-scan cyclic voltammetry (FSCV) allows species identification and concentration measurements but often exhibits a delayed response time due to the time-dependent adsorption/desorption of electroactive species at the electrode. We sought to improve the temporal resolution of FSCV to make it more comparable to CPA by increasing the waveform repetition rate from 10 to 60 Hz with uncoated carbon-fiber electrodes. The faster acquisition led to diminished time delays of the recordings that tracked more closely with CPA measurements. The measurements reveal that FSCV at 10 Hz underestimates the normal rate of dopamine uptake by about 18%. However, FSCV collection at 10 and 60 Hz provide identical results when a dopamine transporter (DAT) blocker such as cocaine is bath applied. To verify further the utility of this method, we used transgenic mice that overexpress DAT. After accounting for the slight adsorption delay time, FSCV at 60 Hz adequately monitored the increased uptake rate that arose from overexpression of DAT and, again, was similar to CPA results. Furthermore, the utility of collecting data at 60 Hz was verified in an anesthetized rat by using a higher scan rate (2400 V/s) to increase sensitivity and the overall signal. © 2012 American Chemical Society.

Authors
Kile, BM; Walsh, PL; McElligott, ZA; Bucher, ES; Guillot, TS; Salahpour, A; Caron, MG; Wightman, RM
MLA Citation
Kile, BM, Walsh, PL, McElligott, ZA, Bucher, ES, Guillot, TS, Salahpour, A, Caron, MG, and Wightman, RM. "Optimizing the temporal resolution of fast-scan cyclic voltammetry." ACS Chemical Neuroscience 3.4 (2012): 285-292.
PMID
22708011
Source
scival
Published In
ACS Chemical Neuroscience
Volume
3
Issue
4
Publish Date
2012
Start Page
285
End Page
292
DOI
10.1021/cn200119u

Dual control of dopamine synthesis and release by presynaptic and postsynaptic dopamine D2 receptors

Dysfunctions of dopaminergic homeostasis leading to either low or high dopamine (DA) levels are causally linked to Parkinson's disease, schizophrenia, and addiction. Major sites of DA synthesis are the mesencephalic neurons originating in the substantia nigra and ventral tegmental area; these structures send major projections to the dorsal striatum (DSt) and nucleus accumbens (NAcc), respectively. DA finely tunes its own synthesis and release by activatingDAD2 receptors (D2R). To date, this critical D2R-dependent function was thought to be solely due to activation of D2Rs on dopaminergic neurons (D2 autoreceptors); instead, using site-specific D2R knock-out mice, we uncover that D2 heteroreceptors located on non-DAergic medium spiny neurons participate in the control of DA levels. This D2 heteroreceptor-mediated mechanism is more efficient in the DSt than in NAcc, indicating that D2R signaling differentially regulates mesolimbic- versus nigrostriatal-mediated functions. This study reveals previously unappreciated control of DA signaling, shedding new light on region-specific regulation of DA-mediated effects. © 2012 the authors.

Authors
Anzalone, A; Lizardi-Ortiz, JE; Ramos, M; Mei, CD; Hopf, FW; Iaccarino, C; Halbout, B; Jacobsen, J; Kinoshita, C; Welter, M; Caron, MG; Bonci, A; Sulzer, D; Borrelli, E
MLA Citation
Anzalone, A, Lizardi-Ortiz, JE, Ramos, M, Mei, CD, Hopf, FW, Iaccarino, C, Halbout, B, Jacobsen, J, Kinoshita, C, Welter, M, Caron, MG, Bonci, A, Sulzer, D, and Borrelli, E. "Dual control of dopamine synthesis and release by presynaptic and postsynaptic dopamine D2 receptors." Journal of Neuroscience 32.26 (2012): 9023-9024.
PMID
22745501
Source
scival
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
32
Issue
26
Publish Date
2012
Start Page
9023
End Page
9024
DOI
10.1523/JNEUROSCI.0918-12.2012

Food for Thought: The Physiological Relevance of Ghrelin and Dopamine D2 Receptor Heterodimerization in the Regulation of Appetite

Establishing whether G protein-coupled receptors (GPCRs) form physiologically relevant functional homo- and heteroligomers invivo has been a major biochemical challenge. In this issue of Neuron, Kern et al. (2012) investigate whether the anorexigenic effects of D2 dopamine receptors may be a direct consequence of allosteric modulation by Apo-ghrelin receptors. © 2012 Elsevier Inc.

Authors
Salahpour, A; Caron, M
MLA Citation
Salahpour, A, and Caron, M. "Food for Thought: The Physiological Relevance of Ghrelin and Dopamine D2 Receptor Heterodimerization in the Regulation of Appetite." Neuron 73.2 (2012): 210-211.
PMID
22284175
Source
scival
Published In
Neuron
Volume
73
Issue
2
Publish Date
2012
Start Page
210
End Page
211
DOI
10.1016/j.neuron.2012.01.004

Consideration of allosterism and interacting proteins in the physiological functions of the serotonin transporter

The serotonin transporter (SERT) functions to transport serotonin (5-HT) from the extracellular space into neurons to maintain homeostatic control of 5-HT. It is the molecular target for selective serotonin reuptake inhibitor (SSRI) antidepressants. Preclinical research has shown that some SERT inhibitors can bind to two distinct binding sites on the SERT, a primary high affinity binding site and a low affinity allosteric binding site. Mutational studies of the SERT and computational modeling methods with escitalopram resulted in the identification of key amino acid residues important for the function of the allosteric binding site. While this allosteric binding site appears to influence the clinical efficacy of escitalopram under physiological conditions, the molecular mechanism of this effect is still poorly understood and may involve a large network of protein-protein interactions with the SERT. Dynamic interfaces between the SERT and the SERT interacting proteins (SIPs) potentially influence not only the SERT on its uptake function, its regulation, and trafficking, but also on known as well as yet to be identified non-canonical signaling pathways through SIPs. In this commentary, we outline approaches in the areas of selective small-molecule allosteric compound discovery, biochemistry, in vivo genetic knock-in mouse models, as well as computational and structural biology. These studies of the intra-molecular allosteric modulation of the SERT in the context of the myriad of potential inter-molecular signaling interactions with SIPs may help uncover unknown physiological functions of the SERT. © 2011 Elsevier Inc. All rights reserved.

Authors
Zhong, H; Sánchez, C; Caron, MG
MLA Citation
Zhong, H, Sánchez, C, and Caron, MG. "Consideration of allosterism and interacting proteins in the physiological functions of the serotonin transporter." Biochemical Pharmacology 83.4 (2012): 435-442.
PMID
21983034
Source
scival
Published In
Biochemical Pharmacology
Volume
83
Issue
4
Publish Date
2012
Start Page
435
End Page
442
DOI
10.1016/j.bcp.2011.09.020

A functional alternative splicing mutation in human tryptophan hydroxylase-2.

The brain serotonergic system has an essential role in the physiological functions of the central nervous system and dysregulation of serotonin (5-HT) homeostasis has been implicated in many neuropsychiatric disorders. The tryptophan hydroxylase-2 (TPH2) gene is the rate-limiting enzyme in brain 5-HT synthesis, and thus is an ideal candidate gene for understanding the role of dysregulation of brain serotonergic homeostasis. Here, we characterized a common, but functional single-nucleotide polymorphism (SNP rs1386493) in the TPH2 gene, which decreases efficiency of normal RNA splicing, resulting in a truncated TPH2 protein (TPH2-TR) by alternative splicing. TPH2-TR, which lacks TPH2 enzyme activity, dominant-negatively affects full-length TPH2 function, causing reduced 5-HT production. The predicted mRNA for TPH2-TR is present in postmortem brain of rs1386493 carriers. The rs13864923 variant does not appear to be overrepresented in either global or multiplex depression cohorts. However, in combination with other gene variants linked to 5-HT homeostasis, this variant may exhibit important epistatic influences.

Authors
Zhang, X; Nicholls, PJ; Laje, G; Sotnikova, TD; Gainetdinov, RR; Albert, PR; Rajkowska, G; Stockmeier, CA; Speer, MC; Steffens, DC; Austin, MC; McMahon, FJ; Krishnan, KRR; Garcia-Blanco, MA; Caron, MG
MLA Citation
Zhang, X, Nicholls, PJ, Laje, G, Sotnikova, TD, Gainetdinov, RR, Albert, PR, Rajkowska, G, Stockmeier, CA, Speer, MC, Steffens, DC, Austin, MC, McMahon, FJ, Krishnan, KRR, Garcia-Blanco, MA, and Caron, MG. "A functional alternative splicing mutation in human tryptophan hydroxylase-2." Mol Psychiatry 16.12 (December 2011): 1169-1176.
PMID
20856248
Source
pubmed
Published In
Molecular Psychiatry
Volume
16
Issue
12
Publish Date
2011
Start Page
1169
End Page
1176
DOI
10.1038/mp.2010.99

Discovery of β-arrestin-biased dopamine D2 ligands for probing signal transduction pathways essential for antipsychotic efficacy.

Elucidating the key signal transduction pathways essential for both antipsychotic efficacy and side-effect profiles is essential for developing safer and more effective therapies. Recent work has highlighted noncanonical modes of dopamine D(2) receptor (D(2)R) signaling via β-arrestins as being important for the therapeutic actions of both antipsychotic and antimanic agents. We thus sought to create unique D(2)R agonists that display signaling bias via β-arrestin-ergic signaling. Through a robust diversity-oriented modification of the scaffold represented by aripiprazole (1), we discovered UNC9975 (2), UNC0006 (3), and UNC9994 (4) as unprecedented β-arrestin-biased D(2)R ligands. These compounds also represent unprecedented β-arrestin-biased ligands for a G(i)-coupled G protein-coupled receptor (GPCR). Significantly, UNC9975, UNC0006, and UNC9994 are simultaneously antagonists of G(i)-regulated cAMP production and partial agonists for D(2)R/β-arrestin-2 interactions. Importantly, UNC9975 displayed potent antipsychotic-like activity without inducing motoric side effects in inbred C57BL/6 mice in vivo. Genetic deletion of β-arrestin-2 simultaneously attenuated the antipsychotic actions of UNC9975 and transformed it into a typical antipsychotic drug with a high propensity to induce catalepsy. Similarly, the antipsychotic-like activity displayed by UNC9994, an extremely β-arrestin-biased D(2)R agonist, in wild-type mice was completely abolished in β-arrestin-2 knockout mice. Taken together, our results suggest that β-arrestin signaling and recruitment can be simultaneously a significant contributor to antipsychotic efficacy and protective against motoric side effects. These functionally selective, β-arrestin-biased D(2)R ligands represent valuable chemical probes for further investigations of D(2)R signaling in health and disease.

Authors
Allen, JA; Yost, JM; Setola, V; Chen, X; Sassano, MF; Chen, M; Peterson, S; Yadav, PN; Huang, X-P; Feng, B; Jensen, NH; Che, X; Bai, X; Frye, SV; Wetsel, WC; Caron, MG; Javitch, JA; Roth, BL; Jin, J
MLA Citation
Allen, JA, Yost, JM, Setola, V, Chen, X, Sassano, MF, Chen, M, Peterson, S, Yadav, PN, Huang, X-P, Feng, B, Jensen, NH, Che, X, Bai, X, Frye, SV, Wetsel, WC, Caron, MG, Javitch, JA, Roth, BL, and Jin, J. "Discovery of β-arrestin-biased dopamine D2 ligands for probing signal transduction pathways essential for antipsychotic efficacy." Proc Natl Acad Sci U S A 108.45 (November 8, 2011): 18488-18493.
PMID
22025698
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
108
Issue
45
Publish Date
2011
Start Page
18488
End Page
18493
DOI
10.1073/pnas.1104807108

Opposite function of dopamine D1 and N-methyl-D-aspartate receptors in striatal cannabinoid-mediated signaling.

It is well established that the cannabinoid and dopamine systems interact at various levels to regulate basal ganglia function. Although it is well known that acute administration of cannabinoids to mice can modify dopamine-dependent behaviors, the intraneuronal signaling pathways employed by these agents in the striatum are not well understood. Here we used knockout mouse models to examine the regulation of striatal extracellular-signal-regulated kinases 1 and 2 (ERK1/2) signaling by behaviorally relevant doses of cannabinoids. This cellular pathway has been implicated as a central mediator of drug reward and synaptic plasticity. In C57BL/6J mice, acute administration of the cannabinoid agonists, (-)-11-hydroxydimethylheptyl-Δ8-tetrahydrocannabinol (HU-210) and delta-9-tetrahydrocannabinol (Δ(9) -THC), promoted a dose- and time-dependent decrease in the phosphorylation of ERK1/2 in dorsal striatum. Co-administration of the CB1 cannabinoid receptor antagonist N-(Piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide(AM251) with HU-210 prevented ERK1/2 inactivation, indicating a requirement for activation of this receptor. In dopamine D1 receptor knockout animals treated with HU-210, the magnitude of the HU-210-dependent decrease in striatal ERK1/2 signaling was greater than in wild-type controls. In contrast, HU-210 administration to N-methyl-D-aspartate receptor knockdown mice was ineffective at promoting striatal ERK1/2 inactivation. Genetic deletion of other potential ERK1/2 mediators, the dopamine D2 receptors or β-arrestin-1 or -2, did not affect the HU-210-induced modulation of ERK1/2 signaling in the striatum. These results support the hypothesis that dopamine D1 receptors and N-methyl-D-aspartate receptors act in an opposite manner to regulate striatal CB1 cannabinoid receptor signal transduction.

Authors
Daigle, TL; Wetsel, WC; Caron, MG
MLA Citation
Daigle, TL, Wetsel, WC, and Caron, MG. "Opposite function of dopamine D1 and N-methyl-D-aspartate receptors in striatal cannabinoid-mediated signaling." Eur J Neurosci 34.9 (November 2011): 1378-1389.
PMID
22034973
Source
pubmed
Published In
European Journal of Neuroscience
Volume
34
Issue
9
Publish Date
2011
Start Page
1378
End Page
1389
DOI
10.1111/j.1460-9568.2011.07874.x

Elimination of the vesicular acetylcholine transporter in the striatum reveals regulation of behaviour by cholinergic-glutamatergic co-transmission.

Cholinergic neurons in the striatum are thought to play major regulatory functions in motor behaviour and reward. These neurons express two vesicular transporters that can load either acetylcholine or glutamate into synaptic vesicles. Consequently cholinergic neurons can release both neurotransmitters, making it difficult to discern their individual contributions for the regulation of striatal functions. Here we have dissected the specific roles of acetylcholine release for striatal-dependent behaviour in mice by selective elimination of the vesicular acetylcholine transporter (VAChT) from striatal cholinergic neurons. Analysis of several behavioural parameters indicates that elimination of VAChT had only marginal consequences in striatum-related tasks and did not affect spontaneous locomotion, cocaine-induced hyperactivity, or its reward properties. However, dopaminergic sensitivity of medium spiny neurons (MSN) and the behavioural outputs in response to direct dopaminergic agonists were enhanced, likely due to increased expression/function of dopamine receptors in the striatum. These observations indicate that previous functions attributed to striatal cholinergic neurons in spontaneous locomotor activity and in the rewarding responses to cocaine are mediated by glutamate and not by acetylcholine release. Our experiments demonstrate how one population of neurons can use two distinct neurotransmitters to differentially regulate a given circuitry. The data also raise the possibility of using VAChT as a target to boost dopaminergic function and decrease high striatal cholinergic activity, common neurochemical alterations in individuals affected with Parkinson's disease.

Authors
Guzman, MS; De Jaeger, X; Raulic, S; Souza, IA; Li, AX; Schmid, S; Menon, RS; Gainetdinov, RR; Caron, MG; Bartha, R; Prado, VF; Prado, MAM
MLA Citation
Guzman, MS, De Jaeger, X, Raulic, S, Souza, IA, Li, AX, Schmid, S, Menon, RS, Gainetdinov, RR, Caron, MG, Bartha, R, Prado, VF, and Prado, MAM. "Elimination of the vesicular acetylcholine transporter in the striatum reveals regulation of behaviour by cholinergic-glutamatergic co-transmission." PLoS Biol 9.11 (November 2011): e1001194-.
PMID
22087075
Source
pubmed
Published In
PLoS biology
Volume
9
Issue
11
Publish Date
2011
Start Page
e1001194
DOI
10.1371/journal.pbio.1001194

Rod vision is controlled by dopamine-dependent sensitization of rod bipolar cells by GABA.

Dark and light adaptation of retinal neurons allow our vision to operate over an enormous light intensity range. Here we report a mechanism that controls the light sensitivity and operational range of rod-driven bipolar cells that mediate dim-light vision. Our data indicate that the light responses of these cells are enhanced by sustained chloride currents via GABA(C) receptor channels. This sensitizing GABAergic input is controlled by dopamine D1 receptors, with horizontal cells serving as a plausible source of GABA release. Our findings expand the role of dopamine in vision from its well-established function of suppressing rod-driven signals in bright light to enhancing the same signals under dim illumination. They further reveal a role for GABA in sensitizing the circuitry for dim-light vision, thereby complementing GABA's traditional role in providing dynamic feedforward and feedback inhibition in the retina.

Authors
Herrmann, R; Heflin, SJ; Hammond, T; Lee, B; Wang, J; Gainetdinov, RR; Caron, MG; Eggers, ED; Frishman, LJ; McCall, MA; Arshavsky, VY
MLA Citation
Herrmann, R, Heflin, SJ, Hammond, T, Lee, B, Wang, J, Gainetdinov, RR, Caron, MG, Eggers, ED, Frishman, LJ, McCall, MA, and Arshavsky, VY. "Rod vision is controlled by dopamine-dependent sensitization of rod bipolar cells by GABA." Neuron 72.1 (October 6, 2011): 101-110.
PMID
21982372
Source
pubmed
Published In
Neuron
Volume
72
Issue
1
Publish Date
2011
Start Page
101
End Page
110
DOI
10.1016/j.neuron.2011.07.030

Functional interaction between trace amine-associated receptor 1 and dopamine D2 receptor.

The ability of dopamine receptors to interact with other receptor subtypes may provide mechanisms for modulating dopamine-related functions and behaviors. In particular, there is evidence suggesting that the trace amine-associated receptor 1 (TAAR1) affects the dopaminergic system by regulating the firing rate of dopaminergic neurons or by altering dopamine D2 receptor (D2R) responsiveness to ligands. TAAR1 is a Gα(s) protein-coupled receptor that is activated by biogenic amines, "trace amines," such as β-phenylethylamine (β-PEA) and tyramine that are normally found at low concentrations in the mammalian brain. In the present study, we investigated the biochemical mechanism of interaction between TAAR1 and D2R and the role this interaction plays in D2R-related signaling and behaviors. Using a bioluminescence resonance energy transfer biosensor for cAMP, we demonstrated that the D2R antagonists haloperidol, raclopride, and amisulpride were able to enhance selectively a TAAR1-mediated β-PEA increase of cAMP. Moreover, TAAR1 and D2R were able to form heterodimers when coexpressed in human embryonic kidney 293 cells, and this direct interaction was disrupted in the presence of haloperidol. In addition, in mice lacking TAAR1, haloperidol-induced striatal c-Fos expression and catalepsy were significantly reduced. Taken together, these data suggest that TAAR1 and D2R have functional and physical interactions that could be critical for the modulation of the dopaminergic system by TAAR1 in vivo.

Authors
Espinoza, S; Salahpour, A; Masri, B; Sotnikova, TD; Messa, M; Barak, LS; Caron, MG; Gainetdinov, RR
MLA Citation
Espinoza, S, Salahpour, A, Masri, B, Sotnikova, TD, Messa, M, Barak, LS, Caron, MG, and Gainetdinov, RR. "Functional interaction between trace amine-associated receptor 1 and dopamine D2 receptor." Mol Pharmacol 80.3 (September 2011): 416-425.
PMID
21670104
Source
pubmed
Published In
Molecular pharmacology
Volume
80
Issue
3
Publish Date
2011
Start Page
416
End Page
425
DOI
10.1124/mol.111.073304

Growth Arrest Specific 8 (Gas8) and G protein-coupled receptor kinase 2 (GRK2) cooperate in the control of Smoothened signaling.

The G protein-coupled receptor (GPCR)-like molecule Smoothened (Smo) undergoes dynamic intracellular trafficking modulated by the microtubule associated kinase GRK2 and recruitment of β-arrestin. Of this trafficking, especially the translocation of Smo into primary cilia and back to the cytoplasm is essential for the activation of Hedgehog (Hh) signaling in vertebrates. The complete mechanism of this bidirectional transport, however, is not completely understood. Here we demonstrate that Growth Arrest Specific 8 (Gas8), a microtubule associated subunit of the Dynein Regulatory Complex (DRC), interacts with Smo to modulate this process. Gas8 knockdown in ciliated cells reduces Smo signaling activity and ciliary localization whereas overexpression stimulates Smo activity in a GRK2-dependent manner. The C terminus of Gas8 is important for both Gas8 interaction with Smo and facilitating Smo signaling. In zebrafish, knocking down Gas8 results in attenuated Hh transcriptional responses and impaired early muscle development. These effects can be reversed by the co-injection of Gas8 mRNA or by constitutive activation of the downstream Gli transcription factors. Furthermore, Gas8 and GRK2 display a synergistic effect on zebrafish early muscle development and some effects of GRK2 knockdown can be rescued by Gas8 mRNA. Interestingly, Gas8 does not interfere with cilia assembly, as the primary cilia architecture is unchanged upon Gas8 knock down or heterologous expression. This is in contrast to cells stably expressing both GRK2 and Smo, in which cilia are significantly elongated. These results identify Gas8 as a positive regulator of Hh signaling that cooperates with GRK2 to control Smo signaling.

Authors
Evron, T; Philipp, M; Lu, J; Meloni, AR; Burkhalter, M; Chen, W; Caron, MG
MLA Citation
Evron, T, Philipp, M, Lu, J, Meloni, AR, Burkhalter, M, Chen, W, and Caron, MG. "Growth Arrest Specific 8 (Gas8) and G protein-coupled receptor kinase 2 (GRK2) cooperate in the control of Smoothened signaling." J Biol Chem 286.31 (August 5, 2011): 27676-27686.
PMID
21659505
Source
pubmed
Published In
The Journal of biological chemistry
Volume
286
Issue
31
Publish Date
2011
Start Page
27676
End Page
27686
DOI
10.1074/jbc.M111.234666

TAAR1 activation modulates monoaminergic neurotransmission, preventing hyperdopaminergic and hypoglutamatergic activity.

The trace amine-associated receptor 1 (TAAR1), activated by endogenous metabolites of amino acids like the trace amines p-tyramine and β-phenylethylamine, has proven to be an important modulator of the dopaminergic system and is considered a promising target for the treatment of neuropsychiatric disorders. To decipher the brain functions of TAAR1, a selective TAAR1 agonist, RO5166017, was engineered. RO5166017 showed high affinity and potent functional activity at mouse, rat, cynomolgus monkey, and human TAAR1 stably expressed in HEK293 cells as well as high selectivity vs. other targets. In mouse brain slices, RO5166017 inhibited the firing frequency of dopaminergic and serotonergic neurons in regions where Taar1 is expressed (i.e., the ventral tegmental area and dorsal raphe nucleus, respectively). In contrast, RO5166017 did not change the firing frequency of noradrenergic neurons in the locus coeruleus, an area devoid of Taar1 expression. Furthermore, modulation of TAAR1 activity altered the desensitization rate and agonist potency at 5-HT(1A) receptors in the dorsal raphe, suggesting that TAAR1 modulates not only dopaminergic but also serotonergic neurotransmission. In WT but not Taar1(-/-) mice, RO5166017 prevented stress-induced hyperthermia and blocked dopamine-dependent hyperlocomotion in cocaine-treated and dopamine transporter knockout mice as well as hyperactivity induced by an NMDA antagonist. These results tie TAAR1 to the control of monoamine-driven behaviors and suggest anxiolytic- and antipsychotic-like properties for agonists such as RO5166017, opening treatment opportunities for psychiatric disorders.

Authors
Revel, FG; Moreau, J-L; Gainetdinov, RR; Bradaia, A; Sotnikova, TD; Mory, R; Durkin, S; Zbinden, KG; Norcross, R; Meyer, CA; Metzler, V; Chaboz, S; Ozmen, L; Trube, G; Pouzet, B; Bettler, B; Caron, MG; Wettstein, JG; Hoener, MC
MLA Citation
Revel, FG, Moreau, J-L, Gainetdinov, RR, Bradaia, A, Sotnikova, TD, Mory, R, Durkin, S, Zbinden, KG, Norcross, R, Meyer, CA, Metzler, V, Chaboz, S, Ozmen, L, Trube, G, Pouzet, B, Bettler, B, Caron, MG, Wettstein, JG, and Hoener, MC. "TAAR1 activation modulates monoaminergic neurotransmission, preventing hyperdopaminergic and hypoglutamatergic activity." Proc Natl Acad Sci U S A 108.20 (May 17, 2011): 8485-8490.
PMID
21525407
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
108
Issue
20
Publish Date
2011
Start Page
8485
End Page
8490
DOI
10.1073/pnas.1103029108

Quantitative label-free phosphoproteomics strategy for multifaceted experimental designs.

Protein phosphorylation is a critical regulator of signaling in nearly all eukaryotic cellular pathways and dysregulated phosphorylation has been implicated in an array of diseases. The majority of MS-based quantitative phosphorylation studies are currently performed from transformed cell lines because of the ability to generate large amounts of starting material with incorporated isotopically labeled amino acids during cell culture. Here we describe a general label-free quantitative phosphoproteomic strategy capable of directly analyzing relatively small amounts of virtually any biological matrix, including human tissue and biological fluids. The strategy utilizes a TiO(2) enrichment protocol in which the selectivity and recovery of phosphopeptides were optimized by assessing a twenty-point condition matrix of binding modifier concentrations and peptide-to-resin capacity ratios. The quantitative reproducibility of the TiO(2) enrichment was determined to be 16% RSD through replicate enrichments of a wild-type Danio rerio (zebrafish) lysate. Measured phosphopeptide fold-changes from alpha-casein spiked into wild-type zebrafish lysate backgrounds were within 5% of the theoretical value. Application to a morpholino induced knock-down of G protein-coupled receptor kinase 5 (GRK5) in zebrafish embryos resulted in the quantitation of 719 phosphorylated peptides corresponding to 449 phosphorylated proteins from 200 μg of zebrafish embryo lysates.

Authors
Soderblom, EJ; Philipp, M; Thompson, JW; Caron, MG; Moseley, MA
MLA Citation
Soderblom, EJ, Philipp, M, Thompson, JW, Caron, MG, and Moseley, MA. "Quantitative label-free phosphoproteomics strategy for multifaceted experimental designs." Anal Chem 83.10 (May 15, 2011): 3758-3764.
PMID
21491946
Source
pubmed
Published In
Analytical Chemistry
Volume
83
Issue
10
Publish Date
2011
Start Page
3758
End Page
3764
DOI
10.1021/ac200213b

Impaired NMDA receptor transmission alters striatal synapses and DISC1 protein in an age-dependent manner.

NMDA receptors are key regulators of synaptic plasticity, and their hypofunction is thought to contribute to the pathophysiology of CNS disorders. Furthermore, NMDA receptors participate in the formation, maintenance, and elimination of synapses. The consequences of NMDA receptor hypofunction on synapse biology were explored in a genetic mouse model, in which the levels of NMDA receptors are reduced to 10% of normal levels (i.e., NR1-knockdown mice). In these mice, synapse number is reduced in an age-dependent manner; reductions are observed at the postpubertal age of 6 wk, but normal at 2 wk of age. Efforts to uncover the biochemical underpinnings of this phenomenon reveal synapse-specific reductions in 14-3-3ε protein and in Disrupted in Schizophrenia-1 (DISC1), two schizophrenia susceptibility factors that have been implicated in the regulation of spine density. Subchronic administration of MK-801, an NMDA receptor antagonist, produces similar synaptic reductions in both spine density and DISC1, indicating that synaptic levels of DISC1 are regulated by NMDA receptor function. The synaptic reduction of DISC1 and 14-3-3ε is developmentally correlated with the age-dependent decrease in striatal spine density.

Authors
Ramsey, AJ; Milenkovic, M; Oliveira, AF; Escobedo-Lozoya, Y; Seshadri, S; Salahpour, A; Sawa, A; Yasuda, R; Caron, MG
MLA Citation
Ramsey, AJ, Milenkovic, M, Oliveira, AF, Escobedo-Lozoya, Y, Seshadri, S, Salahpour, A, Sawa, A, Yasuda, R, and Caron, MG. "Impaired NMDA receptor transmission alters striatal synapses and DISC1 protein in an age-dependent manner." Proc Natl Acad Sci U S A 108.14 (April 5, 2011): 5795-5800.
PMID
21436042
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
108
Issue
14
Publish Date
2011
Start Page
5795
End Page
5800
DOI
10.1073/pnas.1012621108

Paracrine modulation of cholangiocyte serotonin synthesis orchestrates biliary remodeling in adults.

Paracrine signaling between cholangiocytes and stromal cells regulates biliary remodeling. Cholangiocytes have neuroepithelial characteristics and serotonin receptor agonists inhibit their growth, but whether they are capable of serotonin biosynthesis is unknown. We hypothesized that cholangiocytes synthesize serotonin and that cross talk between liver myofibroblasts (MF) and cholangiocytes regulates this process to influence biliary remodeling. Transwell cultures of cholangiocytes ± MF, and tryptophan hydroxylase-2 knockin (TPH2KI) mice with an inactivating mutation of the neuronal tryptophan hydroxylase (TPH) isoform, TPH2, were evaluated. Results in the cell culture models confirm that cholangiocytes have serotonin receptors and demonstrate for the first time that these cells express TPH2 and produce serotonin, which autoinhibits their growth but stimulates MF production of TGF-β(1). Increased TGF-β(1), in turn, counteracts autocrine inhibition of cholangiocyte growth by repressing cholangiocyte TPH2 expression. Studies of TPH2KI mice confirm that TPH2-mediated production of serotonin plays an important role in remodeling damaged bile ducts because mice with decreased TPH2 function have reduced biliary serotonin levels and exhibit excessive cholangiocyte proliferation, accumulation of aberrant ductules and liver progenitors, and increased liver fibrosis after bile duct ligation. This new evidence that cholangiocytes express the so-called neuronal isoform of TPH, synthesize serotonin de novo, and deploy serotonin as an autocrine/paracrine signal to regulate regeneration of the biliary tree complements earlier work that revealed that passive release of serotonin from platelets stimulates hepatocyte proliferation. Given the prevalent use of serotonin-modulating drugs, these findings have potentially important implications for recovery from various types of liver damage.

Authors
Omenetti, A; Yang, L; Gainetdinov, RR; Guy, CD; Choi, SS; Chen, W; Caron, MG; Diehl, AM
MLA Citation
Omenetti, A, Yang, L, Gainetdinov, RR, Guy, CD, Choi, SS, Chen, W, Caron, MG, and Diehl, AM. "Paracrine modulation of cholangiocyte serotonin synthesis orchestrates biliary remodeling in adults." Am J Physiol Gastrointest Liver Physiol 300.2 (February 2011): G303-G315.
PMID
21071507
Source
pubmed
Published In
American journal of physiology. Gastrointestinal and liver physiology
Volume
300
Issue
2
Publish Date
2011
Start Page
G303
End Page
G315
DOI
10.1152/ajpgi.00368.2010

A dopamine D1 receptor-dependent β-arrestin signaling complex potentially regulates morphine-induced psychomotor activation but not reward in mice.

Morphine is a widely used analgesic in humans that is associated with multiple untoward effects, such as addiction and physical dependence. In rodent models, morphine also induces locomotor activity. These effects likely involve functionally selective mechanisms. Indeed, G protein-coupled receptor desensitization and adaptor protein β-arrestin 2 (βarr2) through its interaction with the μ-opioid receptor regulates the analgesic but not the rewarding properties of morphine. However, βarr2 is also required for morphine-induced locomotor activity in mice, but the exact cellular and molecular mechanisms that mediate this arrestin-dependent behavior are not understood. In this study, we show that βarr2 is required for morphine-induced locomotor activity in a dopamine D1 receptor (D1R)-dependent manner and that a βarr2/phospho-ERK (βarr2/pERK) signaling complex may mediate this behavior. Systemic administration of SL327, an MEK inhibitor, inhibits morphine-induced locomotion in wild-type mice in a dose-dependent manner. Acute morphine administration to mice promotes the formation of a βarr2/pERK signaling complex. Morphine-induced locomotor activity and formation of the βarr2/pERK signaling complex is blunted in D1R knockout (D1-KO) mice and is presumably independent of D2 dopamine receptors. However, D1Rs are not required for morphine-induced reward as D1-KO mice show the same conditioned place preference for morphine as do control mice. Taken together, these results suggest a potential role for a D1R-dependent βarr2/pERK signaling complex in selectively mediating the locomotor-stimulating but not the rewarding properties of morphine.

Authors
Urs, NM; Daigle, TL; Caron, MG
MLA Citation
Urs, NM, Daigle, TL, and Caron, MG. "A dopamine D1 receptor-dependent β-arrestin signaling complex potentially regulates morphine-induced psychomotor activation but not reward in mice." Neuropsychopharmacology : official publication of the American College of Neuropsychopharmacology 36.3 (February 2011): 551-558.
PMID
20980993
Source
epmc
Published In
Neuropsychopharmacology (Nature)
Volume
36
Issue
3
Publish Date
2011
Start Page
551
End Page
558
DOI
10.1038/npp.2010.186

Identification of the GPR55 agonist binding site using a novel set of high-potency GPR55 selective ligands

Marijuana is the most widely abused illegal drug, and its spectrum of effects suggests that several receptors are responsible for the activity. Two cannabinoid receptor subtypes, CB1 and CB2, have been identified, but the complex pharmacological properties of exogenous cannabinoids and endocannabinoids are not fully explained by their signaling. The orphan receptor GPR55 binds a subset of CB1 and CB2 ligands and has been proposed as a cannabinoid receptor. This designation, however, is controversial as a result of recent studies in which lysophosphatidylinositol (LPI) was identified as a GPR55 agonist. Defining a biological role for GPR55 requires GPR55 selective ligands that have been unavailable. From a β-arrestin, high-throughput, high-content screen of 300000 compounds run in collaboration with the Molecular Libraries Probe Production Centers Network initiative (PubChem AID1965), we identified potent GPR55 selective agonists. By modeling of the GPR55 activated state, we compared the GPR55 binding conformations of three of the novel agonists obtained from the screen, CID1792197, CID1172084, and CID2440433 (PubChem Compound IDs), with that of LPI. Our modeling indicates the molecular shapes and electrostatic potential distributions of these agonists mimic those of LPI; the GPR55 binding site accommodates ligands that have inverted-L or T shapes with long, thin profiles that can fit vertically deep in the receptor binding pocket while their broad head regions occupy a horizontal binding pocket near the GPR55 extracellular loops. Our results will allow the optimization and design of second-generation GPR55 ligands and provide a means for distinguishing GPR55 selective ligands from those interacting with cannabinoid receptors. © 2011 American Chemical Society.

Authors
Kotsikorou, E; Madrigal, KE; Hurst, DP; Sharir, H; Lynch, DL; Heynen-Genel, S; Milan, LB; Chung, TDY; Seltzman, HH; Bai, Y; Caron, MG; Barak, L; Abood, ME; Reggio, PH
MLA Citation
Kotsikorou, E, Madrigal, KE, Hurst, DP, Sharir, H, Lynch, DL, Heynen-Genel, S, Milan, LB, Chung, TDY, Seltzman, HH, Bai, Y, Caron, MG, Barak, L, Abood, ME, and Reggio, PH. "Identification of the GPR55 agonist binding site using a novel set of high-potency GPR55 selective ligands." Biochemistry 50.25 (2011): 5633-5647.
PMID
24274581
Source
scival
Published In
Biochemistry
Volume
50
Issue
25
Publish Date
2011
Start Page
5633
End Page
5647
DOI
10.1021/bi200010k

Advancing drug discovery for schizophrenia

Sponsored by the New York Academy of Sciences and with support from the National Institute of Mental Health, the Life Technologies Foundation, and the Josiah Macy Jr. Foundation, "Advancing Drug Discovery for Schizophrenia" was held March 9-11 at the New York Academy of Sciences in New York City. The meeting, comprising individual talks and panel discussions, highlighted basic, clinical, and translational research approaches, all of which contribute to the overarching goal of enhancing the pharmaceutical armamentarium for treating schizophrenia. This report surveys work by the vanguard of schizophrenia research in such topics as genetic and epigenetic approaches; small molecule therapeutics; and the relationships between target genes, neuronal function, and symptoms of schizophrenia. © 2011 New York Academy of Sciences..

Authors
Marder, SR; Roth, B; Sullivan, PF; Scolnick, EM; Nestler, EJ; Geyer, MA; Welnberger, DR; Karayiorgou, M; Guidotti, A; Gingrich, J; Akbarian, S; Buchanan, RW; Lieberman, JA; Conn, PJ; Haggarty, SJ; Law, AJ; Campbell, B; Krystal, JH; Moghaddam, B; Saw, A; Caron, MG; George, SR; Allen, JA; Solis, M
MLA Citation
Marder, SR, Roth, B, Sullivan, PF, Scolnick, EM, Nestler, EJ, Geyer, MA, Welnberger, DR, Karayiorgou, M, Guidotti, A, Gingrich, J, Akbarian, S, Buchanan, RW, Lieberman, JA, Conn, PJ, Haggarty, SJ, Law, AJ, Campbell, B, Krystal, JH, Moghaddam, B, Saw, A, Caron, MG, George, SR, Allen, JA, and Solis, M. "Advancing drug discovery for schizophrenia." Annals of the New York Academy of Sciences 1236.1 (2011): 30-43.
PMID
22032400
Source
scival
Published In
Annals of the New York Academy of Sciences
Volume
1236
Issue
1
Publish Date
2011
Start Page
30
End Page
43
DOI
10.1111/j.1749-6632.2011.06216.x

Multiscale imaging characterization of dopamine transporter knockout mice reveals regional alterations in spine density of medium spiny neurons

The dopamine transporter knockout (DAT KO) mouse is a model of chronic hyperdopaminergia used to study a wide range of neuropsychiatric disorders such as schizophrenia, attention deficit hyperactivity disorder (ADHD), drug abuse, depression, and Parkinson's disease (PD). Early studies characterizing this mouse model revealed a subtle, but significant, decrease in the anterior striatal volume of DAT KO mice accompanied by a decrease in neuronal cell body numbers (Cyr et al., 2005). The present studies were conducted to examine medium spiny neuron (MSN) morphology by extending these earlier reports to include multiscale imaging studies using correlated light microscopy (LM) and electron microscopy (EM) techniques. Specifically, we set out to determine if chronic hyperdopaminergia results in quantifiable or qualitative changes in DAT KO mouse MSNs relative to wild-type (WT) littermates. Using Neurolucida Explorer's morphometric analysis, we measured spine density, dendritic length and synapse number at ages that correspond with the previously reported changes in striatal volume and progressive cell loss. Light microscopic analysis using Neurolucida tracings of photoconverted striatal MSNs revealed a highly localized loss of dendritic spines on the proximal portion of the dendrite (30 μm from the soma) in the DAT KO group. Next, thick sections containing MSN dendritic segments located at a distance of 20-60 μm from the cell soma, a region of the dendrite where spine density is reported to be the highest, were analyzed using electron microscope tomography (EMT). Because of the resolution limits of LM, the EM analysis was an extra measure taken to assure that our analysis included nearly all spines. Spine density measurements collected from the EMT data revealed only a modest decrease in the DAT KO group (n = 3 mice) compared to age-matched WT controls (n = 3 mice), a trend that supports the LM findings. Finally, a synaptic quantification using unbiased stereology did not detect a difference between DAT KO mice (n = 6 mice) and WT controls (n = 7 mice) at the EM level, supporting the focal nature of the early synaptic loss. These findings suggest that DAT KO mice have MSNs with highly localized spine loss and not an overall morphologically distinct cell shape. The characterization of morphological changes in DAT KO mice may provide information about the neural substrates underlying altered behaviors in these mice, with relevance for human neurological disorders thought to involve altered dopaminergic homeostasis. Results from this study also indicate the difficulty in correlating structural changes across scales, as the results on fine structure revealed thus far are subtle and non-uniform across striatal MSNs. The complexities associated with multiscale studies are driving the development of shared online informatics resources by gaining access to data where it is being analyzed. © 2011 Elsevier B.V. All rights reserved.

Authors
Berlanga, ML; Price, DL; Phung, BS; Giuly, R; Terada, M; Yamada, N; Cyr, M; Caron, MG; Laakso, A; Martone, ME; Ellisman, MH
MLA Citation
Berlanga, ML, Price, DL, Phung, BS, Giuly, R, Terada, M, Yamada, N, Cyr, M, Caron, MG, Laakso, A, Martone, ME, and Ellisman, MH. "Multiscale imaging characterization of dopamine transporter knockout mice reveals regional alterations in spine density of medium spiny neurons." Brain Research 1390 (2011): 41-49.
PMID
21439946
Source
scival
Published In
Brain Research
Volume
1390
Publish Date
2011
Start Page
41
End Page
49
DOI
10.1016/j.brainres.2011.03.044

Dissecting the contribution of individual receptor subunits to the enhancement of N-methyl-d-aspartate currents by dopamine D1 receptor activation in striatum

Dopamine, via activation of D1 receptors, enhances N-methyl-d-aspartate (NMDA) receptor-mediated responses in striatal medium-sized spiny neurons. However, the role of specific NMDA receptor subunits in this enhancement remains unknown. Here we used genetic and pharmacological tools to dissect the contribution of NR1 and NR2A/B subunits to NMDA responses and their modulation by dopamine receptors. We demonstrate that D1 enhancement of NMDA responses does not occur or is significantly reduced in mice with genetic knock-down of NR1 subunits, indicating a critical role of these subunits. Interestingly, spontaneous and evoked α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionic acid (AMPA) receptor-mediated responses were significantly enhanced in NR1 knock-down animals, probably as a compensatory mechanism for the marked reduction in NMDA receptor function. The NMDA receptor subunits NR2A and NR2B played differential roles in D1 modulation. Whereas genetic deletion or pharmacological blockade of NR2A subunits enhanced D1 potentiation of NMDA responses, blockade of NR2B subunits reduced this potentiation, suggesting that these regulatory subunits of the NMDA receptor counterbalance their respective functions. In addition, using D1 and D2 receptor EGFP-expressing mice, we demonstrate that NR2A subunits contribute more to NMDA responses in D1-MSSNs, whereas NR2B subunits contribute more to NMDA responses in D2 cells. The differential contribution of discrete receptor subunits to NMDA responses and dopamine modulation in the striatum has important implications for synaptic plasticity and selective neuronal vulnerability in disease states. © 2011 Jocoy, André, Cummings, Rao, Wu, Ramsey, Caron, Cepeda and Levine.

Authors
Jocoy, EL; André, VM; Cummings, DM; Rao, SP; Wu, N; Ramsey, AJ; Caron, MG; Cepeda, C; Levine, MS
MLA Citation
Jocoy, EL, André, VM, Cummings, DM, Rao, SP, Wu, N, Ramsey, AJ, Caron, MG, Cepeda, C, and Levine, MS. "Dissecting the contribution of individual receptor subunits to the enhancement of N-methyl-d-aspartate currents by dopamine D1 receptor activation in striatum." Frontiers in Systems Neuroscience MAY 2011 (2011).
Source
scival
Published In
Frontiers in Systems Neuroscience
Issue
MAY 2011
Publish Date
2011
DOI
10.3389/fnsys.2011.00028

A greater role for the norepinephrine transporter than the serotonin transporter in murine nociception

Norepinephrine and serotonin involvement in nociceptive functions is supported by observations of analgesic effects of norepinephrine transporter (NET) and serotonin transporter (SERT) inhibitors such as amitriptyline. However, the relative contribution of NET and SERT to baseline nociception, as well as amitriptyline analgesia, is unclear. Amitriptyline and morphine analgesia in wild-type (WT) mice and littermates with gene knockout (KO) of SERT, NET or both transporters was conducted using the hotplate and tail-flick tests. Hypoalgesia was observed in NET KO mice, and to a lesser extent in SERT KO mice. The magnitude of this hypoalgesia in NET KO mice was so profound that it limited the assessment of drug-induced analgesia. Nonetheless, the necessary exclusion of these subjects because of profound baseline hypoalgesia strongly supports the role of norepinephrine and NET in basal nociceptive behavior while indicating a much smaller role for serotonin and SERT. To further clarify the role of NET and SERT in basal nociceptive sensitivity further experiments were conducted in SERT KO and NET KO mice across a range of temperatures. NET KO mice were again found to have pronounced thermal hypoalgesia compared to WT mice in both the hotplate and tail-flick tests, while only limited effects were observed in SERT KO mice. Furthermore, in the acetic acid writhing test of visceral nociception pronounced hypoalgesia was again found in NET KO mice, but no change in SERT KO mice. As some of these effects may have resulted from developmental consequences of NET KO, the effects of the selective NET blocker nisoxetine and the selective SERT blocker fluoxetine were also examined in WT mice: only nisoxetine produced analgesia in these mice. Collectively these data suggest that NET has a far greater role in determining baseline analgesia, and perhaps other analgesic effects, than SERT in mice. © 2011.

Authors
Hall, FS; Schwarzbaum, JM; Perona, MTG; Templin, JS; Caron, MG; Lesch, K-P; Murphy, DL; Uhl, GR
MLA Citation
Hall, FS, Schwarzbaum, JM, Perona, MTG, Templin, JS, Caron, MG, Lesch, K-P, Murphy, DL, and Uhl, GR. "A greater role for the norepinephrine transporter than the serotonin transporter in murine nociception." Neuroscience 175 (2011): 315-327.
PMID
21129446
Source
scival
Published In
Neuroscience
Volume
175
Publish Date
2011
Start Page
315
End Page
327
DOI
10.1016/j.neuroscience.2010.11.057

An interaction between L-prostaglandin D synthase and arrestin increases PGD2 production

L-type prostaglandin synthase (L-PGDS) produces PGD2, a lipid mediator involved in neuromodulation and inflammation. Here, we show that L-PGDS and arrestin-3 (Arr3) interact directly and can be co-immunoprecipitated endogenously from MG-63 osteoblasts. Perinuclear L-PGDS/Arr3 co-localization is observed in PGD2-producing MG-63 cells and is induced by the addition of the L-PGDS substrate or co-expression of COX-2 in HEK293 cells. Inhibition of L-PGDS activity in MG-63 cells triggers redistribution of Arr3 and L-PGDS to the cytoplasm. Perinuclear localization of L-PGDS is detected in wild-type mouse embryonic fibroblasts (MEFs) but is more diffused in MEFs-arr-2 -/--arr-3-/-. Arrestin-3 promotes PGD2 production by L-PGDS in vitro. IL-1β-induced PGD2 production is significantly lower in MEFs-arr-2-/--arr-3-/- than in wild-type MEFs but can be rescued by expressing Arr2 or Arr3. A peptide corresponding to amino acids 86-100 of arrestin-3 derived from its L-PGDS binding domain stimulates L-PGDS-mediated PGD2 production in vitro and in MG-63 cells. We report the first characterization of an interactor/ modulator of a PGD2 synthase and the identification of a new function for arrestin, which may open new opportunities for improving therapies for the treatment of inflammatory diseases. © 2011 by The American Society for Biochemistry and Molecular Biology, Inc.

Authors
Mathurin, K; Gallant, MA; Germain, P; Allard-Chamard, H; Brisson, J; Iorio-Morin, C; Fernandes, ADB; Caron, MG; Laporte, SA; Parent, J-L
MLA Citation
Mathurin, K, Gallant, MA, Germain, P, Allard-Chamard, H, Brisson, J, Iorio-Morin, C, Fernandes, ADB, Caron, MG, Laporte, SA, and Parent, J-L. "An interaction between L-prostaglandin D synthase and arrestin increases PGD2 production." Journal of Biological Chemistry 286.4 (2011): 2696-2706.
PMID
21112970
Source
scival
Published In
The Journal of biological chemistry
Volume
286
Issue
4
Publish Date
2011
Start Page
2696
End Page
2706
DOI
10.1074/jbc.M110.178277

Novel strains of mice deficient for the vesicular acetylcholine transporter: Insights on transcriptional regulation and control of locomotor behavior

Defining the contribution of acetylcholine to specific behaviors has been challenging, mainly because of the difficulty in generating suitable animal models of cholinergic dysfunction. We have recently shown that, by targeting the vesicular acetylcholine transporter (VAChT) gene, it is possible to generate genetically modified mice with cholinergic deficiency. Here we describe novel VAChT mutant lines. VAChT gene is embedded within the first intron of the choline acetyltransferase (ChAT) gene, which provides a unique arrangement and regulation for these two genes. We generated a VAChT allele that is flanked by loxP sequences and carries the resistance cassette placed in a ChAT intronic region (FloxNeo allele). We show that mice with the FloxNeo allele exhibit differential VAChT expression in distinct neuronal populations. These mice show relatively intact VAChT expression in somatomotor cholinergic neurons, but pronounced decrease in other cholinergic neurons in the brain. VAChT mutant mice present preserved neuromuscular function, but altered brain cholinergic function and are hyperactive. Genetic removal of the resistance cassette rescues VAChT expression and the hyperactivity phenotype. These results suggest that release of ACh in the brain is normally required to "turn down" neuronal circuits controlling locomotion. © 2011 Martins-Silva et al.

Authors
Martins-Silva, C; Jaeger, XD; Guzman, MS; Lima, RDF; Santos, MS; Kushmerick, C; Gomez, MV; Caron, MG; Prado, MAM; Prado, VF
MLA Citation
Martins-Silva, C, Jaeger, XD, Guzman, MS, Lima, RDF, Santos, MS, Kushmerick, C, Gomez, MV, Caron, MG, Prado, MAM, and Prado, VF. "Novel strains of mice deficient for the vesicular acetylcholine transporter: Insights on transcriptional regulation and control of locomotor behavior." PLoS ONE 6.3 (2011).
PMID
21423695
Source
scival
Published In
PloS one
Volume
6
Issue
3
Publish Date
2011
DOI
10.1371/journal.pone.0017611

The dopamine metabolite 3-methoxytyramine is a neuromodulator.

Dopamine (3-hydroxytyramine) is a well-known catecholamine neurotransmitter involved in multiple physiological functions including movement control. Here we report that the major extracellular metabolite of dopamine, 3-methoxytyramine (3-MT), can induce behavioral effects in a dopamine-independent manner and these effects are partially mediated by the trace amine associated receptor 1 (TAAR1). Unbiased in vivo screening of putative trace amine receptor ligands for potential effects on the movement control revealed that 3-MT infused in the brain is able to induce a complex set of abnormal involuntary movements in mice acutely depleted of dopamine. In normal mice, the central administration of 3-MT caused a temporary mild hyperactivity with a concomitant set of abnormal movements. Furthermore, 3-MT induced significant ERK and CREB phosphorylation in the mouse striatum, signaling events generally related to PKA-mediated cAMP accumulation. In mice lacking TAAR1, both behavioral and signaling effects of 3-MT were partially attenuated, consistent with the ability of 3-MT to activate TAAR1 receptors and cause cAMP accumulation as well as ERK and CREB phosphorylation in cellular assays. Thus, 3-MT is not just an inactive metabolite of DA, but a novel neuromodulator that in certain situations may be involved in movement control. Further characterization of the physiological functions mediated by 3-MT may advance understanding of the pathophysiology and pharmacology of brain disorders involving abnormal dopaminergic transmission, such as Parkinson's disease, dyskinesia and schizophrenia.

Authors
Sotnikova, TD; Beaulieu, J-M; Espinoza, S; Masri, B; Zhang, X; Salahpour, A; Barak, LS; Caron, MG; Gainetdinov, RR
MLA Citation
Sotnikova, TD, Beaulieu, J-M, Espinoza, S, Masri, B, Zhang, X, Salahpour, A, Barak, LS, Caron, MG, and Gainetdinov, RR. "The dopamine metabolite 3-methoxytyramine is a neuromodulator. (Published online)" PLoS One 5.10 (October 18, 2010): e13452-.
Website
http://hdl.handle.net/10161/4579
PMID
20976142
Source
pubmed
Published In
PloS one
Volume
5
Issue
10
Publish Date
2010
Start Page
e13452
DOI
10.1371/journal.pone.0013452

Targeting of the orphan receptor GPR35 by pamoic acid: a potent activator of extracellular signal-regulated kinase and β-arrestin2 with antinociceptive activity.

Known agonists of the orphan receptor GPR35 are kynurenic acid, zaprinast, 5-nitro-2-(3-phenylproplyamino) benzoic acid, and lysophosphatidic acids. Their relatively low affinities for GPR35 and prominent off-target effects at other pathways, however, diminish their utility for understanding GPR35 signaling and for identifying potential therapeutic uses of GPR35. In a screen of the Prestwick Library of drugs and drug-like compounds, we have found that pamoic acid is a potent GPR35 agonist. Pamoic acid is considered by the Food and Drug Administration as an inactive compound that enables long-acting formulations of numerous drugs, such as the antihelminthics oxantel pamoate and pyrantel pamoate; the psychoactive compounds hydroxyzine pamoate (Vistaril) and imipramine pamoate (Tofranil-PM); and the peptide hormones triptorelin pamoate (Trelstar) and octreotide pamoate (OncoLar). We have found that pamoic acid induces a G(i/o)-linked, GPR35-mediated increase in the phosphorylation of extracellular signal-regulated kinase 1/2, recruitment of β-arrestin2 to GPR35, and internalization of GPR35. In mice, it attenuates visceral pain perception, indicating an antinociceptive effect, possibly through GPR35 receptors. We have also identified in collaboration with the Sanford-Burnham Institute Molecular Libraries Probe Production Center new classes of GPR35 antagonist compounds, including the nanomolar potency antagonist methyl-5-[(tert-butylcarbamothioylhydrazinylidene)methyl]-1-(2,4-difluorophenyl)pyrazole-4-carboxylate (CID2745687). Pamoic acid and potent antagonists such as CID2745687 present novel opportunities for expanding the chemical space of GPR35, elucidating GPR35 pharmacology, and stimulating GPR35-associated drug development. Our results indicate that the unexpected biological functions of pamoic acid may yield potential new uses for a common drug constituent.

Authors
Zhao, P; Sharir, H; Kapur, A; Cowan, A; Geller, EB; Adler, MW; Seltzman, HH; Reggio, PH; Heynen-Genel, S; Sauer, M; Chung, TDY; Bai, Y; Chen, W; Caron, MG; Barak, LS; Abood, ME
MLA Citation
Zhao, P, Sharir, H, Kapur, A, Cowan, A, Geller, EB, Adler, MW, Seltzman, HH, Reggio, PH, Heynen-Genel, S, Sauer, M, Chung, TDY, Bai, Y, Chen, W, Caron, MG, Barak, LS, and Abood, ME. "Targeting of the orphan receptor GPR35 by pamoic acid: a potent activator of extracellular signal-regulated kinase and β-arrestin2 with antinociceptive activity." Mol Pharmacol 78.4 (October 2010): 560-568.
PMID
20826425
Source
pubmed
Published In
Molecular pharmacology
Volume
78
Issue
4
Publish Date
2010
Start Page
560
End Page
568
DOI
10.1124/mol.110.066746

Tryptophan hydroxylase 2 genotype determines brain serotonin synthesis but not tissue content in C57Bl/6 and BALB/c congenic mice.

Tryptophan hydroxylase 2 (TPH2) catalyzes the rate-limiting step in the synthesis of brain serotonin (5-HT). In a previous report, a single nucleotide polymorphism in mTph2 (C1473G) reduced 5-HT synthesis by 55%. Mouse strains expressing the 1473C allele, such as C57Bl/6, have higher 5-HT synthesis rates than strains expressing the 1473G allele, such as BALB/c. Many studies have attributed strain differences to Tph2 genotype without ruling out the potential role of alterations in other genes. To test the role of the C1473G polymorphism in strain differences, we generated C57Bl/6 and BALB/c mice congenic for the Tph2 locus. We found that the 1473G allele reduced 5-HT synthesis in C57Bl/6 mice but had no effect on 5-HT tissue content except for a slight reduction (15%) in the frontal cortex. In BALB/c mice, the 1473C allele increased 5-HT synthesis but again did not affect 5-HT tissue content. At the same time, 5-hydroxyindoleacetic acid (5-HIAA) was significantly elevated in BALB/c congenic mice. In C57Bl/6 mice, there was no effect of genotype on 5-HIAA levels. BALB/c mice had lower expression of monoamine oxidase A and B than C57Bl/6 mice, but there was no effect of Tph2 genotype. On the tail suspension test, escitalopram treatment reduced immobility regardless of genotype. These data demonstrate that the C1473G polymorphism determines differences in 5-HT synthesis rates among strains but only minimally affects 5-HT tissue levels.

Authors
Siesser, WB; Zhang, X; Jacobsen, JPR; Sotnikova, TD; Gainetdinov, RR; Caron, MG
MLA Citation
Siesser, WB, Zhang, X, Jacobsen, JPR, Sotnikova, TD, Gainetdinov, RR, and Caron, MG. "Tryptophan hydroxylase 2 genotype determines brain serotonin synthesis but not tissue content in C57Bl/6 and BALB/c congenic mice." Neurosci Lett 481.1 (August 30, 2010): 6-11.
PMID
20600620
Source
pubmed
Published In
Neuroscience Letters
Volume
481
Issue
1
Publish Date
2010
Start Page
6
End Page
11
DOI
10.1016/j.neulet.2010.06.035

Noradrenergic control of cortico-striato-thalamic and mesolimbic cross-structural synchrony.

Although normal dopaminergic tone has been shown to be essential for the induction of cortico-striatal and mesolimbic theta oscillatory activity, the influence of norepinephrine on these brain networks remains relatively unknown. To address this question, we simultaneously recorded local field potentials and single-neuron activity across 10 interconnected brain areas (ventral striatum, frontal association cortex, hippocampus, primary motor cortex, orbital frontal cortex, prelimbic cortex, dorsal lateral striatum, medial dorsal nucleus of thalamus, substantia nigra pars reticularis, and ventral tegmental area) in a combined genetically and pharmacologically induced mouse model of hyponoradrenergia. Our results show that norepinephrine (NE) depletion induces a novel state in male mice characterized by a profound disruption of coherence across multiple cortico-striatal circuits and an increase in mesolimbic cross-structural coherence. Moreover, this brain state is accompanied by a complex behavioral phenotype consisting of transient hyperactivity, stereotypic behaviors, and an acute 12-fold increase in grooming. Notably, treatment with a norepinephrine precursors (l-3,4-dihydroxyphenylalanine at 100 mg/kg or l-threo-dihydroxyphenylserine at 5 mg/kg) or a selective serotonin reuptake inhibitor (fluoxetine at 20 mg/kg) attenuates the abnormal behaviors and selectively reverses the circuit changes observed in NE-depleted mice. Together, our results demonstrate that norepinephrine modulates the dynamic tuning of coherence across cortico-striato-thalamic circuits, and they suggest that changes in coherence across these circuits mediate the abnormal generation of hyperactivity and repetitive behaviors.

Authors
Dzirasa, K; Phillips, HW; Sotnikova, TD; Salahpour, A; Kumar, S; Gainetdinov, RR; Caron, MG; Nicolelis, MAL
MLA Citation
Dzirasa, K, Phillips, HW, Sotnikova, TD, Salahpour, A, Kumar, S, Gainetdinov, RR, Caron, MG, and Nicolelis, MAL. "Noradrenergic control of cortico-striato-thalamic and mesolimbic cross-structural synchrony." J Neurosci 30.18 (May 5, 2010): 6387-6397.
PMID
20445065
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
30
Issue
18
Publish Date
2010
Start Page
6387
End Page
6397
DOI
10.1523/JNEUROSCI.0764-10.2010

Role of Dopamine Transporters in Neuronal Homeostasis

© 2010 by Oxford University Press, Inc. All rights reserved. Dopamine (DA) neurotransmission is controlled by several critical processes. A complex homeostatic balance between the amount of DA synthesized, packaged into vesicles, released, reuptaken via plasma membrane transporter and metabolized, determines the overall status of dopaminergic signaling. The plasma membrane dopamine transporter (DAT) provides effective control of both the extracellular and intracellular concentrations of DA by recapturing released neurotransmitters in the presynaptic terminals. The vesicular monoamine transporter 2 (VMAT2) directly controls vesicular storage and release capacity by pumping monoamines from the cytoplasm of neurons into synaptic vesicles. These transporters are primary targets of many psychotropic drugs that potently affect synaptic DA and related physiological processes. This chapter summarizes recent advances in the understanding of the molecular and cellular mechanisms involved in the DAT and VMAT2 functions. It discusses the role of these transporters in the action of psychostimulant drugs and neurotoxins, as revealed in studies using mutant mice.

Authors
Caron, MG; Gainetdinov, RR
MLA Citation
Caron, MG, and Gainetdinov, RR. "Role of Dopamine Transporters in Neuronal Homeostasis." Dopamine Handbook. February 1, 2010.
Source
scopus
Publish Date
2010
DOI
10.1093/acprof:oso/9780195373035.003.0007

Dependence of serotonergic and other nonadrenergic enteric neurons on norepinephrine transporter expression

The norepinephrine transporter (NET), which is expressed on the plasma membranes of noradrenergic neurons, is important in terminating neurotransmission. The noradrenergic sympathetic neurons that innervate the bowel express NET, but they are extrinsic and their cell bodies are not components of the enteric nervous system (ENS). Subsets of neurons were nevertheless found in the murine ENS that express transcripts encoding NET, NET protein, and dopamine β-hydroxylase; these neurons lack tyrosine hydroxylase (TH) and thus are not catecholaminergic. Enteric NET expression, moreover, preceded the in growth of sympathetic axons during development and did not disappear when the gut was extrinsically denervated. Transiently catecholaminegic (TC), neural crest-derived precursors of enteric neurons expressed NET at embryonic day 10 (E10) and NET expression in the fetal gut peaked coincidentally with early neurogenesis at E12. Serotonergic neurons, which are born early from TC progenitors, were found to express NET in the adult ENS, as did also other early-born neurons containing calretinin or neuronal nitric oxide synthase (nNOS) immunoreactivities. NET was not expressed in TH-immunoreactive dopaminergic neurons, which are born perinatally. Genetic deletion of NET almost eliminated tryptophan hydroxylase 2 expression and significantly reduced the numbers of total, 5-HT- and calretinin-immunoreactive enteric neurons, without affecting the immunoreactivities of nNOS or TH. These observations indicate that TC precursors of subsets of noncatecholaminergic enteric neurons express NET that persists in the successors of these cells despite their loss of TH. NET expression is essential for development and/or survival of some (5-HT- and calretinin-expressing), but not all (nNOS-expressing), of these neurons. Copyright © 2010 the authors.

Authors
Li, Z; Caron, MG; Blakely, RD; Margolis, KG; Gershon, MD
MLA Citation
Li, Z, Caron, MG, Blakely, RD, Margolis, KG, and Gershon, MD. "Dependence of serotonergic and other nonadrenergic enteric neurons on norepinephrine transporter expression." Journal of Neuroscience 30.49 (2010): 16730-16740.
PMID
21148012
Source
scival
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
30
Issue
49
Publish Date
2010
Start Page
16730
End Page
16740
DOI
10.1523/JNEUROSCI.2276-10.2010

International workshop at the nobel forum, karolinska institutet on g protein-coupled receptors: Finding the words to describe monomers, oligomers, and their molecular mechanisms and defining their meaning. Can a consensus be reached

A meeting was held May 19, 2010 at the Karolinski Institute on Nomenclature in Pharmacology. This meeting occurred in conjunction with the Symposium The Changing World of G Protein Coupled Receptors: From Monomers to Dimers and Receptor Mosaics (Higher-order Oligomers) held the previous day at the Royal Swedish Academy of Science. Two broad topics of nomenclature were discussed; ligand nomenclature and the definition of 'receptorreceptor' interactions. This paper summarizes discussions on these topics along with a consensus definition of the term 'receptorreceptor' interaction. © 2010 Informa Healthcare USA, Inc.

Authors
Kenakin, T; Agnati, LF; Caron, M; Fredholm, B; Guidoli, D; Kobilka, B; Lefkowitz, RW; Lohse, M; Woods, A; Fuxe, K
MLA Citation
Kenakin, T, Agnati, LF, Caron, M, Fredholm, B, Guidoli, D, Kobilka, B, Lefkowitz, RW, Lohse, M, Woods, A, and Fuxe, K. "International workshop at the nobel forum, karolinska institutet on g protein-coupled receptors: Finding the words to describe monomers, oligomers, and their molecular mechanisms and defining their meaning. Can a consensus be reached." Journal of Receptors and Signal Transduction 30.5 (2010): 284-286.
PMID
20858022
Source
scival
Published In
Journal of Receptors and Signal Transduction (Informa)
Volume
30
Issue
5
Publish Date
2010
Start Page
284
End Page
286
DOI
10.3109/10799893.2010.512438

Dysregulation of dopamine transporters via dopamine D2 autoreceptors triggers anomalous dopamine efflux associated with attention-deficit hyperactivity disorder

The neurotransmitter dopamine (DA) modulates brain circuits involved in attention, reward, and motor activity. Synaptic DA homeostasis is primarily controlled via two presynaptic regulatory mechanisms, DA D2 receptor (D2R)-mediated inhibition of DA synthesis and release, and DA transporter (DAT)-mediated DA clearance. D2Rs can physically associate with DAT and regulate DAT function, linking DA release and reuptake to a common mechanism. We have established that the attention-deficit hyperactivity disorder-associated human DAT coding variant Ala559Val (hDAT A559V) results in anomalous DA efflux (ADE) similar to that caused by amphetamine-like psychostimulants. Here, we show that tonic activation of D 2R provides support for hDAT A559V-mediated ADE. We determine in hDAT A559V a pertussis toxin-sensitive, CaMKII-dependent phosphorylation mechanism that supports D2R-driven DA efflux. These studies identify a signaling network downstream of D2R activation, normally constraining DA action at synapses, that may be altered by DAT mutation to impact risk for DA-related disorders. Copyright © 2010 the authors.

Authors
Bowton, E; Saunders, C; Erreger, K; Sakrikar, D; Matthies, HJ; Sen, N; Jessen, T; Colbran, RJ; Caron, MG; Javitch, JA; Blakely, RD; Galli, A
MLA Citation
Bowton, E, Saunders, C, Erreger, K, Sakrikar, D, Matthies, HJ, Sen, N, Jessen, T, Colbran, RJ, Caron, MG, Javitch, JA, Blakely, RD, and Galli, A. "Dysregulation of dopamine transporters via dopamine D2 autoreceptors triggers anomalous dopamine efflux associated with attention-deficit hyperactivity disorder." Journal of Neuroscience 30.17 (2010): 6048-6057.
PMID
20427663
Source
scival
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
30
Issue
17
Publish Date
2010
Start Page
6048
End Page
6057
DOI
10.1523/JNEUROSCI.5094-09.2010

Dysautonomia due to reduced cholinergic neurotransmission causes cardiac remodeling and heart failure

Overwhelming evidence supports the importance of the sympathetic nervous system in heart failure. In contrast, much less is known about the role of failing cholinergic neurotransmission in cardiac disease. By using a unique genetically modified mouse line with reduced expression of the vesicular acetylcholine transporter (VAChT) and consequently decreased release of acetylcholine, we investigated the consequences of altered cholinergic tone for cardiac function. M-mode echocardiography, hemodynamic experiments, analysis of isolated perfused hearts, and measurements of cardiomyocyte contraction indicated that VAChT mutant mice have decreased left ventricle function associated with altered calcium handling. Gene expression was analyzed by quantitative reverse transcriptase PCR and Western blotting, and the results indicated that VAChT mutant mice have profound cardiac remodeling and reactivation of the fetal gene program. This phenotype was attributable to reduced cholinergic tone, since administration of the cholinesterase inhibitor pyridostigmine for 2 weeks reversed the cardiac phenotype in mutant mice. Our findings provide direct evidence that decreased cholinergic neurotransmission and underlying autonomic imbalance cause plastic alterations that contribute to heart dysfunction. Copyright © 2010, American Society for Microbiology. All Rights Reserved.

Authors
Lara, A; Damasceno, DD; Pires, R; Gros, R; Gomes, ER; Gavioli, M; Lima, RF; Guimarães, D; Lima, P; Jr, CRB; Vasconcelos, A; Roman-Campos, D; Menezes, CAS; Sirvente, RA; Salemi, VM; Mady, C; Caron, MG; Ferreira, AJ; Brum, PC; Resende, RR; Cruz, JS; Gomez, MV; Prado, VF; Almeida, APD; Prado, MAM; Guatimosim, S
MLA Citation
Lara, A, Damasceno, DD, Pires, R, Gros, R, Gomes, ER, Gavioli, M, Lima, RF, Guimarães, D, Lima, P, Jr, CRB, Vasconcelos, A, Roman-Campos, D, Menezes, CAS, Sirvente, RA, Salemi, VM, Mady, C, Caron, MG, Ferreira, AJ, Brum, PC, Resende, RR, Cruz, JS, Gomez, MV, Prado, VF, Almeida, APD, Prado, MAM, and Guatimosim, S. "Dysautonomia due to reduced cholinergic neurotransmission causes cardiac remodeling and heart failure." Molecular and Cellular Biology 30.7 (2010): 1746-1756.
PMID
20123977
Source
scival
Published In
Molecular and Cellular Biology
Volume
30
Issue
7
Publish Date
2010
Start Page
1746
End Page
1756
DOI
10.1128/MCB.00996-09

G Protein-coupled receptor kinases phosphorylate LRP6 in the Wnt pathway.

Wnt ligands conduct their functions in canonical Wnt signaling by binding to two receptors, the single transmembrane low density lipoprotein receptor-related proteins 5 and 6 (LRP5/6) and seven transmembrane (7TM) Frizzled receptors. Subsequently, phosphorylation of serine/threonine residues within five repeating signature PPPSP motifs on LRP6 is responsible for LRP6 activation. GSK3beta, a cytosolic kinase for phosphorylation of a downstream effector beta-catenin, was proposed to participate in such LRP6 phosphorylation. Here, we report a new class of membrane-associated kinases for LRP6 phosphorylation. We found that G protein-coupled receptor kinases 5 and 6 (GRK5/6), traditionally known to phosphorylate and desensitize 7TM G protein-coupled receptors, directly phosphorylate the PPPSP motifs on single transmembrane LRP6 and regulate Wnt/LRP6 signaling. GRK5/6-induced LRP6 activation is inhibited by the LRP6 antagonist Dickkopf. Depletion of GRK5 markedly reduces Wnt3A-stimulated LRP6 phosphorylation in cells. In zebrafish, functional knock-down of GRK5 results in reduced Wnt signaling, analogous to LRP6 knock-down, as assessed by decreased abundance of beta-catenin and lowered expression of the Wnt target genes cdx4, vent, and axin2. Expression of GRK5 rescues the diminished beta-catenin and axin2 response caused by GRK5 depletion. Thus, our findings identify GRK5/6 as novel kinases for the single transmembrane receptor LRP6 during Wnt signaling.

Authors
Chen, M; Philipp, M; Wang, J; Premont, RT; Garrison, TR; Caron, MG; Lefkowitz, RJ; Chen, W
MLA Citation
Chen, M, Philipp, M, Wang, J, Premont, RT, Garrison, TR, Caron, MG, Lefkowitz, RJ, and Chen, W. "G Protein-coupled receptor kinases phosphorylate LRP6 in the Wnt pathway." J Biol Chem 284.50 (December 11, 2009): 35040-35048.
PMID
19801552
Source
pubmed
Published In
The Journal of biological chemistry
Volume
284
Issue
50
Publish Date
2009
Start Page
35040
End Page
35048
DOI
10.1074/jbc.M109.047456

Atypical responsiveness of the orphan receptor GPR55 to cannabinoid ligands.

The cannabinoid receptor 1 (CB(1)) and CB(2) cannabinoid receptors, associated with drugs of abuse, may provide a means to treat pain, mood, and addiction disorders affecting widespread segments of society. Whether the orphan G-protein coupled receptor GPR55 is also a cannabinoid receptor remains unclear as a result of conflicting pharmacological studies. GPR55 has been reported to be activated by exogenous and endogenous cannabinoid compounds but surprisingly also by the endogenous non-cannabinoid mediator lysophosphatidylinositol (LPI). We examined the effects of a representative panel of cannabinoid ligands and LPI on GPR55 using a beta-arrestin-green fluorescent protein biosensor as a direct readout of agonist-mediated receptor activation. Our data demonstrate that AM251 and SR141716A (rimonabant), which are cannabinoid antagonists, and the lipid LPI, which is not a cannabinoid receptor ligand, are GPR55 agonists. They possess comparable efficacy in inducing beta-arrestin trafficking and, moreover, activate the G-protein-dependent signaling of protein kinase CbetaII. Conversely, the potent synthetic cannabinoid agonist CP55,940 acts as a GPR55 antagonist/partial agonist. CP55,940 blocks GPR55 internalization, the formation of beta-arrestin GPR55 complexes, and the phosphorylation of ERK1/2; CP55,940 produces only a slight amount of protein kinase CbetaII membrane recruitment but does not stimulate membrane remodeling like LPI, AM251, or rimonabant. Our studies provide a paradigm for measuring the responsiveness of GPR55 to a variety of ligand scaffolds comprising cannabinoid and novel compounds and suggest that at best GPR55 is an atypical cannabinoid responder. The activation of GPR55 by rimonabant may be responsible for some of the off-target effects that led to its removal as a potential obesity therapy.

Authors
Kapur, A; Zhao, P; Sharir, H; Bai, Y; Caron, MG; Barak, LS; Abood, ME
MLA Citation
Kapur, A, Zhao, P, Sharir, H, Bai, Y, Caron, MG, Barak, LS, and Abood, ME. "Atypical responsiveness of the orphan receptor GPR55 to cannabinoid ligands." J Biol Chem 284.43 (October 23, 2009): 29817-29827.
PMID
19723626
Source
pubmed
Published In
The Journal of biological chemistry
Volume
284
Issue
43
Publish Date
2009
Start Page
29817
End Page
29827
DOI
10.1074/jbc.M109.050187

Trace amine-associated receptors as emerging therapeutic targets.

Endogenous trace amines (TAs) of unknown biological function are structurally related to classic monoaminergic neurotransmitters and found at low concentrations in the mammalian brain. Their recently discovered group of G protein-coupled receptors, trace amine-associated receptors (TAARs), may represent putative targets not only for trace and other amines but also for a variety of monoaminergic compounds, including amphetamines and monoamine metabolites. The trace amine-associated receptor 1 (TAAR1), which is in part associated with the monoaminergic neuronal circuitry controlling various functions, including movement, is the best characterized of the class, although little is known about its regulation and function. Here we review the pharmacology and biochemical properties of the TAAR1 and its physiological functions as revealed in studies involving knockout mice lacking this receptor. Potential therapeutic applications of future selective TAAR1 agonists and antagonists are also discussed. Although understanding of biology and functions mediated by other TAARs is still in its infancy, it is expected that further characterization of the functional roles and biochemical properties of TAARs and identification of endogenous and exogenous ligands will eventually promote these receptors as an attractive class of targets to correct monoaminergic processes that could be dysfunctional in a host of disorders of brain and periphery.

Authors
Sotnikova, TD; Caron, MG; Gainetdinov, RR
MLA Citation
Sotnikova, TD, Caron, MG, and Gainetdinov, RR. "Trace amine-associated receptors as emerging therapeutic targets." Mol Pharmacol 76.2 (August 2009): 229-235. (Review)
PMID
19389919
Source
pubmed
Published In
Molecular pharmacology
Volume
76
Issue
2
Publish Date
2009
Start Page
229
End Page
235
DOI
10.1124/mol.109.055970

Hyperdopaminergia and NMDA receptor hypofunction disrupt neural phase signaling.

Neural phase signaling has gained attention as a putative coding mechanism through which the brain binds the activity of neurons across distributed brain areas to generate thoughts, percepts, and behaviors. Neural phase signaling has been shown to play a role in various cognitive processes, and it has been suggested that altered phase signaling may play a role in mediating the cognitive deficits observed across neuropsychiatric illness. Here, we investigated neural phase signaling in two mouse models of cognitive dysfunction: mice with genetically induced hyperdopaminergia [dopamine transporter knock-out (DAT-KO) mice] and mice with genetically induced NMDA receptor hypofunction [NMDA receptor subunit-1 knockdown (NR1-KD) mice]. Cognitive function in these mice was assessed using a radial-arm maze task, and local field potentials were recorded from dorsal hippocampus and prefrontal cortex as DAT-KO mice, NR1-KD mice, and their littermate controls engaged in behavioral exploration. Our results demonstrate that both DAT-KO and NR1-KD mice display deficits in spatial cognitive performance. Moreover, we show that persistent hyperdopaminergia alters interstructural phase signaling, whereas NMDA receptor hypofunction alters interstructural and intrastructural phase signaling. These results demonstrate that dopamine and NMDA receptor dependent glutamate signaling play a critical role in coordinating neural phase signaling, and encourage further studies to investigate the role that deficits in phase signaling play in mediating cognitive dysfunction.

Authors
Dzirasa, K; Ramsey, AJ; Takahashi, DY; Stapleton, J; Potes, JM; Williams, JK; Gainetdinov, RR; Sameshima, K; Caron, MG; Nicolelis, MAL
MLA Citation
Dzirasa, K, Ramsey, AJ, Takahashi, DY, Stapleton, J, Potes, JM, Williams, JK, Gainetdinov, RR, Sameshima, K, Caron, MG, and Nicolelis, MAL. "Hyperdopaminergia and NMDA receptor hypofunction disrupt neural phase signaling." J Neurosci 29.25 (June 24, 2009): 8215-8224.
PMID
19553461
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
29
Issue
25
Publish Date
2009
Start Page
8215
End Page
8224
DOI
10.1523/JNEUROSCI.1773-09.2009

"To learn, you must pay attention." Molecular insights into teachers' wisdom.

Authors
Caron, MG; Wightman, RM
MLA Citation
Caron, MG, and Wightman, RM. ""To learn, you must pay attention." Molecular insights into teachers' wisdom." Proc Natl Acad Sci U S A 106.18 (May 5, 2009): 7267-7268.
PMID
19416916
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
106
Issue
18
Publish Date
2009
Start Page
7267
End Page
7268
DOI
10.1073/pnas.0903306106

Spinal cord stimulation restores locomotion in animal models of Parkinson's disease.

Dopamine replacement therapy is useful for treating motor symptoms in the early phase of Parkinson's disease, but it is less effective in the long term. Electrical deep-brain stimulation is a valuable complement to pharmacological treatment but involves a highly invasive surgical procedure. We found that epidural electrical stimulation of the dorsal columns in the spinal cord restores locomotion in both acute pharmacologically induced dopamine-depleted mice and in chronic 6-hydroxydopamine-lesioned rats. The functional recovery was paralleled by a disruption of aberrant low-frequency synchronous corticostriatal oscillations, leading to the emergence of neuronal activity patterns that resemble the state normally preceding spontaneous initiation of locomotion. We propose that dorsal column stimulation might become an efficient and less invasive alternative for treatment of Parkinson's disease in the future.

Authors
Fuentes, R; Petersson, P; Siesser, WB; Caron, MG; Nicolelis, MAL
MLA Citation
Fuentes, R, Petersson, P, Siesser, WB, Caron, MG, and Nicolelis, MAL. "Spinal cord stimulation restores locomotion in animal models of Parkinson's disease." Science 323.5921 (March 20, 2009): 1578-1582.
PMID
19299613
Source
pubmed
Published In
Science
Volume
323
Issue
5921
Publish Date
2009
Start Page
1578
End Page
1582
DOI
10.1126/science.1164901

Hedgehog signaling: is Smo a G protein-coupled receptor?

The Hedgehog signal transducer Smoothened is structurally similar to G protein-coupled receptors. Now there is direct evidence that Smoothened relies on heterotrimeric G proteins in order to transduce the Hedgehog signal.

Authors
Philipp, M; Caron, MG
MLA Citation
Philipp, M, and Caron, MG. "Hedgehog signaling: is Smo a G protein-coupled receptor?." Curr Biol 19.3 (February 10, 2009): R125-R127. (Review)
PMID
19211052
Source
pubmed
Published In
Current Biology
Volume
19
Issue
3
Publish Date
2009
Start Page
R125
End Page
R127
DOI
10.1016/j.cub.2008.12.010

Reduced expression of the vesicular acetylcholine transporter causes learning deficits in mice.

Storage of acetylcholine in synaptic vesicles plays a key role in maintaining cholinergic function. Here we used mice with a targeted mutation in the vesicular acetylcholine transporter (VAChT) gene that reduces transporter expression by 40% to investigate cognitive processing under conditions of VAChT deficiency. Motor skill learning in the rotarod revealed that VAChT mutant mice were slower to learn this task, but once they reached maximum performance they were indistinguishable from wild-type mice. Interestingly, motor skill performance maintenance after 10 days was unaffected in these mutant mice. We also tested whether reduced VAChT levels affected learning in an object recognition memory task. We found that VAChT mutant mice presented a deficit in memory encoding necessary for the temporal order version of the object recognition memory, but showed no alteration in spatial working memory, or spatial memory in general when tested in the Morris water maze test. The memory deficit in object recognition memory observed in VAChT mutant mice could be reversed by cholinesterase inhibitors, suggesting that learning deficits caused by reduced VAChT expression can be ameliorated by restoring ACh levels in the synapse. These data indicate an important role for cholinergic tone in motor learning and object recognition memory.

Authors
de Castro, BM; Pereira, GS; Magalhães, V; Rossato, JI; De Jaeger, X; Martins-Silva, C; Leles, B; Lima, P; Gomez, MV; Gainetdinov, RR; Caron, MG; Izquierdo, I; Cammarota, M; Prado, VF; Prado, MAM
MLA Citation
de Castro, BM, Pereira, GS, Magalhães, V, Rossato, JI, De Jaeger, X, Martins-Silva, C, Leles, B, Lima, P, Gomez, MV, Gainetdinov, RR, Caron, MG, Izquierdo, I, Cammarota, M, Prado, VF, and Prado, MAM. "Reduced expression of the vesicular acetylcholine transporter causes learning deficits in mice." Genes Brain Behav 8.1 (February 2009): 23-35.
PMID
18778400
Source
pubmed
Published In
Genes, Brain and Behavior
Volume
8
Issue
1
Publish Date
2009
Start Page
23
End Page
35
DOI
10.1111/j.1601-183X.2008.00439.x

Reduced D2-mediated signaling activity and trans-synaptic upregulation of D1 and D2 dopamine receptors in mice overexpressing the dopamine transporter.

The dopamine transporter (DAT) regulates the temporal and spatial actions of dopamine by reuptaking this neurotransmitter into the presynaptic neurons. We recently generated transgenic mice overexpressing DAT (DAT-tg) that have a 3-fold increase in DAT protein levels which results in a 40% reduction of the extracellular DA concentration in the striatum. The aim of this study was to examine the effect of this reduction in dopaminergic tone on postsynaptic responses mediated by dopamine receptors. We report here that DAT-tg mice have increased levels of striatal D1 (30%) and D2 (approximately 60%) dopamine receptors with D1 receptor signaling components not significantly altered, as evidenced by unaffected basal or stimulated levels of phospho-GluR1 (Ser845) and phospho-ERK2. However, the novel D2 mediated Akt signaling is markedly altered in DAT-tg animals. In particular, there is a 300% increase in the basal levels of phospho-Akt in the striatum of DAT-tg, reflecting the reduced extracellular dopamine tone in these animals. This increase in basal pAkt levels can be pharmacologically recapitulated by partial dopamine depletion in WT mice treated with the selective tyrosine hydroxylase inhibitor alpha-methyl-para-tyrosine (alpha-MPT). Behaviorally, DAT-tg animals demonstrate an augmented synergistic interaction between up-regulated D1 and D2 receptors, which results in increased climbing behavior in transgenic mice after stimulation with either apomorphine or a co-administration of selective D1 and D2 receptor agonists. In sum, our study reveals that hypodopaminegia caused by up-regulation of DAT results in significant alterations at postsynaptic receptor function with most notable dysregulation at the level of D2 receptor signaling.

Authors
Ghisi, V; Ramsey, AJ; Masri, B; Gainetdinov, RR; Caron, MG; Salahpour, A
MLA Citation
Ghisi, V, Ramsey, AJ, Masri, B, Gainetdinov, RR, Caron, MG, and Salahpour, A. "Reduced D2-mediated signaling activity and trans-synaptic upregulation of D1 and D2 dopamine receptors in mice overexpressing the dopamine transporter." Cell Signal 21.1 (January 2009): 87-94.
PMID
18929645
Source
pubmed
Published In
Cellular Signalling
Volume
21
Issue
1
Publish Date
2009
Start Page
87
End Page
94
DOI
10.1016/j.cellsig.2008.09.011

Knockout of the norepinephrine transporter and pharmacologically diverse antidepressants prevent behavioral and brain neurotrophin alterations in two chronic stress models of depression

Diverse factors such as changes in neurotrophins and brain plasticity have been proposed to be involved in the actions of antidepressant drugs (ADs). However, in mouse models of depression based on chronic stress, it is still unclear whether simultaneous changes in behavior and neurotrophin expression occur and whether these changes can be corrected or prevented comparably by chronic administration of ADs or genetic manipulations that produce antidepressant-like effects such as the knockout of the norepinephrine transporter (NET) gene. Here we show that chronic restraint or social defeat stress induce comparable effects on behavior and changes in the expression of neurotrophins in depression-related brain regions. Chronic stress caused down-regulation of BDNF, nerve growth factor, and neurotrophin-3 in hippocampus and cerebral cortex and up-regulation of these targets in striatal regions. In wild-type mice, these effects could be prevented by concomitant chronic administration of five pharmacologically diverse ADs. In contrast, NET knock out (NETKO) mice were resistant to stress-induced depressive-like changes in behavior and brain neurotrophin expression. Thus, the resistance of the NETKO mice to the stress-induced depression-associated behaviors and biochemical changes highlight the importance of noradrenergic pathways in the maintenance of mood. In addition, these mice represent a useful model to study depression-resistant behaviors, and they might help to provide deeper insights into the identification of downstream targets involved in the mechanisms of antidepressants. © 2009 International Society for Neurochemistry.

Authors
Haenisch, B; Bilkei-Gorzo, A; Caron, MG; Bönisch, H
MLA Citation
Haenisch, B, Bilkei-Gorzo, A, Caron, MG, and Bönisch, H. "Knockout of the norepinephrine transporter and pharmacologically diverse antidepressants prevent behavioral and brain neurotrophin alterations in two chronic stress models of depression." Journal of Neurochemistry 111.2 (2009): 403-416.
PMID
19694905
Source
scival
Published In
Journal of Neurochemistry
Volume
111
Issue
2
Publish Date
2009
Start Page
403
End Page
416
DOI
10.1111/j.1471-4159.2009.06345.x

The vesicular acetylcholine transporter is required for neuromuscular development and function

The vesicular acetylcholine (ACh) transporter (VAChT) mediates ACh storage by synaptic vesicles. However, the VAChT-independent release of ACh is believed to be important during development. Here we generated VAChT knockout mice and tested the physiological relevance of the VAChT-independent release of ACh. Homozygous VAChT knockout mice died shortly after birth, indicating that VAChT-mediated storage of ACh is essential for life. Indeed, synaptosomes obtained from brains of homozygous knockouts were incapable of releasing ACh in response to depolarization. Surprisingly, electrophysiological recordings at the skeletal-neuromuscular junction show that VAChT knockout mice present spontaneous miniature end-plate potentials with reduced amplitude and frequency, which are likely the result of a passive transport of ACh into synaptic vesicles. Interestingly, VAChT knockouts exhibit substantial increases in amounts of choline acetyltransferase, high-affinity choline transporter, and ACh. However, the development of the neuromuscular junction in these mice is severely affected. Mutant VAChT mice show increases in motoneuron and nerve terminal numbers. End plates are large, nerves exhibit abnormal sprouting, and muscle is necrotic. The abnormalities are similar to those of mice that cannot synthesize ACh due to a lack of choline acetyltransferase. Our results indicate that VAChT is essential to the normal development of motor neurons and the release of ACh. Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Authors
Castro, BMD; Jaeger, XD; Martins-Silva, C; Lima, RDF; Amaral, E; Menezes, C; Lima, P; Neves, CML; Pires, RG; Gould, TW; Welch, I; Kushmerick, C; Guatimosim, C; Izquierdo, I; Cammarota, M; Rylett, RJ; Gomez, MV; Caron, MG; Oppenheim, RW; Prado, MAM; Prado, VF
MLA Citation
Castro, BMD, Jaeger, XD, Martins-Silva, C, Lima, RDF, Amaral, E, Menezes, C, Lima, P, Neves, CML, Pires, RG, Gould, TW, Welch, I, Kushmerick, C, Guatimosim, C, Izquierdo, I, Cammarota, M, Rylett, RJ, Gomez, MV, Caron, MG, Oppenheim, RW, Prado, MAM, and Prado, VF. "The vesicular acetylcholine transporter is required for neuromuscular development and function." Molecular and Cellular Biology 29.19 (2009): 5238-5250.
PMID
19635813
Source
scival
Published In
Molecular and Cellular Biology
Volume
29
Issue
19
Publish Date
2009
Start Page
5238
End Page
5250
DOI
10.1128/MCB.00245-09

Cocaine-conditioned locomotion in dopamine transporter, norepinephrine transporter and 5-HT transporter knockout mice

The behavioral effects of cocaine are affected by gene knockout (KO) of the dopamine transporter (DAT), the serotonin transporter (SERT) and the norepinephrine transporter (NET). The relative involvement of each of these transporters varies depending on the particular behavioral response to cocaine considered, as well as on other factors such as genetic background of the subjects. Interestingly, the effects of these gene knockouts on cocaine-induced locomotion are quite different from those on reward assessed in the conditioned place preference paradigm. To further explore the role of these genes in the rewarding effects of cocaine, the ability of five daily injections of cocaine to induce conditioned locomotion was assessed in DAT, SERT and NET KO mice. Cocaine increased locomotor activity acutely during the initial conditioning session in SERT KO and NET KO, but not DAT KO, mice. Surprisingly, locomotor responses in the cocaine-paired subjects diminished over the five conditioning sessions in SERT KO mice, while locomotor responses increased in DAT KO mice, despite the fact that they did not demonstrate any initial locomotor responses to cocaine. Cocaine-induced locomotion was unchanged over the course of conditioning in NET KO mice. In the post-conditioning assessment, conditioned locomotion was not observed in DAT KO mice, and was reduced in SERT KO and NET KO mice. These data reaffirm the central role of dopamine and DAT in the behavioral effects of cocaine. Furthermore, they emphasize the polygenic basis of cocaine-mediated behavior and the non-unitary nature of drug reward mechanisms, particularly in the context of previous studies that have shown normal cocaine-conditioned place preference in DAT KO mice. © 2009.

Authors
Hall, FS; Li, X-F; Randall-Thompson, J; Sora, I; Murphy, DL; Lesch, K-P; Caron, M; Uhl, GR
MLA Citation
Hall, FS, Li, X-F, Randall-Thompson, J, Sora, I, Murphy, DL, Lesch, K-P, Caron, M, and Uhl, GR. "Cocaine-conditioned locomotion in dopamine transporter, norepinephrine transporter and 5-HT transporter knockout mice." Neuroscience 162.4 (2009): 870-880.
PMID
19482066
Source
scival
Published In
Neuroscience
Volume
162
Issue
4
Publish Date
2009
Start Page
870
End Page
880
DOI
10.1016/j.neuroscience.2009.05.058

Orphan transporter SLC6A18 is renal neutral amino acid transporter B0AT3

The orphan transporter Slc6a18 (XT2) is highly expressed at the luminal membrane of kidney proximal tubules and displays ∼50% identity with Slc6a19 (B0AT1), which is the main neutral amino acid transporter in both kidney and small intestine. As yet, the amino acid transport function of XT2 has only been experimentally supported by the urinary glycine loss observed in xt2 null mice. We report here that in Xenopus laevis oocytes, co-expressed ACE2 (angiotensin-converting enzyme 2) associates with XT2 and reveals its function as a Na+ - and Cl--dependent neutral amino acid transporter. In contrast to its association with ACE2 observed in Xenopus laevis oocytes, our experiments with ace2 and collectrin null mice demonstrate that in vivo it is Collectrin, a smaller homologue of ACE2, that is required for functional expression of XT2 in kidney. To assess the function of XT2 in vivo, we reanalyzed its knock-out mouse model after more than 10 generations of backcrossing into C57BL/6 background. In addition to the previously published glycinuria, we observed a urinary loss of several other amino acids, in particular β-branched and small neutral ones. Using telemetry, we confirmed the previously described link of XT2 absence with hypertension but only in physically restrained animals. Taken together, our data indicate that the formerly orphan transporter XT2 functions as a sodium and chloride-dependent neutral amino acid transporter that we propose to rename B0AT3. © 2009 by The American Society for Biochemistry and Molecular Biology, Inc.

Authors
Singer, D; Camargo, SMR; Huggel, K; Romeo, E; Danilczyk, U; Kuba, K; Chesnov, S; Caron, MG; Penninger, JM; Verrey, F
MLA Citation
Singer, D, Camargo, SMR, Huggel, K, Romeo, E, Danilczyk, U, Kuba, K, Chesnov, S, Caron, MG, Penninger, JM, and Verrey, F. "Orphan transporter SLC6A18 is renal neutral amino acid transporter B0AT3." Journal of Biological Chemistry 284.30 (2009): 19953-19960.
PMID
19478081
Source
scival
Published In
The Journal of biological chemistry
Volume
284
Issue
30
Publish Date
2009
Start Page
19953
End Page
19960
DOI
10.1074/jbc.M109.011171

PSD-95 is essential for hallucinogen and atypical antipsychotic drug actions at serotonin receptors

Here, we report that postsynaptic density protein of 95 kDa (PSD-95), a postsynaptic density scaffolding protein, classically conceptualized as being essential for the regulation of ionotropic glutamatergic signaling at the postsynaptic membrane, plays an unanticipated and essential role in mediating the actions of hallucinogens and atypical antipsychotic drugs at 5-HT 2A and 5-HT2C serotonergic G-protein-coupled receptors. We show that PSD-95 is crucial for normal 5-HT2A and 5-HT2C expression in vivo and that PSD-95 maintains normal receptor expression by promoting apical dendritic targeting and stabilizing receptor turnover in vivo. Significantly, 5-HT2A-and 5-HT2Cmediated downstream signaling is impaired in PSD-95null mice, and the 5-HT 2A-mediated head-twitch response is abnormal. Furthermore, the ability of 5-HT2A inverse agonists to normalize behavioral changes induced by glutamate receptor antagonists is abolished in the absence of PSD-95 in vivo. These results demonstrate that PSD-95, in addition to the well known role it plays in scaffolding macromolecular glutamatergic signaling complexes, profoundly modulates metabotropic 5-HT2A and 5-HT2C receptor function. Copyright © 2009 Society for Neuroscience.

Authors
Abbas, AI; Yadav, PN; Yao, W-D; Arbuckle, MI; Grant, SGN; Caron, MG; Roth, BL
MLA Citation
Abbas, AI, Yadav, PN, Yao, W-D, Arbuckle, MI, Grant, SGN, Caron, MG, and Roth, BL. "PSD-95 is essential for hallucinogen and atypical antipsychotic drug actions at serotonin receptors." Journal of Neuroscience 29.22 (2009): 7124-7136.
PMID
19494135
Source
scival
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
29
Issue
22
Publish Date
2009
Start Page
7124
End Page
7136
DOI
10.1523/JNEUROSCI.1090-09.2009

Norepinephrine transport-mediated gene expression in noradrenergic neurogenesis

Background: We have identified a differential gene expression profile in neural crest stem cells that is due to deletion of the norepinephrine transporter (NET) gene. NET is the target of psychotropic substances, such as tricyclic antidepressants and the drug of abuse, cocaine. NET mutations have been implicated in depression, anxiety, orthostatic intolerance and attention deficit hyperactivity disorder (ADHD). NET function in adult noradrenergic neurons of the peripheral and central nervous systems is to internalize norepinephrine from the synaptic cleft. By contrast, during embryogenesis norepinephrine (NE) transport promotes differentiation of neural crest stem cells and locus ceruleus progenitors into noradrenergic neurons, whereas NET inhibitors block noradrenergic differentiation. While the structure of NET und the regulation of NET function are well described, little is known about downstream target genes of norepinephrine (NE) transport. Results: We have prepared gene expression profiles of in vitro differentiating wild type and norepinephrine transporter-deficient (NETKO) mouse neural crest cells using long serial analysis of gene expression (LongSAGE). Comparison analyses have identified a number of important differentially expressed genes, including genes relevant to neural crest formation, noradrenergic neuron differentiation and the phenotype of NETKO mice. Examples of differentially expressed genes that affect noradrenergic cell differentiation include genes in the bone morphogenetic protein (BMP) signaling pathway, the Phox2b binding partner Tlx2, the ubiquitin ligase Praja2, and the inhibitor of Notch signaling, Numbl. Differentially expressed genes that are likely to contribute to the NETKO phenotype include dopamine-β-hydroxylase (Dbh), tyrosine hydroxylase (Th), the peptide transmitter 'cocaine and amphetamine regulated transcript' (Cart), and the serotonin receptor subunit Htr3a. Real-time PCR confirmed differential expression of key genes not only in neural crest cells, but also in the adult superior cervical ganglion and locus ceruleus. In addition to known genes we have identified novel differentially expressed genes and thus provide a valuable database for future studies. Conclusion: Loss of NET function during embryonic development in the mouse deregulates signaling pathways that are critically involved in neural crest formation and noradrenergic cell differentiation. The data further suggest deregulation of signaling pathways in the development and/or function of the NET-deficient peripheral, central and enteric nervous systems. © 2009 Hu et al; licensee BioMed Central Ltd.

Authors
Hu, YF; Caron, MG; Sieber-Blum, M
MLA Citation
Hu, YF, Caron, MG, and Sieber-Blum, M. "Norepinephrine transport-mediated gene expression in noradrenergic neurogenesis." BMC Genomics 10 (2009).
PMID
19356247
Source
scival
Published In
BMC Genomics
Volume
10
Publish Date
2009
DOI
10.1186/1471-2164-10-151

Persistent hyperdopaminergia decreases the peak frequency of hippocampal theta oscillations during quiet waking and REM sleep.

Long-term changes in dopaminergic signaling are thought to underlie the pathophysiology of a number of psychiatric disorders. Several conditions are associated with cognitive deficits such as disturbances in attention processes and learning and memory, suggesting that persistent changes in dopaminergic signaling may alter neural mechanisms underlying these processes. Dopamine transporter knockout (DAT-KO) mice exhibit a persistent five-fold increase in extracellular dopamine levels. Here, we demonstrate that DAT-KO mice display lower hippocampal theta oscillation frequencies during baseline periods of waking and rapid-eye movement sleep. These altered theta oscillations are not reversed via treatment with the antidopaminergic agent haloperidol. Thus, we propose that persistent hyperdopaminergia, together with secondary alterations in other neuromodulatory systems, results in lower frequency activity in neural systems responsible for various cognitive processes.

Authors
Dzirasa, K; Santos, LM; Ribeiro, S; Stapleton, J; Gainetdinov, RR; Caron, MG; Nicolelis, MAL
MLA Citation
Dzirasa, K, Santos, LM, Ribeiro, S, Stapleton, J, Gainetdinov, RR, Caron, MG, and Nicolelis, MAL. "Persistent hyperdopaminergia decreases the peak frequency of hippocampal theta oscillations during quiet waking and REM sleep." PLoS One 4.4 (2009): e5238-.
PMID
19381303
Source
pubmed
Published In
PloS one
Volume
4
Issue
4
Publish Date
2009
Start Page
e5238
DOI
10.1371/journal.pone.0005238

MicroRNA-219 modulates NMDA receptor-mediated neurobehavioral dysfunction

N-methyl-D-aspartate (NMDA) glutamate receptors are regulators of fast neurotransmission and synaptic plasticity in the brain. Disruption of NMDA-mediated glutamate signaling has been linked to behavioral deficits displayed in psychiatric disorders such as schizophrenia. Recently, noncoding RNA molecules such as microRNAs (miRNAs) have emerged as critical regulators of neuronal functions. Here we show that pharmacological (dizocilpine) or genetic (NR1 hypomorphism) disruption of NMDA receptor signaling reduces levels of a brain-specific miRNA, miR-219, in the prefrontal cortex (PFC) of mice. Consistent with a role for miR-219 in NMDA receptor signaling, we identify calcium/calmodulin-dependent protein kinase II γ subunit (CaMKIIγ), a component of the NMDA receptor signaling cascade, as a target of miR-219. In vivo inhibition of miR-219 by specific antimiR in the murine brain significantly modulated behavioral responses associated with disrupted NMDA receptor transmission. Furthermore, pretreatment with the antipsychotic drugs haloperidol and clozapine prevented dizocilpine-induced effects on miR-219. Taken together, these data support an integral role for miR-219 in the expression of behavioral aberrations associated with NMDA receptor hypofunction. © 2009 by The National Academy of Sciences of the USA.

Authors
Kocerh, J; Faghihi, MA; Lopez-Toledano, MA; Huang, J; Ramsey, AJ; Caron, MG; Sales, N; Willoughby, D; Elmen, J; Hansen, HF; Orum, H; Kauppinen, S; Kenny, PJ; Wahlestedt, C
MLA Citation
Kocerh, J, Faghihi, MA, Lopez-Toledano, MA, Huang, J, Ramsey, AJ, Caron, MG, Sales, N, Willoughby, D, Elmen, J, Hansen, HF, Orum, H, Kauppinen, S, Kenny, PJ, and Wahlestedt, C. "MicroRNA-219 modulates NMDA receptor-mediated neurobehavioral dysfunction." Proceedings of the National Academy of Sciences of the United States of America 106.9 (2009): 3507-3512.
PMID
19196972
Source
scival
Published In
Proceedings of the National Academy of Sciences of USA
Volume
106
Issue
9
Publish Date
2009
Start Page
3507
End Page
3512
DOI
10.1073/pnas.0805854106

Building a new conceptual framework for receptor heteromers

Receptor heteromers constitute a new area of research that is reshaping our thinking about biochemistry, cell biology, pharmacology and drug discovery. In this commentary, we recommend clear definitions that should facilitate both information exchange and research on this growing class of transmembrane signal transduction units and their complex properties. We also consider research questions underlying the proposed nomenclature, with recommendations for receptor heteromer identification in native tissues and their use as targets for drug development. © 2009 Nature America, Inc. All rights reserved.

Authors
Ferré, S; Baler, R; Bouvier, M; Caron, MG; Devi, LA; Durroux, T; Fuxe, K; George, SR; Javitch, JA; Lohse, MJ; Mackie, K; Milligan, G; Pfleger, KDG; Pin, J-P; Volkow, ND; Waldhoer, M; Woods, AS; Franco, R
MLA Citation
Ferré, S, Baler, R, Bouvier, M, Caron, MG, Devi, LA, Durroux, T, Fuxe, K, George, SR, Javitch, JA, Lohse, MJ, Mackie, K, Milligan, G, Pfleger, KDG, Pin, J-P, Volkow, ND, Waldhoer, M, Woods, AS, and Franco, R. "Building a new conceptual framework for receptor heteromers." Nature Chemical Biology 5.3 (2009): 131-134.
PMID
19219011
Source
scival
Published In
Nature Chemical Biology
Volume
5
Issue
3
Publish Date
2009
Start Page
131
End Page
134
DOI
10.1038/nchembio0309-131

Akt/GSK3 signaling in the action of psychotropic drugs.

Psychotropic drugs acting on monoamine neurotransmission are major pharmacological treatments for neuropsychiatric conditions such as schizophrenia, depression, bipolar disorder, Tourette syndrome, ADHD, and Alzheimer disease. Independent lines of research involving biochemical and behavioral approaches in normal and/or genetically modified mice provide converging evidence for an involvement of the signaling molecules Akt and glycogen synthase kinase-3 (GSK3) in the regulation of behavior by dopamine and serotonin (5-HT). These signaling molecules have also received attention for their role in the actions of psychoactive drugs such as antidepressants, antipsychotics, lithium, and other mood stabilizers. Furthermore, investigations of the mechanism by which D2 dopamine receptors regulate Akt/GSK3 signaling strongly support the physiological relevance of a new modality of G protein-coupled receptor (GPCR) signaling involving the multifunctional scaffolding protein beta-arrestin 2. Elucidation of the contribution of multiple signaling pathways to the action of psychotropic drugs may provide a better biological understanding of psychiatric disorders and lead to more efficient therapeutics.

Authors
Beaulieu, J-M; Gainetdinov, RR; Caron, MG
MLA Citation
Beaulieu, J-M, Gainetdinov, RR, and Caron, MG. "Akt/GSK3 signaling in the action of psychotropic drugs." Annu Rev Pharmacol Toxicol 49 (2009): 327-347. (Review)
PMID
18928402
Source
pubmed
Published In
Annual Review of Pharmacology and Toxicology
Volume
49
Publish Date
2009
Start Page
327
End Page
347
DOI
10.1146/annurev.pharmtox.011008.145634

Smoothened signaling in vertebrates is facilitated by a G protein-coupled receptor kinase.

Smoothened, a heptahelical membrane protein, functions as the transducer of Hedgehog signaling. The kinases that modulate Smoothened have been thoroughly analyzed in flies. However, little is known about how phosphorylation affects Smoothened in vertebrates, mainly, because the residues, where Smoothened is phosphorylated are not conserved from Drosophila to vertebrates. Given its molecular architecture, Smoothened signaling is likely to be regulated in a manner analogous to G protein-coupled receptors (GPCRs). Previously, it has been shown, that arrestins and GPCR kinases, (GRKs) not only desensitize G protein-dependent receptor signaling but also function as triggers for GPCR trafficking and formation of signaling complexes. Here we describe that a GRK contributes to Smoothened-mediated signaling in vertebrates. Knockdown of the zebrafish homolog of mammalian GRK2/3 results in lowered Hedgehog transcriptional responses, impaired muscle development, and neural patterning. Results obtained in zebrafish are corroborated both in cell culture, where zGRK2/3 phosphorylates Smoothened and promotes Smoothened signal transduction and in mice where deletion of GRK2 interferes with neural tube patterning. Together, these data suggest that a GRK functions as a vertebrate kinase for Smoothened, promoting Hedgehog signal transduction during early development.

Authors
Philipp, M; Fralish, GB; Meloni, AR; Chen, W; MacInnes, AW; Barak, LS; Caron, MG
MLA Citation
Philipp, M, Fralish, GB, Meloni, AR, Chen, W, MacInnes, AW, Barak, LS, and Caron, MG. "Smoothened signaling in vertebrates is facilitated by a G protein-coupled receptor kinase." Mol Biol Cell 19.12 (December 2008): 5478-5489.
PMID
18815277
Source
pubmed
Published In
Molecular Biology of the Cell
Volume
19
Issue
12
Publish Date
2008
Start Page
5478
End Page
5489
DOI
10.1091/mbc.E08-05-0448

Implanted reuptake-deficient or wild-type dopaminergic neurons improve ON L-dopa dyskinesias without OFF-dyskinesias in a rat model of Parkinson's disease.

OFF-L-dopa dyskinesias have been a surprising side-effect of intrastriatal foetal ventral mesencephalic transplantation in patients with Parkinson's disease. It has been proposed that excessive and unregulated dopaminergic stimulation of host post-synaptic striatal neurons by the grafts could be responsible for these dyskinesias. To address this issue we transplanted foetal dopaminergic neurons from mice lacking the dopamine transporter (DATKO) or from wild-type mice, into a rat model of Parkinson's disease and L-dopa-induced dyskinesias. Both wild-type and DATKO grafts reinnervated the host striatum to a similar extent, but DATKO grafts produced a greater and more diffuse increase in extra-cellular striatal dopamine levels. Interestingly, grafts containing wild-type dopaminergic neurons improved parkinsonian signs to a similar extent as DATKO grafts, but provided a more complete reduction of L-dopa induced dyskinesias. Neither DATKO nor wild-type grafts induced OFF-L-dopa dyskinesias. Behavioural and receptor autoradiography analyses demonstrated that DATKO grafts induced a greater normalization of striatal dopaminergic receptor supersensitivity than wild-type grafts. Both graft types induced a similar downregulation and normalization of PEnk and fosb/Deltafosb in striatal neurons. In summary, DATKO grafts causing high and diffuse extra-cellular dompamine levels do not per se alter graft-induced recovery or produce OFF-L-dopa dyskinesias. Wild-type dopaminergic neurons appear to be the most effective neuronal type to restore function and reduce L-dopa-induced dyskinesias.

Authors
Vinuela, A; Hallett, PJ; Reske-Nielsen, C; Patterson, M; Sotnikova, TD; Caron, MG; Gainetdinov, RR; Isacson, O
MLA Citation
Vinuela, A, Hallett, PJ, Reske-Nielsen, C, Patterson, M, Sotnikova, TD, Caron, MG, Gainetdinov, RR, and Isacson, O. "Implanted reuptake-deficient or wild-type dopaminergic neurons improve ON L-dopa dyskinesias without OFF-dyskinesias in a rat model of Parkinson's disease." Brain 131.Pt 12 (December 2008): 3361-3379.
PMID
18988638
Source
pubmed
Published In
Brain
Volume
131
Issue
Pt 12
Publish Date
2008
Start Page
3361
End Page
3379
DOI
10.1093/brain/awn192

Activity-dependent expression of ELAV/Hu RBPs and neuronal mRNAs in seizure and cocaine brain.

Growing evidence indicates that both seizure (glutamate) and cocaine (dopamine) treatment modulate synaptic plasticity within the mesolimbic region of the CNS. Activation of glutamatergic neurons depends on the localized translation of neuronal mRNA products involved in modulating synaptic plasticity. In this study, we demonstrate the dendritic localization of HuR and HuD RNA-binding proteins (RBPs) and their association with neuronal mRNAs following these two paradigms of seizure and cocaine treatment. Both the ubiquitously expressed HuR and neuronal HuD RBPs were detected in different regions as well as within dendrites of the brain and in dissociated neurons. Quantitative analysis revealed an increase in HuR, HuD and p-glycogen synthase kinase 3beta (GSK3beta) protein levels as well as neuronal mRNAs encoding Homer, CaMKIIalpha, vascular early response gene, GAP-43, neuritin, and neuroligin protein products following either seizure or cocaine treatment. Inhibition of the Akt/GSK3beta signaling pathway by acute or chronic LiCl treatment revealed changes in HuR, HuD, pGSK3beta, p-Akt, and beta-catenin protein levels. In addition, a genetically engineered hyperdopaminergic mouse model (dopamine transporter knockout) revealed decreased expression of HuR protein levels, but no significant change was observed in HuD or fragile-X mental retardation protein RBPs. Finally, our data suggest that HuR and HuD RBPs potentially interact directly with neuronal mRNAs important for differentiation and synaptic plasticity.

Authors
Tiruchinapalli, DM; Caron, MG; Keene, JD
MLA Citation
Tiruchinapalli, DM, Caron, MG, and Keene, JD. "Activity-dependent expression of ELAV/Hu RBPs and neuronal mRNAs in seizure and cocaine brain." J Neurochem 107.6 (December 2008): 1529-1543.
PMID
19014379
Source
pubmed
Published In
Journal of Neurochemistry
Volume
107
Issue
6
Publish Date
2008
Start Page
1529
End Page
1543
DOI
10.1111/j.1471-4159.2008.05718.x

Genetic NMDA receptor deficiency disrupts acute and chronic effects of cocaine but not amphetamine.

NMDA receptor-mediated glutamate transmission is required for several forms of neuronal plasticity. Its role in the neuronal responses to addictive drugs is an ongoing subject of investigation. We report here that the acute locomotor-stimulating effect of cocaine is absent in NMDA receptor-deficient mice (NR1-KD). In contrast, their acute responses to amphetamine and to direct dopamine receptor agonists are not significantly altered. The striking attenuation of cocaine's acute effects is not likely explained by alterations in the dopaminergic system of NR1-KD mice, since most parameters of pre- and postsynaptic dopamine function are unchanged. Consistent with the behavioral findings, cocaine induces less c-Fos expression in the striatum of these mice, while amphetamine-induced c-Fos expression is intact. Furthermore, chronic cocaine-induced sensitization and conditioned place preference are attenuated and develop more slowly in mutant animals, but amphetamine's effects are not altered significantly. Our results highlight the importance of NMDA receptor-mediated glutamatergic transmission specifically in cocaine actions, and support a hypothesis that cocaine and amphetamine elicit their effects through differential actions on signaling pathways.

Authors
Ramsey, AJ; Laakso, A; Cyr, M; Sotnikova, TD; Salahpour, A; Medvedev, IO; Dykstra, LA; Gainetdinov, RR; Caron, MG
MLA Citation
Ramsey, AJ, Laakso, A, Cyr, M, Sotnikova, TD, Salahpour, A, Medvedev, IO, Dykstra, LA, Gainetdinov, RR, and Caron, MG. "Genetic NMDA receptor deficiency disrupts acute and chronic effects of cocaine but not amphetamine." Neuropsychopharmacology 33.11 (October 2008): 2701-2714.
PMID
18185498
Source
pubmed
Published In
Neuropsychopharmacology
Volume
33
Issue
11
Publish Date
2008
Start Page
2701
End Page
2714
DOI
10.1038/sj.npp.1301663

Antagonism of dopamine D2 receptor/beta-arrestin 2 interaction is a common property of clinically effective antipsychotics.

Since the unexpected discovery of the antipsychotic activity of chlorpromazine, a variety of therapeutic agents have been developed for the treatment of schizophrenia. Despite differences in their activities at various neurotransmitter systems, all clinically effective antipsychotics share the ability to interact with D2 class dopamine receptors (D2R). D2R mediate their physiological effects via both G protein-dependent and independent (beta-arrestin 2-dependent) signaling, but the role of these D2R-mediated signaling events in the actions of antipsychotics remains unclear. We demonstrate here that while different classes of antipsychotics have complex pharmacological profiles at G protein-dependent D2R long isoform (D2(L)R) signaling, they share the common property of antagonizing dopamine-mediated interaction of D2(L)R with beta-arrestin 2. Using two cellular assays based on a bioluminescence resonance energy transfer (BRET) approach, we demonstrate that a series of antipsychotics including haloperidol, clozapine, aripiprazole, chlorpromazine, quetiapine, olanzapine, risperidone, and ziprasidone all potently antagonize the beta-arrestin 2 recruitment to D2(L)R induced by quinpirole. However, these antipsychotics have various effects on D2(L)R mediated G(i/o) protein activation ranging from inverse to partial agonists and antagonists with highly variable efficacies and potencies at quinpirole-induced cAMP inhibition. These results suggest that the different classes of clinically effective antipsychotics share a common molecular mechanism involving inhibition of D2(L)R/beta-arrestin 2 mediated signaling. Thus, selective targeting of D2(L)R/beta-arrestin 2 interaction and related signaling pathways may provide new opportunities for antipsychotic development.

Authors
Masri, B; Salahpour, A; Didriksen, M; Ghisi, V; Beaulieu, J-M; Gainetdinov, RR; Caron, MG
MLA Citation
Masri, B, Salahpour, A, Didriksen, M, Ghisi, V, Beaulieu, J-M, Gainetdinov, RR, and Caron, MG. "Antagonism of dopamine D2 receptor/beta-arrestin 2 interaction is a common property of clinically effective antipsychotics." Proc Natl Acad Sci U S A 105.36 (September 9, 2008): 13656-13661.
PMID
18768802
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
105
Issue
36
Publish Date
2008
Start Page
13656
End Page
13661
DOI
10.1073/pnas.0803522105

N-desalkylquetiapine, a potent norepinephrine reuptake inhibitor and partial 5-HT1A agonist, as a putative mediator of quetiapine's antidepressant activity.

Quetiapine is an atypical antipsychotic drug that is also US FDA approved for treating bipolar depression, albeit by an unknown mechanism. To discover the potential mechanism for this apparently unique action, we screened quetiapine, its metabolite N-Desalkylquetiapine, and dibenzo[b,f][1,4]thiazepine-11(10-H)-one (DBTO) against a large panel of G-protein-coupled receptors, ion channels, and neurotransmitter transporters. DBTO was inactive at all tested molecular targets. N-Desalkylquetiapine had a high affinity (3.4 nM) for the histamine H(1) receptor and moderate affinities (10-100 nM) for the norepinephrine reuptake transporter (NET), the serotonin 5-HT(1A), 5-HT(1E), 5-HT(2A), 5-HT(2B), 5-HT(7) receptors, the alpha(1B)-adrenergic receptor, and the M(1), M(3), and M(5) muscarinic receptors. The compound had low affinities (100-1000 nM) for the 5-HT(1D), 5-HT(2C), 5-HT(3), 5-HT(5), 5-HT(6), alpha(1A), alpha(2A), alpha(2B), alpha(2C), H(2), M(2), M(4), and dopamine D(1), D(2), D(3), and D(4) receptors. N-Desalkylquetiapine potently inhibited human NE transporter with a K(i) of 12 nM, about 100-fold more potent than quetiapine itself. N-Desalkylquetiapine was also 10-fold more potent and more efficacious than quetiapine at the 5-HT(1A) receptor. N-Desalkylquetiapine was an antagonist at 5-HT(2A), 5-HT(2B), 5-HT(2C), alpha(1A), alpha(1D), alpha(2A), alpha(2C), H(1), M(1), M(3), and M(5) receptors. In the mouse tail suspension test, N-Desalkylquetiapine displayed potent antidepressant-like activity in VMAT2 heterozygous mice at doses as low as 0.1 mg/kg. These data strongly suggest that the antidepressant activity of quetiapine is mediated, at least in part, by its metabolite N-Desalkylquetiapine through NET inhibition and partial 5-HT(1A) agonism. Possible contributions of this metabolite to the side effects of quetiapine are discussed.

Authors
Jensen, NH; Rodriguiz, RM; Caron, MG; Wetsel, WC; Rothman, RB; Roth, BL
MLA Citation
Jensen, NH, Rodriguiz, RM, Caron, MG, Wetsel, WC, Rothman, RB, and Roth, BL. "N-desalkylquetiapine, a potent norepinephrine reuptake inhibitor and partial 5-HT1A agonist, as a putative mediator of quetiapine's antidepressant activity." Neuropsychopharmacology 33.10 (September 2008): 2303-2312.
PMID
18059438
Source
pubmed
Published In
Neuropsychopharmacology
Volume
33
Issue
10
Publish Date
2008
Start Page
2303
End Page
2312
DOI
10.1038/sj.npp.1301646

Pharmacological characterization of membrane-expressed human trace amine-associated receptor 1 (TAAR1) by a bioluminescence resonance energy transfer cAMP biosensor.

Trace amines are neurotransmitters whose role in regulating invertebrate physiology has been appreciated for many decades. Recent studies indicate that trace amines may also play a role in mammalian physiology by binding to a novel family of G protein-coupled receptors (GPCRs) that are found throughout the central nervous system. A major obstacle impeding the careful pharmacological characterization of trace amine associated receptors (TAARs) is their extremely poor membrane expression in model cell systems, and a molecular basis for this phenomenon has not been determined. In the present study, we show that the addition of an asparagine-linked glycosylation site to the N terminus of the human trace amine associated receptor 1 (TAAR1) is sufficient to enable its plasma membrane expression, and thus its pharmacological characterization with a novel cAMP EPAC (exchange protein directly activated by cAMP) protein based bioluminescence resonance energy transfer (BRET) biosensor. We applied this novel cAMP BRET biosensor to evaluate the activity of putative TAAR1 ligands. This study represents the first comprehensive investigation of the membrane-expressed human TAAR1 pharmacology. Our strategy to express TAARs and to identify their ligands using a cAMP BRET assay could provide a foundation for characterizing the functional role of trace amines in vivo and suggests a strategy to apply to groups of poorly expressing GPCRs that have remained difficult to investigate in model systems.

Authors
Barak, LS; Salahpour, A; Zhang, X; Masri, B; Sotnikova, TD; Ramsey, AJ; Violin, JD; Lefkowitz, RJ; Caron, MG; Gainetdinov, RR
MLA Citation
Barak, LS, Salahpour, A, Zhang, X, Masri, B, Sotnikova, TD, Ramsey, AJ, Violin, JD, Lefkowitz, RJ, Caron, MG, and Gainetdinov, RR. "Pharmacological characterization of membrane-expressed human trace amine-associated receptor 1 (TAAR1) by a bioluminescence resonance energy transfer cAMP biosensor." Mol Pharmacol 74.3 (September 2008): 585-594.
PMID
18524885
Source
pubmed
Published In
Molecular pharmacology
Volume
74
Issue
3
Publish Date
2008
Start Page
585
End Page
594
DOI
10.1124/mol.108.048884

A regulatory domain in the N terminus of tryptophan hydroxylase 2 controls enzyme expression.

Serotonin is involved in a variety of physiological processes in the central nervous system and the periphery. As the rate-limiting enzyme in serotonin synthesis, tryptophan hydroxylase plays an important role in modulating these processes. Of the two variants of tryptophan hydroxylase, tryptophan hydroxylase 2 (TPH2) is expressed predominantly in the central nervous system, whereas tryptophan hydroxylase 1 (TPH1) is expressed mostly in peripheral tissues. Although the two enzymes share considerable sequence homology, the regulatory domain of TPH2 contains an additional 41 amino acids at the N terminus that TPH1 lacks. Here we show that the extended TPH2 N-terminal domain contains a unique sequence involved in the regulation of enzyme expression. When expressed in cultured mammalian cells, TPH2 is synthesized less efficiently and is also less stable than TPH1. Removal of the unique portion of the N terminus of TPH2 results in expression of the enzyme at a level similar to that of TPH1, whereas protein chimeras containing this fragment are expressed at lower levels than their wild-type counterparts. We identify a region centered on amino acids 10-20 that mediates the bulk of this effect. We also demonstrate that phosphorylation of serine 19, a protein kinase A consensus site located in this N-terminal domain, results in increased TPH2 stability and consequent increases in enzyme output in cell culture systems. Because this domain is unique to TPH2, these data provide evidence for selective regulation of brain serotonin synthesis.

Authors
Murphy, KL; Zhang, X; Gainetdinov, RR; Beaulieu, J-M; Caron, MG
MLA Citation
Murphy, KL, Zhang, X, Gainetdinov, RR, Beaulieu, J-M, and Caron, MG. "A regulatory domain in the N terminus of tryptophan hydroxylase 2 controls enzyme expression." J Biol Chem 283.19 (May 9, 2008): 13216-13224.
PMID
18339632
Source
pubmed
Published In
The Journal of biological chemistry
Volume
283
Issue
19
Publish Date
2008
Start Page
13216
End Page
13224
DOI
10.1074/jbc.M706749200

Food reward in the absence of taste receptor signaling.

Food palatability and hedonic value play central roles in nutrient intake. However, postingestive effects can influence food preferences independently of palatability, although the neurobiological bases of such mechanisms remain poorly understood. Of central interest is whether the same brain reward circuitry that is responsive to palatable rewards also encodes metabolic value independently of taste signaling. Here we show that trpm5-/- mice, which lack the cellular machinery required for sweet taste transduction, can develop a robust preference for sucrose solutions based solely on caloric content. Sucrose intake induced dopamine release in the ventral striatum of these sweet-blind mice, a pattern usually associated with receipt of palatable rewards. Furthermore, single neurons in this same ventral striatal region showed increased sensitivity to caloric intake even in the absence of gustatory inputs. Our findings suggest that calorie-rich nutrients can directly influence brain reward circuits that control food intake independently of palatability or functional taste transduction.

Authors
de Araujo, IE; Oliveira-Maia, AJ; Sotnikova, TD; Gainetdinov, RR; Caron, MG; Nicolelis, MAL; Simon, SA
MLA Citation
de Araujo, IE, Oliveira-Maia, AJ, Sotnikova, TD, Gainetdinov, RR, Caron, MG, Nicolelis, MAL, and Simon, SA. "Food reward in the absence of taste receptor signaling." Neuron 57.6 (March 27, 2008): 930-941.
PMID
18367093
Source
pubmed
Published In
Neuron
Volume
57
Issue
6
Publish Date
2008
Start Page
930
End Page
941
DOI
10.1016/j.neuron.2008.01.032

Increased amphetamine-induced hyperactivity and reward in mice overexpressing the dopamine transporter.

The dopamine transporter (DAT) plays a key role in the regulation of dopaminergic signaling wherein it controls both the spatial and temporal actions of dopamine. Here we evaluated the behavioral and neurochemical consequences of increased DAT function by generating DAT transgenic mice (DAT-tg) that overexpress the transporter. These mice were generated by pronuclear injection of a bacterial artificial chromosome containing the mouse DAT locus, yielding an anatomical expression pattern of DAT-tg identical to WT. In DAT-tg mice there is a 3-fold increase in the levels of total and membrane-expressed DAT, but synaptic plasma membrane fractions of DAT-tg mice show only a 30% increase in transporter levels. Functional studies reveal that in the DAT-tg animals there is a 50% increase in the rate of dopamine (DA) uptake resulting in extracellular levels of DA that are decreased by approximately 40%. Behaviorally, DAT-tg animals display similar locomotor stimulation when treated with DAT blockers such as GBR12909, methylphenidate, and cocaine. However, these mice demonstrate markedly increased locomotor responses to amphetamine compared with WT animals. Furthermore, compared with controls, there is a 3-fold greater increase in the amount of DA released by amphetamine in DAT-tg mice that correlates with the 3-fold increase in protein expression. Finally, DAT-tg animals show reduced operant responding for natural reward while displaying preference for amphetamine at much lower doses (0.2 and 0.5 mg/kg) than WT mice (2 mg/kg). These results suggest that overexpression of DAT leads to a marked increase in sensitivity to psychomotor and rewarding properties of amphetamine.

Authors
Salahpour, A; Ramsey, AJ; Medvedev, IO; Kile, B; Sotnikova, TD; Holmstrand, E; Ghisi, V; Nicholls, PJ; Wong, L; Murphy, K; Sesack, SR; Wightman, RM; Gainetdinov, RR; Caron, MG
MLA Citation
Salahpour, A, Ramsey, AJ, Medvedev, IO, Kile, B, Sotnikova, TD, Holmstrand, E, Ghisi, V, Nicholls, PJ, Wong, L, Murphy, K, Sesack, SR, Wightman, RM, Gainetdinov, RR, and Caron, MG. "Increased amphetamine-induced hyperactivity and reward in mice overexpressing the dopamine transporter." Proc Natl Acad Sci U S A 105.11 (March 18, 2008): 4405-4410.
PMID
18347339
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
105
Issue
11
Publish Date
2008
Start Page
4405
End Page
4410
DOI
10.1073/pnas.0707646105

Role of GSK3 beta in behavioral abnormalities induced by serotonin deficiency.

Dysregulation of brain serotonin (5-HT) neurotransmission is thought to underlie mental conditions as diverse as depression, anxiety disorders, bipolar disorder, autism, and schizophrenia. Despite treatment of these conditions with serotonergic drugs, the molecular mechanisms by which 5-HT is involved in the regulation of aberrant emotional behaviors are poorly understood. Here, we generated knockin mice expressing a mutant form of the brain 5-HT synthesis enzyme, tryptophan hydroxylase 2 (Tph2). This mutant is equivalent to a rare human variant (R441H) identified in few individuals with unipolar major depression. Expression of mutant Tph2 in mice results in markedly reduced ( approximately 80%) brain 5-HT production and leads to behavioral abnormalities in tests assessing 5-HT-mediated emotional states. This reduction in brain 5-HT levels is accompanied by activation of glycogen synthase kinase 3beta (GSK3beta), a signaling molecule modulated by many psychiatric therapeutic agents. Importantly, inactivation of GSK3beta in Tph2 knockin mice, using pharmacological or genetic approaches, alleviates the aberrant behaviors produced by 5-HT deficiency. These findings establish a critical role of Tph2 in the maintenance of brain serotonin homeostasis and identify GSK3beta signaling as an important pathway through which brain 5-HT deficiency induces abnormal behaviors. Targeting GSK3beta and related signaling events may afford therapeutic advantages for the management of certain 5-HT-related psychiatric conditions.

Authors
Beaulieu, J-M; Zhang, X; Rodriguiz, RM; Sotnikova, TD; Cools, MJ; Wetsel, WC; Gainetdinov, RR; Caron, MG
MLA Citation
Beaulieu, J-M, Zhang, X, Rodriguiz, RM, Sotnikova, TD, Cools, MJ, Wetsel, WC, Gainetdinov, RR, and Caron, MG. "Role of GSK3 beta in behavioral abnormalities induced by serotonin deficiency." Proc Natl Acad Sci U S A 105.4 (January 29, 2008): 1333-1338.
PMID
18212115
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
105
Issue
4
Publish Date
2008
Start Page
1333
End Page
1338
DOI
10.1073/pnas.0711496105

A beta-arrestin 2 signaling complex mediates lithium action on behavior.

Besides their role in desensitization, beta-arrestin 1 and 2 promote the formation of signaling complexes allowing G protein-coupled receptors (GPCR) to signal independently from G proteins. Here we show that lithium, a pharmacological agent used for the management of psychiatric disorders such as bipolar disorder, schizophrenia, and depression, regulates Akt/glycogen synthase kinase 3 (GSK3) signaling and related behaviors in mice by disrupting a signaling complex composed of Akt, beta-arrestin 2, and protein phosphatase 2A. When administered to beta-arrestin 2 knockout mice, lithium fails to affect Akt/GSK3 signaling and induce behavioral changes associated with GSK3 inhibition as it does in normal animals. These results point toward a pharmacological approach to modulating GPCR function that affects the formation of beta-arrestin-mediated signaling complexes.

Authors
Beaulieu, J-M; Marion, S; Rodriguiz, RM; Medvedev, IO; Sotnikova, TD; Ghisi, V; Wetsel, WC; Lefkowitz, RJ; Gainetdinov, RR; Caron, MG
MLA Citation
Beaulieu, J-M, Marion, S, Rodriguiz, RM, Medvedev, IO, Sotnikova, TD, Ghisi, V, Wetsel, WC, Lefkowitz, RJ, Gainetdinov, RR, and Caron, MG. "A beta-arrestin 2 signaling complex mediates lithium action on behavior." Cell 132.1 (January 11, 2008): 125-136.
PMID
18191226
Source
pubmed
Published In
Cell
Volume
132
Issue
1
Publish Date
2008
Start Page
125
End Page
136
DOI
10.1016/j.cell.2007.11.041

Looking at lithium: Molecular moods and complex behavior

Lithium and other mood-stabilizing drugs are used for the management of bipolar mood disorders and, to a lesser extent, for augmentation of other psychoactive drugs. Lithium also has neuroprotective properties that may be useful for treatment of neurodegenerative diseases such as Alzheimer's disease and amyotrophic lateral sclerosis. Over the years, lithium has been shown to inhibit inositol monophosphatases and glycogen synthase kinase 3, but the relevance of such enzyme inhibition to the therapeutic effects of lithium has remained difficult to assess. Here, we provide an overview of recent advances in the identification of molecular mechanisms involved in the regulation of behavior by lithium. We also highlight recent findings suggesting that lithium could exert some of its behavioral effects by acting on a dopamine receptor-regulated signaling complex composed of Akt, protein phosphatase 2A, and the multifunctional protein scaffold beta-arrestin 2.

Authors
Beaulieu, J-M; Caron, MG
MLA Citation
Beaulieu, J-M, and Caron, MG. "Looking at lithium: Molecular moods and complex behavior." Molecular Interventions 8.5 (2008): 230-241.
PMID
19015387
Source
scival
Published In
Molecular interventions
Volume
8
Issue
5
Publish Date
2008
Start Page
230
End Page
241
DOI
10.1124/mi.8.5.8

Animal models of depression in dopamine, serotonin, and norepinephrine transporter knockout mice: Prominent effects of dopamine transporter deletions

Antidepressant drugs produce therapeutic actions and many of their side effects via blockade of the plasma membrane transporters for serotonin (SERT/SLC6A2), norepinephrine (NET/SLC6A1), and dopamine (DAT/SLC6A3). Many antidepressants block several of these transporters; some are more selective. Mouse gene knockouts of these transporters provide interesting models for possible effects of chronic antidepressant treatments. To examine the role of monoamine transporters in models of depression DAT, NET, and SERT knockout (KO) mice and wild-type littermates were studied in the forced swim test (FST), the tail suspension test, and for sucrose consumption. To dissociate general activity from potential antidepressant effects three types of behavior were assessed in the FST: immobility, climbing, and swimming. In confirmation of earlier reports, both DAT KO and NET KO mice exhibited less immobility than wild-type littermates whereas SERT KO mice did not. Effects of DAT deletion were not simply because of hyperactivity, as decreased immobility was observed in DAT+/- mice that were not hyperactive as well as in DAT-/- mice that displayed profound hyperactivity. Climbing was increased, whereas swimming was almost eliminated in DAT-/- mice, and a modest but similar effect was seen in NET KO mice, which showed a modest decrease in locomotor activity. Combined increases in climbing and decreases in immobility are characteristic of FST results in antidepressant animal models, whereas selective effects on swimming are associated with the effects of stimulant drugs. Therefore, an effect on climbing is thought to more specifically reflect antidepressant effects, as has been observed in several other proposed animal models of reduced depressive phenotypes. A similar profile was observed in the tail suspension test, where DAT, NET, and SERT knockouts were all found to reduce immobility, but much greater effects were observed in DAT KO mice. However, to further determine whether these effects of DAT KO in animal models of depression may be because of the confounding effects of hyperactivity, mice were also assessed in a sucrose consumption test. Sucrose consumption was increased in DAT KO mice consistent with reduced anhedonia, and inconsistent with competitive hyperactivity; no increases were observed in SERT KO or NET KO mice. In summary, the effects of DAT KO in animal models of depression are larger than those produced by NET or SERT KO, and unlikely to be simply the result of the confounding effects of locomotor hyperactivity; thus, these data support reevaluation of the role that DAT expression could play in depression and the potential antidepressant effects of DAT blockade. © 2008 Wolters Kluwer Health | Lippincott Williams & Wilkins.

Authors
Perona, MTG; Waters, S; Hall, FS; Sora, I; Lesch, K-P; Murphy, DL; Caron, M; Uhl, GR
MLA Citation
Perona, MTG, Waters, S, Hall, FS, Sora, I, Lesch, K-P, Murphy, DL, Caron, M, and Uhl, GR. "Animal models of depression in dopamine, serotonin, and norepinephrine transporter knockout mice: Prominent effects of dopamine transporter deletions." Behavioural Pharmacology 19.5-6 (2008): 566-574.
PMID
18690111
Source
scival
Published In
Behavioural Pharmacology
Volume
19
Issue
5-6
Publish Date
2008
Start Page
566
End Page
574
DOI
10.1097/FBP.0b013e32830cd80f

Atypical antipsychotics clozapine and quetiapine attenuate prepulse inhibition deficits in dopamine transporter knockout mice

Sensorimotor gating disruptions are seen in various psychiatric illnesses with putatively different pathologies, including schizophrenia and bipolar disorder. Interestingly, mice lacking the dopamine (DA) transporter (DAT) gene display markedly increased levels of DA, deficits in sensorimotor gating, and hyperactivity relative to wild-type mice. Atypical antipsychotics are effective treatments of schizophrenia and manic symptoms, presumably in part by antagonizing DA receptors. Here we report that treatment with clozapine (3 mg/kg) or quetiapine (2.5mg/kg) attenuated prepulse inhibition deficits in male DAT knockout mice. Thus male DAT knockout mice may provide a useful animal model for predicting the efficacy of novel drugs in treating psychiatric illnesses characterized by a dysregulated DA system. © 2008 Wolters Kluwer Health | Lippincott Williams & Wilkins.

Authors
Powell, SB; Young, JW; Ong, JC; Caron, MG; Geyer, MA
MLA Citation
Powell, SB, Young, JW, Ong, JC, Caron, MG, and Geyer, MA. "Atypical antipsychotics clozapine and quetiapine attenuate prepulse inhibition deficits in dopamine transporter knockout mice." Behavioural Pharmacology 19.5-6 (2008): 562-565.
PMID
18690110
Source
scival
Published In
Behavioural Pharmacology
Volume
19
Issue
5-6
Publish Date
2008
Start Page
562
End Page
565
DOI
10.1097/FBP.0b013e32830dc110

Trace amine associated receptor 1 and movement control.

The recently discovered trace amine associated receptors (TAARs) represent attractive potential mediators of certain aspects of movement control. The TAAR that is best characterized, TAAR1, is particularly interesting because it can be activated by a variety of monoaminergic compounds including trace amines, amphetamines and dopamine metabolites. By using an experimental paradigm developed in our laboratory that involves a novel model of acute dopamine deficiency (DDD mice), and mice lacking TAAR1 (TAAR1 knockout mice), we explored the role of TAAR1 in movement control and actions of antiparkinsonian drugs. These investigations may eventually bring novel approaches to the pharmacology of Parkinson's disease.

Authors
Sotnikova, TD; Zorina, OI; Ghisi, V; Caron, MG; Gainetdinov, RR
MLA Citation
Sotnikova, TD, Zorina, OI, Ghisi, V, Caron, MG, and Gainetdinov, RR. "Trace amine associated receptor 1 and movement control." Parkinsonism Relat Disord 14 Suppl 2 (2008): S99-102. (Review)
PMID
18585080
Source
pubmed
Published In
Parkinsonism & Related Disorders
Volume
14 Suppl 2
Publish Date
2008
Start Page
S99
End Page
102
DOI
10.1016/j.parkreldis.2008.04.006

Reply to Belmaker et al.: GSK3β haploinsufficiency results in lithium-like effects in the forced-swim test

Authors
Beaulieu, J-M; Zhang, X; Rodriguiz, RM; Sotnikova, TD; Wetsel, WC; Gainetdinov, RR; Caron, MG
MLA Citation
Beaulieu, J-M, Zhang, X, Rodriguiz, RM, Sotnikova, TD, Wetsel, WC, Gainetdinov, RR, and Caron, MG. "Reply to Belmaker et al.: GSK3β haploinsufficiency results in lithium-like effects in the forced-swim test." Proceedings of the National Academy of Sciences of the United States of America 105.20 (2008): E24-.
Source
scival
Published In
Proceedings of the National Academy of Sciences of USA
Volume
105
Issue
20
Publish Date
2008
Start Page
E24
DOI
10.1073/pnas.0803026105

Food Reward in the Absence of Taste Receptor Signaling (DOI:10.1016/j.neuron.2008.01.032)

Authors
Araujo, IED; Oliveira-Maia, AJ; Sotnikova, TD; Gainetdinov, RR; Caron, MG; Nicolelis, MAL; Simon, SA
MLA Citation
Araujo, IED, Oliveira-Maia, AJ, Sotnikova, TD, Gainetdinov, RR, Caron, MG, Nicolelis, MAL, and Simon, SA. "Food Reward in the Absence of Taste Receptor Signaling (DOI:10.1016/j.neuron.2008.01.032)." Neuron 58.2 (2008): 295--.
Source
scival
Published In
Neuron
Volume
58
Issue
2
Publish Date
2008
Start Page
295-
DOI
10.1016/j.neuron.2008.04.003

Complementary roles of the DRY motif and C-terminus tail of GPCRS for G protein coupling and β-arrestin interaction

β-arrestin mediates the desensitization of GPCRs and acts as an adaptor molecule to recruit the receptor complex to clathrin-rich regions. Class-A GPCRs subsequently dissociate from β-arrestin but class-B GPCRs internalize with β-arrestin in the endocytic vesicles. Here the dopamine D2 and D3 receptors, which have similar structural features but different intracellular trafficking properties, were used in an attempt to better understand the structural requirements for the classification of GPCRs. The C-terminus tail of the vasopressin type-2 receptor was added to the ends of D2R and D3R to increase their affinity to β-arrestin. A point mutation was introduced into the DRY motif to change their basal activation levels. Among a battery of constructs in which the C-terminus tail and/or DRY motif was altered, class-B behavior was observed with the constructs whose affinities for β-arrestin were increased complementarily and their signaling was either maintained or regained. In conclusion, the DRY motif and C-terminal tail of the GPCRs determine complementarily their intracellular trafficking behavior by regulating the affinity to β-arrestin and G protein coupling. © 2007 Elsevier Inc. All rights reserved.

Authors
Kim, K-M; Caron, MG
MLA Citation
Kim, K-M, and Caron, MG. "Complementary roles of the DRY motif and C-terminus tail of GPCRS for G protein coupling and β-arrestin interaction." Biochemical and Biophysical Research Communications 366.1 (2008): 42-47.
PMID
18036556
Source
scival
Published In
Biochemical and Biophysical Research Communications
Volume
366
Issue
1
Publish Date
2008
Start Page
42
End Page
47
DOI
10.1016/j.bbrc.2007.11.055

Vmat2 heterozygous mutant mice display a depressive-like phenotype.

The vesicular monoamine transporter 2 (VMAT2) is localized primarily within the CNS and is responsible for transporting monoamines from the cytoplasm into secretory vesicles. Because reserpine (a VMAT inhibitor) can precipitate depressive-like symptoms in humans, we investigated whether Vmat2 heterozygous (HET) mice present with depressive-like behaviors. The mutants showed locomotor and rearing retardation in the open field and appeared anhedonic to 1 and 1.5% sucrose solutions. Immobility times for Vmat2 heterozygotes were prolonged in forced swim and imipramine normalized this behavior. HET animals also showed enhanced immobility in tail suspension and this response was alleviated by fluoxetine, reboxetine, and bupropion. Stimulated GTPgammaS binding indicated that alpha2-adrenergic receptors in HET hippocampus were more sensitive to UK 14,304 (5-bromo-N-(4,5-dihydro-1-H-imidazol-2-yl)-6-quinoxalinamine) stimulation than in wild type (WT) mice. In learned helplessness, mice were exposed to a shuttle box for 4 d or were given inescapable foot-shocks for the same time period. On day 5, all animals were tested in shock escape. Failure rates and the latency to escape were similar for WT and HET mice that were only pre-exposed to the test apparatus. In foot-shock groups, learned helplessness was more robust in heterozygotes than in WT controls. Basal secretion of serum corticosterone was not distinguished by genotype; however, corticosterone levels in mutants were more responsive to stress. Anxiety-like responses of WT and HET animals in the open field, light-dark exploration, zero maze, and novelty-suppressed feeding tests were indistinguishable. Collectively, these findings suggest that Vmat2 heterozygotes display a depressive-like phenotype that is devoid of anxiety-like behavior.

Authors
Fukui, M; Rodriguiz, RM; Zhou, J; Jiang, SX; Phillips, LE; Caron, MG; Wetsel, WC
MLA Citation
Fukui, M, Rodriguiz, RM, Zhou, J, Jiang, SX, Phillips, LE, Caron, MG, and Wetsel, WC. "Vmat2 heterozygous mutant mice display a depressive-like phenotype." J Neurosci 27.39 (September 26, 2007): 10520-10529.
PMID
17898223
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
27
Issue
39
Publish Date
2007
Start Page
10520
End Page
10529
DOI
10.1523/JNEUROSCI.4388-06.2007

Effects of stimulant medication on growth rates across 3 years in the MTA follow-up.

OBJECTIVE: To evaluate the hypothesis of stimulant medication effect on physical growth in the follow-up phase of the Multimodal Treatment Study of Children With ADHD. METHOD: Naturalistic subgroups were established based on patterns of treatment with stimulant medication at baseline, 14-, 24-, and 36-month assessments: not medicated (n = 65), newly medicated (n = 88), consistently medicated (n = 70), and inconsistently medicated (n = 147). Analysis of variance was used to evaluate effects of subgroup and assessment time on measures of relative size (z scores) obtained from growth norms. RESULTS: The subgroup x assessment time interaction was significant for z height (p <.005) and z weight (p <.0001), due primarily to divergence of the newly medicated and the not medicated subgroups. These initially stimulant-naïve subgroups had z scores significantly >0 at baseline. The newly medicated subgroup showed decreases in relative size that reached asymptotes by the 36-month assessment, when this group showed average growth of 2.0 cm and 2.7 kg less than the not medicated subgroup, which showed slight increases in relative size. CONCLUSIONS: Stimulant-naïve school-age children with Combined type attention-deficit/hyperactivity disorder were, as a group, larger than expected from norms before treatment but show stimulant-related decreases in growth rates after initiation of treatment, which appeared to reach asymptotes within 3 years without evidence of growth rebound.

Authors
Swanson, JM; Elliott, GR; Greenhill, LL; Wigal, T; Arnold, LE; Vitiello, B; Hechtman, L; Epstein, JN; Pelham, WE; Abikoff, HB; Newcorn, JH; Molina, BSG; Hinshaw, SP; Wells, KC; Hoza, B; Jensen, PS; Gibbons, RD; Hur, K; Stehli, A; Davies, M; March, JS; Conners, CK; Caron, M; Volkow, ND
MLA Citation
Swanson, JM, Elliott, GR, Greenhill, LL, Wigal, T, Arnold, LE, Vitiello, B, Hechtman, L, Epstein, JN, Pelham, WE, Abikoff, HB, Newcorn, JH, Molina, BSG, Hinshaw, SP, Wells, KC, Hoza, B, Jensen, PS, Gibbons, RD, Hur, K, Stehli, A, Davies, M, March, JS, Conners, CK, Caron, M, and Volkow, ND. "Effects of stimulant medication on growth rates across 3 years in the MTA follow-up." J Am Acad Child Adolesc Psychiatry 46.8 (August 2007): 1015-1027.
PMID
17667480
Source
pubmed
Published In
Journal of the American Academy of Child and Adolescent Psychiatry
Volume
46
Issue
8
Publish Date
2007
Start Page
1015
End Page
1027
DOI
10.1097/chi.0b013e3180686d7e

N-terminal tyrosine modulation of the endocytic adaptor function of the beta-arrestins.

The highly homologous beta-arrestin1 and -2 adaptor proteins play important roles in the function of G protein-coupled receptors. Either beta-arrestin variant can function as a molecular chaperone for clathrin-mediated receptor internalization. This role depends primarily upon two distinct, contiguous C-terminal beta-arrestin motifs recognizing clathrin and the beta-adaptin subunit of AP2. However, a molecular basis is lacking to explain the different endocytic efficacies of the two beta-arrestin isoforms and the observation that beta-arrestin N-terminal substitution mutants can act as dominant negative inhibitors of receptor endocytosis. Despite the near identity of the beta-arrestins throughout their N termini, sequence variability is present at a small number of residues and includes tyrosine to phenylalanine substitutions. Here we show that corresponding N-terminal (Y/F)VTL sequences in beta-arrestin1 and -2 differentially regulate mu-adaptin binding. Our results indicate that the beta-arrestin1 Tyr-54 lessens the interaction with mu-adaptin and moreover is a Src phosphorylation site. A gain of endocytic function is obtained with the beta-arrestin1 Y54F substitution, which improves both the beta-arrestin1 interaction with mu-adaptin and the ability to enhance beta2-adrenergic receptor internalization. These data indicate that beta-arrestin2 utilizes mu-adaptin as an endocytic partner, and that the inability of beta-arrestin1 to sustain a similar degree of interaction with mu-adaptin may result from coordination of Tyr-54 by neighboring residues or its modification by Src kinase. Additionally, these naturally occurring variations in beta-arrestins may also differentially regulate the composition of the signaling complexes organized on the receptor.

Authors
Marion, S; Fralish, GB; Laporte, S; Caron, MG; Barak, LS
MLA Citation
Marion, S, Fralish, GB, Laporte, S, Caron, MG, and Barak, LS. "N-terminal tyrosine modulation of the endocytic adaptor function of the beta-arrestins." J Biol Chem 282.26 (June 29, 2007): 18937-18944.
PMID
17456469
Source
pubmed
Published In
The Journal of biological chemistry
Volume
282
Issue
26
Publish Date
2007
Start Page
18937
End Page
18944
DOI
10.1074/jbc.M700090200

Dopamine levels modulate the updating of tastant values.

To survive, animals must constantly update the internal value of stimuli they encounter; a process referred to as incentive learning. Although there have been many studies investigating whether dopamine is necessary for reward, or for the association between stimuli and actions with rewards, less is known about the role of dopamine in the updating of the internal value of stimuli per se. We used a single-bottle forced-choice task to investigate the role of dopamine in learning the value of tastants. We show that dopamine transporter knock-out mice (DAT-KO), which have constitutively elevated dopamine levels, develop a more positive bias towards a hedonically positive tastant (sucrose 400 mM) than their wild-type littermates. Furthermore, when compared to wild-type littermates, DAT-KO mice develop a less negative bias towards a hedonically negative tastant (quinine HCl 10 mM). Importantly, these effects develop with training, because at the onset of training DAT-KO and wild-type mice display similar biases towards sucrose and quinine. These data suggest that dopamine levels can modulate the updating of tastant values, a finding with implications for understanding sensory-specific motivation and reward seeking.

Authors
Costa, RM; Gutierrez, R; de Araujo, IE; Coelho, MRP; Kloth, AD; Gainetdinov, RR; Caron, MG; Nicolelis, MAL; Simon, SA
MLA Citation
Costa, RM, Gutierrez, R, de Araujo, IE, Coelho, MRP, Kloth, AD, Gainetdinov, RR, Caron, MG, Nicolelis, MAL, and Simon, SA. "Dopamine levels modulate the updating of tastant values." Genes Brain Behav 6.4 (June 2007): 314-320.
PMID
16848782
Source
pubmed
Published In
Genes, Brain and Behavior
Volume
6
Issue
4
Publish Date
2007
Start Page
314
End Page
320
DOI
10.1111/j.1601-183X.2006.00257.x

A murine model of hyperdopaminergic state displays altered respiratory control.

The dopamine transporter (DAT) protein plays an important role in the termination of dopamine signaling. We addressed the hypothesis that loss of DAT function would result in a distinctive cardiorespiratory phenotype due to the significant role of dopamine in the control of breathing, especially with respect to chemical control, metabolism, and thermoregulation. The DAT knockout mouse (DAT-/-) displays a state of functional hyperdopaminergia characterized by marked novelty driven hyperactivity. Certain behavioral and drug responses in these mice are reminiscent of endophenotypes of individuals with attention deficit hyperactivity disorders (ADHD). We performed experiments on conscious, unrestrained DAT-/- mice (KO) and littermate DAT+/+ wild-type (WT) controls. Ventilation was measured by the barometric technique during normoxia, hypoxia, or hypercapnia. We measured core body temperature and CO2 production as an index of metabolism. DAT-/- mice displayed a significantly lower respiratory frequency than WT mice, reflecting a prolonged inspiratory time. DAT-/- mice exhibited a reduced ventilatory response to hypoxia characterized by an attenuation of both the respiratory frequency and tidal volume responses. Both groups showed similar metabolic responses to hypoxia. Circadian measurements of body temperature were significantly lower in DAT-/- mice than WT mice during inactive periods. We conclude that loss of the DAT protein in this murine model of altered dopaminergic neurotransmission results in a significant respiratory and thermal phenotype that has possible implications for understanding of conditions associated with altered dopamine regulation.

Authors
Vincent, SG; Waddell, AE; Caron, MG; Walker, JKL; Fisher, JT
MLA Citation
Vincent, SG, Waddell, AE, Caron, MG, Walker, JKL, and Fisher, JT. "A murine model of hyperdopaminergic state displays altered respiratory control." FASEB J 21.7 (May 2007): 1463-1471.
PMID
17255472
Source
pubmed
Published In
The FASEB journal : official publication of the Federation of American Societies for Experimental Biology
Volume
21
Issue
7
Publish Date
2007
Start Page
1463
End Page
1471
DOI
10.1096/fj.06-7248com

The Akt-GSK-3 signaling cascade in the actions of dopamine.

Drugs that act on dopamine neurotransmission are important tools for the management of multiple neuropsychiatric disorders. Classically, dopamine receptors have been shown to regulate cAMP-PKA (protein kinase A) and Ca(2+) pathways through G-protein-mediated signaling. However, it has become apparent that, in addition to this canonical action, D(2)-class dopamine receptors can function through a protein kinase B (Akt)-GSK-3 (glycogen synthase kinase 3) signaling cascade. This novel signaling mode involves the multifunctional scaffolding protein beta-arrestin 2, which has a role in G-protein-coupled receptor (GPCR) desensitization. In this article, we provide an overview of how this dual function of components of the GPCR desensitization machinery relates to dopamine-receptor-mediated responses and we summarize recent insights into the relevance of the Akt-GSK-3 signaling cascade for the expression of dopamine-associated behaviors and the actions of dopaminergic drugs.

Authors
Beaulieu, J-M; Gainetdinov, RR; Caron, MG
MLA Citation
Beaulieu, J-M, Gainetdinov, RR, and Caron, MG. "The Akt-GSK-3 signaling cascade in the actions of dopamine." Trends Pharmacol Sci 28.4 (April 2007): 166-172. (Review)
PMID
17349698
Source
pubmed
Published In
Trends in Pharmacological Sciences
Volume
28
Issue
4
Publish Date
2007
Start Page
166
End Page
172
DOI
10.1016/j.tips.2007.02.006

Regulation of Akt signaling by D2 and D3 dopamine receptors in vivo.

The serine/threonine kinase Akt is a downstream target of dopamine receptor signaling that is inhibited/dephosphorylated in response to direct and indirect dopamine receptor agonists. Although pharmacological studies uncovered the involvement of D2-class dopamine receptors in Akt regulation, they did not identify the role of individual receptor subtypes in this process. Here we used knock-out mice lacking the D1, D2, D2 long, or D3 dopamine receptors as well as a D4 receptor-selective antagonist to address the function of each of these receptors in the regulation of Akt in vivo. Under basal conditions, D2, D2 long, and D3 knock-out mice display enhanced striatal Akt activation, whereas D1 knock-out mice and mice treated with the D4 receptor antagonist L745870 (3-[[4-(4-chlorophenyl)piperazin-1-yl]methyl]-1H-pyrrolo[2,3-b]pyridine trihydrochloride) have phospho-Akt levels comparable with those of normal control animals. Furthermore, both amphetamine and apomorphine lose their ability to inhibit Akt in D2 knock-out mice but retain their normal effect on this signaling molecule in D1 knock-out animals. Finally, D3 knock-out mice show a reduced sensitivity of Akt-mediated signaling to dopaminergic drugs but retain the action of these drugs on Akt at high dose regimens. These results indicate that D2 receptors are essential for the inhibition of Akt by dopamine and that D3 receptors also participate in this signaling potentially by enhancing D2 receptor response. Identification of the functions of individual dopamine receptor subtypes in Akt regulation may help the development of new pharmaceutical approaches for mental disorders related to abnormal dopamine transmission such as bipolar disorder and schizophrenia.

Authors
Beaulieu, J-M; Tirotta, E; Sotnikova, TD; Masri, B; Salahpour, A; Gainetdinov, RR; Borrelli, E; Caron, MG
MLA Citation
Beaulieu, J-M, Tirotta, E, Sotnikova, TD, Masri, B, Salahpour, A, Gainetdinov, RR, Borrelli, E, and Caron, MG. "Regulation of Akt signaling by D2 and D3 dopamine receptors in vivo." J Neurosci 27.4 (January 24, 2007): 881-885.
PMID
17251429
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
27
Issue
4
Publish Date
2007
Start Page
881
End Page
885
DOI
10.1523/JNEUROSCI.5074-06.2007

Local knockdown of genes in the brain using small interfering RNA: a phenotypic comparison with knockout animals.

BACKGROUND: Recent reports have suggested effectiveness of RNA interference (RNAi) for the analysis of gene functions in the brain. This study sought to determine the efficiency of local small interfering RNA (siRNA) injections, comparing this approach with animals generated through classical gene targeting. METHODS: Small interfering RNA against dopamine transporter (DAT) (35 microg/14 days) or tyrosine hydroxylase (TH) (15 microg/3 days) was injected into the ventral tegmental/substantia nigra areas of the brain of adult wildtype or DAT-knockout mice, respectively. RESULTS: Local injections of siRNA resulted in a 35% to 40% reduction of DAT and TH protein levels in the striatum, respectively. Despite negligible effect of DAT knockdown on novelty-induced locomotion, the locomotor response of DAT siRNA treated animals to amphetamine was blunted similar to what is observed in the DAT heterozygote animals. Since incomplete reduction of TH levels in normal mice does not produce behavioral effects, TH siRNA experiments were carried out in DAT-knockout animals that show increased dependence on newly synthesized dopamine. Knockdown of TH in these animals resulted in reduced basal locomotion. CONCLUSIONS: Local injection of siRNA in the brain reduced gene expression by 40% to 50%, suggesting that siRNA-mediated knockdown of genes in the brain can be a complementary tool to classical transgenesis for the analysis of gene functions.

Authors
Salahpour, A; Medvedev, IO; Beaulieu, J-M; Gainetdinov, RR; Caron, MG
MLA Citation
Salahpour, A, Medvedev, IO, Beaulieu, J-M, Gainetdinov, RR, and Caron, MG. "Local knockdown of genes in the brain using small interfering RNA: a phenotypic comparison with knockout animals." Biol Psychiatry 61.1 (January 1, 2007): 65-69.
PMID
16712807
Source
pubmed
Published In
Biological Psychiatry
Volume
61
Issue
1
Publish Date
2007
Start Page
65
End Page
69
DOI
10.1016/j.biopsych.2006.03.020

Roles of protein kinase C and actin-binding protein 280 in the regulation of intracellular trafficking of dopamine D3 receptor

D3 dopamine receptor (D3R) is expressed mainly in parts of the brain that control the emotional behaviors. It is believed that the improper regulation of D3R is involved in the etiology of schizophrenia. Desensitization of D3R is weakly associated with G protein-coupled receptor kinase (GRK)/β-arrestin-directed internalization. This suggests that there might be an alternative pathway that regulates D 3R signaling. This report shows that D3R undergoes robust protein kinase C (PKC)-dependent sequestration that is accompanied by receptor phosphorylation and the desensitization of signaling. PKC-dependent D 3R sequestration, which was enhanced by PKC-β or -δ, was dynamin dependent but independent of GRK, β-arrestin, or caveolin 1. Site-directed mutagenesis of all possible phosphorylation sites within the intracellular loops of D3R identified serine residues at positions 229 and 257 as the critical amino acids responsible for phorbol-12-myristate-13- acetate (PMA)-induced D3R phosphorylation, sequestration, and desensitization. In addition, the LxxY endocytosis motif, which is located between residues 252 and 255, was found to play accommodating roles for PMA-induced D3R sequestration. A continuous interaction with the actin-binding protein 280 (filamin A), which was previously known to interact with D3R, is required for PMA-induced D3R sequestration. In conclusion, the PKC-dependent but GRK-/β-arrestin-independent phosphorylation of D3R is the main pathway responsible for the sequestration and desensitization of D3R. Filamin A is essential for both the efficient signaling and sequestration of D3R. Copyright © 2007 by The Endocrine Society.

Authors
Cho, E-Y; Cho, D-I; Park, JH; Kurose, H; Caron, MG; Kim, K-M
MLA Citation
Cho, E-Y, Cho, D-I, Park, JH, Kurose, H, Caron, MG, and Kim, K-M. "Roles of protein kinase C and actin-binding protein 280 in the regulation of intracellular trafficking of dopamine D3 receptor." Molecular Endocrinology 21.9 (2007): 2242-2254.
PMID
17536008
Source
scival
Published In
Molecular endocrinology (Baltimore, Md.)
Volume
21
Issue
9
Publish Date
2007
Start Page
2242
End Page
2254
DOI
10.1210/me.2007-0202

Inhibition of the dopamine D1 receptor signaling by PSD-95

Dopamine D1 receptors play an important role in movement, reward, and learning and are implicated in a number of neurological and psychiatric disorders. These receptors are concentrated in dendritic spines of neurons, including the spine head and the postsynaptic density. D1 within spines is thought to modulate the local channels and receptors to control the excitability and synaptic properties of spines. The molecular mechanisms mediating D1 trafficking, anchorage, and function in spines remain elusive. Here we show that the synaptic scaffolding protein PSD-95 thought to play a role in stabilizing glutamate receptors in the postsynaptic density, interacts with D1 and regulates its trafficking and function. Interestingly, the D1-PSD-95 interaction does not require the well characterized domains of PSD-95 but is mediated by the carboxyl-terminal tail of D1 and the NH2 terminus of PSD-95, a region that is recognized only recently to participate in protein-protein interaction. Co-expression of PSD-95 with D1 in mammalian cells inhibits the D1-mediated cAMP accumulation without altering the total expression level or the agonist binding properties of the receptor. The diminished D1 signaling is mediated by reduced D1 expression at the cell surface as a consequence of an enhanced constitutive, dynamin-dependent endocytosis. In addition, genetically engineered mice lacking PSD-95 show a heightened behavioral response to either a D1 agonist or the psychostimulant amphetamine. These studies demonstrate a role for a glutamatergic scaffold in dopamine receptor signaling and trafficking and identify a new potential target for the modulation of abnormal dopaminergic function. © 2007 by The American Society for Biochemistry and Molecular Biology, Inc.

Authors
Zhang, J; Vinuela, A; Neely, MH; Hallett, PJ; Grant, SGN; Miller, GM; Isacson, O; Caron, MG; Yao, W-D
MLA Citation
Zhang, J, Vinuela, A, Neely, MH, Hallett, PJ, Grant, SGN, Miller, GM, Isacson, O, Caron, MG, and Yao, W-D. "Inhibition of the dopamine D1 receptor signaling by PSD-95." Journal of Biological Chemistry 282.21 (2007): 15778-15789.
PMID
17369255
Source
scival
Published In
The Journal of biological chemistry
Volume
282
Issue
21
Publish Date
2007
Start Page
15778
End Page
15789
DOI
10.1074/jbc.M611485200

Dopamine enhances motor and neuropathological consequences of polyglutamine expanded huntingtin.

An expansion in the CAG repeat of the IT15 (huntingtin) gene underlies the development of Huntington's disease (HD), but the basis for the specific vulnerability of dopamine-receptive striatal neurons remains unclear. To examine the potential role of the dopamine system in the emergence of pathological conditions in HD, we generated a double mutant mouse strain with both enhanced dopamine transmission and endogenous expression of a mutant huntingtin gene. This strain was generated by crossing the dopamine transporter knock-out mouse, which exhibits a 5-fold elevation in extracellular dopamine levels in the striatum and locomotor hyperactivity, to a knock-in mouse model of HD containing 92 CAG repeats. These double mutant mice exhibited an increased stereotypic activity at 6 months of age, followed by a progressive decline of their locomotor hyperactivity. Expression of the mutated huntingtin did not alter dopamine or its metabolite levels in normal or dopamine transporter knock-out mice. However, the mutant huntingtin protein aggregated much earlier and to a greater extent in the striatum and other dopaminergic brain regions in the hyperdopaminergic mouse model of HD. Furthermore, the formation of neuropil aggregates in the striatum and other regions of hyperdopaminergic HD mice was observed at 4 months of age, well before similar events occurred in normal HD mice (12 months). These findings indicate that dopamine contributes to the deleterious effects of mutated huntingtin on striatal function, and this is accompanied by enhanced formation of huntingtin aggregates.

Authors
Cyr, M; Sotnikova, TD; Gainetdinov, RR; Caron, MG
MLA Citation
Cyr, M, Sotnikova, TD, Gainetdinov, RR, and Caron, MG. "Dopamine enhances motor and neuropathological consequences of polyglutamine expanded huntingtin." FASEB J 20.14 (December 2006): 2541-2543.
PMID
17065224
Source
pubmed
Published In
The FASEB journal : official publication of the Federation of American Societies for Experimental Biology
Volume
20
Issue
14
Publish Date
2006
Start Page
2541
End Page
2543
DOI
10.1096/fj.06-6533fje

Paradoxical striatal cellular signaling responses to psychostimulants in hyperactive mice.

Recent investigations have shown that three major striatal-signaling pathways (protein kinase A/DARPP-32, Akt/glycogen synthase kinase 3, and ERK) are involved in the regulation of locomotor activity by the monoaminergic neurotransmitter dopamine. Here we used dopamine transporter knock-out mice to examine which particular changes in the regulation of these cell signaling mechanisms are associated with distinct behavioral responses to psychostimulants. In normal animals, amphetamine and methylphenidate increase extracellular levels of dopamine, leading to an enhancement of locomotor activity. However, in dopamine transporter knock-out mice that display a hyperactivity phenotype resulting from a persistent hyperdopaminergic state, these drugs antagonize hyperactivity. Under basal conditions, dopamine transporter knock-out mice show enhanced striatal DARPP-32 phosphorylation, activation of ERK, and inactivation of Akt as compared with wild-type littermates. However, administration of amphetamine or methylphenidate to these mice reveals that inhibition of ERK signaling is a common determinant for the ability of these drugs to antagonize hyperactivity. In contrast, psychostimulants activate ERK and induce hyperactivity in normal animals. In hyperactive mice psychostimulant-mediated behavioral inhibition and ERK regulation are also mimicked by the serotonergic drugs fluoxetine and 5-carboxamidotryptamine, thereby revealing the involvement of serotonin-dependent inhibition of striatal ERK signaling. Furthermore, direct inhibition of the ERK signaling cascade in vivo using the MEK inhibitor SL327 recapitulates the actions of psychostimulants in hyperactive mice and prevents the locomotor-enhancing effects of amphetamine in normal animals. These data suggest that the inhibitory action of psychostimulants on dopamine-dependent hyperactivity results from altered regulation of striatal ERK signaling. In addition, these results illustrate how altered homeostatic state of neurotransmission can influence in vivo signaling responses and biological actions of pharmacological agents used to manage psychiatric conditions such as Attention Deficit Hyperactivity Disorder (ADHD).

Authors
Beaulieu, J-M; Sotnikova, TD; Gainetdinov, RR; Caron, MG
MLA Citation
Beaulieu, J-M, Sotnikova, TD, Gainetdinov, RR, and Caron, MG. "Paradoxical striatal cellular signaling responses to psychostimulants in hyperactive mice." J Biol Chem 281.43 (October 27, 2006): 32072-32080.
PMID
16954211
Source
pubmed
Published In
The Journal of biological chemistry
Volume
281
Issue
43
Publish Date
2006
Start Page
32072
End Page
32080
DOI
10.1074/jbc.M606062200

Rapid alterations in corticostriatal ensemble coordination during acute dopamine-dependent motor dysfunction.

Dopaminergic dysregulation can cause motor dysfunction, but the mechanisms underlying dopamine-related motor disorders remain under debate. We used an inducible and reversible pharmacogenetic approach in dopamine transporter knockout mice to investigate the simultaneous activity of neuronal ensembles in the dorsolateral striatum and primary motor cortex during hyperdopaminergia ( approximately 500% of controls) with hyperkinesia, and after rapid and profound dopamine depletion (<0.2%) with akinesia in the same animal. Surprisingly, although most cortical and striatal neurons ( approximately 70%) changed firing rate during the transition between dopamine-related hyperkinesia and akinesia, the overall cortical firing rate remained unchanged. Conversely, neuronal oscillations and ensemble activity coordination within and between cortex and striatum did change rapidly between these periods. During hyperkinesia, corticostriatal activity became largely asynchronous, while during dopamine-depletion the synchronicity increased. Thus, dopamine-related disorders like Parkinson's disease may not stem from changes in the overall levels of cortical activity, but from dysfunctional activity coordination in corticostriatal circuits.

Authors
Costa, RM; Lin, S-C; Sotnikova, TD; Cyr, M; Gainetdinov, RR; Caron, MG; Nicolelis, MAL
MLA Citation
Costa, RM, Lin, S-C, Sotnikova, TD, Cyr, M, Gainetdinov, RR, Caron, MG, and Nicolelis, MAL. "Rapid alterations in corticostriatal ensemble coordination during acute dopamine-dependent motor dysfunction." Neuron 52.2 (October 19, 2006): 359-369.
PMID
17046697
Source
pubmed
Published In
Neuron
Volume
52
Issue
2
Publish Date
2006
Start Page
359
End Page
369
DOI
10.1016/j.neuron.2006.07.030

Dopaminergic control of sleep-wake states.

Dopamine depletion is involved in the pathophysiology of Parkinson's disease, whereas hyperdopaminergia may play a fundamental role in generating endophenotypes associated with schizophrenia. Sleep disturbances are known to occur in both schizophrenia and Parkinson's disease, suggesting that dopamine plays a role in regulating the sleep-wake cycle. Here, we show that novelty-exposed hyperdopaminergic mice enter a novel awake state characterized by spectral patterns of hippocampal local field potentials that resemble electrophysiological activity observed during rapid-eye-movement (REM) sleep. Treatment with haloperidol, a D2 dopamine receptor antagonist, reduces this abnormal intrusion of REM-like activity during wakefulness. Conversely, mice acutely depleted of dopamine enter a different novel awake state characterized by spectral patterns of hippocampal local field potentials that resemble electrophysiological activity observed during slow-wave sleep (SWS). This dopamine-depleted state is marked by an apparent suppression of SWS and a complete suppression of REM sleep. Treatment with D2 (but not D1) dopamine receptor agonists recovers REM sleep in these mice. Altogether, these results indicate that dopamine regulates the generation of sleep-wake states. We propose that psychosis and the sleep disturbances experienced by Parkinsonian patients result from dopamine-mediated disturbances of REM sleep.

Authors
Dzirasa, K; Ribeiro, S; Costa, R; Santos, LM; Lin, S-C; Grosmark, A; Sotnikova, TD; Gainetdinov, RR; Caron, MG; Nicolelis, MAL
MLA Citation
Dzirasa, K, Ribeiro, S, Costa, R, Santos, LM, Lin, S-C, Grosmark, A, Sotnikova, TD, Gainetdinov, RR, Caron, MG, and Nicolelis, MAL. "Dopaminergic control of sleep-wake states." J Neurosci 26.41 (October 11, 2006): 10577-10589.
PMID
17035544
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
26
Issue
41
Publish Date
2006
Start Page
10577
End Page
10589
DOI
10.1523/JNEUROSCI.1767-06.2006

Smoothened signal transduction is promoted by G protein-coupled receptor kinase 2.

Deregulation of the Sonic hedgehog pathway has been implicated in an increasing number of human cancers. In this pathway, the seven-transmembrane (7TM) signaling protein Smoothened regulates cellular proliferation and differentiation through activation of the transcription factor Gli. The activity of mammalian Smoothened is controlled by three different hedgehog proteins, Indian, Desert, and Sonic hedgehog, through their interaction with the Smoothened inhibitor Patched. However, the mechanisms of signal transduction from Smoothened are poorly understood. We show that a kinase which regulates signaling by many "conventional" 7TM G-protein-coupled receptors, G protein-coupled receptor kinase 2 (GRK2), participates in Smoothened signaling. Expression of GRK2, but not catalytically inactive GRK2, synergizes with active Smoothened to mediate Gli-dependent transcription. Moreover, knockdown of endogenous GRK2 by short hairpin RNA (shRNA) significantly reduces signaling in response to the Smoothened agonist SAG and also inhibits signaling induced by an oncogenic Smoothened mutant, Smo M2. We find that GRK2 promotes the association between active Smoothened and beta-arrestin 2. Indeed, Gli-dependent signaling, mediated by coexpression of Smoothened and GRK2, is diminished by beta-arrestin 2 knockdown with shRNA. Together, these data suggest that GRK2 plays a positive role in Smoothened signaling, at least in part, through the promotion of an association between beta-arrestin 2 and Smoothened.

Authors
Meloni, AR; Fralish, GB; Kelly, P; Salahpour, A; Chen, JK; Wechsler-Reya, RJ; Lefkowitz, RJ; Caron, MG
MLA Citation
Meloni, AR, Fralish, GB, Kelly, P, Salahpour, A, Chen, JK, Wechsler-Reya, RJ, Lefkowitz, RJ, and Caron, MG. "Smoothened signal transduction is promoted by G protein-coupled receptor kinase 2." Mol Cell Biol 26.20 (October 2006): 7550-7560.
Website
http://hdl.handle.net/10161/7792
PMID
16908539
Source
pubmed
Published In
Molecular and Cellular Biology
Volume
26
Issue
20
Publish Date
2006
Start Page
7550
End Page
7560
DOI
10.1128/MCB.00546-06

Dopamine: from pharmacology to molecular biology and back.

Authors
Caron, MG; Gainetdinov, RR
MLA Citation
Caron, MG, and Gainetdinov, RR. "Dopamine: from pharmacology to molecular biology and back." Wien Klin Wochenschr 118.19-20 (October 2006): 565-568.
PMID
17136328
Source
pubmed
Published In
Wiener klinische Wochenschrift
Volume
118
Issue
19-20
Publish Date
2006
Start Page
565
End Page
568
DOI
10.1007/s00508-006-0735-6

Mice deficient for the vesicular acetylcholine transporter are myasthenic and have deficits in object and social recognition.

An important step for cholinergic transmission involves the vesicular storage of acetylcholine (ACh), a process mediated by the vesicular acetylcholine transporter (VAChT). In order to understand the physiological roles of the VAChT, we developed a genetically altered strain of mice with reduced expression of this transporter. Heterozygous and homozygous VAChT knockdown mice have a 45% and 65% decrease in VAChT protein expression, respectively. VAChT deficiency alters synaptic vesicle filling and affects ACh release. Whereas VAChT homozygous mutant mice demonstrate major neuromuscular deficits, VAChT heterozygous mice appear normal in that respect and could be used for analysis of central cholinergic function. Behavioral analyses revealed that aversive learning and memory are not altered in mutant mice; however, performance in cognitive tasks involving object and social recognition is severely impaired. These observations suggest a critical role of VAChT in the regulation of ACh release and physiological functions in the peripheral and central nervous system.

Authors
Prado, VF; Martins-Silva, C; de Castro, BM; Lima, RF; Barros, DM; Amaral, E; Ramsey, AJ; Sotnikova, TD; Ramirez, MR; Kim, H-G; Rossato, JI; Koenen, J; Quan, H; Cota, VR; Moraes, MFD; Gomez, MV; Guatimosim, C; Wetsel, WC; Kushmerick, C; Pereira, GS; Gainetdinov, RR; Izquierdo, I; Caron, MG; Prado, MAM
MLA Citation
Prado, VF, Martins-Silva, C, de Castro, BM, Lima, RF, Barros, DM, Amaral, E, Ramsey, AJ, Sotnikova, TD, Ramirez, MR, Kim, H-G, Rossato, JI, Koenen, J, Quan, H, Cota, VR, Moraes, MFD, Gomez, MV, Guatimosim, C, Wetsel, WC, Kushmerick, C, Pereira, GS, Gainetdinov, RR, Izquierdo, I, Caron, MG, and Prado, MAM. "Mice deficient for the vesicular acetylcholine transporter are myasthenic and have deficits in object and social recognition." Neuron 51.5 (September 7, 2006): 601-612.
PMID
16950158
Source
pubmed
Published In
Neuron
Volume
51
Issue
5
Publish Date
2006
Start Page
601
End Page
612
DOI
10.1016/j.neuron.2006.08.005

A beta-arrestin binding determinant common to the second intracellular loops of rhodopsin family G protein-coupled receptors.

beta-Arrestins have been shown to inhibit competitively G protein-dependent signaling and to mediate endocytosis for many of the hundreds of nonvisual rhodopsin family G protein-coupled receptors (GPCR). An open question of fundamental importance concerning the regulation of signal transduction of several hundred rhodopsin-like GPCRs is how these receptors of limited sequence homology, when considered in toto, can all recruit and activate the two highly conserved beta-arrestin proteins as part of their signaling/desensitization process. Although the serine and threonine residues that form GPCR kinase phosphorylation sites are common beta-arrestin-associated receptor determinants regulating receptor desensitization and internalization, the agonist-activated conformation of a GPCR probably reveals the most fundamental determinant mediating the GPCR and arrestin interaction. Here we identified a beta-arrestin binding determinant common to the rhodopsin family GPCRs formed from the proximal 10 residues of the second intracellular loop. We demonstrated by both gain and loss of function studies for the serotonin 2C, beta2-adrenergic, alpha2a)adrenergic, and neuropeptide Y type 2 receptors that the highly conserved amino acids, proline and alanine, naturally occurring in rhodopsin family receptors six residues distal to the highly conserved second loop DRY motif regulate beta-arrestin binding and beta-arrestin-mediated internalization. In particular, as demonstrated for the beta2 AR, this occurs independently of changes in GPCR kinase phosphorylation. These results suggest that a GPCR conformation directed by the second intracellular loop, likely using the loop itself as a binding patch, may function as a switch for transitioning beta-arrestin from its inactive form to its active receptor-binding state.

Authors
Marion, S; Oakley, RH; Kim, K-M; Caron, MG; Barak, LS
MLA Citation
Marion, S, Oakley, RH, Kim, K-M, Caron, MG, and Barak, LS. "A beta-arrestin binding determinant common to the second intracellular loops of rhodopsin family G protein-coupled receptors." J Biol Chem 281.5 (February 3, 2006): 2932-2938.
PMID
16319069
Source
pubmed
Published In
The Journal of biological chemistry
Volume
281
Issue
5
Publish Date
2006
Start Page
2932
End Page
2938
DOI
10.1074/jbc.M508074200

Molecular biology, pharmacology and functional role of the plasma membrane dopamine transporter.

The plasma membrane dopamine transporter (DAT) tightly regulates the extracellular concentrations of dopamine (DA) by re-capturing released neurotransmitter back into the presynaptic neuronal terminals and/or neighboring DA projections thereby providing an effective way to regulate synaptic and extrasynaptic DA levels. This transporter is a primary target of many potent psychotropic drugs and neurotoxins, such as cocaine, amphetamines and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). In this review we summarize recent advances in understanding the structure, regulation, and functional roles of DAT in normal DA physiology and pathological conditions, such as attention deficit hyperactivity disorder (ADHD) and neurodegenerative processes, as well as their contribution to the pharmacology of psychostimulant drugs. Significant new insights on these issues have been gained using mice with genetic deletion of DAT.

Authors
Sotnikova, TD; Beaulieu, J-M; Gainetdinov, RR; Caron, MG
MLA Citation
Sotnikova, TD, Beaulieu, J-M, Gainetdinov, RR, and Caron, MG. "Molecular biology, pharmacology and functional role of the plasma membrane dopamine transporter." CNS Neurol Disord Drug Targets 5.1 (February 2006): 45-56. (Review)
PMID
16613553
Source
pubmed
Published In
CNS and Neurological Disorders - Drug Targets
Volume
5
Issue
1
Publish Date
2006
Start Page
45
End Page
56

Functional polymorphisms of the brain serotonin synthesizing enzyme tryptophan hydroxylase-2.

Many neuropsychiatric disorders are considered to be related to the dysregulation of brain serotonergic neurotransmission. Tryptophan hydroxylase-2 (TPH2) is the neuronal-specific enzyme that controls brain serotonin synthesis. There is growing genetic evidence for the possible involvement of TPH2 in serotonin-related neuropsychiatric disorders; however, the degree of genetic variation in TPH2 and, in particular, its possible functional consequences remain unknown. In this short review, we will summarize some recent findings with respect to the functional analysis of TPH2.

Authors
Zhang, X; Beaulieu, J-M; Gainetdinov, RR; Caron, MG
MLA Citation
Zhang, X, Beaulieu, J-M, Gainetdinov, RR, and Caron, MG. "Functional polymorphisms of the brain serotonin synthesizing enzyme tryptophan hydroxylase-2." Cell Mol Life Sci 63.1 (January 2006): 6-11. (Review)
PMID
16378243
Source
pubmed
Published In
Cellular and Molecular Life Sciences
Volume
63
Issue
1
Publish Date
2006
Start Page
6
End Page
11
DOI
10.1007/s00018-005-5417-4

Characterization of the desensitization properties of five dopamine receptor subtypes and alternatively spliced variants of dopamine D2 and D4 receptors

Proper regulation of brain dopaminergic activity is essential for maintaining normal mental functions. In this study, the regulatory properties of five different dopamine receptor subtypes and alternative splicing variants of dopamine D2 and D4 were examined. The stimulation of D1R, D2R, D5R but not D3R, D4R caused the robust translocation of β-arrestin to the plasma membrane. When D1R or D3R were co-expressed with D2R, D1R significantly inhibited the sequestration of D2R, suggesting that the inhibitory effects of D1R on the D2R sequestration could explain the synergistic activity between two receptors. The sequestration of alternatively spliced isoforms of D2R was differently regulated by GRKs and β-arrestins. Three alternative splicing variants of D4R produced a similar level of β-arrestin translocation, and the studies with the deletion mutants of D4R within the third cytoplasmic loop revealed that the regions containing the SH3-binding domains are responsible for the β-arrestin translocation. © 2006 Elsevier Inc. All rights reserved.

Authors
Cho, D-I; Beom, S; Tol, HHMV; Caron, MG; Kim, K-M
MLA Citation
Cho, D-I, Beom, S, Tol, HHMV, Caron, MG, and Kim, K-M. "Characterization of the desensitization properties of five dopamine receptor subtypes and alternatively spliced variants of dopamine D2 and D4 receptors." Biochemical and Biophysical Research Communications 350.3 (2006): 634-640.
PMID
17022946
Source
scival
Published In
Biochemical and Biophysical Research Communications
Volume
350
Issue
3
Publish Date
2006
Start Page
634
End Page
640
DOI
10.1016/j.bbrc.2006.09.090

Dopamine enhances motor and neuropathological consequences of polyglutamine expanded huntingtin.

An expansion in the CAG repeat of the IT15 (huntingtin) gene underlies the development of Huntington's disease (HD), but the basis for the specific vulnerability of dopamine-receptive striatal neurons remains unclear. To examine the potential role of the dopamine system in the emergence of pathological conditions in HD, we generated a double mutant mouse strain with both enhanced dopamine transmission and endogenous expression of a mutant huntingtin gene. This strain was generated by crossing the dopamine transporter knock-out mouse, which exhibits a 5-fold elevation in extracellular dopamine levels in the striatum and locomotor hyperactivity, to a knock-in mouse model of HD containing 92 CAG repeats. These double mutant mice exhibited an increased stereotypic activity at 6 months of age, followed by a progressive decline of their locomotor hyperactivity. Expression of the mutated huntingtin did not alter dopamine or its metabolite levels in normal or dopamine transporter knock-out mice. However, the mutant huntingtin protein aggregated much earlier and to a greater extent in the striatum and other dopaminergic brain regions in the hyperdopaminergic mouse model of HD. Furthermore, the formation of neuropil aggregates in the striatum and other regions of hyperdopaminergic HD mice was observed at 4 months of age, well before similar events occurred in normal HD mice (12 months). These findings indicate that dopamine contributes to the deleterious effects of mutated huntingtin on striatal function, and this is accompanied by enhanced formation of huntingtin aggregates.

Authors
Cyr, M; Sotnikova, TD; Gainetdinov, RR; Caron, MG
MLA Citation
Cyr, M, Sotnikova, TD, Gainetdinov, RR, and Caron, MG. "Dopamine enhances motor and neuropathological consequences of polyglutamine expanded huntingtin." The FASEB journal : official publication of the Federation of American Societies for Experimental Biology 20.14 (2006): 2541-2543.
Source
scival
Published In
The FASEB journal : official publication of the Federation of American Societies for Experimental Biology
Volume
20
Issue
14
Publish Date
2006
Start Page
2541
End Page
2543
DOI
10.1096/fj.06-6533fje

Erratum: Dopaminergic Control of Sleep-Wake States (Journal of Neuroscience (October 11, 2006) (10577-10589))

Authors
Dzirasa, K; Ribeiro, S; Costa, R; Santos, LM; Lin, S-C; Grosmark, A; Sotnikova, TD; Gainetdinov, RR; Caron, MG; Nicolelis, MAL
MLA Citation
Dzirasa, K, Ribeiro, S, Costa, R, Santos, LM, Lin, S-C, Grosmark, A, Sotnikova, TD, Gainetdinov, RR, Caron, MG, and Nicolelis, MAL. "Erratum: Dopaminergic Control of Sleep-Wake States (Journal of Neuroscience (October 11, 2006) (10577-10589))." Journal of Neuroscience 26.47 (2006).
Source
scival
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
26
Issue
47
Publish Date
2006

Effect of antidepressant drugs in mice lacking the norepinephrine transporter

One of the main theories concerning the mechanism of action of antidepressant drugs (ADs) is based on the notion that the neurochemical background of depression involves an impairment of central noradrenergic transmission with a concomitant decrease of the norepinephrine (NE) in the synaptic gap. Many ADs increase synaptic NE availability by inhibition of the reuptake of NE. Using mice lacking NE transporter (NET-/-) we examined their baseline phenotype as well as the response in the forced swim test (FST) and in the tail suspension test (TST) upon treatment with ADs that display different pharmacological profiles. In both tests, the NET-/- mice behaved like wild-type (WT) mice acutely treated with ADs. Autoradiographic studies showed decreased binding of the β-adrenergic ligand [3H]CGP12177 in the cerebral cortex of NET-/- mice, indicating the changes at the level of β-adrenergic receptors similar to those obtained with ADs treatment. The binding of [3H]prazosin to α1-adrenergic receptors in the cerebral cortex of NET -/- mice was also decreased, most probably as an adaptive response to the sustained elevation of extracellular NE levels observed in these mice. A pronounced NET knockout-induced shortening of the immobility time in the TST (by ca 50%) compared to WT mice was not reduced any further by NET-inhibiting ADs such as reboxetine, desipramine, and imipramine. Citalopram, which is devoid of affinity for the NET, exerted a significant reduction of immobility time in the NET-/- mice. In the FST, reboxetine, desipramine, imipramine, and citalopram administered acutely did not reduce any further the immobility time shortened by NET knockout itself (ca 25%); however, antidepressant-like action of repeatedly (7 days) administered desipramine was observed in NET -/- mice, indicating that the chronic presence of this drug may also affect other neurochemical targets involved in the behavioral reactions monitored by this test. From the present study, it may be concluded that mice lacking the NET may represent a good model of some aspects of depression-resistant behavior, paralleled with alterations in the expression of adrenergic receptors, which result as an adaptation to elevated levels of extracellular NE. © 2006 Nature Publishing Group All rights reserved.

Authors
Dziedzicka-Wasylewska, M; Faron-Górecka, A; Kuśmider, M; Drozdowska, E; Rogóz, Z; Siwanowicz, J; Caron, MG; Bönisch, H
MLA Citation
Dziedzicka-Wasylewska, M, Faron-Górecka, A, Kuśmider, M, Drozdowska, E, Rogóz, Z, Siwanowicz, J, Caron, MG, and Bönisch, H. "Effect of antidepressant drugs in mice lacking the norepinephrine transporter." Neuropsychopharmacology 31.11 (2006): 2424-2432.
PMID
16554743
Source
scival
Published In
Neuropsychopharmacology (Nature)
Volume
31
Issue
11
Publish Date
2006
Start Page
2424
End Page
2432
DOI
10.1038/sj.npp.1301064

DDD mice, a novel acute mouse model of Parkinson's disease

We describe the development of a novel animal model of acute severe dopamine (DA) deficiency by using genetically altered mice lacking the DA transporter (DAT-KO mice). In the absence of a DAT-mediated recycling mechanism in these mice, striatal DA concentrations become entirely dependent on its de novo synthesis, and acute pharmacologic inhibition of tyrosine hydroxylase induces transient (up to 16 hours) elimination of brain DA. Dopamine-deficient DAT-KO mice (DDD mice) demonstrate a striking behavioral phenotype manifested as severe akinesia, rigidity, tremor, and ptosis. We propose that DDD mice represent a novel acute model of severe DA deficiency that might be used to identify compounds with potential therapeutic use for the treatment of Parkinson's disease (PD). This model is particularly promising as a tool for evaluating the efficacy of compounds that may induce movement independently of DA. The advantages and limitations of DDD mice in comparison to other rodent PD models are discussed. ©2006AAN Enterprises, Inc.

Authors
Sotnikova, TD; Caron, MG; Gainetdinov, RR
MLA Citation
Sotnikova, TD, Caron, MG, and Gainetdinov, RR. "DDD mice, a novel acute mouse model of Parkinson's disease." Neurology 67.7 SUPPL. 2 (2006): S12-S17.
PMID
17030735
Source
scival
Published In
Neurology
Volume
67
Issue
7 SUPPL. 2
Publish Date
2006
Start Page
S12
End Page
S17

Norepinephrine transporter knockout-induced up-regulation of brain alpha2A/C-adrenergic receptors

The norepinephrine transporter (NET) is responsible for the rapid removal of norepinephrine released from sympathetic neurons; this release is controlled by inhibitory α2-adrenergic receptors (α2ARs). Long-term inhibition of the NET by antidepressants has been reported to change the density and function of pre- and postsynaptic ARs, which may contribute to the antidepressant effects of NET inhibitors such as desipramine. NET-deficient (NET-KO) mice have been described to behave like antidepressant-treated mice. By means of quantitative real-time PCR we show that mRNAs encoding the α2A-adrenergic receptor (α2AAR) and the α2C-adrenergic receptor (α2CAR) are up-regulated in the brainstem, and that α2CAR mRNA is also elevated in the hippocampus and striatum of NET-KO mice. These results were confirmed at the protein level by quantitative autoradiography. The NET-KO mice showed enhanced binding of the selective α2AR antagonist [ 3H]RX821002 in several brain regions. Most robust increases (20-25%) in α2AR expression were observed in the hippocampus and in the striatum. Significant increases (16%) were also seen in the extended amygdala and thalamic structures. In an 'in vivo' test, the α2AR agonist clonidine (0.1 mg/kg) caused a significantly greater reduction of locomotor activity in NET-KO mice than in wild-type mice, showing the relevance of our findings at the functional level. © 2006 International Society for Neurochemistry.

Authors
Gilsbach, R; Faron-Górecka, A; Rogóz, Z; Brüss, M; Caron, MG; Dziedzicka-Wasylewska, M; Bönisch, H
MLA Citation
Gilsbach, R, Faron-Górecka, A, Rogóz, Z, Brüss, M, Caron, MG, Dziedzicka-Wasylewska, M, and Bönisch, H. "Norepinephrine transporter knockout-induced up-regulation of brain alpha2A/C-adrenergic receptors." Journal of Neurochemistry 96.4 (2006): 1111-1120.
PMID
16417582
Source
scival
Published In
Journal of Neurochemistry
Volume
96
Issue
4
Publish Date
2006
Start Page
1111
End Page
1120
DOI
10.1111/j.1471-4159.2005.03598.x

Norepinephrine transporter-deficient mice respond to anxiety producing and fearful environments with bradycardia and hypotension

The study of anxiety and fear involves complex interrelationships between psychiatry and the autonomic nervous system. Altered noradrenergic signaling is linked to certain types of depression and anxiety disorders, and treatment often includes specific transporter blockade. The norepinephrine transporter is crucial in limiting catecholaminergic signaling. Norepinephrine transporter-deficient mice have increased circulating catecholamines and elevated heart rate and blood pressure. We hypothesized, therefore, that reduced norepinephrine clearance would heighten the autonomic cardiovascular response to anxiety and fear. In separate experiments, norepinephrine transporter-deficient (norepinephrine transporter-/-) mice underwent tactile startle and trace fear conditioning to measure hemodynamic responses. A dramatic tachycardia was observed in norepinephrine transporter-/- mice compared with controls following both airpuff or footshock stimuli, and pressure changes were also greater. Interestingly, in contrast to normally elevated home cage levels in norepinephrine transporter-deficient mice, prestimulus heart rate and blood pressure were actually higher in norepinephrine transporter+/+ animals throughout behavioral testing. Upon placement in the behavioral chamber, norepinephrine transporter-deficient mice demonstrated a notable bradycardia and depressor effect that was more pronounced in females. Power spectral analysis indicated an increase in low frequency oscillations of heart rate variability; in mice, suggesting increased parasympathetic tone. Finally, norepinephrine transporter-/- mice exhibited sexual dimorphism in freeze behavior, which was greatest in females. Therefore, while reduced catecholamine clearance amplifies immediate cardiovascular responses to anxiety- or fear-inducing stimuli in norepinephrine transporter-/- mice, norepinephrine transporter deficiency apparently prevents protracted hemodynamic escalation in a fearful environment. Conceivably, chronic norepinephrine transporter blockade with transporter-specific drugs might attenuate recognition of autonomic and somatic distress signals in individuals with anxiety disorders, possibly lessening their behavioral reactivity, and reducing the cardiovascular risk factors associated with persistent emotional arousal. © 2006 IBRO.

Authors
Keller, NR; Diedrich, A; Appalsamy, M; Miller, LC; Caron, MG; McDonald, MP; Shelton, RC; Blakely, RD; Robertson, D
MLA Citation
Keller, NR, Diedrich, A, Appalsamy, M, Miller, LC, Caron, MG, McDonald, MP, Shelton, RC, Blakely, RD, and Robertson, D. "Norepinephrine transporter-deficient mice respond to anxiety producing and fearful environments with bradycardia and hypotension." Neuroscience 139.3 (2006): 931-946.
PMID
16515844
Source
scival
Published In
Neuroscience
Volume
139
Issue
3
Publish Date
2006
Start Page
931
End Page
946
DOI
10.1016/j.neuroscience.2006.01.008

Beta-arrestin goes nuclear.

Arrestins have important roles in the cytoplasm and at the plasma membrane, including the desensitization and internalization of G protein-coupled receptors (GPCRs). In this issue of Cell, Kang et al. (2005) provide evidence that beta-arrestin 1 moves to the nucleus in response to GPCR stimulation, where it regulates gene expression by facilitating histone acetylation at specific gene promoters.

Authors
Beaulieu, J-M; Caron, MG
MLA Citation
Beaulieu, J-M, and Caron, MG. "Beta-arrestin goes nuclear." Cell 123.5 (December 2, 2005): 755-757.
PMID
16325568
Source
pubmed
Published In
Cell
Volume
123
Issue
5
Publish Date
2005
Start Page
755
End Page
757
DOI
10.1016/j.cell.2005.11.010

Novelty seeking and stereotypic activation of behavior in mice with disruption of the Dat1 gene.

Disruption of the dopamine (DA) transporter (Dat1) gene in mice leads to a 50% reduction or complete elimination of Dat1 expression in striatum of respective heterozygous (HZ) and knockout (KO) mice. Compared to wild-type (WT) controls, extracellular DA is increased approximately two- and five-fold in the mutants. Although open field (OF) activity is similar for WT and HZ animals, it is enhanced for KO mice. The purpose of the present investigations was to study spontaneously emitted behaviors and to determine the behavioral and neurochemical mechanisms that may contribute to the hyperactivity of KO animals. Heterozygotes are less anxious than other genotypes and they engage in novelty-seeking behaviors that include increased time spent in the center of the OF, enhanced investigation of objects, and augmented free exploration of a novel environment. By comparison, KO mice display neophobia when initially exposed to novel conditions. Over time the anxiety-like response habituates and behaviors become activated and stereotyped; these responses are unrelated to exploration or novelty seeking. No alterations in extracellular DA levels or tissue contents from several brain regions are detected at the time of stereotypic activation of KO mice. By contrast, this behavior is accompanied by changes in serotonin metabolism in basal ganglia. This feature may contribute to the behavioral inflexibility of KO mice in different experimental contexts. Collectively, these findings suggest that disruption of the Dat1 gene in mice leads to two different phenotypes; one related to anxiety-reducing and novelty seeking, while the other has some homology to disorders with a stereotypical-perseverative spectrum.

Authors
Pogorelov, VM; Rodriguiz, RM; Insco, ML; Caron, MG; Wetsel, WC
MLA Citation
Pogorelov, VM, Rodriguiz, RM, Insco, ML, Caron, MG, and Wetsel, WC. "Novelty seeking and stereotypic activation of behavior in mice with disruption of the Dat1 gene." Neuropsychopharmacology 30.10 (October 2005): 1818-1831.
PMID
15856082
Source
pubmed
Published In
Neuropsychopharmacology (Nature)
Volume
30
Issue
10
Publish Date
2005
Start Page
1818
End Page
1831
DOI
10.1038/sj.npp.1300724

Dopamine-independent locomotor actions of amphetamines in a novel acute mouse model of Parkinson disease.

Brain dopamine is critically involved in movement control, and its deficiency is the primary cause of motor symptoms in Parkinson disease. Here we report development of an animal model of acute severe dopamine deficiency by using mice lacking the dopamine transporter. In the absence of transporter-mediated recycling mechanisms, dopamine levels become entirely dependent on de novo synthesis. Acute pharmacological inhibition of dopamine synthesis in these mice induces transient elimination of striatal dopamine accompanied by the development of a striking behavioral phenotype manifested as severe akinesia, rigidity, tremor, and ptosis. This phenotype can be reversed by administration of the dopamine precursor, L-DOPA, or by nonselective dopamine agonists. Surprisingly, several amphetamine derivatives were also effective in reversing these behavioral abnormalities in a dopamine-independent manner. Identification of dopamine transporter- and dopamine-independent locomotor actions of amphetamines suggests a novel paradigm in the search for prospective anti-Parkinsonian drugs.

Authors
Sotnikova, TD; Beaulieu, J-M; Barak, LS; Wetsel, WC; Caron, MG; Gainetdinov, RR
MLA Citation
Sotnikova, TD, Beaulieu, J-M, Barak, LS, Wetsel, WC, Caron, MG, and Gainetdinov, RR. "Dopamine-independent locomotor actions of amphetamines in a novel acute mouse model of Parkinson disease." PLoS Biol 3.8 (August 2005): e271-.
PMID
16050778
Source
pubmed
Published In
PLoS biology
Volume
3
Issue
8
Publish Date
2005
Start Page
e271
DOI
10.1371/journal.pbio.0030271

An Akt/beta-arrestin 2/PP2A signaling complex mediates dopaminergic neurotransmission and behavior.

Dopamine plays an important role in the etiology of schizophrenia, and D2 class dopamine receptors are the best-established target of antipsychotic drugs. Here we show that D2 class-receptor-mediated Akt regulation involves the formation of signaling complexes containing beta-arrestin 2, PP2A, and Akt. beta-arrestin 2 deficiency in mice results in reduction of dopamine-dependent behaviors, loss of Akt regulation by dopamine in the striatum, and disruption of the dopamine-dependent interaction of Akt with its negative regulator, protein phosphatase 2A. Importantly, canonical cAMP-mediated dopamine-receptor signaling is not inhibited in the absence of beta-arrestin 2. These results demonstrate that, apart from its classical function in receptor desensitization, beta-arrestin 2 also acts as a signaling intermediate through a kinase/phosphatase scaffold. Furthermore, this function of beta-arrestin 2 is important for the expression of dopamine-associated behaviors, thus implicating beta-arrestin 2 as a positive mediator of dopaminergic synaptic transmission and a potential pharmacological target for dopamine-related psychiatric disorders.

Authors
Beaulieu, J-M; Sotnikova, TD; Marion, S; Lefkowitz, RJ; Gainetdinov, RR; Caron, MG
MLA Citation
Beaulieu, J-M, Sotnikova, TD, Marion, S, Lefkowitz, RJ, Gainetdinov, RR, and Caron, MG. "An Akt/beta-arrestin 2/PP2A signaling complex mediates dopaminergic neurotransmission and behavior." Cell 122.2 (July 29, 2005): 261-273.
PMID
16051150
Source
pubmed
Published In
Cell
Volume
122
Issue
2
Publish Date
2005
Start Page
261
End Page
273
DOI
10.1016/j.cell.2005.05.012

Characterization of conditioned place preference to cocaine in congenic dopamine transporter knockout female mice.

RATIONALE: The dopamine transporter (DAT) is thought to play a major role in the rewarding effects of cocaine. Therefore, it is surprising that cocaine reveals conditioned effects in DAT knockout (DAT-KO) mice. OBJECTIVES: To examine these findings further, we obtained complete dose-effect curves for DAT-KO and DAT wild-type (DAT-WT) mice in a cocaine conditioned place preference (CPP) procedure. METHODS: Congenic C57BL6 background female DAT-KO and DAT-WT mice were conditioned in a three-compartment place preference apparatus. Conditioning consisted of three 30-min sessions with cocaine (2.5, 5.0, 10.0, 20.0, or 40.0 mg/kg) and three 30-min sessions with saline. The distribution of time in each choice compartment was determined after each pair of conditioning sessions (one cocaine and one saline session). RESULTS: DAT-WT mice revealed CPP over a wide range of cocaine doses (5.0-40 mg/kg), whereas DAT-KO mice revealed CPP over a more restricted range of doses, with consistent CPP only occurring with 10 mg/kg of cocaine. CONCLUSIONS: CPP for cocaine develops in both DAT-KO and DAT-WT mice; however, the dose range at which CPP develops is much more restricted in DAT-KO mice than in DAT-WT mice. These observations corroborate the significant role of DAT inhibition in cocaine's conditioned effects.

Authors
Medvedev, IO; Gainetdinov, RR; Sotnikova, TD; Bohn, LM; Caron, MG; Dykstra, LA
MLA Citation
Medvedev, IO, Gainetdinov, RR, Sotnikova, TD, Bohn, LM, Caron, MG, and Dykstra, LA. "Characterization of conditioned place preference to cocaine in congenic dopamine transporter knockout female mice." Psychopharmacology (Berl) 180.3 (July 2005): 408-413.
PMID
15719221
Source
pubmed
Published In
Psychopharmacology
Volume
180
Issue
3
Publish Date
2005
Start Page
408
End Page
413
DOI
10.1007/s00213-005-2173-y

Magnetic resonance imaging at microscopic resolution reveals subtle morphological changes in a mouse model of dopaminergic hyperfunction.

Structural abnormalities of the basal ganglia have been documented in several neuropsychiatric conditions associated with dysregulation of the dopamine system. However, the histological nature underlying these changes is largely unknown. Using magnetic resonance imaging at microscopic resolution (MRI, 9.4 T with 43 microm isotropic spatial resolution) and stereological techniques, we have investigated the effect of increased dopamine neurotransmission on brain morphology in mice with elevated extracellular dopamine, the dopamine transporter knockout (DAT-KO) mice. We first demonstrate the usefulness of MRI at microscopic resolution for the accurate identification and measurement of volumes of specific subregions, accounting for less than 0.03% (0.16 mm(3)) of the volume of a mouse brain. Furthermore, the MRI analysis reveals a significantly lower volume (-9%) of the anterior striatum of DAT-KO mice, while the volume of other dopamine-related structures such as the posterior striatum and the substantia nigra pars reticulata is unchanged in comparison to wild type littermates. Stereological analysis performed in the same brains reveals that one important structural factor accounting for this selective change in volume is a reduction of 18% in the absolute number of neuronal cell bodies. The feasibility of assessing accurately small morphological alterations in mouse models, where the molecular and histological pathologies can be easily compared in a controlled manner, provides a paradigm to examine the relevance of selective brain volumetric changes associated with a number of neuropathological conditions.

Authors
Cyr, M; Caron, MG; Johnson, GA; Laakso, A
MLA Citation
Cyr, M, Caron, MG, Johnson, GA, and Laakso, A. "Magnetic resonance imaging at microscopic resolution reveals subtle morphological changes in a mouse model of dopaminergic hyperfunction." Neuroimage 26.1 (May 15, 2005): 83-90.
PMID
15862208
Source
pubmed
Published In
NeuroImage
Volume
26
Issue
1
Publish Date
2005
Start Page
83
End Page
90
DOI
10.1016/j.neuroimage.2005.01.039

Approaches to identify monoamine transporter interacting proteins.

Authors
Torres, GE; Caron, MG
MLA Citation
Torres, GE, and Caron, MG. "Approaches to identify monoamine transporter interacting proteins." J Neurosci Methods 143.1 (April 15, 2005): 63-68. (Review)
PMID
15763137
Source
pubmed
Published In
Journal of Neuroscience Methods
Volume
143
Issue
1
Publish Date
2005
Start Page
63
End Page
68
DOI
10.1016/j.jneumeth.2004.09.012

G protein-coupled receptor kinase regulates dopamine D3 receptor signaling by modulating the stability of a receptor-filamin-beta-arrestin complex. A case of autoreceptor regulation.

In addition to its postsynaptic role, the dopamine D3 receptor (D3R) serves as a presynaptic autoreceptor, where it provides continuous feedback regulation of dopamine release at nerve terminals for processes as diverse as emotional tone and locomotion. D3R signaling ability is supported by an association with filamin (actin-binding protein 280), which localizes the receptor with G proteins in plasma membrane lipid rafts but is not appreciably antagonized in a classical sense by the ligand-mediated activation of G protein-coupled receptor kinases (GRKs) and beta-arrestins. In this study, we investigate GRK-mediated regulation of D3R.filamin complex stability and its effect on D3R.G protein signaling potential. Studies in HEK-293 cells show that in the absence of agonist the D3R immunoprecipitates in a complex containing both filamin A and beta-arrestin2. Moreover, the filamin directly interacts with beta-arrestin2 as assessed by immunoprecipitation and yeast two-hybrid studies. With reductions in basal GRK2/3 activity, an increase in the basal association of filamin A and beta-arrestin2 with D3R is observed. Conversely, increases in the basal GRK2/3 activity result in a reduction in the interaction between the D3R and filamin but a relative increase in the agonist-mediated interaction between beta-arrestin2 and the D3R. Our data suggest that the D3R, filamin A, and beta-arrestin form a signaling complex that is destabilized by agonist- or expression-mediated increases in GRK2/3 activity. These findings provide a novel GRK-based mechanism for regulating D3R signaling potential and provide insight for interpreting D3R autoreceptor behavior.

Authors
Kim, K-M; Gainetdinov, RR; Laporte, SA; Caron, MG; Barak, LS
MLA Citation
Kim, K-M, Gainetdinov, RR, Laporte, SA, Caron, MG, and Barak, LS. "G protein-coupled receptor kinase regulates dopamine D3 receptor signaling by modulating the stability of a receptor-filamin-beta-arrestin complex. A case of autoreceptor regulation." J Biol Chem 280.13 (April 1, 2005): 12774-12780.
PMID
15687500
Source
pubmed
Published In
The Journal of biological chemistry
Volume
280
Issue
13
Publish Date
2005
Start Page
12774
End Page
12780
DOI
10.1074/jbc.M408901200

Loss-of-function mutation in tryptophan hydroxylase-2 identified in unipolar major depression.

Dysregulation of central serotonin neurotransmission has been widely suspected as an important contributor to major depression. Here, we identify a (G1463A) single nucleotide polymorphism (SNP) in the rate-limiting enzyme of neuronal serotonin synthesis, human tryptophan hydroxylase-2 (hTPH2). The functional SNP in hTPH2 replaces the highly conserved Arg441 with His, which results in approximately 80% loss of function in serotonin production when hTPH2 is expressed in PC12 cells. Strikingly, SNP analysis in a cohort of 87 patients with unipolar major depression revealed that nine patients carried the mutant (1463A) allele, while among 219 controls, three subjects carried this mutation. In addition, this functional SNP was not found in a cohort of 60 bipolar disorder patients. Identification of a loss-of-function mutation in hTPH2 suggests that defect in brain serotonin synthesis may represent an important risk factor for unipolar major depression.

Authors
Zhang, X; Gainetdinov, RR; Beaulieu, J-M; Sotnikova, TD; Burch, LH; Williams, RB; Schwartz, DA; Krishnan, KRR; Caron, MG
MLA Citation
Zhang, X, Gainetdinov, RR, Beaulieu, J-M, Sotnikova, TD, Burch, LH, Williams, RB, Schwartz, DA, Krishnan, KRR, and Caron, MG. "Loss-of-function mutation in tryptophan hydroxylase-2 identified in unipolar major depression." Neuron 45.1 (January 6, 2005): 11-16.
PMID
15629698
Source
pubmed
Published In
Neuron
Volume
45
Issue
1
Publish Date
2005
Start Page
11
End Page
16
DOI
10.1016/j.neuron.2004.12.014

Response to Zhang et al. (2005) loss-of-function mutation in tryptophan hydroxylase-2 identified in unipolar major depression. Neuron 45, 11-16 [2] (multiple letters)

Authors
Zhou, Z; Peters, EJ; Hamilton, SP; McMahon, F; Thomas, C; McGrath, PJ; Rush, J; Trivedi, MH; Charney, DS; Roy, A; Wisniewski, S; Lipsky, R; Goldman, D; Bogaert, AVD; Zutter, SD; Heyrman, L; Mendlewicz, J; Adolfsson, R; Broeckhoven, CV; Del-Favero, J; Glatt, CE; Carlson, E; Taylor, TR; Risch, N; Reus, VI; Schaefer, CA; Zhang, X; Gainetdinov, RR; Beaulieu, J-M; Sotnikova, TD; Burch, LH; Williams, RB; Schwartz, DA; Krishnan, KRR; Caron, MG
MLA Citation
Zhou, Z, Peters, EJ, Hamilton, SP, McMahon, F, Thomas, C, McGrath, PJ, Rush, J, Trivedi, MH, Charney, DS, Roy, A, Wisniewski, S, Lipsky, R, Goldman, D, Bogaert, AVD, Zutter, SD, Heyrman, L, Mendlewicz, J, Adolfsson, R, Broeckhoven, CV, Del-Favero, J, Glatt, CE, Carlson, E, Taylor, TR, Risch, N, Reus, VI, Schaefer, CA, Zhang, X, Gainetdinov, RR, Beaulieu, J-M, Sotnikova, TD, Burch, LH, Williams, RB, Schwartz, DA, Krishnan, KRR, and Caron, MG. "Response to Zhang et al. (2005) loss-of-function mutation in tryptophan hydroxylase-2 identified in unipolar major depression. Neuron 45, 11-16 [2] (multiple letters)." Neuron 48.5 (2005): 702-706.
PMID
16337901
Source
scival
Published In
Neuron
Volume
48
Issue
5
Publish Date
2005
Start Page
702
End Page
706
DOI
10.1016/j.neuron.2005.11.018

Erratum: Novelty seeking and stereotypic activation of behavior in mice with disruption of the DatI gene (Neuropsychopharmacology (April 27, 2005), DOI:10.1038/sj.npp.1300724)

Authors
Pogorelov, VM; Rodriguiz, RM; Insco, ML; Caron, MG; Wetsel, WC
MLA Citation
Pogorelov, VM, Rodriguiz, RM, Insco, ML, Caron, MG, and Wetsel, WC. "Erratum: Novelty seeking and stereotypic activation of behavior in mice with disruption of the DatI gene (Neuropsychopharmacology (April 27, 2005), DOI:10.1038/sj.npp.1300724)." Neuropsychopharmacology 30.9 (2005): 1772--.
Source
scival
Published In
Neuropsychopharmacology (Nature)
Volume
30
Issue
9
Publish Date
2005
Start Page
1772-
DOI
10.1038/sj.npp.1300797

G protein-coupled receptor kinase 2 in multiple sclerosis and experimental autoimmune encephalomyelitis

Many modulators of inflammation, including chemokines, neuropeptides, and neurotransmitters signal via G protein-coupled receptors (GPCR). GPCR kinases (GRK) can phosphorylate agonist-activated GPCR thereby promoting receptor desensitization. Here we describe that in leukocytes from patients with active relapsing-remitting multiple sclerosis (MS) or with secondary progressive MS, GRK2 levels are significantly reduced. Unexpectedly, cells from patients during remission express even lower levels of GRK2. The level of GRK2 in leukocytes of patients after stroke, a neurological disorder with paralysis but without an autoimmune component, was similar to GRK2 levels in cells from healthy individuals. In addition, we demonstrate that the course of recombinant myelin oligodendrocyte glycoprotein (1-125)-induced experimental autoimmune encephalomyelitis (EAE), an animal model for MS, is markedly different in GRK2+/- mice that express 50% of the GRK2 protein in comparison with wild-type mice. Onset of EAE was significantly advanced by 5 days in GRK2 +/- mice. The earlier onset of EAE was associated with increased early infiltration of the CNS by T cells and macrophages. Although disease scores in the first phase of EAE were similar in both groups, GRK2+/- animals did not develop relapses, whereas wild-type animals did. The absence of relapses in GRK2+/- mice was associated with a marked reduction in inflammatory infiltrates in the CNS. Recombinant myelin oligodendrocyte glycoprotein-induced T cell proliferation and cytokine production were normal in GRK2+/- animals. We conclude that down-regulation of GRK2 expression may have important consequences for the onset and progression of MS. Copyright © 2005 by The American Association of Immunologists, Inc.

Authors
Vroon, A; Kavelaars, A; Limmroth, V; Lombardi, MS; Goebel, MU; Dam, A-MV; Caron, MG; Schedlowski, M; Heijnen, CJ
MLA Citation
Vroon, A, Kavelaars, A, Limmroth, V, Lombardi, MS, Goebel, MU, Dam, A-MV, Caron, MG, Schedlowski, M, and Heijnen, CJ. "G protein-coupled receptor kinase 2 in multiple sclerosis and experimental autoimmune encephalomyelitis." Journal of Immunology 174.7 (2005): 4400-4406.
PMID
15778405
Source
scival
Published In
Journal of Immunology
Volume
174
Issue
7
Publish Date
2005
Start Page
4400
End Page
4406

Beta-arrestin 2 regulates zebrafish development through the hedgehog signaling pathway.

beta-arrestins are multifunctional proteins that act as scaffolds and transducers of intracellular signals from heptahelical transmembrane-spanning receptors (7TMR). Hedgehog (Hh) signaling, which uses the putative 7TMR, Smoothened, is established as a fundamental pathway in development, and unregulated Hh signaling is associated with certain malignancies. Here, we show that the functional knockdown of beta-arrestin 2 in zebrafish embryos recapitulates the many phenotypes of Hh pathway mutants. Expression of wild-type beta-arrestin 2, or constitutive activation of the Hh pathway downstream of Smoothened, rescues the phenotypes caused by beta-arrestin 2 deficiency. These results suggest that a functional interaction between beta-arrestin 2 and Smoothened may be critical to regulate Hh signaling in zebrafish development.

Authors
Wilbanks, AM; Fralish, GB; Kirby, ML; Barak, LS; Li, Y-X; Caron, MG
MLA Citation
Wilbanks, AM, Fralish, GB, Kirby, ML, Barak, LS, Li, Y-X, and Caron, MG. "Beta-arrestin 2 regulates zebrafish development through the hedgehog signaling pathway." Science 306.5705 (December 24, 2004): 2264-2267.
PMID
15618520
Source
pubmed
Published In
Science
Volume
306
Issue
5705
Publish Date
2004
Start Page
2264
End Page
2267
DOI
10.1126/science.1104193

Dopamine transporter-dependent and -independent actions of trace amine beta-phenylethylamine.

Beta-phenylethylamine (beta-PEA) is an endogenous amine that is found in trace amounts in the brain. It is believed that the locomotor-stimulating action of beta-PEA, much like amphetamine, depends on its ability to increase extracellular dopamine (DA) concentrations owing to reversal of the direction of dopamine transporter (DAT)-mediated DA transport. beta-PEA can also bind directly to the recently identified G protein-coupled receptors, but the physiological significance of this interaction is unclear. To assess the mechanism by which beta-PEA mediates its effects, we compared the neurochemical and behavioral effects of this amine in wild type (WT), heterozygous and 'null' DAT mutant mice. In microdialysis studies, beta-PEA, administered either systemically or locally via intrastriatal infusion, produced a pronounced outflow of striatal DA in WT mice whereas no increase was detected in mice lacking the DAT (DAT-KO mice). Similarly, in fast-scan voltammetry studies beta-PEA did not alter DA release and clearance rate in striatal slices from DAT-KO mice. In behavioral studies beta-PEA produced a robust but transient increase in locomotor activity in WT and heterozygous mice. In DAT-KO mice, whose locomotor activity and stereotypy are increased in a novel environment, beta-PEA (10-100 mg/kg) exerted a potent inhibitory action. At high doses, beta-PEA induced stereotypies in WT and heterozygous mice; some manifestations of stereotypy were also observed in the DAT-KO mice. These data demonstrate that the DAT is required for the striatal DA-releasing and hyperlocomotor actions of beta-PEA. The inhibitory action on hyperactivity and certain stereotypies induced by beta-PEA in DAT-KO mice indicate that targets other than the DAT are responsible for these effects.

Authors
Sotnikova, TD; Budygin, EA; Jones, SR; Dykstra, LA; Caron, MG; Gainetdinov, RR
MLA Citation
Sotnikova, TD, Budygin, EA, Jones, SR, Dykstra, LA, Caron, MG, and Gainetdinov, RR. "Dopamine transporter-dependent and -independent actions of trace amine beta-phenylethylamine." J Neurochem 91.2 (October 2004): 362-373.
PMID
15447669
Source
pubmed
Published In
Journal of Neurochemistry
Volume
91
Issue
2
Publish Date
2004
Start Page
362
End Page
373
DOI
10.1111/j.1471-4159.2004.02721.x

Tryptophan hydroxylase-2 controls brain serotonin synthesis.

Dysregulation of brain serotonin contributes to many psychiatric disorders. Tryptophan hydroxylase-2 (Tph2), rather than Tph1, is preferentially expressed in the brain. We report a functional (C1473G) single-nucleotide polymorphism in mouse Tph2 that results in the substitution of Pro447 with Arg447 and leads to decreased serotonin levels in PC12 cells. Moreover, in BALB/cJ and DBA/2 mice that are homozygous for the 1473G allele, brain serotonin tissue content and synthesis are reduced in comparison to C57Bl/6 and 129X1/SvJ mice that are homozygous for the 1473C allele. Our data provide direct evidence for a fundamental role of Tph2 in brain serotonin synthesis.

Authors
Zhang, X; Beaulieu, J-M; Sotnikova, TD; Gainetdinov, RR; Caron, MG
MLA Citation
Zhang, X, Beaulieu, J-M, Sotnikova, TD, Gainetdinov, RR, and Caron, MG. "Tryptophan hydroxylase-2 controls brain serotonin synthesis." Science 305.5681 (July 9, 2004): 217-.
PMID
15247473
Source
pubmed
Published In
Science
Volume
305
Issue
5681
Publish Date
2004
Start Page
217
DOI
10.1126/science.1097540

Relative opioid efficacy is determined by the complements of the G protein-coupled receptor desensitization machinery.

G protein-coupled receptor regulation by G protein-coupled receptor kinases and beta-arrestins can lead to desensitization and subsequent internalization of the receptor. In in vitro and cellular systems, beta-arrestins do not seem to play a major role in regulating micro opioid receptor (microOR) responsiveness. Removal of the betaarrestin2 (betaarr2) gene in mice leads paradoxically to enhanced and prolonged microOR-mediated antinociception. The betaarr2 knockout (betaarr2-KO) mice also fail to develop morphine antinociceptive tolerance in the hot-plate test, further indicating that the betaarr2 protein plays an essential role in microOR regulation in vivo. In this study, the contribution of betaarr2 to the regulation of the microOR was examined in both human embryonic kidney 293 cells and in betaarr2-KO mice after treatment with several opiate agonists. A green fluorescent protein tagged betaarr2 was used to assess receptor-betaarr2 interactions in living cells. Opiate agonists that induced robust betaarr2-green fluorescent protein translocation produced similar analgesia profiles in wild-type and betaarr2-KO mice, whereas those that do not promote robust betaarr2 recruitment, such as morphine and heroin, produce enhanced analgesia in vivo. In this report, we present a rationale to explain the seemingly paradoxical relationship between beta-arrestins and microOR regulation wherein morphine-like agonists fail to promote efficient internalization and resensitization of the receptor.

Authors
Bohn, LM; Dykstra, LA; Lefkowitz, RJ; Caron, MG; Barak, LS
MLA Citation
Bohn, LM, Dykstra, LA, Lefkowitz, RJ, Caron, MG, and Barak, LS. "Relative opioid efficacy is determined by the complements of the G protein-coupled receptor desensitization machinery." Mol Pharmacol 66.1 (July 2004): 106-112.
PMID
15213301
Source
pubmed
Published In
Molecular pharmacology
Volume
66
Issue
1
Publish Date
2004
Start Page
106
End Page
112
DOI
10.1124/mol.66.1.106

Dopamine Receptors

© 2004 Elsevier Inc. All rights reserved. Dopamine (DA) released from presynaptic nerve terminals elicits its effect through a family of G protein-coupled DA receptors. In the 1970s, it became evident that at least two classes of DA receptors (D1 and D2) were required to explain all the physiologic effects of DA and the pharmacologic profile of drugs blocking its actions. This traditional classification was mainly based on the ability of D1 receptors to stimulate the signal transduction enzyme adenylate cyclase, whereas D2 receptors either did not have an effect or inhibited it. The genomic organization of DA-receptor genes supports the idea that DA receptors have evolved from two distinct gene families, giving rise to D1-like and D2-like subfamilies. All DA receptors belong to a superfamily of G protein-coupled receptors (GPCRs) with seven transmembrane regions. D1-like receptors have a relatively short third cytoplasmic loop and long C-terminal tail, whereas D2-like receptors have a considerably longer third cytoplasmic loop and short C-terminal tail. These structural properties are common in many G protein-coupled receptors that stimulate and inhibit adenylate cyclase, respectively.

Authors
Laakso, A; Caron, MG
MLA Citation
Laakso, A, and Caron, MG. "Dopamine Receptors." Primer on the Autonomic Nervous System: Second Edition. May 5, 2004. 39-43.
Source
scopus
Publish Date
2004
Start Page
39
End Page
43
DOI
10.1016/B978-012589762-4/50010-4

MDMA "ecstasy" alters hyperactive and perseverative behaviors in dopamine transporter knockout mice.

RATIONALE: Mice lacking the gene for the dopamine transporter (DAT) show a unique behavioral phenotype characterized by locomotor hyperactivity and repetitive circling in a novel environment. The hyperactivity of DAT (-/-) mice can be attenuated by psychostimulants and by serotonin uptake inhibitors, suggesting an important role for serotonin in the attenuation of locomotor hyperactivity in these mice. OBJECTIVES: These studies characterized the effects of 3,4-methylenedioxy-N-methylamphetamine (MDMA), a serotonin releaser, on the amount and patterns of locomotor activity in DAT (+/+) and (-/-) mice. We compared the locomotor patterns produced by MDMA to those observed in DAT (-/-) mice, and examined whether MDMA altered the hyperactivity and perseverative locomotor patterns in DAT (-/-) mice. METHODS: The effects of MDMA (10, 30 mg/kg) on locomotor activity in DAT (+/+) mice were measured for 90 min in a video tracker system to determine the optimal dose for subsequent studies in DAT (+/+) and (-/-) mice. The effects of 20 mg/kg MDMA on patterns of locomotor activity in DAT (+/+) and (-/-) mice were measured for 90 min. RESULTS: In DAT (+/+) mice, MDMA increased locomotor activity and induced repetitive straight movement patterns. In DAT (-/-) mice, however, MDMA (20 mg/kg) attenuated the characteristic locomotor hyperactivity seen in these mice. In contrast, MDMA potentiated the thigmotaxis and decreased entropy observed in the DAT (-/-) mice. CONCLUSIONS: The effects of MDMA observed here demonstrate that the different aspects of the abnormal locomotor behavior exhibited by DAT (-/-) mice can be independently manipulated by pharmacological treatments.

Authors
Powell, SB; Lehmann-Masten, VD; Paulus, MP; Gainetdinov, RR; Caron, MG; Geyer, MA
MLA Citation
Powell, SB, Lehmann-Masten, VD, Paulus, MP, Gainetdinov, RR, Caron, MG, and Geyer, MA. "MDMA "ecstasy" alters hyperactive and perseverative behaviors in dopamine transporter knockout mice." Psychopharmacology (Berl) 173.3-4 (May 2004): 310-317.
PMID
14747902
Source
pubmed
Published In
Psychopharmacology
Volume
173
Issue
3-4
Publish Date
2004
Start Page
310
End Page
317
DOI
10.1007/s00213-003-1765-7

Hypertension and impaired glycine handling in mice lacking the orphan transporter XT2.

A family of orphan transporters has been discovered that are structurally related to the Na(+)-Cl(-)-dependent neurotransmitter transporters, including the dopamine transporter. One member of this family, the mouse XT2 gene, is predominantly expressed in the kidney and has 95% homology to rat ROSIT (renal osmotic stress-induced Na(+)-Cl(-) organic solute cotransporter). To study the physiological functions of this transporter, we generated XT2-knockout mice by gene targeting. XT2(-/-) mice develop and survive normally with no apparent abnormalities. To attempt to identify potential substrates for XT2, we screened urine from XT2-knockout mice by high-pressure liquid chromatography and mass spectrometry and found significantly elevated concentrations of glycine. To study glycine handling, XT2(+/+) and XT2(-/-) mice were injected with radiolabeled glycine, and urine samples were collected to monitor glycine excretion. After 2 h, XT2(-/-) mice were found to excrete almost twice as much glycine as the XT2(+/+) controls (P = 0.03). To determine whether the absence of the XT2 transporter affected sodium and fluid homeostasis, we measured systolic blood pressure by computerized tail-cuff manometry. Systolic blood pressure was significantly higher in XT2(-/-) mice (127 +/- 3 mmHg) than in wild-type controls (114 +/- 2 mmHg; P < 0.001). This difference in systolic blood pressure was maintained on high and low salt feeding. To examine whether the alteration in blood pressure and the defect in glycine handling were related, we measured systolic blood pressure in the XT2(-/-) mice during dietary glycine supplementation. Glycine loading caused systolic blood pressure to fall in the XT2(-/-) mice from 127 +/- 3 to 115 +/- 3 mmHg (P < 0.001), a level virtually identical to that of the wild-type controls. These data suggest that the XT2 orphan transporter is involved in glycine reabsorption and that the absence of this transporter is sufficient to cause hypertension.

Authors
Quan, H; Athirakul, K; Wetsel, WC; Torres, GE; Stevens, R; Chen, YT; Coffman, TM; Caron, MG
MLA Citation
Quan, H, Athirakul, K, Wetsel, WC, Torres, GE, Stevens, R, Chen, YT, Coffman, TM, and Caron, MG. "Hypertension and impaired glycine handling in mice lacking the orphan transporter XT2." Mol Cell Biol 24.10 (May 2004): 4166-4173.
PMID
15121838
Source
pubmed
Published In
Molecular and Cellular Biology
Volume
24
Issue
10
Publish Date
2004
Start Page
4166
End Page
4173

Lithium antagonizes dopamine-dependent behaviors mediated by an AKT/glycogen synthase kinase 3 signaling cascade.

Dopamine (DA) is a neurotransmitter involved in the control of locomotion, emotion, cognition, and reward. Administration of lithium salts is known to inhibit DA-associated behaviors in experimental animal models through unknown mechanisms. Here, we used a pharmacogenetic approach to show that DA can exert its behavioral effects by acting on a lithium-sensitive signaling cascade involving Akt/PKB and glycogen synthase kinase 3 (GSK-3). In the mouse striatum, increased DA neurotransmission arising either from administration of amphetamine or from the lack of the DA transporter results in inactivation of Akt and concomitant activation of GSK-3alpha and GSK-3beta. These biochemical changes are not affected by activation of the cAMP pathway but are effectively reversed either by inhibition of DA synthesis, D2 receptor blockade, or administration of lithium salts. Furthermore, pharmacological or genetic inhibition of GSK-3 significantly reduces DA-dependent locomotor behaviors. These data support the involvement of GSK-3 as an important mediator of DA and lithium action in vivo and suggest that modulation of the Akt/GSK-3 pathway might be relevant to DA-related disorders, such as attention deficit hyperactivity disorder and schizophrenia.

Authors
Beaulieu, J-M; Sotnikova, TD; Yao, W-D; Kockeritz, L; Woodgett, JR; Gainetdinov, RR; Caron, MG
MLA Citation
Beaulieu, J-M, Sotnikova, TD, Yao, W-D, Kockeritz, L, Woodgett, JR, Gainetdinov, RR, and Caron, MG. "Lithium antagonizes dopamine-dependent behaviors mediated by an AKT/glycogen synthase kinase 3 signaling cascade." Proc Natl Acad Sci U S A 101.14 (April 6, 2004): 5099-5104.
PMID
15044694
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
101
Issue
14
Publish Date
2004
Start Page
5099
End Page
5104
DOI
10.1073/pnas.0307921101

Identification of PSD-95 as a regulator of dopamine-mediated synaptic and behavioral plasticity.

To identify the molecular mechanisms underlying psychostimulant-elicited plasticity in the brain reward system, we undertook a phenotype-driven approach using genome-wide microarray profiling of striatal transcripts from three genetic and one pharmacological mouse models of psychostimulant or dopamine supersensitivity. A small set of co-affected genes was identified. One of these genes encoding the synaptic scaffolding protein PSD-95 is downregulated in the striatum of all three mutants and in chronically, but not acutely, cocaine-treated mice. At the synaptic level, enhanced long-term potentiation (LTP) of the frontocortico-accumbal glutamatergic synapses correlates with PSD-95 reduction in every case. Finally, targeted deletion of PSD-95 in an independent line of mice enhances LTP, augments the acute locomotor-stimulating effects of cocaine, but leads to no further behavioral plasticity in response to chronic cocaine. Our findings uncover a previously unappreciated role of PSD-95 in psychostimulant action and identify a molecular and cellular mechanism shared between drug-related plasticity and learning.

Authors
Yao, W-D; Gainetdinov, RR; Arbuckle, MI; Sotnikova, TD; Cyr, M; Beaulieu, J-M; Torres, GE; Grant, SGN; Caron, MG
MLA Citation
Yao, W-D, Gainetdinov, RR, Arbuckle, MI, Sotnikova, TD, Cyr, M, Beaulieu, J-M, Torres, GE, Grant, SGN, and Caron, MG. "Identification of PSD-95 as a regulator of dopamine-mediated synaptic and behavioral plasticity." Neuron 41.4 (February 19, 2004): 625-638.
PMID
14980210
Source
pubmed
Published In
Neuron
Volume
41
Issue
4
Publish Date
2004
Start Page
625
End Page
638

The selective serotonin-2A receptor antagonist M100907 reverses behavioral deficits in dopamine transporter knockout mice.

A hyperdopaminergic state in humans has been hypothesized to contribute to the pathology of a number of psychiatric illnesses, including schizophrenia, bipolar disorder, and attention deficit hyperactivity disorder. Mice that display elevated synaptic levels of dopamine due to a genetically engineered deletion of the dopamine transporter (DAT) model behavioral deficits that simulate the above conditions. As novel treatment strategies for these disorders have focused on the serotonin (5-HT) 2A receptor, we determined the capacity of the highly selective 5-HT(2A) receptor antagonist M100907 to reverse behavioral deficits in DAT knockout (KO) mice. Prior to drug treatment, DAT KO mice exhibited increased levels of locomotor activity and highly linearized movement in a novel environment, as well as reduced prepulse inhibition (PPI) of acoustic startle, compared to wild-type littermates. Treatment with M100907 (0.3-1.0 mg/kg, but not 0.1 mg/kg) reversed locomotor deficits in DAT KO mice. Similarly, treatment with 1.0 mg/kg M100907 reversed the PPI deficits in DAT KO mice. These data indicate that selective 5-HT(2A) receptor antagonists, such as M100907, may represent a class of drugs that can be used to treat conditions in which a chronic, elevated dopaminergic tone is present and contributes to abnormal behavior and sensorimotor gating deficits.

Authors
Barr, AM; Lehmann-Masten, V; Paulus, M; Gainetdinov, RR; Caron, MG; Geyer, MA
MLA Citation
Barr, AM, Lehmann-Masten, V, Paulus, M, Gainetdinov, RR, Caron, MG, and Geyer, MA. "The selective serotonin-2A receptor antagonist M100907 reverses behavioral deficits in dopamine transporter knockout mice." Neuropsychopharmacology 29.2 (February 2004): 221-228.
PMID
14603268
Source
pubmed
Published In
Neuropsychopharmacology (Nature)
Volume
29
Issue
2
Publish Date
2004
Start Page
221
End Page
228
DOI
10.1038/sj.npp.1300343

RNA editing induces variation in desensitization and trafficking of 5-hydroxytryptamine 2c receptor isoforms.

The 5-hydroxytryptamine2c receptor (5-HT2cR) is subjected to RNA editing, in the second intracellular loop, generating 14 different isoforms in human brain. This post-transcriptional event markedly alters the signaling properties of the receptor by reducing its ability to couple to G-proteins. Although the non-edited form of the receptor is essentially fully constitutively active, edited forms show lesser degrees of constitutive activity. We have used two extensively edited receptor isoforms, VGV and VSV, and the non-edited INI isoform to investigate how variations in constitutive receptor activity affect the trafficking and the interaction of these isoforms with components of the desensitization machinery in HEK 293 cells. We found that cell surface expression of the 5-HT2cR decreased in parallel with increased constitutive activity of the isoforms. The subcellular distribution of the various isoforms was dependent of their ability to interact with betaarrestin2, which correlated with the constitutive activity level of each isoform. We observed that the agonist-independent interaction of betaarrestin2 with constitutively active 5-HT2cR isoforms was reversed by inverse agonist treatments promoting receptor redistribution to the cell surface. Overexpression of a G-protein-coupled receptor kinase (GRK2) was able to stabilize the interaction of betaarrestin2 with constitutively active 5-HT2cR isoforms even in the presence of inverse agonists. Taken together, our observations indicate that the constitutively active 5-HT2cR isoforms are spontaneously internalized in an agonist-independent manner. This endocytosis process is mediated by a GRK/betaarrestin-dependent mechanism and is directly correlated with the constitutive activity status of the RNA edited receptor variants. Thus the ultimate physiological output of constitutively active receptors may be determined not only by their agonist-independent activity but also by their interactions with GRKs and betaarrestin.

Authors
Marion, S; Weiner, DM; Caron, MG
MLA Citation
Marion, S, Weiner, DM, and Caron, MG. "RNA editing induces variation in desensitization and trafficking of 5-hydroxytryptamine 2c receptor isoforms." J Biol Chem 279.4 (January 23, 2004): 2945-2954.
PMID
14602721
Source
pubmed
Published In
The Journal of biological chemistry
Volume
279
Issue
4
Publish Date
2004
Start Page
2945
End Page
2954
DOI
10.1074/jbc.M308742200

Aberrant responses in social interaction of dopamine transporter knockout mice.

The dopamine (DA) transporter (DAT) controls the temporal and spatial resolution of dopaminergic neurotransmission. Disruption of the Dat1 gene in mice leads to increased extracellular DA concentrations and reduced expression of D1- and D2-like receptors in striatum. The mutants are hyperactive in the open field and they display deficits in learning and memory. In humans, dopaminergic dysfunction has been associated with a number of different psychiatric disorders and some of these conditions are accompanied by abnormal social responses. To determine whether social responses were also impaired in DAT knockout (KO) mice, behaviors of group- and isolation-housed animals were compared. All group-housed animals readily established hierarchies. However, the social organizations of the mutants were changed over time. Under both group- and isolation-housed conditions, mutants exhibited increased rates of reactivity and aggression following mild social contact. In isolation, exposure to a novel environment exacerbated these abnormal responses. Regardless of housing context, stereotyped and perseverative patterns of social responses were a common feature of the KO repertoire. In fact, many abnormal behaviors were due to the emergence and predominance of these inflexible behaviors. These data suggest that KO mice may serve as a useful animal model for understanding not only how DA dysfunction contributes to social abnormalities, but also how behavioral inflexibility distorts their social responses.

Authors
Rodriguiz, RM; Chu, R; Caron, MG; Wetsel, WC
MLA Citation
Rodriguiz, RM, Chu, R, Caron, MG, and Wetsel, WC. "Aberrant responses in social interaction of dopamine transporter knockout mice." Behav Brain Res 148.1-2 (January 5, 2004): 185-198.
PMID
14684259
Source
pubmed
Published In
Behavioural Brain Research
Volume
148
Issue
1-2
Publish Date
2004
Start Page
185
End Page
198

G protein-coupled receptor kinase/beta-arrestin systems and drugs of abuse: psychostimulant and opiate studies in knockout mice.

G protein-coupled receptors (GPCRs) currently represent pharmaceutical targets for numerous medicinal compounds that are used to treat conditions ranging from blood pressure dysregulation to depression to pain, demonstrating the wide range of functions mediated by this receptor family. GPCR activation is determined not only by the initiation of signaling cascades but also by regulatory mechanisms that control the extent and duration of their signals. The balance of activation and desensitization dictate the ultimate physiological response to both endogenous and exogenous receptor stimuli. Therefore, these mechanisms may play a particularly relevant role during chronic exposure to agonists such as in conditions when drugs are abused. Two major classes of drugs of abuse, opiates and psychostimulants, both use either direct or indirect GPCR signaling mechanisms to mediate their effects. Therefore, the regulation of GPCRs may have bearing on the neuronal adaptations that underlie the reinforcing properties of drugs of abuse.

Authors
Bohn, LM; Gainetdinov, RR; Caron, MG
MLA Citation
Bohn, LM, Gainetdinov, RR, and Caron, MG. "G protein-coupled receptor kinase/beta-arrestin systems and drugs of abuse: psychostimulant and opiate studies in knockout mice." Neuromolecular Med 5.1 (2004): 41-50. (Review)
PMID
15001811
Source
pubmed
Published In
NeuroMolecular Medicine
Volume
5
Issue
1
Publish Date
2004
Start Page
41
End Page
50
DOI
10.1385/NMM:5:1:041

Desensitization of G protein-coupled receptors and neuronal functions.

G protein-coupled receptors (GPCRs) have proven to be the most highly favorable class of drug targets in modern pharmacology. Over 90% of nonsensory GPCRs are expressed in the brain, where they play important roles in numerous neuronal functions. GPCRs can be desensitized following activation by agonists by becoming phosphorylated by members of the family of G protein-coupled receptor kinases (GRKs). Phosphorylated receptors are then bound by arrestins, which prevent further stimulation of G proteins and downstream signaling pathways. Discussed in this review are recent progress in understanding basics of GPCR desensitization, novel functional roles, patterns of brain expression, and receptor specificity of GRKs and beta arrestins in major brain functions. In particular, screening of genetically modified mice lacking individual GRKs or beta arrestins for alterations in behavioral and biochemical responses to cocaine and morphine has revealed a functional specificity in dopamine and mu-opioid receptor regulation of locomotion and analgesia. An important and specific role of GRKs and beta arrestins in regulating physiological responsiveness to psychostimulants and morphine suggests potential involvement of these molecules in certain brain disorders, such as addiction, Parkinson's disease, mood disorders, and schizophrenia. Furthermore, the utility of a pharmacological strategy aimed at targeting this GPCR desensitization machinery to regulate brain functions can be envisaged.

Authors
Gainetdinov, RR; Premont, RT; Bohn, LM; Lefkowitz, RJ; Caron, MG
MLA Citation
Gainetdinov, RR, Premont, RT, Bohn, LM, Lefkowitz, RJ, and Caron, MG. "Desensitization of G protein-coupled receptors and neuronal functions." Annu Rev Neurosci 27 (2004): 107-144. (Review)
PMID
15217328
Source
pubmed
Published In
Annual Review of Neuroscience
Volume
27
Publish Date
2004
Start Page
107
End Page
144
DOI
10.1146/annurev.neuro.27.070203.144206

Uptake and release of norepinephrine by serotonergic terminals in norepinephrine transporter knock-out mice: Implications for the action of selective serotonin reuptake inhibitors

Our aim was to investigate the functional properties of the noradrenergic system in genetically modified mice lacking the norepinephrine transporter (NET). We measured the uptake and release of [3H]norepinephrine ([3H]NE) from hippocampal and cortical slices of NET-/- knock-out (KO) and NET+/+ wild-type (WT) mice and investigated the presynaptic α2-adenoceptor-mediated modulation of NE release in vitro and in vivo. The [3H]NE uptake was reduced to 12.6% (hippocampus) and 33.5% (frontal cortex) of WT control in KO mice. The neuronal component of this residual uptake was decreased by 79.4 and 100%, respectively, when a selective serotonin reuptake inhibitor (SSRI) citalopram was present during the loading. The more preserved neuronal release of [3H]NE (hippocampus, 28.1%; frontal cortex, 74.4%; compared with WT) almost completely disappeared in both regions (94.1 and 95.3% decrease compared with KO, respectively) in the presence of citalopram, suggesting that [3H]NE was taken up and released by serotonergic varicosities. This was further supported by the finding that the release of [3H]NE from hippocampal slices of KO mice was not modulated by the α2-adrenoceptor antagonist 7,8-(methylenedioxy)-14- α-hydroxyalloberbane HCl, whereas the endogenous release of NE measured by microdialysis was even more efficiently enhanced by this drug in NET-deficient mice. These experiments indicate that serotonergic varicosities can accumulate and release NE as a result of the heterologous uptake of transmitters. Because the diffusion of NE may be spatially limited by serotonin transporters, the SSRIs, despite their selectivity, might enhance not only serotonergic but also noradrenergic neurotransmission, which might contribute to their antidepressant action.

Authors
Vizi, ES; Zsilla, G; Caron, MG; Kiss, JP
MLA Citation
Vizi, ES, Zsilla, G, Caron, MG, and Kiss, JP. "Uptake and release of norepinephrine by serotonergic terminals in norepinephrine transporter knock-out mice: Implications for the action of selective serotonin reuptake inhibitors." Journal of Neuroscience 24.36 (2004): 7888-7894.
PMID
15356201
Source
scival
Published In
Journal of Neuroscience
Volume
24
Issue
36
Publish Date
2004
Start Page
7888
End Page
7894
DOI
10.1523/JNEUROSCI.1506-04.2004

Norepinephrine transporter-deficient mice exhibit excessive tachycardia and elevated blood pressure with wakefulness and activity

Background-Norepinephrine (NE) is a primary neurotransmitter of central autonomic regulation and sympathetic nerve conduction, and the norepinephrine transporter (NET) is crucial in limiting catecholaminergic signaling. NET is sensitive to antidepressants, cocaine, and amphetamine. NET blockade often is associated with cardiovascular side effects, and NET deficiency is linked to tachycardia in familial orthostatic intolerance. Methods and Results-We telemetrically monitored NET-deficient (NET-/-) mice to determine the cardiovascular effects of reduced NE reuptake. Mean arterial pressure was elevated in resting NET-/- mice compared with NET+/+ controls (103±0.6 versus 99±0.4 mm Hg; P<0.01), and corresponding pressures increased to 122±0.3 and 116±0.3 mm Hg (P<0.0001) with activity. Heart rate was also greater in resting NET -/- mice (565±5 versus 551±3 bpm; P<0.05), and genotypic differences were highly significant during the active phase (640±5 versus 607±3 bpm; P<0.0001). Conversely, the respiratory rate of resting NET-/- mice was dramatically reduced, whereas increases after the day/night shift surpassed those of controls. Plasma catecholamines in NET-/- and NET+/+ mice were as follows: NE, 69±8 and 32±7; dihydroxyphenylglycol, 2+0.4 and 17±3; epinephrine, 15±3 and 4±0.6; and dopamine, 13±4 and 4±1 pmol/mL. Catechols in urine, brain, and heart also were determined. Conclusions-Resting mean arterial pressure and heart rate are maintained at nearly normal levels in NET-deficient mice, most likely as a result of increased central sympathoinhibition. However, sympathetic activation with wakefulness and activity apparently overwhelms central modulation, amplifying peripheral catecholaminergic signaling, particularly in the heart.

Authors
Keller, NR; Diedrich, A; Appalsamy, M; Tuntrakool, S; Lonce, S; Finney, C; Caron, MG; Robertson, D
MLA Citation
Keller, NR, Diedrich, A, Appalsamy, M, Tuntrakool, S, Lonce, S, Finney, C, Caron, MG, and Robertson, D. "Norepinephrine transporter-deficient mice exhibit excessive tachycardia and elevated blood pressure with wakefulness and activity." Circulation 110.10 (2004): 1191-1196.
PMID
15337696
Source
scival
Published In
Circulation
Volume
110
Issue
10
Publish Date
2004
Start Page
1191
End Page
1196
DOI
10.1161/01.CIR.0000141804.90845.E6

Comparative studies of molecular mechanisms of dopamine D2 and D3 receptors for the activation of extracellular signal-regulated kinase

Dopamine D2 and D3 receptors (D2R/D 3R), which have similar structural architecture as well as functional similarities, are expressed in the same brain dopaminergic neurons. It is intriguing that two receptor proteins with virtually the same functional roles are expressed in the same neuron. Recently we have shown that D2R and D3R possess different regulatory processes including intracellular trafficking properties, which implies that they might employ different signaling mechanisms for regulation of the same cellular processes. Here we studied the signaling pathways of ERK activation mediated by D2R and D 3R in HEK-293 cells and corroborated them with concomitant studies in COS-7 cells and C6 cells. Our results show that Src, phosphatidylinositol 3-kinase, and atypical protein kinase C were commonly involved in D 2R-/D3R-mediated ERK activation. However, β-arrestin and sequestration of D2R/D3R were found not to be involved. ERK activations mediated by D3R, but not D2R, were blocked by βARK-CT, AG1478 epidermal growth factor receptor (EGFR) inhibitor, and by dominant negative mutants of Ras and Raf, suggesting the involvement of the Gβγi pathway. The α-subunit of Go (Gαo) was able to couple with D3R to mediate ERK activation. We conclude that D3R mainly utilizes the βγ pathway of Gi protein, which involves the transactivation of EGFR in HEK-293 cells. In contrast, the α-subunit of the Gi protein plays a main role in D2R-mediated ERK activation. Our study suggests one example of intricate cellular regulations in the brain, that is, dopaminergic neurons could regulate ERK activity more flexibly through alternative usage of either the D2R or D 3R pathway depending on the cellular situation.

Authors
Beom, S; Cheong, D; Torres, G; Caron, MG; Kim, K-M
MLA Citation
Beom, S, Cheong, D, Torres, G, Caron, MG, and Kim, K-M. "Comparative studies of molecular mechanisms of dopamine D2 and D3 receptors for the activation of extracellular signal-regulated kinase." Journal of Biological Chemistry 279.27 (2004): 28304-28314.
PMID
15102843
Source
scival
Published In
Journal of Biological Chemistry
Volume
279
Issue
27
Publish Date
2004
Start Page
28304
End Page
28314
DOI
10.1074/jbc.M403899200

Genetic and genomic approaches to reward and addiction.

Drug addiction is recognized as a mental disease affecting the brain's natural reward system. Drugs of abuse strongly activate reward structures in the brain and induce lasting changes in behavior that reflect changes in neuron physiology and biochemistry. With the ultimate goal of developing therapeutic interventions, it is of interest to determine the molecular and cellular components of motivation and reward, and identify those gene products that contribute to the process of drug addiction. Our laboratory has chosen three general genetic approaches to examine reward and addiction: reverse genetics to assess the role of candidate genes in drug responsiveness, forward genetics to discover novel regulators of dopamine transmission, and gene expression profiling to define gene sets in different brain structures that contribute to the molecular and neurobiological basis of reward.

Authors
Mohn, AR; Yao, W-D; Caron, MG
MLA Citation
Mohn, AR, Yao, W-D, and Caron, MG. "Genetic and genomic approaches to reward and addiction." Neuropharmacology 47 Suppl 1 (2004): 101-110. (Review)
PMID
15464129
Source
pubmed
Published In
Neuropharmacology
Volume
47 Suppl 1
Publish Date
2004
Start Page
101
End Page
110
DOI
10.1016/j.neuropharm.2004.07.025

Green fluorescent protein-tagged beta-arrestin translocation as a measure of G protein-coupled receptor activation.

The G protein-coupled receptor (GPCR) superfamily is the largest family of integral membrane proteins. GPCRs respond to a wide variety of sensory and chemical stimuli and contribute directly to the regulation of all major organ systems. As such, GPCRs represent primary drug targets for therapeutic intervention. Although GPCRs respond to a diverse range of ligands and signal through multiple heterotrimeric G proteins, the inactivation of GPCR signaling is mediated by a limited set of proteins. In particular, the desensitization of the majority of GPCRs is mediated by the binding of two arrestin isoforms, beta-arrestin1 and beta-arrestin2, that exhibit overlapping substrate specificity. In response to GPCR activation and phosphorylation by GPCR kinases, beta-arrestins redistribute from the cytosol to the plasma membrane to bind GPCRs and subsequently target the receptors for internalization via clathrin-coated vesicles. This property of beta-arrestins has allowed the development of a green fluorescent protein (GFP)-based assay for detecting GPCR activation by confocal microscopy. This beta-arrestin-GFP translocation methodology is described in detail in this chapter.

Authors
Ferguson, SSG; Caron, MG
MLA Citation
Ferguson, SSG, and Caron, MG. "Green fluorescent protein-tagged beta-arrestin translocation as a measure of G protein-coupled receptor activation." Methods in molecular biology (Clifton, N.J.) 237 (2004): 121-126.
PMID
14501044
Source
scival
Published In
Methods in molecular biology (Clifton, N.J.)
Volume
237
Publish Date
2004
Start Page
121
End Page
126

Clozapine-, but not haloperidol-, induced increases in ΔFosB-like immunoreactivity are completely blocked in the striatum of mice lacking D3 dopamine receptors

On the basis of anatomical and pharmacological evidence, we have proposed that D3 receptor antagonism plays a role in the mediation of clozapine-, but not haloperidol-, induced immediate-early gene expression in the striatum. To test this hypothesis directly, we compared the effects of repeated administration of vehicle (8 mL/kg/day), clozapine (20 mg/kg/day) and haloperidol (2 mg/kg/day) for 17 days on expression of ΔFosB-like immunoreactivity (ΔFosB-Ir) in the island of Calleja major, nucleus accumbens and caudate-putamen of wild-type C57BI6 (WT) and D3 receptor knockout (D3KO) mice. In vehicle-treated mice, the number of ΔFosB-Ir neurons in the nucleus accumbens was greater in D3KO than in WT mice. This finding is consistent with results implicating D3 receptor activation in the tonic inhibition of this limbic structure. Unlike rats, clozapine significantly increased the number of ΔFosB-Ir neurons in both the nucleus accumbens and the caudate-putamen of WT mice albeit to a lesser extent in the caudate-putamen than nucleus accumbens. Similar to rats, however, ΔFosB-Ir in the island of Calleja major of WT mice was elevated by clozapine but not by haloperidol. In the nucleus accumbens and caudate-putamen, haloperidol produced similar increases in ΔFosB-Ir in WT and D3KO mice. By contrast, clozapine-induced increases in ΔFosB-Ir in the island of Calleja major, nucleus accumbens and caudate-putamen of WT mice were absent in D3KO mice. These findings, which indicate that D3 receptor blockade is essential for clozapine-induced increases in striatal ΔFosB-Ir, suggest that D3 receptor antagonism may contribute to the unique therapeutic profile of this atypical antipsychotic.

Authors
Robertson, GS; Lee, CJ; Sridhar, K; Nakabeppu, Y; Cheng, M; Wang, Y-M; Caron, MG
MLA Citation
Robertson, GS, Lee, CJ, Sridhar, K, Nakabeppu, Y, Cheng, M, Wang, Y-M, and Caron, MG. "Clozapine-, but not haloperidol-, induced increases in ΔFosB-like immunoreactivity are completely blocked in the striatum of mice lacking D3 dopamine receptors." European Journal of Neuroscience 20.11 (2004): 3189-3194.
PMID
15579174
Source
scival
Published In
European Journal of Neuroscience
Volume
20
Issue
11
Publish Date
2004
Start Page
3189
End Page
3194
DOI
10.1111/j.1460-9568.2004.03774.x

Effect of torsinA on membrane proteins reveals a loss of function and a dominant-negative phenotype of the dystonia-associated ΔE-torsinA mutant

Most cases of early-onset torsion dystonia (EOTD) are caused by a deletion of one glutamic acid in the carboxyl terminus of a protein named torsinA. The mutation causes the protein to aggregate in perinuclear inclusions as opposed to the endoplasmic reticulum localization of the wild-type protein. Although there is increasing evidence that dysfunction of the dopamine system is implicated in the development of EOTD, the biological function of torsinA and its relation to dopaminergic neurotransmission has remained unexplored. Here, we show that torsinA can regulate the cellular trafficking of the dopamine transporter, as well as other polytopic membrane-bound proteins, including G protein-coupled receptors, transporters, and ion channels. This effect was prevented by mutating the ATP-binding site in torsinA. The dystonia-associated torsinA deletion mutant (ΔE-torsinA) did not have any effect on the cell surface distribution of polytopic membrane-associated proteins, suggesting that the mutation linked with EOTD results in a loss of function. However, a mutation in the ATP-binding site in ΔE-torsinA reversed the aggregate phenotype associated with the mutant. Moreover, the deletion mutant acts as a dominant-negative of wild-type torsinA through a mechanism presumably involving association of wild-type and mutant torsinA. Taken together, our results provide evidence for a functional role for torsinA and a loss of function and a dominant-negative phenotype of the ΔE-torsinA mutation. These properties may contribute to the autosomal dominant nature of the condition.

Authors
Torres, GE; Sweeney, AL; Beaulieu, J-M; Shashidharan, P; Caron, MG
MLA Citation
Torres, GE, Sweeney, AL, Beaulieu, J-M, Shashidharan, P, and Caron, MG. "Effect of torsinA on membrane proteins reveals a loss of function and a dominant-negative phenotype of the dystonia-associated ΔE-torsinA mutant." Proceedings of the National Academy of Sciences of the United States of America 101.44 (2004): 15650-15655.
PMID
15505207
Source
scival
Published In
Proceedings of the National Academy of Sciences of the United States of America
Volume
101
Issue
44
Publish Date
2004
Start Page
15650
End Page
15655
DOI
10.1073/pnas.0308088101

Neuropathic pain activates the endogenous κ opioid system in mouse spinal cord and induces opioid receptor tolerance

Release of endogenous dynorphin opioids within the spinal cord after partial sciatic nerve ligation (pSNL) is known to contribute to the neuropathic pain processes. Using a phosphoselective antibody [κ opioid receptor (KOR-P)] able to detect the serine 369 phosphorylated form of the KOR, we determined possible sites of dynorphin action within the spinal cord after pSNL. KOR-P immunoreactivity (IR) was markedly increased in the L4-L5 spinal dorsal horn of wild-type C57BL/6 mice (7-21 d) after lesion, but not in mice pretreated with the KOR antagonist nor-binaltorphimine (norBNI). In addition, knock-out mice lacking prodynorphin, KOR, or G-protein receptor kinase 3 (GRK3) did not show significant increases in KOR-P IR after pSNL. KOR-P IR was colocalized in both GABAergic neurons and GFAP-positive astrocytes in both ipsilateral and contralateral spinal dorsal horn. Consistent with sustained opioid release, KOR knock-out mice developed significantly increased tactile allodynia and thermal hyperalgesia in both the early (first week) and late (third week) interval after lesion. Similarly, mice pretreated with norBNI showed enhanced hyperalgesia and allodynia during the 3 weeks after pSNL. Because sustained activation of opioid receptors might induce tolerance, we measured the antinociceptive effect of the κ agonist U50,488 using radiant heat applied to the ipsilateral hindpaw, and we found that agonist potency was significantly decreased 7 d after pSNL. In contrast, neither prodynorphin nor GRK3 knock-out mice showed U50,488 tolerance after pSNL. These findings suggest that pSNL induced a sustained release of endogenous prodynorphin-derived opioid peptides that activated an anti-nociceptive KOR system in mouse spinal cord. Thus, endogenous dynorphin had both pronociceptive and antinociceptive actions after nerve injury and induced GRK3-mediated opioid tolerance.

Authors
Xu, M; Petraschka, M; McLaughlin, JP; Westenbroek, RE; Caron, MG; Lefkowitz, RJ; Czyzyk, TA; Pintar, JE; Terman, GW; Chavkin, C
MLA Citation
Xu, M, Petraschka, M, McLaughlin, JP, Westenbroek, RE, Caron, MG, Lefkowitz, RJ, Czyzyk, TA, Pintar, JE, Terman, GW, and Chavkin, C. "Neuropathic pain activates the endogenous κ opioid system in mouse spinal cord and induces opioid receptor tolerance." Journal of Neuroscience 24.19 (2004): 4576-4584.
Website
http://hdl.handle.net/10161/5936
PMID
15140929
Source
scival
Published In
Journal of Neuroscience
Volume
24
Issue
19
Publish Date
2004
Start Page
4576
End Page
4584
DOI
10.1523/JNEUROSCI.5552-03.2004

Reduced GRK2 level in T cells potentiates chemotaxis and signaling in response to CCL4

Chemokine receptors belong to the family of G-protein-coupled receptors (GPCR). Phosphorylation of GPCR by GPCR kinases (GRKs) is considered to play an important role in desensitization of these, receptors. We have recently shown in patients with rheumatoid arthritis that the level of GRK2 in lymphocytes is reduced by ∼50%. However, the physiological relevance of reduced GRK2 levels in lymphocytes is not known. Here, we investigated whether reduced GRK2 expression changes the chemotactic response of T cells to the chemokines CCL3, CCL4, and CCL5. Activated T cells from GRK2+/- mice, which have a 50% reduction in GRK2 protein levels, showed a significant 40% increase in chemotaxis toward the CCR5 ligand CCL4. In addition, chemotaxis toward the CCR1 and CCR5 ligands CCL3 and CCL5 was also increased. Binding of CCL4 to activated T cells from GRK2+/- and wild-type (WT) mice was similar, but agonist-induced CCR5 phosphorylation was attenuated in GRK2+/- cells. Moreover, the calcium response and phosphorylation of protein kinase B and extracellular-regulated kinase in response to CCL4 were significantly increased in GRK2+/- T cells, showing that signaling is increased when the level of GRK2 is reduced. GRK2+/- and WT cells do become refractory to restimulation with CCL4. In conclusion, a 50% decrease in T cell GRK2 expression results in increased responsiveness to CCL3, CCL4, and CCL5, suggesting that the 50% reduction in lymphocyte GRK2 level as observed during inflammation can have functional consequences for the response of these cells to chemokines.

Authors
Vroon, A; Heijnen, CJ; Lombardi, MS; Cobelens, PM; Jr, FM; Caron, MG; Kavelaars, A
MLA Citation
Vroon, A, Heijnen, CJ, Lombardi, MS, Cobelens, PM, Jr, FM, Caron, MG, and Kavelaars, A. "Reduced GRK2 level in T cells potentiates chemotaxis and signaling in response to CCL4." Journal of Leukocyte Biology 75.5 (2004): 901-909.
PMID
14761932
Source
scival
Published In
Journal of Leukocyte Biology
Volume
75
Issue
5
Publish Date
2004
Start Page
901
End Page
909
DOI
10.1189/jlb.0403136

G protein-coupled receptor kinase 5 regulates airway responses induced by muscarinic receptor activation

G protein-coupled receptors (GPCRs) transduce extracellular signals into intracellular events. The waning responsiveness of GPCRs in the face of persistent agonist stimulation, or desensitization, is a necessary event that ensures physiological homeostasis. GPCR kinases (GRKs) are important regulators of GPCR desensitization. GRK5, one member of the GRK family, desensitizes central M2 muscarinic receptors in mice. We questioned whether GRK5 might also be an important regulator of peripheral muscarinic receptor responsiveness in the cardiopulmonary system. Specifically, we wanted to determine the role of GRK5 in regulating muscarinic receptor-mediated control of airway smooth muscle tone or regulation of cholinergic-induced bradycardia. Tracheal pressure, heart rate, and tracheal smooth muscle tension were measured in mice having a targeted deletion of the GRK5 gene (GRK5-/-) and littermate wild-type (WT) control mice. Both in vivo and in vitro results showed that the airway contractile response to a muscarinic receptor agonist was not different between GRK5-/- and WT mice. However, the relaxation component of bilateral vagal stimulation and the airway smooth muscle relaxation resulting from β2-adrenergic receptor activation were diminished in GRK5-/- mice. These data suggest that M2 muscarinic receptor-mediated opposition of airway smooth muscle relaxation is regulated by GRK5 and is, therefore, excessive in GRK5-/- mice. In addition, this study shows that GRK5 regulates pulmonary responses in a tissue- and receptor-specific manner but does not regulate peripheral cardiac muscarinic receptors. GRK5 regulation of airway responses may have implications in obstructive airway diseases such as asthma or chronic obstructive pulmonary disease.

Authors
Walker, JKL; Gainetdinov, RR; Feldman, DS; McFawn, PK; Caron, MG; Lefkowitz, RJ; Premont, RT; Fisher, JT
MLA Citation
Walker, JKL, Gainetdinov, RR, Feldman, DS, McFawn, PK, Caron, MG, Lefkowitz, RJ, Premont, RT, and Fisher, JT. "G protein-coupled receptor kinase 5 regulates airway responses induced by muscarinic receptor activation." American Journal of Physiology - Lung Cellular and Molecular Physiology 286.2 30-2 (2004): L312-L319.
Source
scival
Published In
American Journal of Physiology - Lung Cellular and Molecular Physiology
Volume
286
Issue
2 30-2
Publish Date
2004
Start Page
L312
End Page
L319

Prolonged Kappa Opioid Receptor Phosphorylation Mediated by G-protein Receptor Kinase Underlies Sustained Analgesic Tolerance

Kappa opioid receptor (KOR) desensitization was previously shown to follow agonist-dependent phosphorylation of serine 369 by G-protein receptor kinase (GRK) and β-arrestin binding in transfected cells. To study the in vivo effects induced by phosphorylation of KOR(S369), C57B1/6 mice were administered single or repeated doses of the KOR agonist, U50,488, and isolated brain glycoprotein was probed with an antibody, KOR-P, that specifically recognized phosphoserine 369 KOR. Western blot analysis using KOR-P antibody showed that labeling intensity increased after either single or repeated treatment of mice with U50,488 by 59 ± 22% and 101 ± 29%, respectively. In contrast, there was no change in labeling intensity by nonphosphoselective KOR antibodies following acute or chronic in vivo treatment with kappa agonist. Moreover, mice lacking GRK3 showed no increase in KOR-P labeling and developed significantly less analgesic tolerance following treatment with kappa agonist. The result suggests that tolerance to kappa agonists includes phosphorylation of serine 369 within KOR by GRK3. Recovery of analgesic potency and reduction of elevated KOR-P labeling in wild-type mice both required 2 weeks to return to base line. Consistent with these results, in vitro phosphorylation by GRK3 of KOR isolated from tolerant mice resulted in 46 ± 7% less 32P incorporation than in KOR isolated from untreated mice. In addition, in vitro 32P incorporation returned to base line levels only in KOR isolated from tolerant mice allowed to recover for 2 weeks. The coincident reversal of analgesic tolerance and slow return to a basal phosphorylation state matched the regeneration rate of functional kappa receptors following irreversible antagonism and suggested that receptor replacement rather than dephosphorylation was required to restore sensitivity.

Authors
McLaughlin, JP; Myers, LC; Zarek, PE; Caron, MG; Lefkowitz, RJ; Czyzyk, TA; Pintar, JE; Chavkin, C
MLA Citation
McLaughlin, JP, Myers, LC, Zarek, PE, Caron, MG, Lefkowitz, RJ, Czyzyk, TA, Pintar, JE, and Chavkin, C. "Prolonged Kappa Opioid Receptor Phosphorylation Mediated by G-protein Receptor Kinase Underlies Sustained Analgesic Tolerance." Journal of Biological Chemistry 279.3 (2004): 1810-1818.
PMID
14597630
Source
scival
Published In
Journal of Biological Chemistry
Volume
279
Issue
3
Publish Date
2004
Start Page
1810
End Page
1818
DOI
10.1074/jbc.M305796200

G-protein receptor kinase 3 (GRK3) influences opioid analgesic tolerance but not opioid withdrawal

1. Tolerance to opioids frequently follows repeated drug administration and affects the clinical utility of these analgesics. Studies in simple cellular systems have demonstrated that prolonged activation of opioid receptors produces homologous receptor desensitization by G-protein receptor kinase mediated receptor phosphorylation and subsequent β-arrestin binding. To define the role of this regulatory mechanism in the control of the electrophysiological and behavioral responses to opioids, we used mice having a targeted disruption of the G-protein receptor kinase 3 (GRK3) gene. 2. Mice lacking GRK3 did not differ from wild-type littermates neither in their response latencies to noxious stimuli on the hot-plate test nor in their acute antinociceptive responses to fentanyl or morphine. 3. Tolerance to the electrophysiological response to the opioid fentanyl, measured in vitro in the hippocampus, was blocked by GRK3 deletion. In addition, tolerance to the antinociceptive effects of fentanyl was significantly reduced in GRK3 knockouts compared to wild-type littermate controls. 4. Tolerance to the antinociceptive effects of morphine was not affected by GRK3 deletion although morphine tolerance in hippocampal slices from GRK3 knockout mice was significantly inhibited. Tolerance developed more slowly in vitro to morphine than fentanyl supporting previous work in in vitro systems showing a correlation between agonist efficacy and GRK3-mediated desensitization. 5. The results of these studies suggest that GRK3-mediated mechanisms are important components of both electrophysiologic and behavioral opioid tolerance. Fentanyl, a high efficacy opioid, more effectively produced GRK3-dependent effects than morphine, a low efficacy agonist.

Authors
Terman, GW; Jin, W; Cheong, Y-P; Lowe, J; Caron, MG; Lefkowitz, RJ; Chavkin, C
MLA Citation
Terman, GW, Jin, W, Cheong, Y-P, Lowe, J, Caron, MG, Lefkowitz, RJ, and Chavkin, C. "G-protein receptor kinase 3 (GRK3) influences opioid analgesic tolerance but not opioid withdrawal." British Journal of Pharmacology 141.1 (2004): 55-64.
PMID
14662727
Source
scival
Published In
British Journal of Pharmacology
Volume
141
Issue
1
Publish Date
2004
Start Page
55
End Page
64
DOI
10.1038/sj.bjp.0705595

Identification of Drosophila neuropeptide receptors by G protein-coupled receptors-beta-arrestin2 interactions.

Activation of G protein-coupled receptors (GPCR) leads to the recruitment of beta-arrestins. By tagging the beta-arrestin molecule with a green fluorescent protein, we can visualize the activation of GPCRs in living cells. We have used this approach to de-orphan and study 11 GPCRs for neuropeptide receptors in Drosophila melanogaster. Here we verify the identities of ligands for several recently de-orphaned receptors, including the receptors for the Drosophila neuropeptides proctolin (CG6986), neuropeptide F (CG1147), corazonin (CG10698), dFMRF-amide (CG2114), and allatostatin C (CG7285 and CG13702). We also de-orphan CG6515 and CG7887 by showing these two suspected tachykinin receptor family members respond specifically to a Drosophila tachykinin neuropeptide. Additionally, the translocation assay was used to de-orphan three Drosophila receptors. We show that CG14484, encoding a receptor related to vertebrate bombesin receptors, responds specifically to allatostatin B. Furthermore, the pair of paralogous receptors CG8985 and CG13803 responds specifically to the FMRF-amide-related peptide dromyosuppressin. To corroborate the findings on orphan receptors obtained by the translocation assay, we show that dromyosuppressin also stimulated GTPgammaS binding and inhibited cAMP by CG8985 and CG13803. Together these observations demonstrate the beta-arrestin-green fluorescent protein translocation assay is an important tool in the repertoire of strategies for ligand identification of novel G protein-coupled receptors.

Authors
Johnson, EC; Bohn, LM; Barak, LS; Birse, RT; Nässel, DR; Caron, MG; Taghert, PH
MLA Citation
Johnson, EC, Bohn, LM, Barak, LS, Birse, RT, Nässel, DR, Caron, MG, and Taghert, PH. "Identification of Drosophila neuropeptide receptors by G protein-coupled receptors-beta-arrestin2 interactions." J Biol Chem 278.52 (December 26, 2003): 52172-52178.
PMID
14555656
Source
pubmed
Published In
The Journal of biological chemistry
Volume
278
Issue
52
Publish Date
2003
Start Page
52172
End Page
52178
DOI
10.1074/jbc.M306756200

Enhanced rewarding properties of morphine, but not cocaine, in beta(arrestin)-2 knock-out mice.

The reinforcing and psychomotor effects of morphine involve opiate stimulation of the dopaminergic system via activation of mu-opioid receptors (muOR). Both mu-opioid and dopamine receptors are members of the G-protein-coupled receptor (GPCR) family of proteins. GPCRs are known to undergo desensitization involving phosphorylation of the receptor and the subsequent binding of beta(arrestins), which prevents further receptor-G-protein coupling. Mice lacking beta(arrestin)-2 (beta(arr2)) display enhanced sensitivity to morphine in tests of pain perception attributable to impaired desensitization of muOR. However, whether abrogating muOR desensitization affects the reinforcing and psychomotor properties of morphine has remained unexplored. In the present study, we examined this question by assessing the effects of morphine and cocaine on locomotor activity, behavioral sensitization, conditioned place preference, and striatal dopamine release in beta(arr2) knock-out (beta(arr2)-KO) mice and their wild-type (WT) controls. Cocaine treatment resulted in very similar neurochemical and behavioral responses between the genotypes. However, in the beta(arr2)-KO mice, morphine induced more pronounced increases in striatal extracellular dopamine than in WT mice. Moreover, the rewarding properties of morphine in the conditioned place preference test were greater in the beta(arr2)-KO mice when compared with the WT mice. Thus, beta(arr2) appears to play a more important role in the dopaminergic effects mediated by morphine than those induced by cocaine.

Authors
Bohn, LM; Gainetdinov, RR; Sotnikova, TD; Medvedev, IO; Lefkowitz, RJ; Dykstra, LA; Caron, MG
MLA Citation
Bohn, LM, Gainetdinov, RR, Sotnikova, TD, Medvedev, IO, Lefkowitz, RJ, Dykstra, LA, and Caron, MG. "Enhanced rewarding properties of morphine, but not cocaine, in beta(arrestin)-2 knock-out mice." J Neurosci 23.32 (November 12, 2003): 10265-10273.
Website
http://hdl.handle.net/10161/5935
PMID
14614085
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
23
Issue
32
Publish Date
2003
Start Page
10265
End Page
10273

Sustained elevation of extracellular dopamine causes motor dysfunction and selective degeneration of striatal GABAergic neurons.

Dopamine is believed to contribute to the degeneration of dopamine-containing neurons in the brain. However, whether dopamine affects the survival of other neuronal populations has remained unclear. Here we document that mice with persistently elevated extracellular dopamine, resulting from inactivation of the dopamine transporter gene, sporadically develop severe symptoms of dyskinesia concomitant with apoptotic death of striatal dopamine-responsive gamma-aminobutyric acidergic neurons. Chronic inhibition of dopamine synthesis prevents the appearance of motor dysfunction. The neuronal death is associated with overactivation of dopaminergic signaling as evidenced by the robust up-regulation of striatal DeltaFosB, cyclin-dependent kinase 5, and p35. Moreover, hyperphosphorylation of the tau protein, a phenomenon associated with the activation of cyclin-dependent kinase 5 in several neurodegenerative disorders, is observed in symptomatic mice. These findings provide in vivo evidence that, in addition to its proposed role in the degeneration of dopamine neurons, dopamine can also contribute to the selective death of its target neurons via a previously unappreciated mechanism.

Authors
Cyr, M; Beaulieu, J-M; Laakso, A; Sotnikova, TD; Yao, W-D; Bohn, LM; Gainetdinov, RR; Caron, MG
MLA Citation
Cyr, M, Beaulieu, J-M, Laakso, A, Sotnikova, TD, Yao, W-D, Bohn, LM, Gainetdinov, RR, and Caron, MG. "Sustained elevation of extracellular dopamine causes motor dysfunction and selective degeneration of striatal GABAergic neurons." Proc Natl Acad Sci U S A 100.19 (September 16, 2003): 11035-11040.
PMID
12958210
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
100
Issue
19
Publish Date
2003
Start Page
11035
End Page
11040
DOI
10.1073/pnas.1831768100

Dishevelled 2 recruits beta-arrestin 2 to mediate Wnt5A-stimulated endocytosis of Frizzled 4.

Wnt proteins, regulators of development in many organisms, bind to seven transmembrane-spanning (7TMS) receptors called frizzleds, thereby recruiting the cytoplasmic molecule dishevelled (Dvl) to the plasma membrane.Frizzled-mediated endocytosis of Wg (a Drosophila Wnt protein) and lysosomal degradation may regulate the formation of morphogen gradients. Endocytosis of Frizzled 4 (Fz4) in human embryonic kidney 293 cells was dependent on added Wnt5A protein and was accomplished by the multifunctional adaptor protein beta-arrestin 2 (betaarr2), which was recruited to Fz4 by binding to phosphorylated Dvl2. These findings provide a previously unrecognized mechanism for receptor recruitment of beta-arrestin and demonstrate that Dvl plays an important role in the endocytosis of frizzled, as well as in promoting signaling.

Authors
Chen, W; ten Berge, D; Brown, J; Ahn, S; Hu, LA; Miller, WE; Caron, MG; Barak, LS; Nusse, R; Lefkowitz, RJ
MLA Citation
Chen, W, ten Berge, D, Brown, J, Ahn, S, Hu, LA, Miller, WE, Caron, MG, Barak, LS, Nusse, R, and Lefkowitz, RJ. "Dishevelled 2 recruits beta-arrestin 2 to mediate Wnt5A-stimulated endocytosis of Frizzled 4." Science 301.5638 (September 5, 2003): 1391-1394.
PMID
12958364
Source
pubmed
Published In
Science
Volume
301
Issue
5638
Publish Date
2003
Start Page
1391
End Page
1394
DOI
10.1126/science.1082808

Center stage for the serotonin transporter: a gain-of-function polymorphism in persons with obsessive-compulsive disorder.

Authors
Torres, GE; Caron, MG
MLA Citation
Torres, GE, and Caron, MG. "Center stage for the serotonin transporter: a gain-of-function polymorphism in persons with obsessive-compulsive disorder." Mol Pharmacol 64.2 (August 2003): 196-198.
PMID
12869622
Source
pubmed
Published In
Molecular pharmacology
Volume
64
Issue
2
Publish Date
2003
Start Page
196
End Page
198
DOI
10.1124/mol.64.2.196

Conditional calcineurin knockout mice exhibit multiple abnormal behaviors related to schizophrenia.

Calcineurin (CN), a calcium- and calmodulin-dependent protein phosphatase, plays a significant role in the central nervous system. Previously, we reported that forebrain-specific CN knockout mice (CN mutant mice) have impaired working memory. To further analyze the behavioral effects of CN deficiency, we subjected CN mutant mice to a comprehensive behavioral test battery. Mutant mice showed increased locomotor activity, decreased social interaction, and impairments in prepulse inhibition and latent inhibition. In addition, CN mutant mice displayed an increased response to the locomotor stimulating effects of MK-801. Collectively, the abnormalities of CN mutant mice are strikingly similar to those described for schizophrenia. We propose that alterations affecting CN signaling could comprise a contributing factor in schizophrenia pathogenesis.

Authors
Miyakawa, T; Leiter, LM; Gerber, DJ; Gainetdinov, RR; Sotnikova, TD; Zeng, H; Caron, MG; Tonegawa, S
MLA Citation
Miyakawa, T, Leiter, LM, Gerber, DJ, Gainetdinov, RR, Sotnikova, TD, Zeng, H, Caron, MG, and Tonegawa, S. "Conditional calcineurin knockout mice exhibit multiple abnormal behaviors related to schizophrenia." Proc Natl Acad Sci U S A 100.15 (July 22, 2003): 8987-8992.
PMID
12851457
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
100
Issue
15
Publish Date
2003
Start Page
8987
End Page
8992
DOI
10.1073/pnas.1432926100

Dopaminergic supersensitivity in G protein-coupled receptor kinase 6-deficient mice.

Brain dopaminergic transmission is a critical component in numerous vital functions, and its dysfunction is involved in several disorders, including addiction and Parkinson's disease. Responses to dopamine are mediated via G protein-coupled dopamine receptors (D1-D5). Desensitization of G protein-coupled receptors is mediated via phosphorylation by members of the family of G protein-coupled receptor kinases (GRK1-GRK7). Here we show that GRK6-deficient mice are supersensitive to the locomotor-stimulating effect of psychostimulants, including cocaine and amphetamine. In addition, these mice demonstrate an enhanced coupling of striatal D2-like dopamine receptors to G proteins and augmented locomotor response to direct dopamine agonists both in intact and in dopamine-depleted animals. The present study indicates that postsynaptic D2-like dopamine receptors are physiological targets for GRK6 and suggests that this regulatory mechanism contributes to central dopaminergic supersensitivity.

Authors
Gainetdinov, RR; Bohn, LM; Sotnikova, TD; Cyr, M; Laakso, A; Macrae, AD; Torres, GE; Kim, KM; Lefkowitz, RJ; Caron, MG; Premont, RT
MLA Citation
Gainetdinov, RR, Bohn, LM, Sotnikova, TD, Cyr, M, Laakso, A, Macrae, AD, Torres, GE, Kim, KM, Lefkowitz, RJ, Caron, MG, and Premont, RT. "Dopaminergic supersensitivity in G protein-coupled receptor kinase 6-deficient mice." Neuron 38.2 (April 24, 2003): 291-303.
PMID
12718862
Source
pubmed
Published In
Neuron
Volume
38
Issue
2
Publish Date
2003
Start Page
291
End Page
303

Constitutive desensitization: a new paradigm for g protein-coupled receptor regulation.

GPCRs are a large family of cell-surface proteins that regulate many important biochemical pathways and physiological responses. The isolation and characterization of GPCRs represent one of the more remarkable success stories that occurred during the revolution in biology of the last quarter century. Of the many discoveries that originated in the laboratory of Robert Lefkowitz at Duke University concerning GPCR regulation, none is more fundamental than the elucidation of the families of GRKs and arrestin proteins that terminate GPCR signaling. In this essay, we will discuss how advances in microscopy and biology have made the visualization of GPCR, GRK, and arrestin activity possible in single cells. Additionally, we will discuss how imaging studies using arrestins and a naturally occurring mutant of the vasopressin receptor led to the recognition of a novel phenotypic receptor behavior, in which the receptor desensitizes in the absence of agonist. We have termed this process constitutive desensitization, and this unexpected receptor property suggests that it may be possible to develop novel classes of signal-inhibiting drugs distinct from conventional antagonists.

Authors
Barak, LS; Wilbanks, AM; Caron, MG
MLA Citation
Barak, LS, Wilbanks, AM, and Caron, MG. "Constitutive desensitization: a new paradigm for g protein-coupled receptor regulation." Assay Drug Dev Technol 1.2 (April 2003): 339-346. (Review)
PMID
15090199
Source
pubmed
Published In
ASSAY and Drug Development Technologies
Volume
1
Issue
2
Publish Date
2003
Start Page
339
End Page
346
DOI
10.1089/15406580360545152

The stability of the G protein-coupled receptor-beta-arrestin interaction determines the mechanism and functional consequence of ERK activation.

By binding to agonist-activated G protein-coupled receptors (GPCRs), beta-arrestins mediate homologous receptor desensitization and endocytosis via clathrin-coated pits. Recent data suggest that beta-arrestins also contribute to GPCR signaling by acting as scaffolds for components of the ERK mitogen-activated protein kinase cascade. Because of these dual functions, we hypothesized that the stability of the receptor-beta-arrestin interaction might affect the mechanism and functional consequences of GPCR-stimulated ERK activation. In transfected COS-7 cells, we found that angiotensin AT1a and vasopressin V2 receptors, which form stable receptor-beta-arrestin complexes, activated a beta-arrestin-bound pool of ERK2 more efficiently than alpha 1b and beta2 adrenergic receptors, which form transient receptor-beta-arrestin complexes. We next studied chimeric receptors in which the pattern of beta-arrestin binding was reversed by exchanging the C-terminal tails of the beta2 and V2 receptors. The ability of the V2 beta 2 and beta 2V2 chimeras to activate beta-arrestin-bound ERK2 corresponded to the pattern of beta-arrestin binding, suggesting that the stability of the receptor-beta-arrestin complex determined the mechanism of ERK2 activation. Analysis of covalently cross-linked detergent lysates and cellular fractionation revealed that wild type V2 receptors generated a larger pool of cytosolic phospho-ERK1/2 and less nuclear phospho-ERK1/2 than the chimeric V2 beta 2 receptor, consistent with the cytosolic retention of beta-arrestin-bound ERK. In stably transfected HEK-293 cells, the V2 beta 2 receptor increased ERK1/2-mediated, Elk-1-driven transcription of a luciferase reporter to a greater extent than the wild type V2 receptor. Furthermore, the V2 beta 2, but not the V2 receptor, was capable of eliciting a mitogenic response. These data suggest that the C-terminal tail of a GPCR, by determining the stability of the receptor-beta-arrestin complex, controls the extent of beta-arrestin-bound ERK activation, and influences both the subcellular localization of activated ERK and the physiologic consequences of ERK activation.

Authors
Tohgo, A; Choy, EW; Gesty-Palmer, D; Pierce, KL; Laporte, S; Oakley, RH; Caron, MG; Lefkowitz, RJ; Luttrell, LM
MLA Citation
Tohgo, A, Choy, EW, Gesty-Palmer, D, Pierce, KL, Laporte, S, Oakley, RH, Caron, MG, Lefkowitz, RJ, and Luttrell, LM. "The stability of the G protein-coupled receptor-beta-arrestin interaction determines the mechanism and functional consequence of ERK activation." J Biol Chem 278.8 (February 21, 2003): 6258-6267.
PMID
12473660
Source
pubmed
Published In
The Journal of biological chemistry
Volume
278
Issue
8
Publish Date
2003
Start Page
6258
End Page
6267
DOI
10.1074/jbc.M212231200

Oligomerization and trafficking of the human dopamine transporter. Mutational analysis identifies critical domains important for the functional expression of the transporter.

The dopamine transporter (DAT) is a presynaptic plasma membrane protein responsible for the termination of dopaminergic neurotransmission in the central nervous system. While most studies have focused on structure/function analysis, much less information is available regarding the assembly and the trafficking of this protein. To address this problem, we performed a mutational analysis of the DAT protein, combined with biochemical, immunological, and functional approaches. In mammalian cells co-expressing differentially tagged DAT molecules, HA-tagged DAT co-purified with 6His-tagged DAT demonstrating a physical interaction between transporter proteins. Evidence for the functional oligomerization of DAT was obtained using dominant-negative mutants of DAT. Two loss-of-function mutant transporters (Y335A and D79G) that were targeted to the cell surface inhibited wild-type DAT uptake activity without affecting the membrane targeting of the wild-type transporter. Moreover, non-functional amino and carboxyl termini-truncated mutants of DAT inhibited wild-type DAT function by interfering with the normal processing of the wild-type transporter to the cell membrane. Mutations in the leucine repeat of the second transmembrane domain of the transporter could eliminate the dominant-negative effect of all these mutants. In addition, a small fragment comprising the first two transmembrane domains of DAT inhibited wild-type transporter function but not when the leucine repeat motif was mutated. Taken together, our results suggest that the assembly of DAT monomers plays a critical role in the expression and function of the transporter.

Authors
Torres, GE; Carneiro, A; Seamans, K; Fiorentini, C; Sweeney, A; Yao, W-D; Caron, MG
MLA Citation
Torres, GE, Carneiro, A, Seamans, K, Fiorentini, C, Sweeney, A, Yao, W-D, and Caron, MG. "Oligomerization and trafficking of the human dopamine transporter. Mutational analysis identifies critical domains important for the functional expression of the transporter." J Biol Chem 278.4 (January 24, 2003): 2731-2739.
PMID
12429746
Source
pubmed
Published In
The Journal of biological chemistry
Volume
278
Issue
4
Publish Date
2003
Start Page
2731
End Page
2739
DOI
10.1074/jbc.M201926200

Plasma membrane monoamine transporters: structure, regulation and function.

Authors
Torres, GE; Gainetdinov, RR; Caron, MG
MLA Citation
Torres, GE, Gainetdinov, RR, and Caron, MG. "Plasma membrane monoamine transporters: structure, regulation and function." Nat Rev Neurosci 4.1 (January 2003): 13-25. (Review)
PMID
12511858
Source
pubmed
Published In
Nature Reviews Neuroscience
Volume
4
Issue
1
Publish Date
2003
Start Page
13
End Page
25
DOI
10.1038/nrn1008

Monoamine transporters: from genes to behavior.

Modulation of fast neurotransmission by monoamines is critically involved in numerous physiological functions and pathological conditions. Plasma membrane monoamine transporters provide one of the most efficient mechanisms controlling functional extracellular monoamine concentrations. These transporters for dopamine (DAT), serotonin (SERT), and norepinephrine (NET), which are expressed selectively on the corresponding neurons, are established targets of many psychostimulants, antidepressants, and neurotoxins. Recently, genetic animal models with targeted disruption of these transporters have become available. These mice have provided opportunities to investigate the functional importance of transporters in homeostatic control of monoaminergic transmission and to evaluate, in an in vivo model system, their roles in physiology and pathology. The use of these mice as test subjects has been helpful in resolving several important issues on specificity and mechanisms of action of certain pharmacological agents. In the present review, we summarize recent advances in understanding the physiology and pharmacology of monoamine transporters gained in mice with targeted genetic deletion of DAT, SERT, and NET.

Authors
Gainetdinov, RR; Caron, MG
MLA Citation
Gainetdinov, RR, and Caron, MG. "Monoamine transporters: from genes to behavior." Annu Rev Pharmacol Toxicol 43 (2003): 261-284. (Review)
PMID
12359863
Source
pubmed
Published In
Annual Review of Pharmacology and Toxicology
Volume
43
Publish Date
2003
Start Page
261
End Page
284
DOI
10.1146/annurev.pharmtox.43.050802.112309

Differential psychostimulant-induced activation of neural circuits in dopamine transporter knockout and wild type mice.

Dopamine (DA) is a neurotransmitter that has been implicated in a wide variety of psychiatric disorders that include attention deficit-hyperactivity disorder (ADHD), schizophrenia, and drug abuse. Recently, we have been working with a mouse in which the gene for the DA transporter (DAT) has been disrupted. This mouse is hyperactive in the open field, displays an inability to inhibit ongoing behaviors, and is deficient on learning and memory tasks. Psychostimulants such as amphetamine and methylphenidate attenuate the hyperlocomotion of the mutants, but stimulate activity of the wild type (WT) controls. The objective of the present study is to examine the neural basis for the differential responses to psychostimulants in these mice. WT and DAT knockout (KO) animals were given vehicle or methylphenidate, amphetamine, or cocaine and brain sections were immunostained for Fos. In WT mice, methylphenidate induced Fos-like immunoreactivity (Fos-LI) in the mesostriatal and mesolimbocortical DA pathways that included the anterior olfactory nucleus, frontal association cortex, orbitofrontal cortex, cingulate cortex, caudate-putamen, globus pallidus, claustrum, lateral septum, nucleus accumbens, basolateral and central nuclei of the amygdala, bed nucleus of stria terminalis, subthalamic nucleus, substantia nigra, ventral tegmental area, and dorsal raphe. Additional areas of activation included the granular dentate gyrus, Edinger-Westphal nucleus, and periaqueductal gray. While the mutants showed little response in most of these same areas, the anterior olfactory nucleus, caudal caudate-putamen, lateral septum, basolateral and central nuclei of the amygdala, and bed nucleus of stria terminalis were activated. Amphetamine and cocaine produced similar changes to that for methylphenidate, except these psychostimulants also induced Fos-LI in the nucleus accumbens of the KO animals. Since the DAT gene is disrupted in the KO mouse, these findings suggest that dopaminergic mechanisms may mediate the WT responses, whereas non-dopaminergic systems predominate in the mutant. In the mutants, it appears that limbic areas and non-dopaminergic transmitter systems within these brain regions may mediate responses to psychostimulants. Inasmuch as the KO mouse may represent a useful animal model for ADHD and because psychostimulants such as cocaine are reinforcing to these animals, our results may provide some useful insights into the neural mechanisms-other than DA-that may contribute to the symptoms of ADHD and/or drug abuse in human patients.

Authors
Trinh, JV; Nehrenberg, DL; Jacobsen, JPR; Caron, MG; Wetsel, WC
MLA Citation
Trinh, JV, Nehrenberg, DL, Jacobsen, JPR, Caron, MG, and Wetsel, WC. "Differential psychostimulant-induced activation of neural circuits in dopamine transporter knockout and wild type mice." Neuroscience 118.2 (2003): 297-310.
PMID
12699766
Source
pubmed
Published In
Neuroscience
Volume
118
Issue
2
Publish Date
2003
Start Page
297
End Page
310

Differential mechanisms of morphine antinociceptive tolerance revealed in (beta)arrestin-2 knock-out mice.

Morphine induces antinociception by activating mu opioid receptors (muORs) in spinal and supraspinal regions of the CNS. (Beta)arrestin-2 (beta)arr2), a G-protein-coupled receptor-regulating protein, regulates the muOR in vivo. We have shown previously that mice lacking (beta)arr2 experience enhanced morphine-induced analgesia and do not become tolerant to morphine as determined in the hot-plate test, a paradigm that primarily assesses supraspinal pain responsiveness. To determine the general applicability of the (beta)arr2-muOR interaction in other neuronal systems, we have, in the present study, tested (beta)arr2 knock-out ((beta)arr2-KO) mice using the warm water tail-immersion paradigm, which primarily assesses spinal reflexes to painful thermal stimuli. In this test, the (beta)arr2-KO mice have greater basal nociceptive thresholds and markedly enhanced sensitivity to morphine. Interestingly, however, after a delayed onset, they do ultimately develop morphine tolerance, although to a lesser degree than the wild-type (WT) controls. In the (beta)arr2-KO but not WT mice, morphine tolerance can be completely reversed with a low dose of the classical protein kinase C (PKC) inhibitor chelerythrine. These findings provide in vivo evidence that the muOR is differentially regulated in diverse regions of the CNS. Furthermore, although (beta)arr2 appears to be the most prominent and proximal determinant of muOR desensitization and morphine tolerance, in the absence of this mechanism, the contributions of a PKC-dependent regulatory system become readily apparent.

Authors
Bohn, LM; Lefkowitz, RJ; Caron, MG
MLA Citation
Bohn, LM, Lefkowitz, RJ, and Caron, MG. "Differential mechanisms of morphine antinociceptive tolerance revealed in (beta)arrestin-2 knock-out mice." J Neurosci 22.23 (December 1, 2002): 10494-10500.
Website
http://hdl.handle.net/10161/5920
PMID
12451149
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
22
Issue
23
Publish Date
2002
Start Page
10494
End Page
10500

Experimental genetic approaches to addiction.

Drugs of abuse are able to elicit compulsive drug-seeking behaviors upon repeated administration, which ultimately leads to the phenomenon of addiction. Evidence indicates that the susceptibility to develop addiction is influenced by sources of reinforcement, variable neuroadaptive mechanisms, and neurochemical changes that together lead to altered homeostasis of the brain reward system. Addiction is hypothesized to be a cycle of progressive dysregulation of the brain reward system that results in the compulsive use and loss of control over drug taking and the initiation of behaviors associated with drug seeking. The view that addiction represents a pathological state of reward provides an approach to identifying the factors that contribute to vulnerability, addiction, and relapse in genetic animal models.

Authors
Laakso, A; Mohn, AR; Gainetdinov, RR; Caron, MG
MLA Citation
Laakso, A, Mohn, AR, Gainetdinov, RR, and Caron, MG. "Experimental genetic approaches to addiction." Neuron 36.2 (October 10, 2002): 213-228. (Review)
PMID
12383778
Source
pubmed
Published In
Neuron
Volume
36
Issue
2
Publish Date
2002
Start Page
213
End Page
228

Apparent loss-of-function mutant GPCRs revealed as constitutively desensitized receptors.

The DRY motif is a triplet amino acid sequence (aspartic acid, arginine, and tyrosine) that is highly conserved in G protein-coupled receptors (GPCRs). Recently, we have shown that a molecular determinant for nephrogenic diabetes insipidus, the vasopressin receptor with a substitution at the DRY motif arginine (V2R R137H), is a constitutively desensitized receptor that is unable to couple to G proteins due to its constitutive association with beta-arrestin [Barak, L. S. (2001) Proc. Natl. Acad. Sci. U.S.A. 98, 93-98]. Additionally, the mutant receptors are localized in endocytic vesicles, identical to wild-type receptors stimulated with agonist. In this study, we asked whether the constitutively desensitized phenotype observed in the V2R R137H represents a general paradigm that may be extended to other GPCRs. We show that arginine substitutions in the DRY motifs of the alpha(1B) adrenergic receptor (alpha(1B)-AR) and angiotensin II type 1A receptor (AT(1A)R) result in receptors that are uncoupled from G proteins, associated with beta-arrestins, and found localized in endocytic vesicles rather than at the plasma membrane in the absence of agonists. The localization of the alpha(1B)-ARs and AT(1A)Rs with arginine substitutions can be restored to the plasma membrane by either using selective antagonists or preventing the endocytosis of the beta-arrestin-receptor complexes. These results indicate that the arginine residue of the DRY motif is essential for preserving the localization of the inactive receptor complex. Furthermore, constitutive desensitization may underlie some loss-of-function receptor phenotypes and represent an unappreciated mechanism of hormonal resistance.

Authors
Wilbanks, AM; Laporte, SA; Bohn, LM; Barak, LS; Caron, MG
MLA Citation
Wilbanks, AM, Laporte, SA, Bohn, LM, Barak, LS, and Caron, MG. "Apparent loss-of-function mutant GPCRs revealed as constitutively desensitized receptors." Biochemistry 41.40 (October 8, 2002): 11981-11989.
PMID
12356298
Source
pubmed
Published In
Biochemistry
Volume
41
Issue
40
Publish Date
2002
Start Page
11981
End Page
11989

Trafficking of the vesicular acetylcholine transporter in SN56 cells: a dynamin-sensitive step and interaction with the AP-2 adaptor complex.

The pathways by which synaptic vesicle proteins reach their destination are not completely defined. Here we investigated the traffic of a green fluorescent protein (GFP)-tagged version of the vesicular acetylcholine transporter (VAChT) in cholinergic SN56 cells, a model system for neuronal processing of this cargo. GFP-VAChT accumulates in small vesicular compartments in varicosities, but perturbation of endocytosis with a dominant negative mutant of dynamin I-K44A impaired GFP-VAChT trafficking to these processes. The protein in this condition accumulated in the cell body plasma membrane and in large vesicular patches therein. A VAChT endocytic mutant (L485A/L486A) was also located at the plasma membrane, however, the protein was not sorted to dynamin I-K44A generated vesicles. A fusion protein containing the VAChT C-terminal tail precipitated the AP-2 adaptor protein complex from rat brain, suggesting that VAChT directly interacts with the endocytic complex. In addition, yeast two hybrid experiments indicated that the C-terminal tail of VAChT interacts with the micro subunit of AP-2 in a di-leucine (L485A/L486A) dependent fashion. These observations suggest that the di-leucine motif regulates sorting of VAChT from the soma plasma membrane through a clathrin dependent mechanism prior to the targeting of the transporter to varicosities.

Authors
Barbosa, J; Ferreira, LT; Martins-Silva, C; Santos, MS; Torres, GE; Caron, MG; Gomez, MV; Ferguson, SSG; Prado, MAM; Prado, VF
MLA Citation
Barbosa, J, Ferreira, LT, Martins-Silva, C, Santos, MS, Torres, GE, Caron, MG, Gomez, MV, Ferguson, SSG, Prado, MAM, and Prado, VF. "Trafficking of the vesicular acetylcholine transporter in SN56 cells: a dynamin-sensitive step and interaction with the AP-2 adaptor complex." J Neurochem 82.5 (September 2002): 1221-1228.
PMID
12358769
Source
pubmed
Published In
Journal of Neurochemistry
Volume
82
Issue
5
Publish Date
2002
Start Page
1221
End Page
1228

The multiple LIM domain-containing adaptor protein Hic-5 synaptically colocalizes and interacts with the dopamine transporter.

The Na+/Cl--dependent dopamine transporter (DAT) is critical in terminating dopaminergic transmission by removing the transmitter away from the synapse. Several lines of evidence suggest that transporter-interacting proteins may play a role in DAT function and regulation. In this report, using the yeast two-hybrid system, we have identified a novel interaction between DAT and the multiple Lin-11, Isl-1, and Mec-3 (LIM) domain-containing adaptor protein Hic-5. This association involves the N-terminal portion of the intracellular tail of DAT and the LIM region of Hic-5. In human embryonic kidney 293 cells, Hic-5 colocalizes with DAT at polarized sites and reduces DAT uptake activity through a mechanism involving a decrease in the cell-surface levels of the transporter. A fragment of Hic-5 containing the LIM domains is sufficient to bind DAT but lacks the ability to inhibit transporter activity. In addition, the LIM fragment prevents the effect of the full-length Hic-5 on DAT localization and function. In the brain, Hic-5 protein is expressed in the cerebral cortex, hippocampus, hypothalamus, cerebellum, and striatum, suggesting a role for this protein in the nervous system. The association of the endogenous Hic-5 and DAT proteins was confirmed biochemically by coimmunoprecipitation from brain striatal extracts. Moreover, immunostaining of rat midbrain neurons in culture revealed a presynaptic colocalization of Hic-5 and DAT. Because Hic-5 has been shown to interact with several signaling molecules, including the nonreceptor protein tyrosine kinases focal adhesion kinase and Fyn, this raises the possibility that this adaptor protein may link DAT to intracellular signaling pathways.

Authors
Carneiro, AM; Ingram, SL; Beaulieu, J-M; Sweeney, A; Amara, SG; Thomas, SM; Caron, MG; Torres, GE
MLA Citation
Carneiro, AM, Ingram, SL, Beaulieu, J-M, Sweeney, A, Amara, SG, Thomas, SM, Caron, MG, and Torres, GE. "The multiple LIM domain-containing adaptor protein Hic-5 synaptically colocalizes and interacts with the dopamine transporter." J Neurosci 22.16 (August 15, 2002): 7045-7054.
PMID
12177201
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
22
Issue
16
Publish Date
2002
Start Page
7045
End Page
7054

Phosphoinositide 3-kinase regulates beta2-adrenergic receptor endocytosis by AP-2 recruitment to the receptor/beta-arrestin complex.

Internalization of beta-adrenergic receptors (betaARs) occurs by the sequential binding of beta-arrestin, the clathrin adaptor AP-2, and clathrin. D-3 phosphoinositides, generated by the action of phosphoinositide 3-kinase (PI3K) may regulate the endocytic process; however, the precise molecular mechanism is unknown. Here we demonstrate that betaARKinase1 directly interacts with the PIK domain of PI3K to form a cytosolic complex. Overexpression of the PIK domain displaces endogenous PI3K from betaARK1 and prevents betaARK1-mediated translocation of PI3K to activated beta2ARs. Furthermore, disruption of the betaARK1/PI3K interaction inhibits agonist-stimulated AP-2 adaptor protein recruitment to the beta2AR and receptor endocytosis without affecting the internalization of other clathrin dependent processes such as internalization of the transferrin receptor. In contrast, AP-2 recruitment is enhanced in the presence of D-3 phospholipids, and receptor internalization is blocked in presence of the specific phosphatidylinositol-3,4,5-trisphosphate lipid phosphatase PTEN. These findings provide a molecular mechanism for the agonist-dependent recruitment of PI3K to betaARs, and support a role for the localized generation of D-3 phosphoinositides in regulating the recruitment of the receptor/cargo to clathrin-coated pits.

Authors
Naga Prasad, SV; Laporte, SA; Chamberlain, D; Caron, MG; Barak, L; Rockman, HA
MLA Citation
Naga Prasad, SV, Laporte, SA, Chamberlain, D, Caron, MG, Barak, L, and Rockman, HA. "Phosphoinositide 3-kinase regulates beta2-adrenergic receptor endocytosis by AP-2 recruitment to the receptor/beta-arrestin complex." J Cell Biol 158.3 (August 5, 2002): 563-575.
PMID
12163475
Source
pubmed
Published In
The Journal of Cell Biology
Volume
158
Issue
3
Publish Date
2002
Start Page
563
End Page
575
DOI
10.1083/jcb.200202113

Monoamine transporter pharmacology and mutant mice.

Monoamine transporters, such as the dopamine transporter, 5-HT transporter and noradrenaline transporter, in the plasma membrane provide effective control over the intensity of monoamine-mediated signaling by recapturing neurotransmitters released by presynaptic neurons. These proteins represent established targets for several psychotropic drugs, including psychostimulants and antidepressants; however, important issues regarding the selectivity and mechanisms of action of these drugs remain unresolved. Although monoamine transporter knockout mice have profound changes in neurotransmission, they provide useful in vivo models to analyze the effects of psychotropic drugs. In this review, we summarize recent insights into the pharmacology of psychotropic drugs using mice in which the genes encoding these transporters have been deleted.

Authors
Gainetdinov, RR; Sotnikova, TD; Caron, MG
MLA Citation
Gainetdinov, RR, Sotnikova, TD, and Caron, MG. "Monoamine transporter pharmacology and mutant mice." Trends Pharmacol Sci 23.8 (August 2002): 367-373. (Review)
PMID
12377578
Source
pubmed
Published In
Trends in Pharmacological Sciences
Volume
23
Issue
8
Publish Date
2002
Start Page
367
End Page
373

Behavioral responses to social stress in noradrenaline transporter knockout mice: effects on social behavior and depression.

Noradrenaline has been implicated in the pathogenesis of depression and the noradrenaline transporter (NET) is a target for some antidepressants. Therefore, mice with disrupted NET gene expression (NET-KO) appear especially suitable for studying this behavioral disorder. We have examined the interaction between social stress (an etiological factor of depression) and the resulting depressive behaviors in NET-KO mice. Social stress was induced by daily defeats from larger resident mice while depression was assessed by the behavioral despair model. Animals subjected to repeated social stress showed reduced weight gain and a gradual shift from offensive to defensive behaviors. The latter may be considered a situation-specific depressive-like behavior. NET gene disruption did not prevent these changes that developed in a homotypic situation (i.e., during the repeated application of the same stressor). In contrast, stressed NET-KO mice showed more struggling in the behavioral despair model than stressed wild type (WT) animals. Thus, NET gene disruption inhibited depression-like behavior in chronically stressed animals tested in a situation heterotypic to the original cause of chronic stress. We suggest that the behavioral effects of NET gene disruption were overruled by experience and learning in the homotypic situation but manifested fully in the heterotypic situation. Tentatively, our data suggest that enhanced noradrenergic function does not prevent situation-specific social learning but impedes the generalization of depression to heterotypic circumstances.

Authors
Haller, J; Bakos, N; Rodriguiz, RM; Caron, MG; Wetsel, WC; Liposits, Z
MLA Citation
Haller, J, Bakos, N, Rodriguiz, RM, Caron, MG, Wetsel, WC, and Liposits, Z. "Behavioral responses to social stress in noradrenaline transporter knockout mice: effects on social behavior and depression." Brain Res Bull 58.3 (July 2002): 279-284.
PMID
12128153
Source
pubmed
Published In
Brain Research Bulletin
Volume
58
Issue
3
Publish Date
2002
Start Page
279
End Page
284

Decreased ethanol preference and consumption in dopamine transporter female knock-out mice.

BACKGROUND: It is commonly believed that the mesolimbic dopamine (DA) system participates in the etiology of alcoholism. One of the most important regulators of DA synaptic transmission is the DA transporter (DAT). We examined the effects of the genetic reduction or deletion of DAT on voluntary ethanol consumption and ethanol-induced sedation. METHODS: Ethanol preference and consumption were assessed in the two-bottle choice paradigm, and the depressant effects of ethanol were evaluated by measuring sleep time after acute injection of ethanol. RESULTS: The latent period to lose the righting reflex was shorter in both knock-out mice (DAT-KO) and heterozygote mice (HET) than in wild-type (WT) mice. No significant difference was found among the three genotypes in the ethanol blood concentration at the onset of regaining the righting reflex. Females of all genotypes consumed more fluid than males of the same genotype. HET and DAT-KO females had increased total fluid consumption compared with WT females. DAT-KO males had increased fluid consumption compared with WT and HET males. Ethanol preference and consumption were not different among male mice of different genotypes. WT and HET females demonstrated significantly higher ethanol consumption than males. HET female mice did not differ from WT mice in ethanol preference. There was no difference between HET and WT mice in the preference for saccharin or quinine solutions. DAT-KO females avoided ethanol, and their consumption and preference were lower than in WT and HET females, despite markedly increased total intake. DAT-KO mice also demonstrated altered taste preference for saccharin and quinine. CONCLUSIONS: Partial deletion of DAT results in increased fluid consumption in female mice but does not change ethanol preference in either sex. Complete deletion of DAT reduces ethanol preference in female mice; this may be due to a combination of the pharmacological actions of DAT deletion and alterations in fluid consumption and taste discrimination.

Authors
Savelieva, KV; Caudle, WM; Findlay, GS; Caron, MG; Miller, GW
MLA Citation
Savelieva, KV, Caudle, WM, Findlay, GS, Caron, MG, and Miller, GW. "Decreased ethanol preference and consumption in dopamine transporter female knock-out mice." Alcohol Clin Exp Res 26.6 (June 2002): 758-764.
PMID
12068242
Source
pubmed
Published In
Alcoholism: Clinical and Experimental Research
Volume
26
Issue
6
Publish Date
2002
Start Page
758
End Page
764

Regulation of arrestin-3 phosphorylation by casein kinase II.

Arrestins play an important role in regulating the function of G protein-coupled receptors including receptor desensitization, internalization, down-regulation, and signaling via nonreceptor tyrosine kinases and mitogen-activated protein kinases. Previous studies have revealed that arrestins themselves are also subject to regulation. In the present study, we focused on identifying potential mechanisms involved in regulating the function of arrestin-3. Using metabolic labeling, phosphoamino acid analysis, and mutagenesis studies, we found that arrestin-3 is constitutively phosphorylated at Thr-382 and becomes dephosphorylated upon beta(2)-adrenergic receptor activation in COS-1 cells. Casein kinase II (CKII) appears to be the major kinase mediating arrestin-3 phosphorylation, since 1) Thr-382 is contained within a canonical consensus sequence for CKII phosphorylation and 2) wild type arrestin-3 but not a T382A mutant is phosphorylated by CKII in vitro. Functional analysis reveals that mutants mimicking the phosphorylated (T382E) and dephosphorylated (T382A or T382V) states of arrestin-3 promote beta(2)-adrenergic receptor internalization and bind clathrin, beta-adaptin, and Src to comparable levels as wild type arrestin-3. This suggests that the phosphorylation of arrestin-3 does not directly regulate interaction with endocytic (clathrin, beta-adaptin) or signaling (Src) components and is in contrast to arrestin-2, where phosphorylation appears to regulate interaction with clathrin and Src. However, additional analysis reveals that arrestin-3 phosphorylation may regulate formation of a large arrestin-3-containing protein complex. Differences between the regulatory roles of arrestin-2 and -3 phosphorylation may contribute to the different cellular functions of these proteins in G protein-coupled receptor signaling and regulation.

Authors
Kim, Y-M; Barak, LS; Caron, MG; Benovic, JL
MLA Citation
Kim, Y-M, Barak, LS, Caron, MG, and Benovic, JL. "Regulation of arrestin-3 phosphorylation by casein kinase II." J Biol Chem 277.19 (May 10, 2002): 16837-16846.
PMID
11877451
Source
pubmed
Published In
The Journal of biological chemistry
Volume
277
Issue
19
Publish Date
2002
Start Page
16837
End Page
16846
DOI
10.1074/jbc.M201379200

Behavioral and neurochemical effects of wild-type and mutated human alpha-synuclein in transgenic mice.

Human alpha-synuclein (halpha-SYN) is implicated in the Parkinson's disease phenotype (PDP) based on a variety of studies in man, animal models, and in vitro studies. The normal function of halpha-SYN and the mechanism by which it contributes to the PDP remains unclear. We created transgenic mice expressing either wild-type (hwalpha-SYN) or a doubly mutated (hm2alpha-SYN) form of halpha-SYN under control of the 9-kb rat tyrosine hydroxylase promoter. These mice expressed halpha-SYN in cell bodies, axons, and terminals of the nigrostriatal system. The expression of halpha-SYN in nigrostriatal terminals produced effects in both constructs resulting in increased density of the dopamine transporter and enhanced toxicity to the neurotoxin MPTP. Expression of hm2alpha-SYN reduced locomotor responses to repeated doses of amphetamine and blocked the development of sensitization. Adult hwalpha-SYN-5 transgenic mice had unremarkable dopaminergic axons and terminals, normal age-related measures on two motor coordination screens, and normal age-related measures of dopamine (DA) and its metabolites. Adult hm2alpha-SYN-39 transgenic mice had abnormal axons and terminals, age-related impairments in motor coordination, and age-related reductions in DA and its metabolites. Expression of hm2alpha-SYN adversely affects the integrity of dopaminergic terminals and leads to age-related declines in motor coordination and dopaminergic markers.

Authors
Richfield, EK; Thiruchelvam, MJ; Cory-Slechta, DA; Wuertzer, C; Gainetdinov, RR; Caron, MG; Di Monte, DA; Federoff, HJ
MLA Citation
Richfield, EK, Thiruchelvam, MJ, Cory-Slechta, DA, Wuertzer, C, Gainetdinov, RR, Caron, MG, Di Monte, DA, and Federoff, HJ. "Behavioral and neurochemical effects of wild-type and mutated human alpha-synuclein in transgenic mice." Exp Neurol 175.1 (May 2002): 35-48.
PMID
12009758
Source
pubmed
Published In
Experimental Neurology
Volume
175
Issue
1
Publish Date
2002
Start Page
35
End Page
48
DOI
10.1006/exnr.2002.7882

beta-Arrestin/AP-2 interaction in G protein-coupled receptor internalization: identification of a beta-arrestin binging site in beta 2-adaptin.

beta-Arrestins, proteins involved in the turn-off of G protein-coupled receptor (GPCR) activation, bind to the beta(2)-adaptin subunit of the clathrin adaptor AP-2. The interaction of beta(2)-adaptin with beta-arrestin involves critical arginine residues in the C-terminal domain of beta-arrestin and plays an important role in initiating clathrin-mediated endocytosis of the beta(2)-adrenergic receptor (beta(2)AR) (Laporte, S. A., Oakley, R. H., Holt, J. A., Barak, L. S., and Caron, M. G. (2000) J. Biol. Chem. 275, 23120--23126). However, the beta-arrestin-binding site in beta(2)-adaptin has not been identified, and little is known about the role of beta-arrestin/AP-2 interaction in the endocytosis of other GPCRs. Using in vitro binding assays, we have identified two glutamate residues (Glu-849 and Glu-902) in beta(2)-adaptin that are important in beta-arrestin binding. These residues are located in the platform subdomain of the C terminus of beta(2)-adaptin, where accessory/adapter endocytic proteins for other classes of receptors interact, distinct from the main site where clathrin interacts. The functional significance of the beta-arrestin/AP-2/clathrin complex in the endocytosis of GPCRs such as the beta(2)AR and vasopressin type II receptor was evaluated using mutant constructs of the beta(2)-adaptin C terminus containing either the clathrin and the beta-arrestin binding domains or the beta-arrestin-binding domain alone. When expressed in human embryonic kidney 293 cells, both constructs acted as dominant negatives inhibiting the agonist-induced internalization of the beta(2)AR and the vasopressin type II receptor. In addition, although the beta(2)-adaptin construct containing both the clathrin and beta-arrestin binding domains was able to block the endocytosis of transferrin receptors, a beta(2)-adaptin construct capable of associating with beta-arrestin but lacking its high affinity clathrin interaction did not interfere with transferrin receptor endocytosis. These results suggest that the interaction of beta-arrestin with beta(2)-adaptin represents a selective endocytic trigger for several members of the GPCR family.

Authors
Laporte, SA; Miller, WE; Kim, K-M; Caron, MG
MLA Citation
Laporte, SA, Miller, WE, Kim, K-M, and Caron, MG. "beta-Arrestin/AP-2 interaction in G protein-coupled receptor internalization: identification of a beta-arrestin binging site in beta 2-adaptin." J Biol Chem 277.11 (March 15, 2002): 9247-9254.
PMID
11777907
Source
pubmed
Published In
The Journal of biological chemistry
Volume
277
Issue
11
Publish Date
2002
Start Page
9247
End Page
9254
DOI
10.1074/jbc.M108490200

Endocytosis of G protein-coupled receptors: roles of G protein-coupled receptor kinases and beta-arrestin proteins.

Sequestration of G protein-coupled receptors from the cell surface is a commonly observed phenomenon following agonist-stimulation. This process is now believed to be important for receptor resensitization as well as for signal transduction. Over the years, numerous studies have aimed at understanding the molecular mechanisms underlying internalization. Proteins such as the G protein-coupled receptor kinases (GRKs) and the beta-arrestins, which were initially characterized as desensitizing molecules, have been shown to be important regulators of the endocytic process. Recently, numerous interacting partners have been identified for each of these two classes of proteins. However, the details regarding the sequence of these interactions and the cross-talk between signaling pathways containing the different protein complexes are just beginning to be uncovered. In this review, we summarize these findings and discuss the role of GRKs and beta-arrestins, two families of key regulatory proteins that regulate G protein-coupled receptor endocytosis.

Authors
Claing, A; Laporte, SA; Caron, MG; Lefkowitz, RJ
MLA Citation
Claing, A, Laporte, SA, Caron, MG, and Lefkowitz, RJ. "Endocytosis of G protein-coupled receptors: roles of G protein-coupled receptor kinases and beta-arrestin proteins." Prog Neurobiol 66.2 (February 2002): 61-79. (Review)
PMID
11900882
Source
pubmed
Published In
Progress in Neurobiology
Volume
66
Issue
2
Publish Date
2002
Start Page
61
End Page
79

Rab5 association with the angiotensin II type 1A receptor promotes Rab5 GTP binding and vesicular fusion.

Previous studies have demonstrated that the internalization of the angiotensin II type 1A receptor (AT(1A)R) may be mediated by both beta-arrestin-sensitive and -insensitive mechanisms. Therefore, we have used the AT(1A)R carboxyl-terminal tail to screen a rat brain yeast two-hybrid expression library for novel AT(1A)R-interacting proteins that might contribute to the regulation of AT(1A)R internalization. We have identified Rab5a as an AT(1A)R-binding protein that selectively associates with the AT(1A)R and not with the beta2-adrenergic receptor. A Rab5a-S34N mutant defective in GTP binding does not prevent the internalization of the AT(1A)R but does prevent the trafficking of the AT(1A)R into larger hollow cored vesicular structures. Agonist activation of the AT(1A)R promotes both the formation of Rab5a.AT(1A)R protein complexes and Rab5a GTP binding. Rab5a interactions with the AT(1A)R are mediated in part by the last 10 amino acid residues of the AT(1A)R carboxyl-terminal tail, and although a mutant receptor lacking these residues internalizes normally, it does not redistribute into larger hollow vesicles. Our data suggest that AT(1A)R activation modulates Rab5a activity leading to the homotypic fusion of endocytic vesicles. These observations suggest that vesicular cargo proteins, such as the AT(1A)R, may control their targeting between intracellular compartments by directly regulating the activity of components of the intracellular trafficking machinery such as Rab5a.

Authors
Seachrist, JL; Laporte, SA; Dale, LB; Babwah, AV; Caron, MG; Anborgh, PH; Ferguson, SSG
MLA Citation
Seachrist, JL, Laporte, SA, Dale, LB, Babwah, AV, Caron, MG, Anborgh, PH, and Ferguson, SSG. "Rab5 association with the angiotensin II type 1A receptor promotes Rab5 GTP binding and vesicular fusion." J Biol Chem 277.1 (January 4, 2002): 679-685.
PMID
11682489
Source
pubmed
Published In
The Journal of biological chemistry
Volume
277
Issue
1
Publish Date
2002
Start Page
679
End Page
685
DOI
10.1074/jbc.M109022200

Dopamine autoreceptor regulation of release and uptake in mouse brain slices in the absence of D(3) receptors.

The effects of the dopamine D(3) receptor, a putative autoreceptor, have been investigated by comparing behavioral and neurochemical properties of wild-type mice and mice with a genetic deletion of the D(3) receptor. The D(3) knock-out mice were modestly hyper-responsive to a novel environment relative to wild-type mice, and, consistent with this, quantitative in vivo microdialysis revealed elevated striatal dopamine extracellular levels. The dynamic actions of autoreceptors on electrically evoked dopamine release were examined in striatal brain slices from these animals and monitored with fast scan cyclic voltammetry at carbon-fiber microelectrodes. Quinpirole, a dopamine receptor agonist with potency at both D(2) and D(3) receptors, inhibited evoked dopamine in a dose-dependent manner with a slightly higher dose required in the knock-out animals (EC(50) of 60+/-10 nM in wild-type animals and 130+/-40 in D(3) knock-out animals; both curves had a Hill slope near 2). Dopamine synthesis inhibition with alpha-methyl-p-tyrosine caused released dopamine levels to decrease in each genotype. However, regulation of secretion by autoreceptors was still operant. Dose-response curves to quinpirole were unchanged in D(3) knock-out tissue, but secretion-regulated release exhibited a Hill slope decreased to 1 in the wild-type animals. In both genotypes, similar quinpirole-evoked increases in uptake rate were evident following synthesis inhibition. These data are consistent with the D(3) receptor having a small but significant role as a dopamine autoreceptor that partially regulates secretion, but not synthesis, in the caudate-putamen.

Authors
Joseph, JD; Wang, Y-M; Miles, PR; Budygin, EA; Picetti, R; Gainetdinov, RR; Caron, MG; Wightman, RM
MLA Citation
Joseph, JD, Wang, Y-M, Miles, PR, Budygin, EA, Picetti, R, Gainetdinov, RR, Caron, MG, and Wightman, RM. "Dopamine autoreceptor regulation of release and uptake in mouse brain slices in the absence of D(3) receptors." Neuroscience 112.1 (2002): 39-49.
PMID
12044470
Source
pubmed
Published In
Neuroscience
Volume
112
Issue
1
Publish Date
2002
Start Page
39
End Page
49

Cocaine mechanisms: Enhanced cocaine, fluoxetine and nisoxetine place preferences following monoamine transporter deletions

Cocaine blocks uptake by neuronal plasma membrane transporters for dopamine, serotonin and norepinephrine, producing subjective effects in humans that are both euphoric/rewarding and also fearful, jittery and aversive. Mice with gene knockouts of each of these transporters display cocaine reward, manifest by cocaine place preferences that are at least as great as wildtype values. Norepinephrine and serotonin receptor knockouts even display enhanced cocaine reward. One explanation for these observations could be that cocaine produces aversive or anhedonic effects by serotonin or norepinephrine receptor blockade in wildtype mice that are removed in serotonin or norepinephrine receptor knockouts, increasing net cocaine reward. Adaptations to removing one transporter could also change the rewarding valence of blocking the remaining transporters. To test these ideas, drugs that block serotonin transporter (fluoxetine), norepinephrine transporter (nisoxetine) or all three transporters (cocaine) were examined in single- or multiple-transporter knockout mice. Fluoxetine and nisoxetine acquire rewarding properties in several knockouts that are not observed in wildtype mice. Adding serotonin transporter knockout to norepinephrine transporter knockouts dramatically potentiates cocaine reward. These and previous data provide evidence that serotonin and norepinephrine transporter blockade can contribute to the net rewarding valence of cocaine. They identify neuroadaptations that may help to explain the retention of cocaine reward by dopamine and serotonin transporter knockout mice. They are consistent with emerging hypotheses that actions at the three primary brain molecular targets for cocaine each provide distinct contributions to cocaine reward and cocaine aversion in wildtype mice, and that this balance changes in mice that develop without dopamine, norepinephrine or serotonin transporters.

Authors
Hall, FS; Li, XF; Sora, I; Xu, F; Caron, M; Lesch, KP; Murphy, DL; Uhl, GR
MLA Citation
Hall, FS, Li, XF, Sora, I, Xu, F, Caron, M, Lesch, KP, Murphy, DL, and Uhl, GR. "Cocaine mechanisms: Enhanced cocaine, fluoxetine and nisoxetine place preferences following monoamine transporter deletions." Neuroscience 115.1 (2002): 153-161.
PMID
12401330
Source
scival
Published In
Neuroscience
Volume
115
Issue
1
Publish Date
2002
Start Page
153
End Page
161
DOI
10.1016/S0306-4522(02)00379-2

Erratum: Monoamine transporter pharmacology and mutant mice (Trends in Pharmacological Sciences (August 2002) (371))

Authors
Gainetdinov, RR; Sotnikova, TD; Caron, MG
MLA Citation
Gainetdinov, RR, Sotnikova, TD, and Caron, MG. "Erratum: Monoamine transporter pharmacology and mutant mice (Trends in Pharmacological Sciences (August 2002) (371))." Trends in Pharmacological Sciences 23.9 (2002): 443--.
Source
scival
Published In
Trends in Pharmacological Sciences
Volume
23
Issue
9
Publish Date
2002
Start Page
443-

Hyperactivity, elevated dopaminergic transmission, and response to amphetamine in M1 muscarinic acetylcholine receptor-deficient mice.

Acetylcholine serves an important modulatory role in the central nervous system. Pharmacological evidence has suggested that cholinergic activity can modulate central dopaminergic transmission; however, the nature of this interaction and the receptors involved remain undefined. In this study we have generated mice lacking the M1 muscarinic acetylcholine receptor and examined the effects of M1 deletion on dopaminergic transmission and locomotor behavior. We report that M1 deficiency leads to elevated dopaminergic transmission in the striatum and significantly increased locomotor activity. M1-deficient mice also have an increased response to the stimulatory effects of amphetamine. Our results provide direct evidence for regulation of dopaminergic transmission by the M1 receptor and are consistent with the idea that M1 dysfunction could be a contributing factor in psychiatric disorders in which altered dopaminergic transmission has been implicated.

Authors
Gerber, DJ; Sotnikova, TD; Gainetdinov, RR; Huang, SY; Caron, MG; Tonegawa, S
MLA Citation
Gerber, DJ, Sotnikova, TD, Gainetdinov, RR, Huang, SY, Caron, MG, and Tonegawa, S. "Hyperactivity, elevated dopaminergic transmission, and response to amphetamine in M1 muscarinic acetylcholine receptor-deficient mice." Proc Natl Acad Sci U S A 98.26 (December 18, 2001): 15312-15317.
PMID
11752469
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
98
Issue
26
Publish Date
2001
Start Page
15312
End Page
15317
DOI
10.1073/pnas.261583798

The NH(2)-terminus of norepinephrine transporter contains a basolateral localization signal for epithelial cells.

When expressed in epithelial cells, dopamine transporter (DAT) was detected predominantly in the apical plasma membrane, whereas norepinephrine transporter (NET) was found in the basolateral membrane, despite 67% overall amino acid sequence identity. To identify possible localization signals responsible for this difference, DAT-NET chimeras were expressed in MDCK cells and localized by immunocytochemistry and transport assays. The results suggested that localization of these transporters in MDCK cells depends on their highly divergent NH(2)-terminal regions. Deletion of the first 58 amino acids of DAT (preceding TM1) did not change its apical localization. However, the replacement of that region with corresponding sequence from NET resulted in localization of the chimeric protein to the basolateral membrane, suggesting that the NH(2)-terminus of NET, which contains two dileucine motifs, contains a basolateral localization signal. Mutation of these leucines to alanines in the context of a basolaterally localized NET/DAT chimera restored transporter localization to the apical membrane, indicating that the dileucine motifs are critical to the basolateral localization signal embodied within the NET NH(2)-terminal region. However, the same mutation in the context of wild-type NET did not disrupt basolateral localization, indicating the presence of additional signals in NET directing its basolateral localization within the plasma membrane.

Authors
Gu, HH; Wu, X; Giros, B; Caron, MG; Caplan, MJ; Rudnick, G
MLA Citation
Gu, HH, Wu, X, Giros, B, Caron, MG, Caplan, MJ, and Rudnick, G. "The NH(2)-terminus of norepinephrine transporter contains a basolateral localization signal for epithelial cells." Mol Biol Cell 12.12 (December 2001): 3797-3807.
PMID
11739781
Source
pubmed
Published In
Molecular Biology of the Cell
Volume
12
Issue
12
Publish Date
2001
Start Page
3797
End Page
3807

Differential regulation of the dopamine D2 and D3 receptors by G protein-coupled receptor kinases and beta-arrestins.

The D(2) and D(3) receptors (D(2)R and D(3)R), which are potential targets for antipsychotic drugs, have a similar structural architecture and signaling pathway. Furthermore, in some brain regions they are expressed in the same cells, suggesting that differences between the two receptors might lie in other properties such as their regulation. In this study we investigated, using COS-7 and HEK-293 cells, the mechanism underlying the intracellular trafficking of the D(2)R and D(3)R. Activation of D(2)R caused G protein-coupled receptor kinase-dependent receptor phosphorylation, a robust translocation of beta-arrestin to the cell membrane, and profound receptor internalization. The internalization of the D(2)R was dynamin-dependent, suggesting that a clathrin-coated endocytic pathway is involved. In addition, the D(2)R, upon agonist-mediated internalization, localized to intracellular compartments distinct from those utilized by the beta(2)-adrenergic receptor. However, in the case of the D(3)R, only subtle agonist-mediated receptor phosphorylation, beta-arrestin translocation to the plasma membrane, and receptor internalization were observed. Interchange of the second and third intracellular loops of the D(2)R and D(3)R reversed their phenotypes, implicating these regions in the regulatory properties of the two receptors. Our studies thus indicate that functional distinctions between the D(2)R and D(3)R may be found in their desensitization and cellular trafficking properties. The differences in their regulatory properties suggest that they have distinct physiological roles in the brain.

Authors
Kim, KM; Valenzano, KJ; Robinson, SR; Yao, WD; Barak, LS; Caron, MG
MLA Citation
Kim, KM, Valenzano, KJ, Robinson, SR, Yao, WD, Barak, LS, and Caron, MG. "Differential regulation of the dopamine D2 and D3 receptors by G protein-coupled receptor kinases and beta-arrestins." J Biol Chem 276.40 (October 5, 2001): 37409-37414.
PMID
11473130
Source
pubmed
Published In
The Journal of biological chemistry
Volume
276
Issue
40
Publish Date
2001
Start Page
37409
End Page
37414
DOI
10.1074/jbc.M106728200

Catecholamine release and uptake in the mouse prefrontal cortex.

Monitoring the release and uptake of catecholamines from terminals in weakly innervated brain regions is an important step in understanding their importance in normal brain function. To that end, we have labeled brain slices from transgenic mice that synthesize placental alkaline phosphatase (PLAP) on neurons containing tyrosine hydroxylase with antibody-fluorochrome conjugate, PLAP-Cy5. Excitation of the fluorochrome enables catecholamine neurons to be visualized in living tissue. Immunohistochemical fluorescence with antibodies to tyrosine hydroxylase and dopamine beta-hydroxylase revealed that the PLAP labeling was specific to catecholamine neurons. In the prefrontal cortex (PFC), immunohistochemical fluorescence of the PLAP along with staining for dopamine transporter (DAT) and norepinephrine transporter (NET) revealed that all three exhibit remarkable spatial overlap. Fluorescence from the PLAP antibody was used to position carbon-fiber microelectrodes adjacent to catecholamine neurons in the PFC. Following incubation with L-DOPA, catecholamine release and subsequent uptake was measured and the effect of uptake inhibitors examined. Release and uptake in NET and DAT knockout mice were also monitored. Uptake rates in the cingulate and prelimbic cortex are so slow that catecholamines can exist in the extracellular fluid for sufficient time to travel approximately 100 microm. The results support heterologous uptake of catecholamines and volume transmission in the PFC of mice.

Authors
Mundorf, ML; Joseph, JD; Austin, CM; Caron, MG; Wightman, RM
MLA Citation
Mundorf, ML, Joseph, JD, Austin, CM, Caron, MG, and Wightman, RM. "Catecholamine release and uptake in the mouse prefrontal cortex." J Neurochem 79.1 (October 2001): 130-142.
PMID
11595765
Source
pubmed
Published In
Journal of Neurochemistry
Volume
79
Issue
1
Publish Date
2001
Start Page
130
End Page
142

Glutamatergic modulation of hyperactivity in mice lacking the dopamine transporter.

In the brain, dopamine exerts an important modulatory influence over behaviors such as emotion, cognition, and affect as well as mechanisms of reward and the control of locomotion. The dopamine transporter (DAT), which reuptakes the released neurotransmitter into presynaptic terminals, is a major determinant of the intensity and duration of the dopaminergic signal. Knockout mice lacking the dopamine transporter (DAT-KO mice) display marked changes in dopamine homeostasis that result in elevated dopaminergic tone and pronounced locomotor hyperactivity. A feature of DAT-KO mice is that their hyperactivity can be inhibited by psychostimulants and serotonergic drugs. The pharmacological effect of these drugs occurs without any observable changes in dopaminergic parameters, suggesting that other neurotransmitter systems in addition to dopamine might contribute to the control of locomotion in these mice. We report here that the hyperactivity of DAT-KO mice can be markedly further enhanced when N-methyl-d-aspartate receptor-mediated glutamatergic transmission is blocked. Conversely, drugs that enhance glutamatergic transmission, such as positive modulators of l-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate glutamate receptors, suppress the hyperactivity of DAT-KO mice. Interestingly, blockade of N- methyl-d-aspartate receptors prevented the inhibitory effects of both psychostimulant and serotonergic drugs on hyperactivity. These findings support the concept of a reciprocal functional interaction between dopamine and glutamate in the basal ganglia and suggest that agents modulating glutamatergic transmission may represent an approach to manage conditions associated with dopaminergic dysfunction.

Authors
Gainetdinov, RR; Mohn, AR; Bohn, LM; Caron, MG
MLA Citation
Gainetdinov, RR, Mohn, AR, Bohn, LM, and Caron, MG. "Glutamatergic modulation of hyperactivity in mice lacking the dopamine transporter." Proc Natl Acad Sci U S A 98.20 (September 25, 2001): 11047-11054.
PMID
11572967
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
98
Issue
20
Publish Date
2001
Start Page
11047
End Page
11054
DOI
10.1073/pnas.191353298

Genetic animal models: focus on schizophrenia.

The neurobiology of schizophrenia remains poorly understood. Symptoms of schizophrenia are classically thought to be associated with an imbalance of the dopaminergic system. However, the contribution of other neurotransmitters, in particular glutamate, has been increasingly appreciated. The role of individual components of neurotransmitter systems in aberrant behaviors can be experimentally tested in transgenic animals. Dopamine transporter knockout mice display persistently elevated dopaminergic tone and therefore might be appropriate substrates to evaluate the dopamine hypothesis. Similarly, NMDA receptor-deficient mice can be used to evaluate the glutamate hypothesis of schizophrenia. In this review we discuss how such animal models might be relevant for understanding the neurochemical underpinnings of certain manifestations of schizophrenia.

Authors
Gainetdinov, RR; Mohn, AR; Caron, MG
MLA Citation
Gainetdinov, RR, Mohn, AR, and Caron, MG. "Genetic animal models: focus on schizophrenia." Trends Neurosci 24.9 (September 2001): 527-533. (Review)
PMID
11506886
Source
pubmed
Published In
Trends in Neurosciences
Volume
24
Issue
9
Publish Date
2001
Start Page
527
End Page
533

Following the trace of elusive amines.

Authors
Premont, RT; Gainetdinov, RR; Caron, MG
MLA Citation
Premont, RT, Gainetdinov, RR, and Caron, MG. "Following the trace of elusive amines." Proc Natl Acad Sci U S A 98.17 (August 14, 2001): 9474-9475.
PMID
11504935
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
98
Issue
17
Publish Date
2001
Start Page
9474
End Page
9475
DOI
10.1073/pnas.181356198

Signal transduction. Bringing channels closer to the action!

Authors
Laporte, SA; Oakley, RH; Caron, MG
MLA Citation
Laporte, SA, Oakley, RH, and Caron, MG. "Signal transduction. Bringing channels closer to the action!." Science 293.5527 (July 6, 2001): 62-63.
PMID
11441172
Source
pubmed
Published In
Science
Volume
293
Issue
5527
Publish Date
2001
Start Page
62
End Page
63
DOI
10.1126/science.1063104

Antiproliferative action of dopamine and norepinephrine in neuroblastoma cells expressing the human dopamine transporter.

Authors
Pifl, C; Zezula, J; Spittler, A; Kattinger, A; Reither, H; Caron, MG; Hornykiewicz, O
MLA Citation
Pifl, C, Zezula, J, Spittler, A, Kattinger, A, Reither, H, Caron, MG, and Hornykiewicz, O. "Antiproliferative action of dopamine and norepinephrine in neuroblastoma cells expressing the human dopamine transporter." FASEB J 15.9 (July 2001): 1607-1609.
PMID
11427501
Source
pubmed
Published In
The FASEB journal : official publication of the Federation of American Societies for Experimental Biology
Volume
15
Issue
9
Publish Date
2001
Start Page
1607
End Page
1609

Molecular determinants underlying the formation of stable intracellular G protein-coupled receptor-beta-arrestin complexes after receptor endocytosis*.

beta-Arrestins bind agonist-activated G protein-coupled receptors (GPCRs) and mediate their desensitization and internalization. Although beta-arrestins dissociate from some receptors at the plasma membrane, such as the beta2 adrenergic receptor, they remain associated with other GPCRs and internalize with them into endocytic vesicles. Formation of stable receptor-beta-arrestin complexes that persist inside the cell impedes receptor resensitization, and the aberrant formation of these complexes may play a role in GPCR-based diseases (Barak, L. S., Oakley, R. H., Laporte, S. A., and Caron, M. G. (2001) Proc. Natl. Acad. Sci. U. S. A. 98, 93-98). Here, we investigate the molecular determinants responsible for sustained receptor/beta-arrestin interactions. We show in real time and in live human embryonic kidney (HEK-293) cells that a beta-arrestin-2-green fluorescent protein conjugate internalizes into endocytic vesicles with agonist-activated neurotensin-1 receptor, oxytocin receptor, angiotensin II type 1A receptor, and substance P receptor. Using receptor mutagenesis, we demonstrate that the ability of beta-arrestin to remain associated with these receptors is mediated by specific clusters of serine and threonine residues located in the receptor carboxyl-terminal tail. These clusters are remarkably conserved in their position within the carboxyl-terminal domain and serve as primary sites of agonist-dependent receptor phosphorylation. In addition, we identify a beta-arrestin mutant with enhanced affinity for the agonist-activated beta2-adrenergic receptor that traffics into endocytic vesicles with receptors that lack serine/threonine clusters and normally dissociate from wild-type beta-arrestin at the plasma membrane. By identifying receptor and beta-arrestin residues critical for the formation of stable receptor-beta-arrestin complexes, these studies provide novel targets for regulating GPCR responsiveness and treating diseases resulting from abnormal GPCR/beta-arrestin interactions.

Authors
Oakley, RH; Laporte, SA; Holt, JA; Barak, LS; Caron, MG
MLA Citation
Oakley, RH, Laporte, SA, Holt, JA, Barak, LS, and Caron, MG. "Molecular determinants underlying the formation of stable intracellular G protein-coupled receptor-beta-arrestin complexes after receptor endocytosis*." J Biol Chem 276.22 (June 1, 2001): 19452-19460.
PMID
11279203
Source
pubmed
Published In
The Journal of biological chemistry
Volume
276
Issue
22
Publish Date
2001
Start Page
19452
End Page
19460
DOI
10.1074/jbc.M101450200

Agonist-dependent recruitment of phosphoinositide 3-kinase to the membrane by beta-adrenergic receptor kinase 1. A role in receptor sequestration.

Agonist-dependent desensitization of the beta-adrenergic receptor requires translocation and activation of the beta-adrenergic receptor kinase1 by liberated Gbetagamma subunits. Subsequent internalization of agonist-occupied receptors occurs as a result of the binding of beta-arrestin to the phosphorylated receptor followed by interaction with the AP2 adaptor and clathrin proteins. Receptor internalization is known to require D-3 phosphoinositides that are generated by the action of phosphoinositide 3-kinase. Phosphoinositide 3-kinases form a family of lipid kinases that couple signals via receptor tyrosine kinases and G-protein-coupled receptors. The molecular mechanism by which phosphoinositide 3-kinase acts to promote beta-adrenergic receptor internalization is not well understood. In the present investigation we demonstrate a novel finding that beta-adrenergic receptor kinase 1 and phosphoinositide 3-kinase form a cytosolic complex, which leads to beta-adrenergic receptor kinase 1-mediated translocation of phosphoinositide 3-kinase to the membrane in an agonist-dependent manner. Furthermore, agonist-induced translocation of phosphoinositide 3-kinase results in rapid interaction with the receptor, which is of functional importance, since inhibition of phosphoinositide 3-kinase activity attenuates beta-adrenergic receptor sequestration. Therefore, agonist-dependent recruitment of phosphoinositide 3-kinase to the membrane is an important step in the process of receptor sequestration and links phosphoinositide 3-kinase to G-protein-coupled receptor activation and sequestration.

Authors
Naga Prasad, SV; Barak, LS; Rapacciuolo, A; Caron, MG; Rockman, HA
MLA Citation
Naga Prasad, SV, Barak, LS, Rapacciuolo, A, Caron, MG, and Rockman, HA. "Agonist-dependent recruitment of phosphoinositide 3-kinase to the membrane by beta-adrenergic receptor kinase 1. A role in receptor sequestration." J Biol Chem 276.22 (June 1, 2001): 18953-18959.
PMID
11259422
Source
pubmed
Published In
The Journal of biological chemistry
Volume
276
Issue
22
Publish Date
2001
Start Page
18953
End Page
18959
DOI
10.1074/jbc.M102376200

Functional interaction between monoamine plasma membrane transporters and the synaptic PDZ domain-containing protein PICK1.

PDZ domain-containing proteins play an important role in the targeting and localization of synaptic membrane proteins. Here, we report an interaction between the PDZ domain-containing protein PICK1 and monoamine neurotransmitter transporters in vitro and in vivo. In dopaminergic neurons, PICK1 colocalizes with the dopamine transporter (DAT) and forms a stable protein complex. Coexpression of PICK1 with DAT in mammalian cells and neurons in culture results in colocalization of the two proteins in a cluster pattern and an enhancement of DAT uptake activity through an increase in the number of plasma membrane DAT. Deletion of the PDZ binding site at the carboxyl terminus of DAT abolishes its association with PICK1 and impairs the localization of the transporter in neurons. These findings indicate a role for PDZ-mediated protein interactions in the localization, expression, and function of monoamine transporters.

Authors
Torres, GE; Yao, WD; Mohn, AR; Quan, H; Kim, KM; Levey, AI; Staudinger, J; Caron, MG
MLA Citation
Torres, GE, Yao, WD, Mohn, AR, Quan, H, Kim, KM, Levey, AI, Staudinger, J, and Caron, MG. "Functional interaction between monoamine plasma membrane transporters and the synaptic PDZ domain-containing protein PICK1." Neuron 30.1 (April 2001): 121-134.
PMID
11343649
Source
pubmed
Published In
Neuron
Volume
30
Issue
1
Publish Date
2001
Start Page
121
End Page
134

Role of the sphingosine-1-phosphate receptor EDG-1 in PDGF-induced cell motility.

EDG-1 is a heterotrimeric guanine nucleotide binding protein-coupled receptor (GPCR) for sphingosine-1-phosphate (SPP). Cell migration toward platelet-derived growth factor (PDGF), which stimulates sphingosine kinase and increases intracellular SPP, was dependent on expression of EDG-1. Deletion of edg-1 or inhibition of sphingosine kinase suppressed chemotaxis toward PDGF and also activation of the small guanosine triphosphatase Rac, which is essential for protrusion of lamellipodia and forward movement. Moreover, PDGF activated EDG-1, as measured by translocation of beta-arrestin and phosphorylation of EDG-1. Our results reveal a role for receptor cross-communication in which activation of a GPCR by a receptor tyrosine kinase is critical for cell motility.

Authors
Hobson, JP; Rosenfeldt, HM; Barak, LS; Olivera, A; Poulton, S; Caron, MG; Milstien, S; Spiegel, S
MLA Citation
Hobson, JP, Rosenfeldt, HM, Barak, LS, Olivera, A, Poulton, S, Caron, MG, Milstien, S, and Spiegel, S. "Role of the sphingosine-1-phosphate receptor EDG-1 in PDGF-induced cell motility." Science 291.5509 (March 2, 2001): 1800-1803.
PMID
11230698
Source
pubmed
Published In
Science
Volume
291
Issue
5509
Publish Date
2001
Start Page
1800
End Page
1803
DOI
10.1126/science.1057559

Genetics of childhood disorders: XXIV. ADHD, part 8: hyperdopaminergic mice as an animal model of ADHD.

Authors
Gainetdinov, RR; Caron, MG
MLA Citation
Gainetdinov, RR, and Caron, MG. "Genetics of childhood disorders: XXIV. ADHD, part 8: hyperdopaminergic mice as an animal model of ADHD." J Am Acad Child Adolesc Psychiatry 40.3 (March 2001): 380-382. (Review)
PMID
11288782
Source
pubmed
Published In
Journal of the American Academy of Child and Adolescent Psychiatry
Volume
40
Issue
3
Publish Date
2001
Start Page
380
End Page
382

Hyperactivity and impaired response habituation in hyperdopaminergic mice.

Abnormal dopaminergic transmission is implicated in schizophrenia, attention deficit hyperactivity disorder, and drug addiction. In an attempt to model aspects of these disorders, we have generated hyperdopaminergic mutant mice by reducing expression of the dopamine transporter (DAT) to 10% of wild-type levels (DAT knockdown). Fast-scan cyclic voltammetry and in vivo microdialysis revealed that released dopamine was cleared at a slow rate in knockdown mice, which resulted in a higher extracellular dopamine concentration. Unlike the DAT knockout mice, the DAT knockdown mice do not display a growth retardation phenotype. They have normal home cage activity but display hyperactivity and impaired response habituation in novel environments. In addition, we show that both the indirect dopamine receptor agonist amphetamine and the direct agonists apomorphine and quinpirole inhibit locomotor activity in the DAT knockdown mice, leading to the hypothesis that a shift in the balance between dopamine auto and heteroreceptor function may contribute to the therapeutic effect of psychostimulants in attention deficit hyperactivity disorder.

Authors
Zhuang, X; Oosting, RS; Jones, SR; Gainetdinov, RR; Miller, GW; Caron, MG; Hen, R
MLA Citation
Zhuang, X, Oosting, RS, Jones, SR, Gainetdinov, RR, Miller, GW, Caron, MG, and Hen, R. "Hyperactivity and impaired response habituation in hyperdopaminergic mice." Proc Natl Acad Sci U S A 98.4 (February 13, 2001): 1982-1987.
PMID
11172062
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
98
Issue
4
Publish Date
2001
Start Page
1982
End Page
1987
DOI
10.1073/pnas.98.4.1982

Constitutive arrestin-mediated desensitization of a human vasopressin receptor mutant associated with nephrogenic diabetes insipidus.

Agonist-dependent desensitization and internalization of G protein-coupled receptors (GPCR) are mediated by the binding of arrestins to phosphorylated receptors. The affinity of arrestins for the phosphorylated GPCR regulates the ability of the internalized receptor to be dephosphorylated and recycled back to the plasma membrane. In this study, we show that the naturally occurring loss of function vasopressin receptor mutation R137H, which is associated with familial nephrogenic diabetes insipidus, induces constitutive arrestin-mediated desensitization. In contrast to the wild-type vasopressin receptor, the nonsignaling R137H receptor is phosphorylated and sequestered in arrestin-associated intracellular vesicles even in the absence of agonist. Eliminating molecular determinants on the receptor that promote high affinity arrestin-receptor interaction reestablishes plasma membrane localization and the ability of the mutated receptors to signal. These findings suggest that unregulated desensitization can contribute to the etiology of a GPCR-based disease, implying that pharmacological targeting of GPCR desensitization may be therapeutically beneficial.

Authors
Barak, LS; Oakley, RH; Laporte, SA; Caron, MG
MLA Citation
Barak, LS, Oakley, RH, Laporte, SA, and Caron, MG. "Constitutive arrestin-mediated desensitization of a human vasopressin receptor mutant associated with nephrogenic diabetes insipidus." Proc Natl Acad Sci U S A 98.1 (January 2, 2001): 93-98.
PMID
11134505
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
98
Issue
1
Publish Date
2001
Start Page
93
End Page
98
DOI
10.1073/pnas.011303698

Prepulse inhibition deficits and perseverative motor patterns in dopamine transporter knock-out mice: differential effects of D1 and D2 receptor antagonists.

Dopamine is known to regulate several behavioral phenomena, including sensorimotor gating and aspects of motor activity. The roles of dopamine D1 and D2 receptors in these behaviors have been documented in the rat literature, but few reports exist on their role in mice. We used dopamine transporter (DAT) (-/-) mice to examine the behavioral consequences of a chronically hyperdopaminergic state, challenging them with the preferential dopamine D2 receptor antagonist raclopride and D1 receptor antagonist SCH23390. At baseline, DAT (-/-) mice exhibited deficient sensorimotor gating as measured by prepulse inhibition (PPI) of the startle response, exhibited nonfocal preservative patterns of locomotion, and were hyperactive in a novel environment. Pretreatment with raclopride significantly increased PPI in the DAT (-/-) mice, whereas SCH23390 had no significant effect. Blockade of D2 receptors did not affect the predominantly straight patterns of motor behavior produced by the DAT (-/-) mice, but antagonism of D1 receptors significantly attenuated the preservative patterns, producing more of a meandering behavior seen in the DAT (+/+) control mice. Both D1 and D2 receptor antagonists decreased the hyperactivity seen in the DAT (-/-) mice. These findings support the role of the D2, but not the D1, receptor in the modulation of PPI in mice. Furthermore, D1 receptor activation appears to be the critical substrate for the expression of preservative patterns of motor behavior, whereas both D1 and D2 receptors appear to regulate the amount of motor activity.

Authors
Ralph, RJ; Paulus, MP; Fumagalli, F; Caron, MG; Geyer, MA
MLA Citation
Ralph, RJ, Paulus, MP, Fumagalli, F, Caron, MG, and Geyer, MA. "Prepulse inhibition deficits and perseverative motor patterns in dopamine transporter knock-out mice: differential effects of D1 and D2 receptor antagonists." J Neurosci 21.1 (January 1, 2001): 305-313.
PMID
11150348
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
21
Issue
1
Publish Date
2001
Start Page
305
End Page
313

The NH2-terminus of norepinephrine transporter contains a basolateral localization signal for epithelial cells

When expressed in epithelial cells, dopamine transporter (DAT) was detected predominantly in the apical plasma membrane, whereas norepinephrine transporter (NET) was found in the basolateral membrane, despite 67% overall amino acid sequence identity. To identify possible localization signals responsible for this difference, DAT-NET chimeras were expressed in MDCK cells and localized by immunocytochemistry and transport assays. The results suggested that localization of these transporters in MDCK cells depends on their highly divergent NH2-terminal regions. Deletion of the first 58 amino acids of DAT (preceding TM1) did not change its apical localization. However, the replacement of that region with corresponding sequence from NET resulted in localization of the chimeric protein to the basolateral membrane, suggesting that the NH2-terminus of NET, which contains two dileucine motifs, contains a basolateral localization signal. Mutation of these leucines to alanines in the context of a basolaterally localized NET/DAT chimera restored transporter localization to the apical membrane, indicating that the dileucine motifs are critical to the basolateral localization signal embodied within the NET NH2-terminal region. However, the same mutation in the context of wild-type NET did not disrupt basolateral localization, indicating the presence of additional signals in NET directing its basolateral localization within the plasma membrane.

Authors
Gu, HH; Wu, X; Giros, B; Caron, MG; Caplan, MJ; Rudnick, G
MLA Citation
Gu, HH, Wu, X, Giros, B, Caron, MG, Caplan, MJ, and Rudnick, G. "The NH2-terminus of norepinephrine transporter contains a basolateral localization signal for epithelial cells." Molecular Biology of the Cell 12.12 (2001): 3797-3807.
Source
scival
Published In
Molecular Biology of the Cell
Volume
12
Issue
12
Publish Date
2001
Start Page
3797
End Page
3807

Potentiated opioid analgesia in norepinephrine transporter knock-out mice.

Several studies have shown that activation of alpha(2)-adrenergic receptors (alpha(2)ARs) leads to mild analgesic effects. Tricyclic antidepressants (TCAs), such as desipramine (DMI), which block norepinephrine transporters (NETs), also produce mild antinociception. The coadministration of either alpha(2)AR agonists or TCAs with opiates produces synergistically potentiated antinociception. It has been postulated that the analgesic effects of TCAs are determined by their ability to inhibit norepinephrine reuptake via interactions with the NET. To test this idea, we studied mice lacking a functional NET in spontaneous and morphine-induced antinociceptive paradigms. Morphine (10 mg/kg, s.c. ) treatment produced greater analgesia, as assayed in the warm water tail-flick assay, in NET-knock-out (-KO) mice than in wild-type (WT) mice. As anticipated, yohimbine, an inhibitor of alpha(2)ARs, blocked this potentiation. Moreover, a warm water swim-stress paradigm, which is known to induce the release of endogenous opioids, produced greater antinociception in NET-KO than in the WT mice. Naloxone, an inhibitor of opioid receptors, blocked the development of the swim-evoked analgesia in both WT and NET-KO mice, confirming the involvement of the endogenous opioid system. In the NET-KO mice, DMI did not further enhance analgesia but was still able to produce inhibitory effects on the locomotor activity of these mutants, suggesting that the effects of this TCA are not exclusively via interactions with the NET. In summary, these results demonstrate in a genetic model that both endogenous and exogenous opiate-mediated analgesia can be enhanced by elimination of the NET, indicating that the interaction of TCAs with NET mediates these effects.

Authors
Bohn, LM; Xu, F; Gainetdinov, RR; Caron, MG
MLA Citation
Bohn, LM, Xu, F, Gainetdinov, RR, and Caron, MG. "Potentiated opioid analgesia in norepinephrine transporter knock-out mice." J Neurosci 20.24 (December 15, 2000): 9040-9045.
PMID
11124980
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
20
Issue
24
Publish Date
2000
Start Page
9040
End Page
9045

Mu-opioid receptor desensitization by beta-arrestin-2 determines morphine tolerance but not dependence.

Morphine is a powerful pain reliever, but also a potent inducer of tolerance and dependence. The development of opiate tolerance occurs on continued use of the drug such that the amount of drug required to elicit pain relief must be increased to compensate for diminished responsiveness. In many systems, decreased responsiveness to agonists has been correlated with the desensitization of G-protein-coupled receptors. In vitro evidence indicates that this process involves phosphorylation of G-protein-coupled receptors and subsequent binding of regulatory proteins called beta-arrestins. Using a knockout mouse lacking beta-arrestin-2 (beta arr2-/-), we have assessed the contribution of desensitization of the mu-opioid receptor to the development of morphine antinociceptive tolerance and the subsequent onset of physical dependence. Here we show that in mice lacking beta-arrestin-2, desensitization of the mu-opioid receptor does not occur after chronic morphine treatment, and that these animals fail to develop antinociceptive tolerance. However, the deletion of beta-arrestin-2 does not prevent the chronic morphine-induced up-regulation of adenylyl cyclase activity, a cellular marker of dependence, and the mutant mice still become physically dependent on the drug.

Authors
Bohn, LM; Gainetdinov, RR; Lin, FT; Lefkowitz, RJ; Caron, MG
MLA Citation
Bohn, LM, Gainetdinov, RR, Lin, FT, Lefkowitz, RJ, and Caron, MG. "Mu-opioid receptor desensitization by beta-arrestin-2 determines morphine tolerance but not dependence." Nature 408.6813 (December 7, 2000): 720-723.
PMID
11130073
Source
pubmed
Published In
Nature
Volume
408
Issue
6813
Publish Date
2000
Start Page
720
End Page
723
DOI
10.1038/35047086

Reply: receptor specificity of G-protein-coupled receptor kinases

Authors
Gainetdinov, RR; Premont, RT; Caron, MG; Lefkowitz, RJ
MLA Citation
Gainetdinov, RR, Premont, RT, Caron, MG, and Lefkowitz, RJ. "Reply: receptor specificity of G-protein-coupled receptor kinases." Trends Pharmacol Sci 21.10 (October 2000): 366-367.
PMID
11050311
Source
pubmed
Published In
Trends in Pharmacological Sciences
Volume
21
Issue
10
Publish Date
2000
Start Page
366
End Page
367

Dopamine D(5) receptor immunolocalization in rat and monkey brain.

Dopamine D(5) receptor localization has been difficult because even the most specific ligands cannot distinguish between molecular subtypes of the D(1)-like receptor subfamily. Antifusion protein rabbit polyclonal antibodies directed against the C-terminus of human D(5) receptor were therefore developed for immunolocalization of the D(5) receptor protein in brain. The antibodies were characterized by immunoblot analysis and immunoprecipitation and used for light microscopic immunocytochemistry in rat and monkey brain. Affinity purified D(5) antibodies were specific for D(5) fusion protein as well as cloned and native D(5) receptor on Western blots, and D(5) antisera specifically immunoprecipitated solubilized, cloned D(5) receptor. Regional distribution of D(5) receptor immunoreactivity was consistent across species and correlated well with D(5) mRNA distribution previously reported in monkey brain. Immunoreactivity was widespread and tended to label perikarya and proximal dendrites of neurons in cerebral cortex, basal ganglia, basal forebrain, hippocampus, diencephalon, brainstem, and cerebellum. Neuropil was immunoreactive in olfactory bulb, islands of Calleja, cerebral cortex, superior colliculus, and molecular layer of cerebellum. The distribution of D(5) in brain was clearly different from that of other dopamine receptor subtypes, including D(1), the other member of the D(1)-like receptor subfamily. This unique distribution corroborates the idea that the D(5) receptor subtype has a distinct role in dopamine neurotransmission.

Authors
Ciliax, BJ; Nash, N; Heilman, C; Sunahara, R; Hartney, A; Tiberi, M; Rye, DB; Caron, MG; Niznik, HB; Levey, AI
MLA Citation
Ciliax, BJ, Nash, N, Heilman, C, Sunahara, R, Hartney, A, Tiberi, M, Rye, DB, Caron, MG, Niznik, HB, and Levey, AI. "Dopamine D(5) receptor immunolocalization in rat and monkey brain." Synapse 37.2 (August 2000): 125-145.
PMID
10881034
Source
pubmed
Published In
Synapse
Volume
37
Issue
2
Publish Date
2000
Start Page
125
End Page
145
DOI
10.1002/1098-2396(200008)37:2<125::AID-SYN7>3.0.CO;2-7

Mice lacking the dopamine transporter display altered regulation of distal colonic motility.

The mechanisms by which dopamine (DA) influences gastrointestinal (GI) tract motility are incompletely understood and complicated by tissue- and species-specific differences in dopaminergic function. To improve the understanding of DA action on GI motility, we used an organ tissue bath system to characterize motor function of distal colonic smooth muscle segments from wild-type and DA transporter knockout (DAT -/-) mice. In wild-type mice, combined blockade of D(1) and D(2) receptors resulted in significant increases in tone (62 +/- 9%), amplitude of spontaneous phasic contractions (167 +/- 24%), and electric field stimulation (EFS)-induced (40 +/- 8%) contractions, suggesting that endogenous DA is inhibitory to mouse distal colonic motility. The amplitudes of spontaneous phasic and EFS-induced contractions were lower in DAT -/- mice relative to wild-type mice. These differences were eliminated by combined D(1) and D(2) receptor blockade, indicating that the inhibitory effects of DA on distal colonic motility are potentiated in DAT -/- mice. Motility index was decreased but spontaneous phasic contraction frequency was enhanced in DAT -/- mice relative to wild-type mice. The fact that spontaneous phasic and EFS-induced contractile activity were altered by the lack of the DA transporter suggests an important role for endogenous DA in modulating motility of mouse distal colon.

Authors
Walker, JK; Gainetdinov, RR; Mangel, AW; Caron, MG; Shetzline, MA
MLA Citation
Walker, JK, Gainetdinov, RR, Mangel, AW, Caron, MG, and Shetzline, MA. "Mice lacking the dopamine transporter display altered regulation of distal colonic motility." Am J Physiol Gastrointest Liver Physiol 279.2 (August 2000): G311-G318.
PMID
10915639
Source
pubmed
Published In
American journal of physiology. Gastrointestinal and liver physiology
Volume
279
Issue
2
Publish Date
2000
Start Page
G311
End Page
G318

The interaction of beta-arrestin with the AP-2 adaptor is required for the clustering of beta 2-adrenergic receptor into clathrin-coated pits.

Beta-arrestins are cytosolic proteins that regulate the signaling and the internalization of G protein-coupled receptors (GPCRs). Although termination of receptor coupling requires beta-arrestin binding to agonist-activated receptors, GPCR endocytosis involves the coordinate interactions between receptor-beta-arrestin complexes and other endocytic proteins such as adaptor protein 2 (AP-2) and clathrin. Clathrin interacts with a conserved motif in the beta-arrestin C-terminal tail; however, the specific molecular determinants in beta-arrestin that bind AP-2 have not been identified. Moreover, the respective contributions of the interactions of beta-arrestin with AP-2 and clathrin toward the targeting of GPCRs to clathrin-coated vesicles have not been established. Here, we identify specific arginine residues (Arg(394) and Arg(396)) in the beta-arrestin 2 C terminus that mediate beta-arrestin binding to AP-2 and show, in vitro, that these domains in beta-arrestin 1 and 2 interact equally well with AP-2 independently of clathrin binding. We demonstrate in HEK 293 cells by fluorescence microscopy that beta(2)-adrenergic receptor-beta-arrestin complexes lacking the beta-arrestin-clathrin binding motif are still targeted to clathrin-coated pits. In marked contrast, receptor-beta-arrestin complexes lacking the beta-arrestin/AP-2 interactions are not effectively compartmentalized in punctated areas of the plasma membrane. These results reveal that the binding of a receptor-beta-arrestin complex to AP-2, not to clathrin, is necessary for the initial targeting of beta(2)-adrenergic receptor to clathrin-coated pits.

Authors
Laporte, SA; Oakley, RH; Holt, JA; Barak, LS; Caron, MG
MLA Citation
Laporte, SA, Oakley, RH, Holt, JA, Barak, LS, and Caron, MG. "The interaction of beta-arrestin with the AP-2 adaptor is required for the clustering of beta 2-adrenergic receptor into clathrin-coated pits." J Biol Chem 275.30 (July 28, 2000): 23120-23126.
PMID
10770944
Source
pubmed
Published In
The Journal of biological chemistry
Volume
275
Issue
30
Publish Date
2000
Start Page
23120
End Page
23126
DOI
10.1074/jbc.M002581200

Differential affinities of visual arrestin, beta arrestin1, and beta arrestin2 for G protein-coupled receptors delineate two major classes of receptors.

Visual arrestin, betaarrestin1, and betaarrestin2 comprise a family of intracellular proteins that desensitize G protein-coupled receptors (GPCRs). In addition, betaarrestin1 and betaarrestin2 target desensitized receptors to clathrin-coated pits for endocytosis. Whether arrestins differ in their ability to interact with GPCRs in cells is not known. In this study, we visualize the interaction of arrestin family members with GPCRs in real time and in live cells using green fluorescent protein-tagged arrestins. In the absence of agonist, visual arrestin and betaarrestin1 were found in both the cytoplasm and nucleus of HEK-293 cells, whereas betaarrestin2 was found only in the cytoplasm. Analysis of agonist-mediated arrestin translocation to multiple GPCRs identified two major classes of receptors. Class A receptors (beta2 adrenergic receptor, mu opioid receptor, endothelin type A receptor, dopamine D1A receptor, and alpha1b adrenergic receptor) bound betaarrestin2 with higher affinity than betaarrestin1 and did not interact with visual arrestin. In contrast, class B receptors (angiotensin II type 1A receptor, neurotensin receptor 1, vasopressin V2 receptor, thyrotropin-releasing hormone receptor, and substance P receptor) bound both betaarrestin isoforms with similar high affinities and also interacted with visual arrestin. Switching the carboxyl-terminal tails of class A and class B receptors completely reversed the affinity of each receptor for the visual and non-visual arrestins. In addition, exchanging the betaarrestin1 and betaarrestin2 carboxyl termini reversed their extent of binding to class A receptors as well as their subcellular distribution. These results reveal for the first time marked differences in the ability of arrestin family members to bind GPCRs at the plasma membrane. Moreover, they show that visual arrestin can interact in cells with GPCRs other than rhodopsin. These findings suggest that GPCR signaling may be differentially regulated depending on the cellular complement of arrestin isoforms and the ability of arrestins to interact with other cellular proteins.

Authors
Oakley, RH; Laporte, SA; Holt, JA; Caron, MG; Barak, LS
MLA Citation
Oakley, RH, Laporte, SA, Holt, JA, Caron, MG, and Barak, LS. "Differential affinities of visual arrestin, beta arrestin1, and beta arrestin2 for G protein-coupled receptors delineate two major classes of receptors." J Biol Chem 275.22 (June 2, 2000): 17201-17210.
PMID
10748214
Source
pubmed
Published In
The Journal of biological chemistry
Volume
275
Issue
22
Publish Date
2000
Start Page
17201
End Page
17210
DOI
10.1074/jbc.M910348199

Increased rewarding properties of morphine in dopamine-transporter knockout mice.

The activation of dopamine (DA) neurotransmission plays a crucial role in the behavioural responses to drugs of abuse. In particular, increased extracellular levels of DA within the mesolimbic pathway have been implicated in the rewarding and locomotor stimulatory properties of morphine. We investigated the behavioural responses to morphine in mice with a genetic disruption of the DA transporter (DAT), resulting in a constitutively high level of extrasynaptic DA. In the conditioned place preference test, DAT-/- mice exhibited a stronger rewarding response to morphine (5 mg/kg, s.c.) compared with control littermates. However, the same dose of morphine failed to increase locomotor activity in DAT-/- mice, whilst enhancing locomotion in DAT+/- and DAT+/+ animals. Morphine-induced analgesia was unaffected in mutant mice, but the behavioural expression of naloxone-induced withdrawal signs was blunted. In vivo voltammetry in the shell of the nucleus accumbens revealed that morphine was able to stimulate DA neurons in DAT-/- mice, resulting in the accumulation of higher extracellular DA levels compared with control animals. Morphine also induced a higher rate of c-fos transcription in the shell of the nucleus accumbens in mutant mice. We conclude that morphine-induced rewarding responses are firmly established in DAT mutant mice despite a DA transmission that is already tonically activated, and independently of any effect on locomotion. These particular behavioural responses to morphine may be associated with the action of the drug on DA release and c-fos expression in the shell of the nucleus accumbens of DAT-/- mice.

Authors
Spielewoy, C; Gonon, F; Roubert, C; Fauchey, V; Jaber, M; Caron, MG; Roques, BP; Hamon, M; Betancur, C; Maldonado, R; Giros, B
MLA Citation
Spielewoy, C, Gonon, F, Roubert, C, Fauchey, V, Jaber, M, Caron, MG, Roques, BP, Hamon, M, Betancur, C, Maldonado, R, and Giros, B. "Increased rewarding properties of morphine in dopamine-transporter knockout mice." Eur J Neurosci 12.5 (May 2000): 1827-1837.
PMID
10792459
Source
pubmed
Published In
European Journal of Neuroscience
Volume
12
Issue
5
Publish Date
2000
Start Page
1827
End Page
1837

Mice lacking the norepinephrine transporter are supersensitive to psychostimulants.

The action of norepinephrine (NE) is terminated, in part, by its uptake into presynaptic noradrenergic neurons by the plasma-membrane NE transporter (NET), which is a target for antidepressants and psychostimulants. Disruption of the NET gene in mice prolonged the clearance of NE and elevated extracellular levels of this catecholamine. In a classical test for antidepressant drugs, the NET-deficient (NET-/-) animals behaved like antidepressant-treated wild-type mice. Mutants were hyper-responsive to locomotor stimulation by cocaine or amphetamine. These responses were accompanied by dopamine D2/D3 receptor supersensitivity. Thus altering NET expression significantly modulates midbrain dopaminergic function, an effect that may be an important component of the actions of antidepressants and psychostimulants.

Authors
Xu, F; Gainetdinov, RR; Wetsel, WC; Jones, SR; Bohn, LM; Miller, GW; Wang, YM; Caron, MG
MLA Citation
Xu, F, Gainetdinov, RR, Wetsel, WC, Jones, SR, Bohn, LM, Miller, GW, Wang, YM, and Caron, MG. "Mice lacking the norepinephrine transporter are supersensitive to psychostimulants." Nat Neurosci 3.5 (May 2000): 465-471.
PMID
10769386
Source
pubmed
Published In
Nature Neuroscience
Volume
3
Issue
5
Publish Date
2000
Start Page
465
End Page
471
DOI
10.1038/74839

Mutational analysis of Gbetagamma and phospholipid interaction with G protein-coupled receptor kinase 2.

Agonist-dependent regulation of G protein-coupled receptors is dependent on their phosphorylation by G protein-coupled receptor kinases (GRKs). GRK2 and GRK3 are selectively regulated in vitro by free Gbetagamma subunits and negatively charged membrane phospholipids through their pleckstrin homology (PH) domains. However, the molecular binding determinants and physiological role for these ligands remain unclear. To address these issues, we generated an array of site-directed mutants within the GRK2 PH domain and characterized their interaction with Gbetagamma and phospholipids in vitro. Mutation of several residues in the loop 1 region of the PH domain, including Lys-567, Trp-576, Arg-578, and Arg-579, resulted in a loss of receptor phosphorylation, likely via disruption of phospholipid binding, that was reversed by Gbetagamma. Alternatively, mutation of residues distal to the C-terminal amphipathic alpha-helix, including Lys-663, Lys-665, Lys-667, and Arg-669, resulted in decreased responsiveness to Gbetagamma. Interestingly, mutation of Arg-587 in beta-sheet 3, a region not previously thought to interact with Gbetagamma, resulted in a specific and profound loss of Gbetagamma responsiveness. To further characterize these effects, two mutants (GRK2(K567E/R578E) and GRK2(R587Q)) were expressed in Sf9 cells and purified. Analysis of these mutants revealed that GRK2(K567E/R578E) was refractory to stimulation by negatively charged phospholipids but bound Gbetagamma similar to wild-type GRK2. In contrast, GRK2(R587Q) was stimulated by acidic phospholipids but failed to bind Gbetagamma. In order to examine the role of phospholipid and Gbetagamma interaction in cells, wild-type and mutant GRK2s were expressed with a beta(2)-adrenergic receptor (beta(2)AR) mutant that is responsive to GRK2 phosphorylation (beta(2)AR(Y326A)). In these cells, GRK2(K567E/R578E) and GRK2(R587Q) were largely defective in promoting agonist-dependent phosphorylation and internalization of beta(2)AR(Y326A). Similarly, wild-type GRK2 but not GRK2(K567E/R578E) or GRK2(R587Q) promoted morphinedependent phosphorylation of the mu-opioid receptor in cells. Thus, we have (i) identified several specific GRK2 binding determinants for Gbetagamma and phospholipids, and (ii) demonstrated that Gbetagamma binding is the limiting step for GRK2-dependent receptor phosphorylation in cells.

Authors
Carman, CV; Barak, LS; Chen, C; Liu-Chen, LY; Onorato, JJ; Kennedy, SP; Caron, MG; Benovic, JL
MLA Citation
Carman, CV, Barak, LS, Chen, C, Liu-Chen, LY, Onorato, JJ, Kennedy, SP, Caron, MG, and Benovic, JL. "Mutational analysis of Gbetagamma and phospholipid interaction with G protein-coupled receptor kinase 2." J Biol Chem 275.14 (April 7, 2000): 10443-10452.
PMID
10744734
Source
pubmed
Published In
The Journal of biological chemistry
Volume
275
Issue
14
Publish Date
2000
Start Page
10443
End Page
10452

Dopamine tone regulates D1 receptor trafficking and delivery in striatal neurons in dopamine transporter-deficient mice.

In vivo, G protein-coupled receptors (GPCR) for neurotransmitters undergo complex intracellular trafficking that contribute to regulate their abundance at the cell surface. Here, we report a previously undescribed alteration in the subcellular localization of D1 dopamine receptor (D1R) that occurs in vivo in striatal dopaminoceptive neurons in response to chronic and constitutive hyperdopaminergia. Indeed, in mice lacking the dopamine transporter, D1R is in abnormally low abundance at the plasma membrane of cell bodies and dendrites and is largely accumulated in rough endoplasmic reticulum and Golgi apparatus. Decrease of striatal extracellular dopamine concentration with 6-hydroxydopamine (6- OHDA) in heterozygous mice restores delivery of the receptor from the cytoplasm to the plasma membrane in cell bodies. These results demonstrate that, in vivo, in the central nervous system, the storage in cytoplasmic compartments involved in synthesis and the membrane delivery contribute to regulate GPCR availability and abundance at the surface of the neurons under control of the neurotransmitter tone. Such regulation may contribute to modulate receptivity of neurons to their endogenous ligands and related exogenous drugs.

Authors
Dumartin, B; Jaber, M; Gonon, F; Caron, MG; Giros, B; Bloch, B
MLA Citation
Dumartin, B, Jaber, M, Gonon, F, Caron, MG, Giros, B, and Bloch, B. "Dopamine tone regulates D1 receptor trafficking and delivery in striatal neurons in dopamine transporter-deficient mice." Proc Natl Acad Sci U S A 97.4 (February 15, 2000): 1879-1884.
PMID
10677550
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
97
Issue
4
Publish Date
2000
Start Page
1879
End Page
1884

Differential quantal release of histamine and 5-hydroxytryptamine from mast cells of vesicular monoamine transporter 2 knockout mice.

The recent availability of mice lacking the neuronal form of the vesicular monoamine transporter 2 (VMAT2) affords the opportunity to study its roles in storage and release. Carbon fiber microelectrodes were used to measure individual secretory events of histamine and 5-hydroxytryptamine (5-HT) from VMAT2-expressing mast cells as a model system for quantal release. VMAT2 is indispensable for monoamine storage because mast cells from homozygous (VMAT2(-/-)) mice, while undergoing granule-cell fusion, do not release monoamines. Cells from heterozygous animals (VMAT2(+/-)) secrete lower amounts of monoamine per granule than cells from wild-type controls. Investigation of corelease of histamine and 5-HT from granules in VMAT2(+/-) cells revealed 5-HT quantal size was reduced more than that of histamine. Thus, although vesicular transport is the limiting factor determining quantal size of 5-HT and histamine release, intragranular association with the heparin matrix also plays a significant role.

Authors
Travis, ER; Wang, YM; Michael, DJ; Caron, MG; Wightman, RM
MLA Citation
Travis, ER, Wang, YM, Michael, DJ, Caron, MG, and Wightman, RM. "Differential quantal release of histamine and 5-hydroxytryptamine from mast cells of vesicular monoamine transporter 2 knockout mice." Proc Natl Acad Sci U S A 97.1 (January 4, 2000): 162-167.
PMID
10618388
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
97
Issue
1
Publish Date
2000
Start Page
162
End Page
167

Bone histomorphometric and biomechanical abnormalities in mice homozygous for deletion of the dopamine transporter gene.

Dopamine (DA) has been reported to have effects on calcium and phosphorus metabolism. The dopamine transporter (DAT) is believed to control the temporal and spatial activity of released DA by rapid uptake of the neurotransmitter into presynaptic terminals. We have evaluated the histologic and biomechanical properties of the skeleton in mice homozygous for deletion of the DA transporter gene (DAT) to help delineate the role of DA in bone biology. We have demonstrated that DAT-/-mice have reduced bone mass and strength. DAT-/- animals had shorter femur length and dry weight. Ash calcium content of the femur was 32% lower in the DAT-/- mice than in the wild-type animals. Cancellous bone volume in the proximal tibial metaphysis was significantly lower in the DAT-/- animals (p < 0.04). There was a 32% reduction in trabecular thickness (p = NS). For the vertebrae, cancellous bone volume was again lower in the DAT-/- animals compared with wild-type as a consequence of increased trabecular spacing (p < 0.05) and reduced trabecular number (p < 0.05). Cortical thickness and bone area in the femoral diaphysis were reduced in the DAT-/-animals. The ultimate bending load (femoral strength) for the DAT-/- mice was 30% lower than the wild-type mice (p = 0.004). Thus, deletion of the DAT gene results in deficiencies in skeletal structure and integrity.

Authors
Bliziotes, M; McLoughlin, S; Gunness, M; Fumagalli, F; Jones, SR; Caron, MG
MLA Citation
Bliziotes, M, McLoughlin, S, Gunness, M, Fumagalli, F, Jones, SR, and Caron, MG. "Bone histomorphometric and biomechanical abnormalities in mice homozygous for deletion of the dopamine transporter gene." Bone 26.1 (January 2000): 15-19.
PMID
10617152
Source
pubmed
Published In
BONE
Volume
26
Issue
1
Publish Date
2000
Start Page
15
End Page
19

Differential regulation of the dopamine D1, D2 and D3 receptor gene expression and changes in the phenotype of the striatal neurons in mice lacking the dopamine transporter.

Mice with a genetic disruption of the dopamine transporter (DAT-/-) exhibit locomotor hyperactivity and profound alterations in the homeostasis of the nigrostriatal system, e.g. a dramatic increase in the extracellular dopamine level. Here, we investigated the adaptive changes in dopamine D1, D2 and D3 receptor gene expression in the caudate putamen and nucleus accumbens of DAT-/- mice. We used quantitative in situ hybridization and found that the constitutive hyperdopaminergia results in opposite regulations in the gene expression for the dopamine receptors. In DAT-/- mice, we observed increased mRNA levels encoding the D3 receptor (caudate putamen, +60-85%; nucleus accumbens, +40-107%), and decreased mRNA levels for both D1 (caudate putamen, -34%; nucleus accumbens, -45%) and D2 receptors (caudate putamen, -36%; nucleus accumbens, -33%). Furthermore, we assessed the phenotypical organization of the striatal efferent neurons by using double in situ hybridization. Our results show that in DAT+/+ mice, D1 and D2 receptor mRNAs are segregated in two different main populations corresponding to substance P and preproenkephalin A mRNA-containing neurons, respectively. The phenotype of D1 or D2 mRNA-containing neurons was unchanged in both the caudate putamen and nucleus accumbens of DAT-/- mice. Interestingly, we found an increased density of preproenkephalin A-negative neurons that express the D3 receptor mRNA in the nucleus accumbens (core, +35%; shell, +46%) of DAT-/- mice. Our data further support the critical role for the D3 receptor in the regulation of D1-D2 interactions, an action being restricted to neurons coexpressing D1 and D3 receptors in the nucleus accumbens.

Authors
Fauchey, V; Jaber, M; Caron, MG; Bloch, B; Le Moine, C
MLA Citation
Fauchey, V, Jaber, M, Caron, MG, Bloch, B, and Le Moine, C. "Differential regulation of the dopamine D1, D2 and D3 receptor gene expression and changes in the phenotype of the striatal neurons in mice lacking the dopamine transporter." Eur J Neurosci 12.1 (January 2000): 19-26.
PMID
10651856
Source
pubmed
Published In
European Journal of Neuroscience
Volume
12
Issue
1
Publish Date
2000
Start Page
19
End Page
26

An animal model of attention deficit hyperactivity disorder.

Authors
Gainetdinov, RR; Caron, MG
MLA Citation
Gainetdinov, RR, and Caron, MG. "An animal model of attention deficit hyperactivity disorder." Mol Med Today 6.1 (January 2000): 43-44.
PMID
10637574
Source
pubmed
Published In
Molecular Medicine Today
Volume
6
Issue
1
Publish Date
2000
Start Page
43
End Page
44

Mutational analysis of Gβγ and phospholipid interaction with kinase G protein-coupled receptor 2

Agonist-dependent regulation of G protein-coupled receptors is dependent on their phosphorylation by G protein-coupled receptor kinases (GRKs). GRK2 and GRK3 are selectively regulated in vitro by free Gβγ subunits and negatively charged membrane phospholipids through their pleckstrin homology (PH) domains. However, the molecular binding determinants and physiological role for these ligands remain unclear. To address these issues, we generated an array of site-directed mutants within the GRK2 PH domain and characterized their interaction with Gβγ and phospholipids in vitro. Mutation of several residues in the loop 1 region of the PH domain, including Lys-567, Trp-576, Arg-578, and Arg-579, resulted in a loss of receptor phosphorylation, likely via disruption of phospholipid binding, that was reversed by Gβγ. Alternatively, mutation of residues distal to the C-terminal amphipathic α- helix, including Lys-663, Lys-665, Lys-667, and Arg-669, resulted in decreased responsiveness to Gβγ. Interestingly, mutation of Arg-587 in β- sheet 3, a region not previously thought to interact with Gβγ, resulted in a specific and profound loss of Gβγ responsiveness. To further characterize these effects, two mutants (GRK2(K567E/R578E) and GRK2(R587Q)) were expressed in Sf9 cells and purified. Analysis of these mutants revealed that GRK2(K567F/R578E) was refractory to stimulation by negatively charged phospholipids but bound Gβγ similar to wild-type GRK2. In contrast, GRK2(R587Q) was stimulated by acidic phospholipids but failed to bind Gβγ. In order to examine the role of phospholipid and Gβγ interaction in cells, wild-type and mutant GRK2s were expressed with a β2-adrenergic receptor (β2AR) mutant that is responsive to GRK2 phosphorylation (β2AR(Y326A)). In these cells, GRK2(K567E/R578E) and GRK2(R587Q) were largely defective in promoting agonist-dependent phosphorylation and internalization of β2AR(Y326A). Similarly, wild-type GRK2 but not GRK2(K567E/R578E) or GRK2(R587Q) promoted morphine-dependent phosphorylation of the μ-opioid receptor in cells. Thus, we have (i) identified several specific GRK2 binding determinants for Gβγ and phospholipids, and (ii) demonstrated that Gβγ binding is the limiting step for GRK2-dependent receptor phosphorylation in cells.

Authors
Carman, CV; Barak, LS; Chen, C; Liu-Chen, L-Y; Onorato, JJ; Kennedy, SP; Caron, MG; Benovic, JL
MLA Citation
Carman, CV, Barak, LS, Chen, C, Liu-Chen, L-Y, Onorato, JJ, Kennedy, SP, Caron, MG, and Benovic, JL. "Mutational analysis of Gβγ and phospholipid interaction with kinase G protein-coupled receptor 2." Journal of Biological Chemistry 275.14 (2000): 10443-10452.
Source
scival
Published In
Journal of Biological Chemistry
Volume
275
Issue
14
Publish Date
2000
Start Page
10443
End Page
10452
DOI
10.1074/jbc.275.14.10443

Protein scaffoldings and modular signaling through seven transmembrane domain-receptors: Beyond the G proteins

G protein coupled receptors (GPCR) represent one of the largest families of proteins encoded by the human genome. They mediate cellular communications important in many fundamental processes. Until recently, GPCR function was thought to be mediated almost exclusively via the obligatory activation of hetero-trimeric G proteins. The development of molecular biological approaches and methods to assess protein-protein interactions have now markedly broadened this view. It is now apparent that proteins such as receptor kinases and arrestins, which quench the signaling function of GPCR, are themselves capable of functioning either as effecters or adaptors that initiate and regulate non-conventional signaling pathways or cell biological processes involved in receptor function. Similarly, the direct interaction of GPCR either through specific sequence motifs, or conformations with small molecular weight G proteins, enzymes, ion channels or scaffolding proteins indicates a larger than expected diversity in the physiological processes affected this receptor family. Finally, realization that homo- and hetero-dimerization of GPCR as well as interactions with escort proteins can influence their processing, trafficking, ligand binding and signaling specificity points to levels of complexity and diversity that were unsuspected. Unraveling the molecular principles of these interactions and their physiological relevance provides new challenges that will undoubtedly reveal novel targets for the development of therapeutic interventions.

Authors
Bouvier, M; Laporte, SA; Lagacé, M; Caron, MG
MLA Citation
Bouvier, M, Laporte, SA, Lagacé, M, and Caron, MG. "Protein scaffoldings and modular signaling through seven transmembrane domain-receptors: Beyond the G proteins." Medecine/Sciences 16.5 (2000): 644-651.
Source
scival
Published In
Medecine/Sciences
Volume
16
Issue
5
Publish Date
2000
Start Page
644
End Page
651

'Model' behavior [4] (multiple letters)

Authors
Sarkis, EH; Gainetdinov, RR; Wetsel, WC; Caron, MG
MLA Citation
Sarkis, EH, Gainetdinov, RR, Wetsel, WC, and Caron, MG. "'Model' behavior [4] (multiple letters)." Science 287.5461 (2000): 2160-2162.
PMID
10744536
Source
scival
Published In
Science
Volume
287
Issue
5461
Publish Date
2000
Start Page
2160
End Page
2162

Endocrine Reviews: Editorial

Authors
Caron, MG
MLA Citation
Caron, MG. "Endocrine Reviews: Editorial." Endocrine Reviews 21.1 (2000): 3--.
Source
scival
Published In
Endocrine Reviews
Volume
21
Issue
1
Publish Date
2000
Start Page
3-
DOI
10.1210/er.21.1.3

Manifestation of dopamine and glutamate systems dysfunction in animal models of schizoid behavior

Authors
Caron, MG; Mohn, AR; Gainetdinov, RR
MLA Citation
Caron, MG, Mohn, AR, and Gainetdinov, RR. "Manifestation of dopamine and glutamate systems dysfunction in animal models of schizoid behavior." Acta Neurologica Scandinavica, Supplement 102.175 (2000): 14-15.
Source
scival
Published In
Acta Neurologica Scandinavica, Supplement
Volume
102
Issue
175
Publish Date
2000
Start Page
14
End Page
15

Mice lacking the dopamine transporter display altered regulation of distal colonic motility

The mechanisms by which dopamine (DA) influences gastrointestinal (GI) tract motility are incompletely understood and complicated by tissue- and species-specific differences in dopaminergic function. To improve the understanding of DA action on GI motility, we used an organ tissue bath system to characterize motor function of distal colonic smooth muscle segments from wild-type and DA transporter knockout (DAT -/-) mice. In wild-type mice, combined blockade of D1 and D2 receptors resulted in significant increases in tone (62 ± 9%), amplitude of spontaneous phasic contractions (167 ± 24%), and electric field stimulation (EFS)-induced (40 ± 8%) contractions, suggesting that endogenous DA is inhibitory to mouse distal colonic motility. The amplitudes of spontaneous phasic and EFS-induced contractions were lower in DAT -/- mice relative to wild-type mice. These differences were eliminated by combined D1 and D2 receptor blockade, indicating that the inhibitory effects of DA on distal colonic motility are potentiated in DAT -/- mice. Motility index was decreased but spontaneous phasic contraction frequency was enhanced in DAT -/- mice relative to wild-type mice. The fact that spontaneous phasic and EFS-induced contractile activity were altered by the lack of the DA transporter suggests an important role for endogenous DA in modulating motility of mouse distal colon.

Authors
Walker, JKL; Gainetdinov, RR; Mangel, AW; Caron, MG; Shetzline, MA
MLA Citation
Walker, JKL, Gainetdinov, RR, Mangel, AW, Caron, MG, and Shetzline, MA. "Mice lacking the dopamine transporter display altered regulation of distal colonic motility." American Journal of Physiology - Gastrointestinal and Liver Physiology 279.2 42-2 (2000): G311-G318.
Source
scival
Published In
American journal of physiology. Gastrointestinal and liver physiology
Volume
279
Issue
2 42-2
Publish Date
2000
Start Page
G311
End Page
G318

Enhanced morphine analgesia in mice lacking beta-arrestin 2.

The ability of morphine to alleviate pain is mediated through a heterotrimeric guanine nucleotide binding protein (G protein)-coupled heptahelical receptor (GPCR), the mu opioid receptor (muOR). The efficiency of GPCR signaling is tightly regulated and ultimately limited by the coordinated phosphorylation of the receptors by specific GPCR kinases and the subsequent interaction of the phosphorylated receptors with beta-arrestin 1 and beta-arrestin 2. Functional deletion of the beta-arrestin 2 gene in mice resulted in remarkable potentiation and prolongation of the analgesic effect of morphine, suggesting that muOR desensitization was impaired. These results provide evidence in vivo for the physiological importance of beta-arrestin 2 in regulating the function of a specific GPCR, the muOR. Moreover, they suggest that inhibition of beta-arrestin 2 function might lead to enhanced analgesic effectiveness of morphine and provide potential new avenues for the study and treatment of pain, narcotic tolerance, and dependence.

Authors
Bohn, LM; Lefkowitz, RJ; Gainetdinov, RR; Peppel, K; Caron, MG; Lin, FT
MLA Citation
Bohn, LM, Lefkowitz, RJ, Gainetdinov, RR, Peppel, K, Caron, MG, and Lin, FT. "Enhanced morphine analgesia in mice lacking beta-arrestin 2." Science 286.5449 (December 24, 1999): 2495-2498.
PMID
10617462
Source
pubmed
Published In
Science
Volume
286
Issue
5449
Publish Date
1999
Start Page
2495
End Page
2498

Dopamine neuronal transport kinetics and effects of amphetamine.

The dopamine (DA) transporter (DAT) regulates DA neurotransmission by recycling DA back into neurons. Drugs that interfere with DAT function, e.g., cocaine and amphetamine, can have profound behavioral effects. The kinetics of DA transport by DAT in isolated synaptosomal or single cell preparations have been previously studied. To investigate how DA transport is regulated in intact tissue and to examine how amphetamine affects the DAT, the kinetics of DA uptake by the DAT were examined in tissue slices of the mouse caudate-putamen with fast-scan cyclic voltammetry. The data demonstrate that inward DA transport is saturable and sodium-dependent. Elevated levels of cytoplasmic DA resulting from disruption of vesicular storage by incubation with 10 microM Ro 4-1284 did not generate DA efflux or decrease its uptake rate. However, incubation with 10 microM amphetamine reduced the net DA uptake rate and increased extracellular DA levels due to DA efflux through the DAT. In addition, a new, elevated steady-state level of extracellular DA was established after electrically stimulated DA release in the presence of amphetamine, norepinephrine, and exogenous DA. These results from intact tissue are consistent with a kinetic model of the DAT established in more purified preparations in which amphetamine and other transported substances make the inwardly facing DAT available for outward transport of intracellular DA.

Authors
Jones, SR; Joseph, JD; Barak, LS; Caron, MG; Wightman, RM
MLA Citation
Jones, SR, Joseph, JD, Barak, LS, Caron, MG, and Wightman, RM. "Dopamine neuronal transport kinetics and effects of amphetamine." J Neurochem 73.6 (December 1999): 2406-2414.
PMID
10582600
Source
pubmed
Published In
Journal of Neurochemistry
Volume
73
Issue
6
Publish Date
1999
Start Page
2406
End Page
2414

Muscarinic supersensitivity and impaired receptor desensitization in G protein-coupled receptor kinase 5-deficient mice.

G protein-coupled receptor kinase 5 (GRK5) is a member of a family of enzymes that phosphorylate activated G protein-coupled receptors (GPCR). To address the physiological importance of GRK5-mediated regulation of GPCRs, mice bearing targeted deletion of the GRK5 gene (GRK5-KO) were generated. GRK5-KO mice exhibited mild spontaneous hypothermia as well as pronounced behavioral supersensitivity upon challenge with the nonselective muscarinic agonist oxotremorine. Classical cholinergic responses such as hypothermia, hypoactivity, tremor, and salivation were enhanced in GRK5-KO animals. The antinociceptive effect of oxotremorine was also potentiated and prolonged. Muscarinic receptors in brains from GRK5-KO mice resisted oxotremorine-induced desensitization, as assessed by oxotremorine-stimulated [5S]GTPgammaS binding. These data demonstrate that elimination of GRK5 results in cholinergic supersensitivity and impaired muscarinic receptor desensitization and suggest that a deficit of GPCR desensitization may be an underlying cause of behavioral supersensitivity.

Authors
Gainetdinov, RR; Bohn, LM; Walker, JK; Laporte, SA; Macrae, AD; Caron, MG; Lefkowitz, RJ; Premont, RT
MLA Citation
Gainetdinov, RR, Bohn, LM, Walker, JK, Laporte, SA, Macrae, AD, Caron, MG, Lefkowitz, RJ, and Premont, RT. "Muscarinic supersensitivity and impaired receptor desensitization in G protein-coupled receptor kinase 5-deficient mice." Neuron 24.4 (December 1999): 1029-1036.
PMID
10624964
Source
pubmed
Published In
Neuron
Volume
24
Issue
4
Publish Date
1999
Start Page
1029
End Page
1036

Association of beta-arrestin with G protein-coupled receptors during clathrin-mediated endocytosis dictates the profile of receptor resensitization.

Resensitization of G protein-coupled receptors (GPCRs) following agonist-mediated desensitization is a necessary step for maintaining physiological responsiveness. However, the molecular mechanisms governing the nature of GPCR resensitization are poorly understood. Here, we examine the role of beta-arrestin in the resensitization of the beta(2) adrenergic receptor (beta(2)AR), known to recycle and resensitize rapidly, and the vasopressin V2 receptor (V2R), known to recycle and resensitize slowly. Upon agonist activation, both receptors recruit beta-arrestin to the plasma membrane and internalize in a beta-arrestin- and clathrin-dependent manner. However, whereas beta-arrestin dissociates from the beta(2)AR at the plasma membrane, it internalizes with the V2R into endosomes. The differential trafficking of beta-arrestin and the ability of these two receptors to dephosphorylate, recycle, and resensitize is completely reversed when the carboxyl-terminal tails of these two receptors are switched. Moreover, the ability of beta-arrestin to remain associated with desensitized GPCRs during clathrin-mediated endocytosis is mediated by a specific cluster of phosphorylated serine residues in the receptor carboxyl-terminal tail. These results demonstrate that the interaction of beta-arrestin with a specific motif in the GPCR carboxyl-terminal tail dictates the rate of receptor dephosphorylation, recycling, and resensitization, and thus provide direct evidence for a novel mechanism by which beta-arrestins regulate the reestablishment of GPCR responsiveness.

Authors
Oakley, RH; Laporte, SA; Holt, JA; Barak, LS; Caron, MG
MLA Citation
Oakley, RH, Laporte, SA, Holt, JA, Barak, LS, and Caron, MG. "Association of beta-arrestin with G protein-coupled receptors during clathrin-mediated endocytosis dictates the profile of receptor resensitization." J Biol Chem 274.45 (November 5, 1999): 32248-32257.
PMID
10542263
Source
pubmed
Published In
The Journal of biological chemistry
Volume
274
Issue
45
Publish Date
1999
Start Page
32248
End Page
32257

Genetic approaches to studying norepinephrine function: knockout of the mouse norepinephrine transporter gene.

Norepinephrine is an important chemical messenger in the nervous system. It regulates affective states, learning and memory, endocrine and autonomic functions. It has been implicated in depression, aggression, and addiction, as well as cardiac and thermal dysregulation. The norepinephrine transporter functions by uptaking norepinephrine back into the cell for cyclic use, and is a direct target of a number of antidepressants and psychostimulants. Functional deletion (knockout) of monamine transporters results in increases in extracellular levels of neurotransmitters, thereby prolonging their actions. For the norepinephrine transporter knockout mice, this altered state of the norepinephrine system should simulate the therapeutic effects of norepinephrine selective antidepressants and some of the effects of psychostimulants. Careful use of such an animal model can hopefully provide valuable insight into the multiple roles norepinephrine plays in normal and pathological physiology.

Authors
Wang, YM; Xu, F; Gainetdinov, RR; Caron, MG
MLA Citation
Wang, YM, Xu, F, Gainetdinov, RR, and Caron, MG. "Genetic approaches to studying norepinephrine function: knockout of the mouse norepinephrine transporter gene." Biol Psychiatry 46.9 (November 1, 1999): 1124-1130. (Review)
PMID
10560019
Source
pubmed
Published In
Biological Psychiatry
Volume
46
Issue
9
Publish Date
1999
Start Page
1124
End Page
1130

Application of microdialysis and voltammetry to assess dopamine functions in genetically altered mice: correlation with locomotor activity.

Authors
Jones, SR; Gainetdinov, RR; Caron, MG
MLA Citation
Jones, SR, Gainetdinov, RR, and Caron, MG. "Application of microdialysis and voltammetry to assess dopamine functions in genetically altered mice: correlation with locomotor activity." Psychopharmacology (Berl) 147.1 (November 1999): 30-32. (Review)
PMID
10591864
Source
pubmed
Published In
Psychopharmacology
Volume
147
Issue
1
Publish Date
1999
Start Page
30
End Page
32

Properties of secretin receptor internalization differ from those of the beta(2)-adrenergic receptor.

The endocytic pathway of the secretin receptor, a class II GPCR, is unknown. Some class I G protein-coupled receptors (GPCRs), such as the beta(2)-adrenergic receptor (beta(2)-AR), internalize in clathrin-coated vesicles and this process is mediated by G protein-coupled receptor kinases (GRKs), beta-arrestin, and dynamin. However, other class I GPCRs, for example, the angiotensin II type 1A receptor (AT(1A)R), exhibit different internalization properties than the beta(2)-AR. The secretin receptor, a class II GPCR, is a GRK substrate, suggesting that like the beta(2)-AR, it may internalize via a beta-arrestin and dynamin directed process. In this paper we characterize the internalization of a wild-type and carboxyl-terminal (COOH-terminal) truncated secretin receptor using flow cytometry and fluorescence imaging, and compare the properties of secretin receptor internalization to that of the beta(2)-AR. In HEK 293 cells, sequestration of both the wild-type and COOH-terminal truncated secretin receptors was unaffected by GRK phosphorylation, whereas inhibition of cAMP-dependent protein kinase mediated phosphorylation markedly decreased sequestration. Addition of secretin to cells resulted in a rapid translocation of beta-arrestin to plasma membrane localized receptors; however, secretin receptor internalization was not reduced by expression of dominant negative beta-arrestin. Thus, like the AT(1A)R, secretin receptor internalization is not inhibited by reagents that interfere with clathrin-coated vesicle-mediated internalization and in accordance with these results, we show that secretin and AT(1A) receptors colocalize in endocytic vesicles. This study demonstrates that the ability of secretin receptor to undergo GRK phosphorylation and beta-arrestin binding is not sufficient to facilitate or mediate its internalization. These results suggest that other receptors may undergo endocytosis by mechanisms used by the secretin and AT(1A) receptors and that kinases other than GRKs may play a greater role in GPCR endocytosis than previously appreciated.

Authors
Walker, JK; Premont, RT; Barak, LS; Caron, MG; Shetzline, MA
MLA Citation
Walker, JK, Premont, RT, Barak, LS, Caron, MG, and Shetzline, MA. "Properties of secretin receptor internalization differ from those of the beta(2)-adrenergic receptor." J Biol Chem 274.44 (October 29, 1999): 31515-31523.
PMID
10531354
Source
pubmed
Published In
The Journal of biological chemistry
Volume
274
Issue
44
Publish Date
1999
Start Page
31515
End Page
31523

Delineating muscarinic receptor functions.

Authors
Gainetdinov, RR; Caron, MG
MLA Citation
Gainetdinov, RR, and Caron, MG. "Delineating muscarinic receptor functions." Proc Natl Acad Sci U S A 96.22 (October 26, 1999): 12222-12223.
PMID
10535900
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
96
Issue
22
Publish Date
1999
Start Page
12222
End Page
12223

Differential regulation of tyrosine hydroxylase in the basal ganglia of mice lacking the dopamine transporter.

Mice lacking the dopamine transporter (DAT) display biochemical and behavioural dopaminergic hyperactivity despite dramatic alteration in dopamine homeostasis. In order to determine the anatomical and functional integrity of the dopaminergic system, we examined the expression of tyrosine hydroxylase (TH), the rate-limiting enzyme of dopamine synthesis as well as DOPA decarboxylase and vesicular monoamine transporter. TH-positive neurons in the substantia nigra were only slightly decreased (-27.6 +/- 4.5%), which can not account for the dramatic decreases in the levels of TH and dopamine that we previously observed in the striatum. TH mRNA levels were decreased by 25% in the ventral midbrain with no modification in the ratio of TH mRNA levels per cell. However, TH protein levels were decreased by 90% in the striatum and 35% in the ventral midbrain. In the striatum, many dopaminergic projections had no detectable TH, while few projections maintained regular labelling as demonstrated using electron microscopy. DOPA decarboxylase levels were not modified and vesicular transporter levels were decreased by only 28.7% which suggests that the loss of TH labelling in the striatum is not due to loss of TH projections. Interestingly, we also observed sporadic TH-positive cell bodies using immunohistochemistry and in situ hybridization in the striatum of homozygote mice, and to some extent that of wild-type animals, which raises interesting possibilities as to their potential contribution to the dopamine hyperactivity and volume transmission previously reported in these animals. In conjunction with our previous findings, these results highlight the complex regulatory mechanisms controlling TH expression at the level of mRNA, protein, activity and distribution. The paradoxical hyperdopaminergia in the DAT KO mice despite a marked decrease in TH and dopamine levels suggests a parallel to Parkinson's disease implying that blockade of DAT may be beneficial in this condition.

Authors
Jaber, M; Dumartin, B; Sagné, C; Haycock, JW; Roubert, C; Giros, B; Bloch, B; Caron, MG
MLA Citation
Jaber, M, Dumartin, B, Sagné, C, Haycock, JW, Roubert, C, Giros, B, Bloch, B, and Caron, MG. "Differential regulation of tyrosine hydroxylase in the basal ganglia of mice lacking the dopamine transporter." Eur J Neurosci 11.10 (October 1999): 3499-3511.
PMID
10564358
Source
pubmed
Published In
European Journal of Neuroscience
Volume
11
Issue
10
Publish Date
1999
Start Page
3499
End Page
3511

Dopamine transporters and neuronal injury.

The plasma membrane dopamine transporter (DAT) and the vesicular monoamine transporter (VMAT2) are essential for normal dopamine neurotransmission. DAT terminates the actions of dopamine by rapidly removing dopamine from the synapse, whereas VMAT2 loads cytoplasmic dopamine into vesicles for storage and subsequent release. Recent data suggest that perturbation of the tightly regulated balance between these two transporters predisposes the neurone to damage by a variety of insults. Most notable is the selective degeneration of DAT- and VMAT2-expressing dopamine nerve terminals in the striatum thought to underlie Parkinson's disease. DAT and VMAT2 expression can predict the selective vulnerability of neuronal populations, which suggests that therapeutic strategies aimed at altering DAT and VMAT2 function could have significant benefits in a variety of disorders.

Authors
Miller, GW; Gainetdinov, RR; Levey, AI; Caron, MG
MLA Citation
Miller, GW, Gainetdinov, RR, Levey, AI, and Caron, MG. "Dopamine transporters and neuronal injury." Trends Pharmacol Sci 20.10 (October 1999): 424-429. (Review)
PMID
10498956
Source
pubmed
Published In
Trends in Pharmacological Sciences
Volume
20
Issue
10
Publish Date
1999
Start Page
424
End Page
429

Mice with reduced NMDA receptor expression display behaviors related to schizophrenia.

N-methyl-D-aspartate receptors (NMDARs) represent a subclass of glutamate receptors that play a critical role in neuronal development and physiology. We report here the generation of mice expressing only 5% of normal levels of the essential NMDAR1 (NR1) subunit. Unlike NR1 null mice, these mice survive to adulthood and display behavioral abnormalities, including increased motor activity and stereotypy and deficits in social and sexual interactions. These behavioral alterations are similar to those observed in pharmacologically induced animal models of schizophrenia and can be ameliorated by treatment with haloperidol or clozapine, antipsychotic drugs that antagonize dopaminergic and serotonergic receptors. These findings support a model in which reduced NMDA receptor activity results in schizophrenic-like behavior and reveals how pharmacological manipulation of monoaminergic pathways can affect this phenotype.

Authors
Mohn, AR; Gainetdinov, RR; Caron, MG; Koller, BH
MLA Citation
Mohn, AR, Gainetdinov, RR, Caron, MG, and Koller, BH. "Mice with reduced NMDA receptor expression display behaviors related to schizophrenia." Cell 98.4 (August 20, 1999): 427-436.
PMID
10481908
Source
pubmed
Published In
Cell
Volume
98
Issue
4
Publish Date
1999
Start Page
427
End Page
436

Functional hyperdopaminergia in dopamine transporter knock-out mice.

Dopamine is an important regulator of many central nervous system functions. Hyperfunction of the dopaminergic system is believed to be related to several pathological conditions. Genetic deletion of the dopamine transporter gene in mice results in a persistent extracellular hyperdopaminergic tone, that is functionally revealed as hyperactivity. The lack of a reuptake mechanism produces a marked increase in functional extracellular dopamine which results in profound plasticity of pre- and postsynaptic parameters of dopamine homeostasis. The mice lacking the dopamine transporter gene may represent an appropriate model to elucidate the molecular adaptive changes accompanying pathological states associated with hyperdopaminergic function.

Authors
Gainetdinov, RR; Jones, SR; Caron, MG
MLA Citation
Gainetdinov, RR, Jones, SR, and Caron, MG. "Functional hyperdopaminergia in dopamine transporter knock-out mice." Biol Psychiatry 46.3 (August 1, 1999): 303-311. (Review)
PMID
10435196
Source
pubmed
Published In
Biological Psychiatry
Volume
46
Issue
3
Publish Date
1999
Start Page
303
End Page
311

Loss of autoreceptor functions in mice lacking the dopamine transporter.

Autoreceptors provide an important inhibitory feedback mechanism for dopamine neurons by altering neuronal functions in response to changes in extracellular levels of dopamine. Elevated dopamine may be a component of several neuropsychiatric disorders. However, evidence concerning the state of autoreceptors in such conditions has remained elusive. The function of dopamine autoreceptors was assessed in mice lacking the dopamine transporter (DAT). Genetic deletion of the DAT gene in mice results in a persistent elevation in levels of extracellular dopamine. Direct assessment of impulse-, synthesis- and release-regulating autoreceptors in these mice reveals a nearly complete loss of function. These findings may provide insight into the neurochemical consequences of hyperdopaminergia.

Authors
Jones, SR; Gainetdinov, RR; Hu, XT; Cooper, DC; Wightman, RM; White, FJ; Caron, MG
MLA Citation
Jones, SR, Gainetdinov, RR, Hu, XT, Cooper, DC, Wightman, RM, White, FJ, and Caron, MG. "Loss of autoreceptor functions in mice lacking the dopamine transporter." Nat Neurosci 2.7 (July 1999): 649-655.
PMID
10404198
Source
pubmed
Published In
Nature Neuroscience
Volume
2
Issue
7
Publish Date
1999
Start Page
649
End Page
655
DOI
10.1038/10204

OCD-Like behaviors caused by a neuropotentiating transgene targeted to cortical and limbic D1+ neurons.

To study the behavioral role of neurons containing the D1 dopamine receptor (D1+), we have used a genetic neurostimulatory approach. We generated transgenic mice that express an intracellular form of cholera toxin (CT), a neuropotentiating enzyme that chronically activates stimulatory G-protein (Gs) signal transduction and cAMP synthesis, under the control of the D1 promoter. Because the D1 promoter, like other CNS-expressed promoters, confers transgene expression that is regionally restricted to different D1+ CNS subsets in different transgenic lines, we observed distinct but related psychomotor disorders in different D1CT-expressing founders. In a D1CT line in which transgene expression was restricted to the following D1+ CNS regions-the piriform cortex layer II, layers II-III of somatosensory cortical areas, and the intercalated nucleus of the amygdala-D1CT mice showed normal CNS and D1+ neural architecture but increased cAMP content in whole extracts of the piriform and somatosensory cortex. These mice also exhibited a constellation of compulsive behavioral abnormalities that strongly resembled human cortical-limbic-induced compulsive disorders such as obsessive-compulsive disorder (OCD). These compulsive behaviors included episodes of perseverance or repetition of any and all normal behaviors, repetitive nonaggressive biting of siblings during grooming, and repetitive leaping. These results suggest that chronic potentiation of cortical and limbic D1+ neurons thought to induce glutamatergic output to the striatum causes behaviors reminiscent of those in human cortical-limbic-induced compulsive disorders.

Authors
Campbell, KM; de Lecea, L; Severynse, DM; Caron, MG; McGrath, MJ; Sparber, SB; Sun, LY; Burton, FH
MLA Citation
Campbell, KM, de Lecea, L, Severynse, DM, Caron, MG, McGrath, MJ, Sparber, SB, Sun, LY, and Burton, FH. "OCD-Like behaviors caused by a neuropotentiating transgene targeted to cortical and limbic D1+ neurons." J Neurosci 19.12 (June 15, 1999): 5044-5053.
PMID
10366637
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
19
Issue
12
Publish Date
1999
Start Page
5044
End Page
5053

The dopamine D2, but not D3 or D4, receptor subtype is essential for the disruption of prepulse inhibition produced by amphetamine in mice.

Brain dopamine (DA) systems are involved in the modulation of the sensorimotor gating phenomenon known as prepulse inhibition (PPI). The class of D2-like receptors, including the D2, D3, and D4 receptor subtypes, have all been implicated in the control of PPI via studies of DA agonists and antagonists in rats. Nevertheless, the functional relevance of each receptor subtype remains unclear because these ligands are not specific. To determine the relevance of each receptor subtype, we used genetically altered strains of "knock-out" mice lacking the DA D2, D3, or D4 receptors. We tested the effects of each knock-out on both the phenotypic expression of PPI and the disruption of PPI produced by the indirect DA agonist d-amphetamine (AMPH). No phenotypic differences in PPI were observed at baseline. AMPH significantly disrupted PPI in the D2 (+/+) mice but had no effect in the D2 (-/-) mice. After AMPH treatment, both DA D3 and D4 receptor (+/+) and (-/-) mice had significant disruptions in PPI. These findings indicate that the AMPH-induced disruption of PPI is mediated via the DA D2 receptor and not the D3 or D4 receptor subtypes. Uncovering the neural mechanisms involved in PPI will further our understanding of the substrates of sensorimotor gating and could lead to better therapeutics to treat gating disorders, such as schizophrenia.

Authors
Ralph, RJ; Varty, GB; Kelly, MA; Wang, YM; Caron, MG; Rubinstein, M; Grandy, DK; Low, MJ; Geyer, MA
MLA Citation
Ralph, RJ, Varty, GB, Kelly, MA, Wang, YM, Caron, MG, Rubinstein, M, Grandy, DK, Low, MJ, and Geyer, MA. "The dopamine D2, but not D3 or D4, receptor subtype is essential for the disruption of prepulse inhibition produced by amphetamine in mice." J Neurosci 19.11 (June 1, 1999): 4627-4633.
PMID
10341260
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
19
Issue
11
Publish Date
1999
Start Page
4627
End Page
4633

Genetic models in pharmacology: present status and future.

Authors
Gainetdinov, RR; Caron, MG
MLA Citation
Gainetdinov, RR, and Caron, MG. "Genetic models in pharmacology: present status and future." Pharmacol Res 39.6 (June 1999): 403-404.
PMID
10373248
Source
pubmed
Published In
Pharmacological Research
Volume
39
Issue
6
Publish Date
1999
Start Page
403
End Page
404
DOI
10.1006/phrs.1999.0498

Cellular trafficking of G protein-coupled receptor/beta-arrestin endocytic complexes.

beta-Arrestins are multifunctional proteins identified on the basis of their ability to bind and uncouple G protein-coupled receptors (GPCR) from heterotrimeric G proteins. In addition, beta-arrestins play a central role in mediating GPCR endocytosis, a key regulatory step in receptor resensitization. In this study, we visualize the intracellular trafficking of beta-arrestin2 in response to activation of several distinct GPCRs including the beta2-adrenergic receptor (beta2AR), angiotensin II type 1A receptor (AT1AR), dopamine D1A receptor (D1AR), endothelin type A receptor (ETAR), and neurotensin receptor (NTR). Our results reveal that in response to beta2AR activation, beta-arrestin2 translocation to the plasma membrane shares the same pharmacological profile as described for receptor activation and sequestration, consistent with a role for beta-arrestin as the agonist-driven switch initiating receptor endocytosis. Whereas redistributed beta-arrestins are confined to the periphery of cells and do not traffic along with activated beta2AR, D1AR, and ETAR in endocytic vesicles, activation of AT1AR and NTR triggers a clear time-dependent redistribution of beta-arrestins to intracellular vesicular compartments where they colocalize with internalized receptors. Activation of a chimeric AT1AR with the beta2AR carboxyl-terminal tail results in a beta-arrestin membrane localization pattern similar to that observed in response to beta2AR activation. In contrast, the corresponding chimeric beta2AR with the AT1AR carboxyl-terminal tail gains the ability to translocate beta-arrestin to intracellular vesicles. These results demonstrate that the cellular trafficking of beta-arrestin proteins is differentially regulated by the activation of distinct GPCRs. Furthermore, they suggest that the carboxyl-tail of the receptors might be involved in determining the stability of receptor/betaarrestin complexes and cellular distribution of beta-arrestins.

Authors
Zhang, J; Barak, LS; Anborgh, PH; Laporte, SA; Caron, MG; Ferguson, SS
MLA Citation
Zhang, J, Barak, LS, Anborgh, PH, Laporte, SA, Caron, MG, and Ferguson, SS. "Cellular trafficking of G protein-coupled receptor/beta-arrestin endocytic complexes." J Biol Chem 274.16 (April 16, 1999): 10999-11006.
PMID
10196181
Source
pubmed
Published In
The Journal of biological chemistry
Volume
274
Issue
16
Publish Date
1999
Start Page
10999
End Page
11006

Increased methamphetamine neurotoxicity in heterozygous vesicular monoamine transporter 2 knock-out mice.

Methamphetamine (METH) is a powerful psychostimulant that is increasingly abused worldwide. Although it is commonly accepted that the dopaminergic system and oxidation of dopamine (DA) play pivotal roles in the neurotoxicity produced by this phenylethylamine, the primary source of DA responsible for this effect has remained elusive. In this study, we used mice heterozygous for vesicular monoamine transporter 2 (VMAT2 +/- mice) to determine whether impaired vesicular function alters the effects of METH. METH-induced dopaminergic neurotoxicity was increased in striatum of VMAT2 +/- mice compared with wild-type mice as revealed by a more consistent DA and metabolite depletion and a greater decrease in dopamine transporter expression. Interestingly, increased METH neurotoxicity in VMAT2 +/- mice was accompanied by less pronounced increase in extracellular DA and indices of free radical formation compared with wild-type mice. These results indicate that disruption of vesicular monoamine transport potentiates METH-induced neurotoxicity in vivo and point, albeit indirectly, to a greater contribution of intraneuronal DA redistribution rather than extraneuronal overflow on mediating this effect.

Authors
Fumagalli, F; Gainetdinov, RR; Wang, YM; Valenzano, KJ; Miller, GW; Caron, MG
MLA Citation
Fumagalli, F, Gainetdinov, RR, Wang, YM, Valenzano, KJ, Miller, GW, and Caron, MG. "Increased methamphetamine neurotoxicity in heterozygous vesicular monoamine transporter 2 knock-out mice." J Neurosci 19.7 (April 1, 1999): 2424-2431.
PMID
10087057
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
19
Issue
7
Publish Date
1999
Start Page
2424
End Page
2431

Altered airway and cardiac responses in mice lacking G protein-coupled receptor kinase 3.

Contraction and relaxation of airway smooth muscles is mediated, in part, by G protein-coupled receptors (GPCRs) and dysfunction of these receptors has been implicated in asthma. Phosphorylation of GPCRs, by G protein-coupled receptor kinase (GRK), is an important mechanism involved in the dampening of GPCR signaling. To determine whether this mechanism might play a role in airway smooth muscle physiology, we examined the airway pressure time index and heart rate (HR) responses to intravenous administration of the cholinergic agonist methacholine (MCh) in genetically altered mice lacking one copy of GRK2 (GRK2 +/-), homozygous GRK3 knockout (GRK3 -/-), and wild-type littermates. (GRK2 -/- mice die in utero.) GRK3 -/- mice demonstrated a significant enhancement in the airway response to 100 and 250 microgram/kg doses of MCh compared with wild-type and GRK2 +/- mice. GRK3 -/- mice also displayed an enhanced sensitivity of the airway smooth muscle response to MCh. In addition, GRK3 -/- mice displayed an altered HR recovery from MCh-induced bradycardia. Although direct stimulation of cardiac muscarinic receptors measured as vagal stimulation-induced bradycardia was similar in GRK3 -/- and wild-type mice, the baroreflex increase in HR associated with sodium nitroprusside-induced hypotension was significantly greater in GRK3 -/- than wild-type mice. Therefore, these data demonstrate that in the mouse, GRK3 may be involved in modulating the cholinergic response of airway smooth muscle and in regulating the chronotropic component of the baroreceptor reflex.

Authors
Walker, JK; Peppel, K; Lefkowitz, RJ; Caron, MG; Fisher, JT
MLA Citation
Walker, JK, Peppel, K, Lefkowitz, RJ, Caron, MG, and Fisher, JT. "Altered airway and cardiac responses in mice lacking G protein-coupled receptor kinase 3." Am J Physiol 276.4 Pt 2 (April 1999): R1214-R1221.
PMID
10198406
Source
pubmed
Published In
The American journal of physiology
Volume
276
Issue
4 Pt 2
Publish Date
1999
Start Page
R1214
End Page
R1221

The beta2-adrenergic receptor/betaarrestin complex recruits the clathrin adaptor AP-2 during endocytosis.

betaarrestins mediate the desensitization of the beta2-adrenergic receptor (beta2AR) and many other G protein-coupled receptors (GPCRs). Additionally, betaarrestins initiate the endocytosis of these receptors via clathrin coated-pits and interact directly with clathrin. Consequently, it has been proposed that betaarrestins serve as clathrin adaptors for the GPCR family by linking these receptors to clathrin lattices. AP-2, the heterotetrameric clathrin adaptor protein, has been demonstrated to mediate the internalization of many types of plasma membrane proteins other than GPCRs. AP-2 interacts with the clathrin heavy chain and cytoplasmic domains of receptors such as those for epidermal growth factor and transferrin. In the present study we demonstrate the formation of an agonist-induced multimeric complex containing a GPCR, betaarrestin 2, and the beta2-adaptin subunit of AP-2. beta2-Adaptin binds betaarrestin 2 in a yeast two-hybrid assay and coimmunoprecipitates with betaarrestins and beta2AR in an agonist-dependent manner in HEK-293 cells. Moreover, beta2-adaptin translocates from the cytosol to the plasma membrane in response to the beta2AR agonist isoproterenol and colocalizes with beta2AR in clathrin-coated pits. Finally, expression of betaarrestin 2 minigene constructs containing the beta2-adaptin interacting region inhibits beta2AR endocytosis. These findings point to a role for AP-2 in GPCR endocytosis, and they suggest that AP-2 functions as a clathrin adaptor for the endocytosis of diverse classes of membrane receptors.

Authors
Laporte, SA; Oakley, RH; Zhang, J; Holt, JA; Ferguson, SS; Caron, MG; Barak, LS
MLA Citation
Laporte, SA, Oakley, RH, Zhang, J, Holt, JA, Ferguson, SS, Caron, MG, and Barak, LS. "The beta2-adrenergic receptor/betaarrestin complex recruits the clathrin adaptor AP-2 during endocytosis." Proc Natl Acad Sci U S A 96.7 (March 30, 1999): 3712-3717.
PMID
10097102
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
96
Issue
7
Publish Date
1999
Start Page
3712
End Page
3717

The β2-adrenergic receptor/βarrestin complex recruits the clathrin adaptor AP-2 during endocytosis

βarrestins mediate the desensitization of the β 2 -adrenergic receptor (β 2 AR) and many other G protein-coupled receptors (GPCRs). Additionally, βarrestins initiate the endocytosis of these receptors via clathrin coated- pits and interact directly with clathrin. Consequently, it has been proposed that βarrestins serve as clathrin adaptors for the GPCR family by linking these receptors to clathrin lattices. AP-2, the heterotetrameric clathrin adaptor protein, has been demonstrated to mediate the internalization of many types of plasma membrane proteins other than GPCRs. AP-2 interacts with the clathrin heavy chain and cytoplasmic domains of receptors such as those for epidermal growth factor and transferrin. In the present study we demonstrate the formation of an agonist-induced multimeric complex containing a GPCR, βarrestin 2, and the β 2 -adaptin subunit of AP-2. β 2 -Adaptin binds βarrestin 2 in a yeast two-hybrid assay and coimmunoprecipitates with βarrestins and β 2 AR in an agonist-dependent manner in HEK-293 cells. Moreover, β2-adaptin translocates from the cytosol to the plasma membrane in response to the β 2 AR agonist isoproterenol and colocalizes with β 2 AR in clathrin-coated pits. Finally, expression of βarrestin 2 minigene constructs containing the β 2 -adaptin interacting region inhibits β 2 AR endocytosis. These findings point to a role for AP-2 in GPCR endocytosis, and they suggest that AP-2 functions as a clathrin adaptor for the endocytosis of diverse classes of membrane receptors.

Authors
Laporte, SA; Oakley, RH; Zhang, J; Holt, JA; Ferguson, SSG; Caron, MG; Barak, LS
MLA Citation
Laporte, SA, Oakley, RH, Zhang, J, Holt, JA, Ferguson, SSG, Caron, MG, and Barak, LS. "The β2-adrenergic receptor/βarrestin complex recruits the clathrin adaptor AP-2 during endocytosis." Proceedings of the National Academy of Sciences of the United States of America 96.7 (March 30, 1999): 3712-3717.
Source
scopus
Published In
Proceedings of the National Academy of Sciences of USA
Volume
96
Issue
7
Publish Date
1999
Start Page
3712
End Page
3717
DOI
10.1073/pnas.96.7.3712

Real-time visualization of the cellular redistribution of G protein-coupled receptor kinase 2 and beta-arrestin 2 during homologous desensitization of the substance P receptor.

The substance P receptor (SPR) is a G protein-coupled receptor (GPCR) that plays a key role in pain regulation. The SPR desensitizes in the continued presence of agonist, presumably via mechanisms that implicate G protein-coupled receptor kinases (GRKs) and beta-arrestins. The temporal relationship of these proposed biochemical events has never been established for any GPCR other than rhodopsin beyond the resolution provided by biochemical assays. We investigate the real-time activation and desensitization of the human SPR in live HEK293 cells using green fluorescent protein conjugates of protein kinase C, GRK2, and beta-arrestin 2. The translocation of protein kinase C betaII-green fluorescent protein to and from the plasma membrane in response to substance P indicates that the human SPR becomes activated within seconds of agonist exposure, and the response desensitizes within 30 s. This desensitization process coincides with a redistribution of GRK2 from the cytosol to the plasma membrane, followed by a robust redistribution of beta-arrestin 2 and a profound change in cell morphology that occurs after 1 min of SPR stimulation. These data establish a role for GRKs and beta-arrestins in homologous desensitization of the SPR and provide the first visual and temporal resolution of the sequence of events underlying homologous desensitization of a GPCR in living cells.

Authors
Barak, LS; Warabi, K; Feng, X; Caron, MG; Kwatra, MM
MLA Citation
Barak, LS, Warabi, K, Feng, X, Caron, MG, and Kwatra, MM. "Real-time visualization of the cellular redistribution of G protein-coupled receptor kinase 2 and beta-arrestin 2 during homologous desensitization of the substance P receptor." J Biol Chem 274.11 (March 12, 1999): 7565-7569.
PMID
10066824
Source
pubmed
Published In
The Journal of biological chemistry
Volume
274
Issue
11
Publish Date
1999
Start Page
7565
End Page
7569

Constitutively active alpha-1b adrenergic receptor mutants display different phosphorylation and internalization features.

We compared the phosphorylation and internalization properties of constitutively active alpha-1b adrenergic receptor (AR) mutants carrying mutations in two distant receptor domains, i.e., at A293 in the distal part of the third intracellular loop and at D142 of the DRY motif lying at the end of the third transmembrane domain. For the A293E and A293I mutants the levels of agonist-independent phosphorylation were 150% and 50% higher than those of the wild-type alpha-1b AR, respectively. On the other hand, for the constitutively active D142A and D142T mutants, the basal levels of phosphorylation were similar to those of the wild-type alpha-1b AR and did not appear to be further stimulated by epinephrine. Overexpression of the guanyl nucleotide binding regulatory protein-coupled receptor kinase GRK2 further increases the basal phosphorylation of the A293E mutant, but not that of D142A mutant. Both the wild-type alpha-1b AR and the A293E mutant could undergo beta-arrestin-mediated internalization. The epinephrine-induced internalization of the constitutively active A293E mutant was significantly higher than that of the wild-type alpha-1b AR. In contrast, the D142A mutant was impaired in its ability to interact with beta-arrestin and to undergo agonist-induced internalization. Interestingly, a double mutant A293E/D142A retained very high constitutive activity and regulatory properties of both the A293E and D142A receptors. These findings demonstrate that two constitutively activating mutations occurring in distant receptor domains of the alpha-1b AR have divergent effects on the regulatory properties of the receptor.

Authors
Mhaouty-Kodja, S; Barak, LS; Scheer, A; Abuin, L; Diviani, D; Caron, MG; Cotecchia, S
MLA Citation
Mhaouty-Kodja, S, Barak, LS, Scheer, A, Abuin, L, Diviani, D, Caron, MG, and Cotecchia, S. "Constitutively active alpha-1b adrenergic receptor mutants display different phosphorylation and internalization features." Mol Pharmacol 55.2 (February 1999): 339-347.
PMID
9927627
Source
pubmed
Published In
Molecular pharmacology
Volume
55
Issue
2
Publish Date
1999
Start Page
339
End Page
347

Beta-arrestin-dependent formation of beta2 adrenergic receptor-Src protein kinase complexes.

The Ras-dependent activation of mitogen-activated protein (MAP) kinase pathways by many receptors coupled to heterotrimeric guanine nucleotide binding proteins (G proteins) requires the activation of Src family tyrosine kinases. Stimulation of beta2 adrenergic receptors resulted in the assembly of a protein complex containing activated c-Src and the receptor. Src recruitment was mediated by beta-arrestin, which functions as an adapter protein, binding both c-Src and the agonist-occupied receptor. beta-Arrestin 1 mutants, impaired either in c-Src binding or in the ability to target receptors to clathrin-coated pits, acted as dominant negative inhibitors of beta2 adrenergic receptor-mediated activation of the MAP kinases Erk1 and Erk2. These data suggest that beta-arrestin binding, which terminates receptor-G protein coupling, also initiates a second wave of signal transduction in which the "desensitized" receptor functions as a critical structural component of a mitogenic signaling complex.

Authors
Luttrell, LM; Ferguson, SS; Daaka, Y; Miller, WE; Maudsley, S; Della Rocca, GJ; Lin, F; Kawakatsu, H; Owada, K; Luttrell, DK; Caron, MG; Lefkowitz, RJ
MLA Citation
Luttrell, LM, Ferguson, SS, Daaka, Y, Miller, WE, Maudsley, S, Della Rocca, GJ, Lin, F, Kawakatsu, H, Owada, K, Luttrell, DK, Caron, MG, and Lefkowitz, RJ. "Beta-arrestin-dependent formation of beta2 adrenergic receptor-Src protein kinase complexes." Science 283.5402 (January 29, 1999): 655-661.
PMID
9924018
Source
pubmed
Published In
Science
Volume
283
Issue
5402
Publish Date
1999
Start Page
655
End Page
661

Role of serotonin in the paradoxical calming effect of psychostimulants on hyperactivity.

The mechanism by which psychostimulants act as calming agents in humans with attention-deficit hyperactivity disorder (ADHD) or hyperkinetic disorder is currently unknown. Mice lacking the gene encoding the plasma membrane dopamine transporter (DAT) have elevated dopaminergic tone and are hyperactive. This activity was exacerbated by exposure to a novel environment. Additionally, these mice were impaired in spatial cognitive function, and they showed a decrease in locomotion in response to psychostimulants. This paradoxical calming effect of psychostimulants depended on serotonergic neurotransmission. The parallels between the DAT knockout mice and individuals with ADHD suggest that common mechanisms may underlie some of their behaviors and responses to psychostimulants.

Authors
Gainetdinov, RR; Wetsel, WC; Jones, SR; Levin, ED; Jaber, M; Caron, MG
MLA Citation
Gainetdinov, RR, Wetsel, WC, Jones, SR, Levin, ED, Jaber, M, and Caron, MG. "Role of serotonin in the paradoxical calming effect of psychostimulants on hyperactivity." Science 283.5400 (January 15, 1999): 397-401.
PMID
9888856
Source
pubmed
Published In
Science
Volume
283
Issue
5400
Publish Date
1999
Start Page
397
End Page
401

Agonist-specific regulation of delta-opioid receptor trafficking by G protein-coupled receptor kinase and beta-arrestin.

Opioid receptors mediate multiple biological functions through their interaction with endogenous opioid peptides as well as opioid alkaloids including morphine and etorphine. Previously we have reported that the ability of distinct opioid agonists to differentially regulate mu-opioid receptor (mu OR) responsiveness is related to their ability to promote G protein-coupled receptor kinase (GRK)-dependent phosphorylation of the receptor (1). In the present study, we further examined the role of GRK and beta-arrestin in agonist-specific regulation of the delta-opioid receptor (delta OR). While both etorphine and morphine effectively activate the delta OR, only etorphine triggers robust delta OR phosphorylation followed by plasma membrane translocation of beta-arrestin and receptor internalization. In contrast, morphine is unable to either elicit delta OR phosphorylation or stimulate beta-arrestin translocation, correlating with its inability to cause delta OR internalization. Unlike for the mu OR, overexpression of GRK2 results in neither the enhancement of delta OR sequestration nor the rescue of delta OR-mediated beta-arrestin translocation. Therefore, our findings not only point to the existence of marked differences in the ability of different opioid agonists to promote delta OR phosphorylation by GRK and binding to beta-arrestin, but also demonstrate differences in the regulation of two opioid receptor subtypes. These observations may have important implications for our understanding of the distinct ability of various opioids in inducing opioid tolerance and addiction.

Authors
Zhang, J; Ferguson, SS; Law, PY; Barak, LS; Caron, MG
MLA Citation
Zhang, J, Ferguson, SS, Law, PY, Barak, LS, and Caron, MG. "Agonist-specific regulation of delta-opioid receptor trafficking by G protein-coupled receptor kinase and beta-arrestin." J Recept Signal Transduct Res 19.1-4 (January 1999): 301-313.
PMID
10071766
Source
pubmed
Published In
Journal of Receptors and Signal Transduction (Informa)
Volume
19
Issue
1-4
Publish Date
1999
Start Page
301
End Page
313
DOI
10.3109/10799899909036653

The β2-adrenergic receptor/βarrestin complex recruits the clathrin adaptor AP-2 during endocytosis

βarrestins mediate the desensitization of the β2-adrenergic receptor (β2AR) and many other G protein-coupled receptors (GPCRs). Additionally, βarrestins initiate the endocytosis of these receptors via clathrin coated- pits and interact directly with clathrin. Consequently, it has been proposed that βarrestins serve as clathrin adaptors for the GPCR family by linking these receptors to clathrin lattices. AP-2, the heterotetrameric clathrin adaptor protein, has been demonstrated to mediate the internalization of many types of plasma membrane proteins other than GPCRs. AP-2 interacts with the clathrin heavy chain and cytoplasmic domains of receptors such as those for epidermal growth factor and transferrin. In the present study we demonstrate the formation of an agonist-induced multimeric complex containing a GPCR, βarrestin 2, and the β2-adaptin subunit of AP-2. β2-Adaptin binds βarrestin 2 in a yeast two-hybrid assay and coimmunoprecipitates with βarrestins and β2AR in an agonist-dependent manner in HEK-293 cells. Moreover, β2-adaptin translocates from the cytosol to the plasma membrane in response to the β2AR agonist isoproterenol and colocalizes with β2AR in clathrin-coated pits. Finally, expression of βarrestin 2 minigene constructs containing the β2-adaptin interacting region inhibits β2AR endocytosis. These findings point to a role for AP-2 in GPCR endocytosis, and they suggest that AP-2 functions as a clathrin adaptor for the endocytosis of diverse classes of membrane receptors.

Authors
Laporte, SA; Oakley, RH; Zhang, J; Holt, JA; Ferguson, SSG; Caron, MG; Barak, LS
MLA Citation
Laporte, SA, Oakley, RH, Zhang, J, Holt, JA, Ferguson, SSG, Caron, MG, and Barak, LS. "The β2-adrenergic receptor/βarrestin complex recruits the clathrin adaptor AP-2 during endocytosis." Proceedings of the National Academy of Sciences of the United States of America 96.7 (1999): 3712-3717.
Source
scival
Published In
Proceedings of the National Academy of Sciences of the United States of America
Volume
96
Issue
7
Publish Date
1999
Start Page
3712
End Page
3717
DOI
10.1073/pnas.96.7.3712

Signaling, desensitization, and trafficking of G protein-coupled receptors revealed by green fluorescent protein conjugates.

Authors
Barak, LS; Zhang, J; Ferguson, SS; Laporte, SA; Caron, MG
MLA Citation
Barak, LS, Zhang, J, Ferguson, SS, Laporte, SA, and Caron, MG. "Signaling, desensitization, and trafficking of G protein-coupled receptors revealed by green fluorescent protein conjugates." Methods Enzymol 302 (1999): 153-171.
PMID
12876769
Source
pubmed
Published In
Methods in Enzymology
Volume
302
Publish Date
1999
Start Page
153
End Page
171

Altered airway and cardiac responses in mice lacking G protein-coupled receptor kinase 3

Contraction and relaxation of airway smooth muscles is mediated, in part, by G protein-coupled receptors (GPCRs) and dysfunction of these receptors has been implicated in asthma. Phosphorylation of GPCRs, by G protein-coupled receptor kinase (GRK), is an important mechanism involved in the dampening of GPCR signaling. To determine whether this mechanism might play a role in airway smooth muscle physiology, we examined the airway pressure time index and heart late (HR) responses to intravenous administration of the cholinergic agonist methacholine (MCh) in genetically altered mice lacking one copy of GRK2 (GRK2 +/-), homozygous GRK3 knockout (GRK3 -/-), and wild-type littermates. (GRK2 -/- mice die in utero.) GRK3 - /- mice demonstrated a significant enhancement in the airway response to 100 and 250 μg/kg doses of MCh compared with wild-type and GRK2 +/- mice. GRK3 - /- mice also displayed an enhanced sensitivity of the airway smooth muscle response to MCh. In addition, GRK3 -/- mice displayed an altered HR recovery from MCh-induced bradycardia. Although direct stimulation of cardiac muscarinic receptors measured as vagal stimulation-induced bradycardia was similar in GRK3 -/- and wild-type mice, the baroreflex increase in HR associated with sodium nitroprusside-induced hypotension was significantly greater in GRK3 -/- than wild-type mice. Therefore, these data demonstrate that in the mouse, GRK3 may be involved in modulating the cholinergic response of airway smooth muscle and in regulating the chronotropic component of the baroreceptor reflex.

Authors
Walker, JKL; Peppel, K; Lefkowitz, RJ; Caron, MG; Fisher, JT
MLA Citation
Walker, JKL, Peppel, K, Lefkowitz, RJ, Caron, MG, and Fisher, JT. "Altered airway and cardiac responses in mice lacking G protein-coupled receptor kinase 3." American Journal of Physiology - Regulatory Integrative and Comparative Physiology 276.4 45-4 (1999): R1214-R1221.
Source
scival
Published In
American journal of physiology. Regulatory, integrative and comparative physiology
Volume
276
Issue
4 45-4
Publish Date
1999
Start Page
R1214
End Page
R1221

β-arrestin-dependent formation of β2 adrenergic receptor-src protein kinase complexes

The Ras-dependent activation of mitogen-activated protein (MAP) kinase pathways by many receptors coupled to heterotrimeric guanine nucleotide binding proteins (G proteins) requires the activation of Src family tyrosine kinases. Stimulation of β2 adrenergic receptors resulted in the assembly of a protein complex containing activated c-Src and the receptor. Src recruitment was mediated by β-arrestin, which functions as an adapter protein, binding both c-Src and the agonist-occupied receptor. β-Arrestin 1 mutants, impaired either in c-Src binding or in the ability to target receptors to clathrin-coated pits, acted as dominant negative inhibitors of β2 adrenergic receptor-mediated activation of the MAP kinases Erk1 and Erk2. These data suggest that β-arrestin binding, which terminates receptor- G protein coupling, also initiates a second wave of signal transduction in which the 'desensitized' receptor functions as a critical structural component of a mitogenic signaling complex.

Authors
Luttrell, LM; Ferguson, SSG; Daaka, Y; Miller, WE; Maudsley, S; Rocca, GJD; Lin, F-T; Kawakatsu, H; Owada, K; Luttrell, DK; Caron, MG; Lefkowitz, RJ
MLA Citation
Luttrell, LM, Ferguson, SSG, Daaka, Y, Miller, WE, Maudsley, S, Rocca, GJD, Lin, F-T, Kawakatsu, H, Owada, K, Luttrell, DK, Caron, MG, and Lefkowitz, RJ. "β-arrestin-dependent formation of β2 adrenergic receptor-src protein kinase complexes." Science 283.5402 (1999): 655-661.
Source
scival
Published In
Science
Volume
283
Issue
5402
Publish Date
1999
Start Page
655
End Page
661
DOI
10.1126/science.283.5402.655

Plasma membrane transporters of serotonin, dopamine, and norepinephrine mediate serotonin accumulation in atypical locations in the developing brain of monoamine oxidase A knock-outs.

Genetic loss or pharmacological inhibition of monoamine oxidase A (MAOA) in mice leads to a large increase in whole-brain levels of serotonin (5-HT). Excess 5-HT in mouse neonates prevents the normal barrel-like clustering of thalamic axons in the somatosensory cortex. Projection fields of other neuron populations may develop abnormally. In the present study, we have analyzed the localization of 5-HT immunolabeling in the developing brain of MAOA knock-out mice. We show numerous atypical locations of 5-HT during embryonic and postnatal development. Catecholaminergic cells of the substantia nigra, ventral tegmental area, hypothalamus, and locus ceruleus display transient 5-HT immunoreactivity. Pharmacological treatments inhibiting specific monoamine plasma membrane transporters and genetic crosses with mice lacking the dopamine plasma membrane transporter show that the accumulation of 5-HT in these catecholaminergic cells is attributable to 5-HT uptake via the dopamine or the norepinephrine plasma membrane transporter. In the telencephalon, transient 5-HT immunolabeling is observed in neurons in the CA1 and CA3 fields of the hippocampus, the central amygdala, the indusium griseum, and the deep layers of the anterior cingulate and retrosplenial cortices. In the diencephalon, primary sensory nuclei, as well as the mediodorsal, centrolateral, oval paracentral, submedial, posterior, and lateral posterior thalamic nuclei, are transiently 5-HT immunolabeled. The cortical projections of these thalamic nuclei are also labeled. In the brainstem, neurons in the lateral superior olivary nucleus and the anteroventral cochlear nucleus are transiently 5-HT immunolabeled. None of these structures appear to express the monoamine biosynthetic enzyme L-aromatic amino acid decarboxylase. The administration of monoamine plasma membrane transporter inhibitors indicates that the 5-HT immunolabeling in these structures is attributable to an uptake of 5-HT by the 5-HT plasma membrane transporter. This points to neuron populations that form highly precise projection maps that could be affected by 5-HT during specific developmental stages.

Authors
Cases, O; Lebrand, C; Giros, B; Vitalis, T; De Maeyer, E; Caron, MG; Price, DJ; Gaspar, P; Seif, I
MLA Citation
Cases, O, Lebrand, C, Giros, B, Vitalis, T, De Maeyer, E, Caron, MG, Price, DJ, Gaspar, P, and Seif, I. "Plasma membrane transporters of serotonin, dopamine, and norepinephrine mediate serotonin accumulation in atypical locations in the developing brain of monoamine oxidase A knock-outs." J Neurosci 18.17 (September 1, 1998): 6914-6927.
PMID
9712661
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
18
Issue
17
Publish Date
1998
Start Page
6914
End Page
6927

Control of myocardial contractile function by the level of beta-adrenergic receptor kinase 1 in gene-targeted mice.

We studied the effect of alterations in the level of myocardial beta-adrenergic receptor kinase betaARK1) in two types of genetically altered mice. The first group is heterozygous for betaARK1 gene ablation, betaARK1(+/-), and the second is not only heterozygous for betaARK1 gene ablation but is also transgenic for cardiac-specific overexpression of a betaARK1 COOH-terminal inhibitor peptide, betaARK1(+/-)betaARKct. In contrast to the embryonic lethal phenotype of the homozygous betaARK1 knockout (Jaber, M., Koch, W. J., Rockman, H. A., Smith, B., Bond, R. A., Sulik, K., Ross, J., Jr., Lefkowitz, R. J., Caron, M. G., and Giros, B. (1996) Proc. Natl. Acad. Sci. U. S. A. 93, 12974-12979), betaARK1(+/-) mice develop normally. Cardiac catheterization was performed in mice and showed a stepwise increase in contractile function in the betaARK1(+/-) and betaARK1(+/-)betaARKct mice with the greatest level observed in the betaARK1(+/-)betaARKct animals. Contractile parameters were measured in adult myocytes isolated from both groups of gene-targeted animals. A significantly greater increase in percent cell shortening and rate of cell shortening following isoproterenol stimulation was observed in the betaARK1(+/-) and betaARK1(+/-)betaARKct myocytes compared with wild-type cells, indicating a progressive increase in intrinsic contractility. These data demonstrate that contractile function can be modulated by the level of betaARK1 activity. This has important implications in disease states such as heart failure (in which betaARK1 activity is increased) and suggests that betaARK1 should be considered as a therapeutic target in this situation. Even partial inhibition of betaARK1 activity enhances beta-adrenergic receptor signaling leading to improved functional catecholamine responsiveness.

Authors
Rockman, HA; Choi, DJ; Akhter, SA; Jaber, M; Giros, B; Lefkowitz, RJ; Caron, MG; Koch, WJ
MLA Citation
Rockman, HA, Choi, DJ, Akhter, SA, Jaber, M, Giros, B, Lefkowitz, RJ, Caron, MG, and Koch, WJ. "Control of myocardial contractile function by the level of beta-adrenergic receptor kinase 1 in gene-targeted mice." J Biol Chem 273.29 (July 17, 1998): 18180-18184.
PMID
9660778
Source
pubmed
Published In
The Journal of biological chemistry
Volume
273
Issue
29
Publish Date
1998
Start Page
18180
End Page
18184

Role of dopamine transporter in methamphetamine-induced neurotoxicity: evidence from mice lacking the transporter.

The role of the dopamine transporter (DAT) in mediating the neurotoxic effects of methamphetamine (METH) was tested in mice lacking DAT. Dopamine (DA) and serotonin (5-HT) content, glial fibrillary acidic protein (GFAP) expression, and free radical formation were assessed as markers of METH neurotoxicity in the striatum and/or hippocampus of wild-type, heterozygote, and homozygote (DAT -/-) mice. Four injections of METH (15 mg/kg, s.c.), each given 2 hr apart, produced 80 and 30% decreases in striatal DA and 5-HT levels, respectively, in wild-type animals 2 d after administration. In addition, GFAP mRNA and protein expression levels, extracellular DA levels, and free radical formation were increased markedly. Hippocampal 5-HT content was decreased significantly as well (43%). Conversely, no significant changes were observed in total DA content, GFAP expression, extracellular DA levels, or free radical formation in the striatum of DAT -/- mice after METH administration. However, modest decreases were observed in striatal and hippocampal 5-HT levels (10 and 17%, respectively). These observations demonstrate that DAT is required for, and DA is an essential mediator of, METH-induced striatal dopaminergic neurotoxicity, whereas serotonergic deficits are only partially dependent on DAT.

Authors
Fumagalli, F; Gainetdinov, RR; Valenzano, KJ; Caron, MG
MLA Citation
Fumagalli, F, Gainetdinov, RR, Valenzano, KJ, and Caron, MG. "Role of dopamine transporter in methamphetamine-induced neurotoxicity: evidence from mice lacking the transporter." J Neurosci 18.13 (July 1, 1998): 4861-4869.
PMID
9634552
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
18
Issue
13
Publish Date
1998
Start Page
4861
End Page
4869

Role for G protein-coupled receptor kinase in agonist-specific regulation of mu-opioid receptor responsiveness.

The G protein-coupled mu-opioid receptor (mu OR) mediates the physiological effects of endogenous opioid peptides as well as the structurally distinct opioid alkaloids morphine and etorphine. An intriguing feature of mu OR signaling is the differential receptor trafficking and desensitization properties following activation by distinct agonists, which have been proposed as possible mechanisms related to opioid tolerance. Here we report that the ability of distinct opioid agonists to differentially regulate mu OR internalization and desensitization is related to their ability to promote G protein-coupled receptor kinase (GRK)-dependent phosphorylation of the mu OR. Although both etorphine and morphine effectively activate the mu OR, only etorphine elicits robust mu OR phosphorylation followed by plasma membrane translocation of beta-arrestin and dynamin-dependent receptor internalization. In contrast, corresponding to its inability to cause mu OR internalization, morphine is unable to either elicit mu OR phosphorylation or stimulate beta-arrestin translocation. However, upon the overexpression of GRK2, morphine gains the capacity to induce mu OR phosphorylation, accompanied by the rescue of beta-arrestin translocation and receptor sequestration. Moreover, overexpression of GRK2 also leads to an attenuation of morphine-mediated inhibition of adenylyl cyclase. These findings point to the existence of marked differences in the ability of different opioid agonists to promote mu OR phosphorylation by GRK. These differences may provide the molecular basis underlying the different analgesic properties of opioid agonists and contribute to the distinct ability of various opioids to induce drug tolerance.

Authors
Zhang, J; Ferguson, SS; Barak, LS; Bodduluri, SR; Laporte, SA; Law, PY; Caron, MG
MLA Citation
Zhang, J, Ferguson, SS, Barak, LS, Bodduluri, SR, Laporte, SA, Law, PY, and Caron, MG. "Role for G protein-coupled receptor kinase in agonist-specific regulation of mu-opioid receptor responsiveness." Proc Natl Acad Sci U S A 95.12 (June 9, 1998): 7157-7162.
PMID
9618555
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
95
Issue
12
Publish Date
1998
Start Page
7157
End Page
7162

Cocaine self-administration in dopamine-transporter knockout mice.

The plasma membrane dopamine transporter (DAT) is responsible for clearing dopamine from the synapse. Cocaine blockade of DAT leads to increased extracellular dopamine, an effect widely considered to be the primary cause of the reinforcing and addictive properties of cocaine. In this study we tested whether these properties are limited to the dopaminergic system in mice lacking DAT. In the absence of DAT, these mice exhibit high levels of extracellular dopamine, but paradoxically still self-administer cocaine. Mapping of the sites of cocaine binding and neuronal activation suggests an involvement of serotonergic brain regions in this response. These results demonstrate that the interaction of cocaine with targets other than DAT, possibly the serotonin transporter, can initiate and sustain cocaine self-administration in these mice.

Authors
Rocha, BA; Fumagalli, F; Gainetdinov, RR; Jones, SR; Ator, R; Giros, B; Miller, GW; Caron, MG
MLA Citation
Rocha, BA, Fumagalli, F, Gainetdinov, RR, Jones, SR, Ator, R, Giros, B, Miller, GW, and Caron, MG. "Cocaine self-administration in dopamine-transporter knockout mice." Nat Neurosci 1.2 (June 1998): 132-137.
PMID
10195128
Source
pubmed
Published In
Nature Neuroscience
Volume
1
Issue
2
Publish Date
1998
Start Page
132
End Page
137
DOI
10.1038/381

The G protein-coupled receptor kinase 2 is a microtubule-associated protein kinase that phosphorylates tubulin.

The G protein-coupled receptor kinase 2 (GRK2) is a serine/threonine kinase that phosphorylates and desensitizes agonist-occupied G protein-coupled receptors (GPCRs). Here we demonstrate that GRK2 is a microtubule-associated protein and identify tubulin as a novel GRK2 substrate. GRK2 is associated with microtubules purified from bovine brain, forms a complex with tubulin in cell extracts, and colocalizes with tubulin in living cells. Furthermore, an endogenous tubulin kinase activity that copurifies with microtubules has properties similar to GRK2 and is inhibited by anti-GRK2 monoclonal antibodies. Indeed, GRK2 phosphorylates tubulin in vitro with kinetic parameters very similar to those for phosphorylation of the agonist-occupied beta2-adrenergic receptor, suggesting a functionally relevant role for this phosphorylation event. In a cellular environment, agonist occupancy of GPCRs, which leads to recruitment of GRK2 to the plasma membrane and its subsequent activation, promotes GRK2-tubulin complex formation and tubulin phosphorylation. These findings suggest a novel role for GRK2 as a GPCR signal transducer mediating the effects of GPCR activation on the cytoskeleton.

Authors
Pitcher, JA; Hall, RA; Daaka, Y; Zhang, J; Ferguson, SS; Hester, S; Miller, S; Caron, MG; Lefkowitz, RJ; Barak, LS
MLA Citation
Pitcher, JA, Hall, RA, Daaka, Y, Zhang, J, Ferguson, SS, Hester, S, Miller, S, Caron, MG, Lefkowitz, RJ, and Barak, LS. "The G protein-coupled receptor kinase 2 is a microtubule-associated protein kinase that phosphorylates tubulin." J Biol Chem 273.20 (May 15, 1998): 12316-12324.
PMID
9575184
Source
pubmed
Published In
The Journal of biological chemistry
Volume
273
Issue
20
Publish Date
1998
Start Page
12316
End Page
12324

Increased MPTP neurotoxicity in vesicular monoamine transporter 2 heterozygote knockout mice.

The neurotoxic action of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) has been proposed to be attenuated by sequestration into intracellular vesicles by the vesicular monoamine transporter (VMAT2). The purpose of this study was to determine if mice with genetically reduced levels of VMAT2 (heterozygote knockout; VMAT2 +/-) were more vulnerable to MPTP. Striatal dopamine (DA) content, the levels of DA transporter (DAT) protein, and the expression of glial fibrillary acidic protein (GFAP) mRNA, a marker of gliosis, were assessed as markers of MPTP neurotoxicity. In all parameters measured VMAT2 +/- mice were more sensitive than their wild-type littermates (VMAT2 +/+). Administration of MPTP (7.5, 15, or 30 mg/kg, b.i.d.) resulted in dose-dependent reductions in striatal DA levels in both VMAT2 +/- and VMAT2 +/+ animals, but the neurotoxic potency of MPTP was approximately doubled in the VMAT2 +/- mice: 59 versus 23% DA loss 7 days after 7.5 mg/kg dose for VMAT2 +/- and VMAT2 +/+ mice, respectively. Dopaminergic nerve terminal integrity, as assessed by DAT protein expression, also revealed more drastic reductions in the VMAT2 +/- mice: 59 versus 35% loss at 7.5 mg/kg and 95 versus 58% loss at 15 mg/kg for VMAT2 +/- and VMAT2 +/+ mice, respectively. Expression of GFAP mRNA 2 days after MPTP was higher in the VMAT2 +/- mice than in the wild-type: 15.8- versus 7.8-fold increase at 7.5 mg/kg and 20.1- versus 9.6-fold at 15 mg/kg for VMAT2 +/- and VMAT2 +/+ mice, respectively. These observations clearly demonstrate that VMAT2 +/- mice are more susceptible to the neurotoxic effects of MPTP, suggesting that VMAT2-mediated sequestration of the neurotoxin into vesicles may play an important role in attenuating MPTP toxicity in vivo.

Authors
Gainetdinov, RR; Fumagalli, F; Wang, YM; Jones, SR; Levey, AI; Miller, GW; Caron, MG
MLA Citation
Gainetdinov, RR, Fumagalli, F, Wang, YM, Jones, SR, Levey, AI, Miller, GW, and Caron, MG. "Increased MPTP neurotoxicity in vesicular monoamine transporter 2 heterozygote knockout mice." J Neurochem 70.5 (May 1998): 1973-1978.
PMID
9572281
Source
pubmed
Published In
Journal of Neurochemistry
Volume
70
Issue
5
Publish Date
1998
Start Page
1973
End Page
1978

Re-evaluation of the role of the dopamine transporter in dopamine system homeostasis.

Mice with a genetic deletion of the dopamine transporter (DAT) were used to assess its role in the function of dopamine (DA) neurons. Profound alterations in the homeostasis of the nigrostriatal DA system were induced by the absence of the DAT. Extracellular levels of DA were elevated and clearance of released DA was 300-times slower than in control mice. This was accompanied by a 20-fold decrease in tissue DA levels and a paradoxical doubling of the rate of DA synthesis. A crucial role is indicated for the DAT in maintenance of DA neuron presynaptic function, particularly in the control of storage mechanisms.

Authors
Gainetdinov, RR; Jones, SR; Fumagalli, F; Wightman, RM; Caron, MG
MLA Citation
Gainetdinov, RR, Jones, SR, Fumagalli, F, Wightman, RM, and Caron, MG. "Re-evaluation of the role of the dopamine transporter in dopamine system homeostasis." Brain Res Brain Res Rev 26.2-3 (May 1998): 148-153. (Review)
PMID
9651511
Source
pubmed
Published In
Brain Res Brain Res Rev
Volume
26
Issue
2-3
Publish Date
1998
Start Page
148
End Page
153

Visualization of dynamic trafficking of a protein kinase C betaII/green fluorescent protein conjugate reveals differences in G protein-coupled receptor activation and desensitization.

Protein kinase C (PKC) links various extracellular signals to intracellular responses and is activated by diverse intracellular factors including diacylglycerol, Ca2+, and arachidonic acid. In this study, using a fully functional green fluorescent protein conjugated PKCbetaII (GFP-PKCbetaII), we demonstrate a novel approach to study the dynamic redistribution of PKC in live cells in response to G protein-coupled receptor activation. Agonist-induced PKC translocation was rapid, transient, and selectively mediated by the activation of Gqalpha- but not Gsalpha- or Gialpha-coupled receptors. Interestingly, although the stimuli were continuously present, only one brief peak of PKC membrane translocation was observed, consistent with rapid desensitization of the signaling pathway. Moreover, when GFP-PKCbetaII was used to examine cross-talk between two Gqalpha-coupled receptors, angiotensin II type 1A receptor (AT1AR) and endothelin A receptor (ETAR), activation of ETARs resulted in a subsequent loss of AT1AR responsiveness, whereas stimulation of AT1ARs did not cause desensitization of the ETAR signaling. The development of GFP-PKCbetaII has allowed not only the real time visualization of the dynamic PKC trafficking in live cells in response to physiological stimuli but has also provided a direct and sensitive means in the assessment of activation and desensitization of receptors implicated in the phospholipase C signaling pathway.

Authors
Feng, X; Zhang, J; Barak, LS; Meyer, T; Caron, MG; Hannun, YA
MLA Citation
Feng, X, Zhang, J, Barak, LS, Meyer, T, Caron, MG, and Hannun, YA. "Visualization of dynamic trafficking of a protein kinase C betaII/green fluorescent protein conjugate reveals differences in G protein-coupled receptor activation and desensitization." J Biol Chem 273.17 (April 24, 1998): 10755-10762.
PMID
9553141
Source
pubmed
Published In
The Journal of biological chemistry
Volume
273
Issue
17
Publish Date
1998
Start Page
10755
End Page
10762

G protein-coupled receptor adaptation mechanisms.

G protein-coupled receptors (GPCRs) transduce extracellular signals that modulate the activity of a wide variety of biological processes, such as neurotransmission, chemoattraction, cardiac function, olfaction, and vision. However, GPCR signalling desensitizes rapidly as the consequence of receptor phosphorylation. G protein-coupled receptor kinase-mediated receptor phosphorylation promotes the binding of beta-arrestin proteins, which not only uncouple GPCRs from their cognate heterotrimeric G protein, but also target them for endocytosis. The sequestration (endocytosis) of desensitized GPCRs to endosomes is required for their dephosphorylation and subsequent resensitization to their pre-ligand exposed state. This review concentrates on the mechanisms underlying GPCR desensitization and resensitization.

Authors
Ferguson, SS; Caron, MG
MLA Citation
Ferguson, SS, and Caron, MG. "G protein-coupled receptor adaptation mechanisms." Semin Cell Dev Biol 9.2 (April 1998): 119-127. (Review)
PMID
9599406
Source
pubmed
Published In
Seminars in Cell and Developmental Biology
Volume
9
Issue
2
Publish Date
1998
Start Page
119
End Page
127
DOI
10.1006/scdb.1997.0216

Profound neuronal plasticity in response to inactivation of the dopamine transporter.

The dopamine transporter (DAT) plays an important role in calibrating the duration and intensity of dopamine neurotransmission in the central nervous system. We have used a strain of mice in which the gene for the DAT has been genetically deleted to identify the DAT's homeostatic role. We find that removal of the DAT dramatically prolongs the lifetime (300 times) of extracellular dopamine. Within the time frame of neurotransmission, no other processes besides diffusion can compensate for the lack of the DAT, and the absence of the DAT produces extensive adaptive changes to control dopamine neurotransmission. Despite the absence of a clearance mechanism, dopamine extracellular levels were only 5 times greater than control animals due to a 95% reduction in content and a 75% reduction in release. Paradoxically, dopamine synthesis rates are doubled despite a decrease of 90% in the levels of tyrosine hydroxylase and degradation is markedly enhanced. Thus, the DAT not only controls the duration of extracellular dopamine signals but also plays a critical role in regulating presynaptic dopamine homeostasis. It is interesting to consider that the switch to a dopamine-deficient, but functionally hyperactive, mode of neurotransmission observed in mice lacking the DAT may represent an extreme example of neuronal plasticity resulting from long-term psychostimulant abuse.

Authors
Jones, SR; Gainetdinov, RR; Jaber, M; Giros, B; Wightman, RM; Caron, MG
MLA Citation
Jones, SR, Gainetdinov, RR, Jaber, M, Giros, B, Wightman, RM, and Caron, MG. "Profound neuronal plasticity in response to inactivation of the dopamine transporter." Proc Natl Acad Sci U S A 95.7 (March 31, 1998): 4029-4034.
PMID
9520487
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
95
Issue
7
Publish Date
1998
Start Page
4029
End Page
4034

A role for receptor kinases in the regulation of class II G protein-coupled receptors. Phosphorylation and desensitization of the secretin receptor.

The secretin receptor is a member of a structurally distinct class of G protein-coupled receptors designated as Class II. The molecular mechanisms of secretin receptor signal termination are unknown. Using transiently transfected HEK 293 cells expressing the secretin receptor, we investigated its mechanisms of desensitization. Binding of [125I]-secretin to plasma membranes of receptor-expressing cells was specific, with a Kd of 2 nM. Secretin evoked an increase in cellular cAMP with an EC50 of 0.4 nM. The response was maximal by 20 min and desensitized rapidly and completely. Immunoprecipitation of a functional, N-terminal epitope-tagged secretin receptor was used to demonstrate agonist-dependent receptor phosphorylation, with an EC50 of 14 nM. Pretreatment with protein kinase A or C inhibitors failed to alter secretin-stimulated cAMP accumulation. G protein-coupled receptor kinases (GRKs) are known to be involved in the desensitization of Class I G protein-coupled receptors; therefore, the effect of cotransfection of GRKs on secretin-stimulated cAMP signaling and phosphorylation was evaluated. GRKs 2 and 5 were the most potent at augmenting desensitization, causing a 40% reduction in the maximal cAMP response to secretin. GRK 5 also caused a shift in the EC50 to the right (p < 0.05). GRK 4 and GRK 6 did not alter dose-dependent signaling, and GRK 3 was intermediate in effect. Receptor phosphorylation correlated with desensitization for each GRK studied, whereas second messenger-dependent kinase phosphorylation appeared to be less important in secretin receptor signal termination. We demonstrate agonist-dependent secretin receptor phosphorylation coincident with profound receptor desensitization of the signaling function in HEK 293 cells, suggesting a role for receptor phosphorylation in this paradigm. Although GRK activity appears important in secretin receptor desensitization in HEK 293 cells, protein kinases A and C appear to play only a minor role. These results demonstrate that the GRK-arrestin system regulates Class II G protein-coupled receptors.

Authors
Shetzline, MA; Premont, RT; Walker, JK; Vigna, SR; Caron, MG
MLA Citation
Shetzline, MA, Premont, RT, Walker, JK, Vigna, SR, and Caron, MG. "A role for receptor kinases in the regulation of class II G protein-coupled receptors. Phosphorylation and desensitization of the secretin receptor." J Biol Chem 273.12 (March 20, 1998): 6756-6762.
PMID
9506976
Source
pubmed
Published In
The Journal of biological chemistry
Volume
273
Issue
12
Publish Date
1998
Start Page
6756
End Page
6762

Mechanisms of amphetamine action revealed in mice lacking the dopamine transporter.

Amphetamine (AMPH) inhibits uptake and causes release of dopamine (DA) from presynaptic terminals. AMPH can act on both vesicular storage of DA and directly on the dopamine transporter (DAT). To assess the relative importance of these two processes, we have examined the releasing actions of AMPH in mice with a genetic deletion of the DAT. The sequence of actions of AMPH has been determined by following the real time changes of DA in the extracellular fluid of intact tissue with fast scan cyclic voltammetry. In striatal slices from wild-type mice, AMPH causes a gradual (approximately 30 min) increase in extracellular DA, with a concomitant disappearance of the pool of DA available for depolarization-evoked release. Conversely, in slices from mice lacking the DAT, although a similar disappearance of electrically stimulated DA release occurs, extracellular DA does not increase. Similarly, microdialysis measurements of DA after AMPH in freely moving animals show no change in mice lacking the DAT, whereas it increases 10-fold in wild-type mice. In contrast, redistribution of DA from vesicles to the cytoplasm by the use of a reserpine-like compound, Ro4-1284, does not increase extracellular DA in slices from wild-type animals; however, subsequent addition of AMPH induces rapid (<5 min) release of DA. Thus, the DAT is required for the releasing action, but not the vesicle-depleting action, of AMPH on DA neurons, and the latter represents the rate-limiting step in the effects of AMPH. Furthermore, these findings suggest that in the absence of pharmacological manipulation, such as the use of amphetamine, endogenous cytoplasmic DA normally does not reach sufficient concentrations to reverse the DAT.

Authors
Jones, SR; Gainetdinov, RR; Wightman, RM; Caron, MG
MLA Citation
Jones, SR, Gainetdinov, RR, Wightman, RM, and Caron, MG. "Mechanisms of amphetamine action revealed in mice lacking the dopamine transporter." J Neurosci 18.6 (March 15, 1998): 1979-1986. (Review)
PMID
9482784
Source
pubmed
Published In
The Journal of neuroscience : the official journal of the Society for Neuroscience
Volume
18
Issue
6
Publish Date
1998
Start Page
1979
End Page
1986

Essential role for G protein-coupled receptor endocytosis in the activation of mitogen-activated protein kinase.

The classical paradigm for G protein-coupled receptor (GPCR) signal transduction involves the agonist-dependent interaction of GPCRs with heterotrimeric G proteins at the plasma membrane and the subsequent generation, by membrane-localized effectors, of soluble second messengers or ion currents. Termination of GPCR signals follows G protein-coupled receptor kinase (GRK)- and beta-arrestin-mediated receptor uncoupling and internalization. Here we show that these paradigms are inadequate to account for GPCR-mediated, Ras-dependent activation of the mitogen-activated protein (MAP) kinases Erk1 and -2. In HEK293 cells expressing dominant suppressor mutants of beta-arrestin or dynamin, beta2-adrenergic receptor-mediated activation of MAP kinase is inhibited. The inhibitors of receptor internalization specifically blocked Raf-mediated activation of MEK. Plasma membrane-delimited steps in the GPCR-mediated activation of the MAP kinase pathway, such as tyrosine phosphorylation of Shc and Raf kinase activation by Ras, are unaffected by inhibitors of receptor internalization. Thus, GRKs and beta-arrestins, which uncouple GPCRs and target them for internalization, function as essential elements in the GPCR-mediated MAP kinase signaling cascade.

Authors
Daaka, Y; Luttrell, LM; Ahn, S; Della Rocca, GJ; Ferguson, SS; Caron, MG; Lefkowitz, RJ
MLA Citation
Daaka, Y, Luttrell, LM, Ahn, S, Della Rocca, GJ, Ferguson, SS, Caron, MG, and Lefkowitz, RJ. "Essential role for G protein-coupled receptor endocytosis in the activation of mitogen-activated protein kinase." J Biol Chem 273.2 (January 9, 1998): 685-688.
PMID
9422717
Source
pubmed
Published In
The Journal of biological chemistry
Volume
273
Issue
2
Publish Date
1998
Start Page
685
End Page
688

Dopamine receptors: from structure to function.

The diverse physiological actions of dopamine are mediated by at least five distinct G protein-coupled receptor subtypes. Two D1-like receptor subtypes (D1 and D5) couple to the G protein Gs and activate adenylyl cyclase. The other receptor subtypes belong to the D2-like subfamily (D2, D3, and D4) and are prototypic of G protein-coupled receptors that inhibit adenylyl cyclase and activate K+ channels. The genes for the D1 and D5 receptors are intronless, but pseudogenes of the D5 exist. The D2 and D3 receptors vary in certain tissues and species as a result of alternative splicing, and the human D4 receptor gene exhibits extensive polymorphic variation. In the central nervous system, dopamine receptors are widely expressed because they are involved in the control of locomotion, cognition, emotion, and affect as well as neuroendocrine secretion. In the periphery, dopamine receptors are present more prominently in kidney, vasculature, and pituitary, where they affect mainly sodium homeostasis, vascular tone, and hormone secretion. Numerous genetic linkage analysis studies have failed so far to reveal unequivocal evidence for the involvement of one of these receptors in the etiology of various central nervous system disorders. However, targeted deletion of several of these dopamine receptor genes in mice should provide valuable information about their physiological functions.

Authors
Missale, C; Nash, SR; Robinson, SW; Jaber, M; Caron, MG
MLA Citation
Missale, C, Nash, SR, Robinson, SW, Jaber, M, and Caron, MG. "Dopamine receptors: from structure to function." Physiol Rev 78.1 (January 1998): 189-225. (Review)
PMID
9457173
Source
pubmed
Published In
Physiological reviews
Volume
78
Issue
1
Publish Date
1998
Start Page
189
End Page
225

Cloning, gene structure and genomic localization of an orphan transporter from mouse kidney with six alternatively-spliced isoforms.

Two genes were identified and characterized that express cDNAs related to previously identified neurotransmitter and/or osmolyte transporters, but which are expressed specifically in the kidney. RNA transcribed from one of these two genes (XT2) was found to undergo an extensive degree of alternative splicing to generate six distinct isoforms. The intron-exon structure of the XT2 gene and the sites of alternative splicing were identified. Expression of the second gene (XT3) was found to be conserved in human kidney, and partial sequence was obtained from a human cDNA library. The expressions of both XT2 and XT3 RNAs were determined in mouse and human tissues, respectively, and the locations of the two genes within the mouse genome were identified. Screening experiments to identify the substrate(s) of these proteins failed to identify specific uptake with any of the tested compounds; however, immunofluorescent microscopy demonstrated that epitope-tagged variants of the protein products of the XT2 and XT3 cDNAs were present on the plasma membrane of transfected cells.

Authors
Nash, SR; Giros, B; Kingsmore, SF; Kim, KM; el-Mestikawy, S; Dong, Q; Fumagalli, F; Seldin, MF; Caron, MG
MLA Citation
Nash, SR, Giros, B, Kingsmore, SF, Kim, KM, el-Mestikawy, S, Dong, Q, Fumagalli, F, Seldin, MF, and Caron, MG. "Cloning, gene structure and genomic localization of an orphan transporter from mouse kidney with six alternatively-spliced isoforms." Receptors Channels 6.2 (1998): 113-128.
PMID
9932288
Source
pubmed
Published In
Receptors & channels
Volume
6
Issue
2
Publish Date
1998
Start Page
113
End Page
128

Role of beta-arrestins in the intracellular trafficking of G-protein-coupled receptors.

Authors
Ferguson, SS; Zhang, J; Barak, LS; Caron, MG
MLA Citation
Ferguson, SS, Zhang, J, Barak, LS, and Caron, MG. "Role of beta-arrestins in the intracellular trafficking of G-protein-coupled receptors." Adv Pharmacol 42 (1998): 420-424.
PMID
9327929
Source
pubmed
Published In
Advances in pharmacology (San Diego, Calif.)
Volume
42
Publish Date
1998
Start Page
420
End Page
424

[Behavioral, cellular and molecular consequences of the dopamine transporter gene inactivation].

Mice lacking the the plasma membrane dopamine transporter (DAT), following gene inactivation or knock out, show an increase in their spontaneous locomotor activity that is of the same magnitude than in normal mice treated with amphetamine or cocaine, known to increase levels of dopamine in the basal ganglia. Many adaptive responses have occurred in these animals than could not compensate for the hyper activity of the dopamine system. Surprisingly, while intracellular dopamine levels were of only 5%, extracellular dopamine levels were increased by 300%. We investigated the regulation of tyrosine hydroxylase (TH), the rate limiting enzyme of dopamine synthesis, and found that this enzyme is regulated at the levels of mRNA, protein, trafficking as well as in its regional, cellular and subcellular organization. Our results establish not only the central importance of the transporter as the key element controlling dopamine levels in the brain, but also its role in the behavioral and biochemical action of amphetamine, cocaine and morphine. In addition, these mice have provided key elements leading to possible clinical and social implications for illnesses such as Parkinson disease, attention deficit disorder and drug addiction.

Authors
Jaber, M; Bloch, B; Caron, MG; Giros, B
MLA Citation
Jaber, M, Bloch, B, Caron, MG, and Giros, B. "[Behavioral, cellular and molecular consequences of the dopamine transporter gene inactivation]." C R Seances Soc Biol Fil 192.6 (1998): 1127-1137. (Review)
PMID
10101608
Source
pubmed
Published In
Journal de la Societe de Biologie
Volume
192
Issue
6
Publish Date
1998
Start Page
1127
End Page
1137

Inactivation of the dopamine transporter reveals essential roles of dopamine in the control of locomotion, psychostimulant response, and pituitary function.

Authors
Fumagalli, F; Jones, S; Bossé, R; Jaber, M; Giros, B; Missale, C; Wightman, RM; Caron, MG
MLA Citation
Fumagalli, F, Jones, S, Bossé, R, Jaber, M, Giros, B, Missale, C, Wightman, RM, and Caron, MG. "Inactivation of the dopamine transporter reveals essential roles of dopamine in the control of locomotion, psychostimulant response, and pituitary function." Adv Pharmacol 42 (1998): 179-182.
PMID
9327873
Source
pubmed
Published In
Advances in pharmacology (San Diego, Calif.)
Volume
42
Publish Date
1998
Start Page
179
End Page
182

Molecular mechanisms of G protein-coupled receptor desensitization and resensitization.

Beta-arrestin proteins play a dual role in regulating G protein-coupled receptor (GPCR) responsiveness by contributing to both receptor desensitization and internalization. Recently, beta-arrestins were also shown to be critical determinants for beta2-adrenergic receptor (beta2AR) resensitization. This was demonstrated by overexpressing wild-type beta-arrestins to rescue the resensitization-defect of a beta2AR (Y326A) mutant (gain of function) and overexpressing a dominant-negative beta-arrestin inhibitor of beta2AR sequestration to impair beta2AR dephosphorylation and resensitization (loss of function). Moreover, the ability of the beta2AR to resensitize in different cell types was shown to be dependent upon beta-arrestin expression levels. To further study the mechanisms underlying beta-arrestin function, green fluorescent protein was coupled to beta-arrestin2 (beta arr2GFP), thus allowing the real-time visualization of the agonist-dependent trafficking of beta-arrestin in living cells. Beta arr2GFP translocation from the cytoplasm to the plasma membrane proceeded with a time course, sensitivity and specificity that was indistinguishable from the most sensitive second messenger readout systems. Beta arr2GFP translocation was GRK-dependent and was demonstrated for 16 different ligand-activated GPCRs. Because beta-arrestin binding is a common divergent step in GPCR signalling, this assay represents a universal methodology for screening orphan receptors, GRK inhibitors and novel GPCR ligands. Moreover, beta arr2GFP provides a valuable new tool to dissect the biological function and regulation of beta-arrestin proteins.

Authors
Ferguson, SS; Zhang, J; Barak, LS; Caron, MG
MLA Citation
Ferguson, SS, Zhang, J, Barak, LS, and Caron, MG. "Molecular mechanisms of G protein-coupled receptor desensitization and resensitization." Life Sci 62.17-18 (1998): 1561-1565. (Review)
PMID
9585136
Source
pubmed
Published In
Life Sciences
Volume
62
Issue
17-18
Publish Date
1998
Start Page
1561
End Page
1565

Cocaine self-administration in dopamine-transporter knockout mice

The plasma membrane dopamine transporter (DAT) is responsible for clearing dopamine from the synapse. Cocaine blockade of DAT leads to increased extracellular dopamine, an effect widely considered to be the primary cause of the reinforcing and addictive properties of cocaine. In this study we tested whether these properties are limited to the dopaminergic system in mice lacking DAT. In the absence of DAT, these mice exhibit high levels of extracellular dopamine, but paradoxically still self-administer cocaine. Mapping of the sites of cocaine binding and neuronal activation suggests an involvement of serotonergic brain regions in this response. These results demonstrate that the interaction of cocaine with targets other than DAT, possibly the serotonin transporter, can initiate and sustain cocaine self-administration in these mice.

Authors
Rocha, BA; Fumagalli, F; Gainetdinov, RR; Jones, SR; Ator, R; Giros, B; Miller, GW; Caron, MG
MLA Citation
Rocha, BA, Fumagalli, F, Gainetdinov, RR, Jones, SR, Ator, R, Giros, B, Miller, GW, and Caron, MG. "Cocaine self-administration in dopamine-transporter knockout mice." Nature Neuroscience 1.2 (1998): 132-137.
Source
scival
Published In
Nature Neuroscience
Volume
1
Issue
2
Publish Date
1998
Start Page
132
End Page
137

Altered cardiac chronotropic responses in βARK 2 knockout mice

G protein-coupled receptor kinases (GRKs) desensitize agonist-activated G protein-coupled receptors (GPCRs) thus attenuating their responses. In cell culture experiments, GRK 3 (βARK 2) phosphorylates and desensitizes multiple GPCRs, including muscarinic acetylcholine receptors (mAChR). Genetically altered mice, lacking βARK 2 (βARK 2 -/-), display a more rapid heart rate recovery after an i.v. injection of methacholine, than wildtype mice (WT). Since this observation is contrary to that expected (assuming βARK 2 desensitizes cardiac m2AChR in vivo) and given that i.v. methacholine induces hypotension, we hypothesized that BARK 2 may desensitize a central component of cardiovascular control, the baroreceptor reflex. To test this hypothesis we examined baroreflexes as well as the impact of direct cardiac m2AChR activation with vagal stimulation. During vagal nerve stimulation (10, 15 and 20 Hz) heart rate decreased, and recovered, similarly in βARK 2 -/- and WT mice. However, the increase in heart rate associated with sodium nitroprusside-induced hypotension was significantly greater in βARK 2 -/- (88 ± 12 beats/min) than WT (31 ± 16 beats/min) mice. Interestingly, propranolol significantly decreased resting heart rate in WT (486 ± 28 to 419 ± 22 beats/min), but not βARK 2 -/-mice (443 ±26 to 426 ± 25 beats/min). In conclusion, the cardiac component of the baroreceptor reflex is likely responsible for the chronotropic differences observed between βARK 2 -/- and WT mice.

Authors
Walker, JKL; Peppel, K; Lefkowitz, RJ; Caron, MG; Fisher, JT
MLA Citation
Walker, JKL, Peppel, K, Lefkowitz, RJ, Caron, MG, and Fisher, JT. "Altered cardiac chronotropic responses in βARK 2 knockout mice." FASEB Journal 12.4 (1998): A396-.
Source
scival
Published In
The FASEB journal : official publication of the Federation of American Societies for Experimental Biology
Volume
12
Issue
4
Publish Date
1998
Start Page
A396

Knockout of the vesicular monoamine transporter 2 gene results in neonatal death and supersensitivity to cocaine and amphetamine.

Vesicular monoamine transporters are known to transport monoamines from the cytoplasm into secretory vesicles. We have used homologous recombination to generate mutant mice lacking the vesicular monoamine transporter 2 (VMAT2), the predominant form expressed in the brain. Newborn homozygotes die within a few days after birth, manifesting severely impaired monoamine storage and vesicular release. In heterozygous adult mice, extracellular striatal dopamine levels, as well as K+- and amphetamine-evoked dopamine release, are diminished. The observed changes in presynaptic homeostasis are accompanied by a pronounced supersensitivity of the mice to the locomotor effects of the dopamine agonist apomorphine, the psychostimulants cocaine and amphetamine, and ethanol. Importantly, VMAT2 heterozygous mice do not develop further sensitization to repeated cocaine administration. These observations stress the importance of VMAT2 in the maintenance of presynaptic function and suggest that these mice may provide an animal model for delineating the mechanisms of vesicular release, monoamine function, and postsynaptic sensitization associated with drug abuse.

Authors
Wang, YM; Gainetdinov, RR; Fumagalli, F; Xu, F; Jones, SR; Bock, CB; Miller, GW; Wightman, RM; Caron, MG
MLA Citation
Wang, YM, Gainetdinov, RR, Fumagalli, F, Xu, F, Jones, SR, Bock, CB, Miller, GW, Wightman, RM, and Caron, MG. "Knockout of the vesicular monoamine transporter 2 gene results in neonatal death and supersensitivity to cocaine and amphetamine." Neuron 19.6 (December 1997): 1285-1296.
PMID
9427251
Source
pubmed
Published In
Neuron
Volume
19
Issue
6
Publish Date
1997
Start Page
1285
End Page
1296

A beta-arrestin/green fluorescent protein biosensor for detecting G protein-coupled receptor activation.

G protein-coupled receptors (GPCR) represent the single most important drug targets for medical therapy, and information from genome sequencing and genomic data bases has substantially accelerated their discovery. The lack of a systematic approach either to identify the function of a new GPCR or to associate it with a cognate ligand has added to the growing number of orphan receptors. In this work we provide a novel approach to this problem using a beta-arrestin2/green fluorescent protein conjugate (betaarr2-GFP). It provides a real-time and single cell based assay to monitor GPCR activation and GPCR-G protein-coupled receptor kinase or GPCR-arrestin interactions. Confocal microscopy demonstrates the translocation of betaarr2-GFP to more than 15 different ligand-activated GPCRs. These data clearly support the common hypothesis that the beta-arrestin binding of an activated receptor is a convergent step of GPCR signaling, increase by 5-fold the number of GPCRs known to interact with beta-arrestins, demonstrate that the cytosol is the predominant reservoir of biologically active beta-arrestins, and provide the first direct demonstration of the critical importance of G protein-coupled receptor kinase phosphorylation to the biological regulation of beta-arrestin activity and GPCR signal transduction in living cells. The use of betaarr2-GFP as a biosensor to recognize the activation of pharmacologically distinct GPCRs should accelerate the identification of orphan receptors and permit the optical study of their signal transduction biology intractable to ordinary biochemical methods.

Authors
Barak, LS; Ferguson, SS; Zhang, J; Caron, MG
MLA Citation
Barak, LS, Ferguson, SS, Zhang, J, and Caron, MG. "A beta-arrestin/green fluorescent protein biosensor for detecting G protein-coupled receptor activation." J Biol Chem 272.44 (October 31, 1997): 27497-27500.
PMID
9346876
Source
pubmed
Published In
The Journal of biological chemistry
Volume
272
Issue
44
Publish Date
1997
Start Page
27497
End Page
27500

A central role for beta-arrestins and clathrin-coated vesicle-mediated endocytosis in beta2-adrenergic receptor resensitization. Differential regulation of receptor resensitization in two distinct cell types.

G protein-coupled receptor (GPCR) sequestration to endosomes is proposed to be the mechanism by which G protein-coupled receptor kinase (GRK)-phosphorylated receptors are dephosphorylated and resensitized. The identification of beta-arrestins as GPCR trafficking molecules suggested that beta-arrestins might represent critical determinants for GPCR resensitization. Therefore, we tested whether beta2-adrenergic receptor (beta2AR) resensitization was dependent upon beta-arrestins and an intact clathrin-coated vesicle endocytic pathway. The overexpression of either the beta-arrestin 1-V53D dominant negative inhibitor of beta2AR sequestration or dynamin I-K44A to block clathrin-coated vesicle-mediated endocytosis impaired both beta2AR dephosphorylation and resensitization. In contrast, resensitization of a sequestration-impaired beta2AR mutant (Y326A) was reestablished following the overexpression of either GRK2 or beta-arrestin 1. Moreover, beta2ARs did not resensitize in COS-7 cells as the consequence of impaired sequestration and dephosphorylation. However, beta2AR resensitization was restored in these cells following the overexpression of beta-arrestin 2. These findings demonstrate, using both loss and gain of function paradigms, that beta2AR dephosphorylation and resensitization are dependent upon an intact sequestration pathway. These studies also indicate that beta-arrestins play an integral role in regulating not only the desensitization and intracellular trafficking of GPCRs but their ability to resensitize. beta-Arrestin expression levels appear to underlie cell type-specific differences in the regulation of GPCR resensitization.

Authors
Zhang, J; Barak, LS; Winkler, KE; Caron, MG; Ferguson, SS
MLA Citation
Zhang, J, Barak, LS, Winkler, KE, Caron, MG, and Ferguson, SS. "A central role for beta-arrestins and clathrin-coated vesicle-mediated endocytosis in beta2-adrenergic receptor resensitization. Differential regulation of receptor resensitization in two distinct cell types." J Biol Chem 272.43 (October 24, 1997): 27005-27014.
PMID
9341139
Source
pubmed
Published In
The Journal of biological chemistry
Volume
272
Issue
43
Publish Date
1997
Start Page
27005
End Page
27014

G protein-coupled receptor kinase 3 (GRK3) gene disruption leads to loss of odorant receptor desensitization.

G protein-coupled receptor kinases (GRKs) 2 and 3 (beta-adrenergic receptor kinases 1 and 2 (betaARK1 and -2)) mediate the agonist-dependent phosphorylation and uncoupling of many G protein-coupled receptors. These two members of the GRK family share a high degree of sequence homology and show overlapping patterns of substrate specificity in vitro. To define their physiological roles in vivo we have generated mice that carry targeted disruption of these genes. In contrast to GRK2-deficient mice, which die in utero (Jaber, M., Koch, W. J., Rockman, H., Smith, B., Bond, R. A., Sulik, K. K., Ross, J. JR., Lefkowitz, R. J. Caron, M. G., and Giros, B. (1996) Proc. Natl. Acad. Sci. U. S. A. 93, 12974-12979), GRK3 deletion allows for normal embryonic and postnatal development. GRK3 is expressed to a high degree in the olfactory epithelium, where GRK2 is absent. Here we report that cilia preparations derived from GRK3-deficient mice lack the fast agonist-induced desensitization normally seen after odorant stimulation. Moreover, total second messenger (cAMP) generation in these cilia preparations following odorant stimulation is markedly reduced when compared with preparations from wild-type littermates. This reduction in the ability to generate cAMP is evident even in the presence of nonodorant receptor stimuli (GTPgammaS and forskolin), suggesting a compensatory dampening of the G protein-adenylyl cyclase system in the GRK3 (-/-) mice in the olfactory epithelium. These findings demonstrate the requirement of GRK3 for odorant-induced desensitization of cAMP responses.

Authors
Peppel, K; Boekhoff, I; McDonald, P; Breer, H; Caron, MG; Lefkowitz, RJ
MLA Citation
Peppel, K, Boekhoff, I, McDonald, P, Breer, H, Caron, MG, and Lefkowitz, RJ. "G protein-coupled receptor kinase 3 (GRK3) gene disruption leads to loss of odorant receptor desensitization." J Biol Chem 272.41 (October 10, 1997): 25425-25428.
PMID
9325250
Source
pubmed
Published In
The Journal of biological chemistry
Volume
272
Issue
41
Publish Date
1997
Start Page
25425
End Page
25428

The dopamine transporter: a crucial component regulating dopamine transmission.

The dopamine system is implicated in the control of locomotion, cognition, and endocrine function. The relative contribution of the various dopamine-related components is not well established mainly because drugs that target the dopaminergic system often lack selectivity. The in vivo gene inactivation procedure, or knockout, enables the creation of new strains of mice lacking a specific gene. This technique has been applied recently to inactivate the expression of the plasma membrane dopamine transporter. Here we summarize the main findings obtained with these transgenic mice carrying this "genetic defect," leading to a better understanding of the relative contribution of the dopamine transporter regarding locomotor activity, regulation of the expression of peptides under the control of dopaminergic activity, and responses to various drugs targeting the dopamine system. Our results establish not only the central importance of the transporter as the key element controlling dopamine levels in the brain, but also its role as an obligatory target for the behavioral and biochemical action of amphetamine and cocaine. In addition, the genetically altered mice offer a unique model to test the specificity and selectivity of dopamine transporter-acting drugs and may provide important new concepts related to the clinical and social implications of conditions such as Parkinson's disease, schizophrenia, and drug addiction.

Authors
Jaber, M; Jones, S; Giros, B; Caron, MG
MLA Citation
Jaber, M, Jones, S, Giros, B, and Caron, MG. "The dopamine transporter: a crucial component regulating dopamine transmission." Mov Disord 12.5 (September 1997): 629-633. (Review)
PMID
9380041
Source
pubmed
Published In
Movement Disorders
Volume
12
Issue
5
Publish Date
1997
Start Page
629
End Page
633
DOI
10.1002/mds.870120502

Selective inhibition of adenylyl cyclase type V by the dopamine D3 receptor.

Despite a great deal of research, the second messenger coupling of the dopamine D3 receptor has not yet been clearly established. The closely related D2 and D4 receptors have been shown to inhibit adenylyl cyclase activity in a variety of cell types, but the D3 receptor has little or no effect on this second messenger system. We now demonstrate that when the D3 receptor and adenylyl cyclase type V are coexpressed in 293 cells, the agonist quinpirole causes 70% inhibition of forskolin-stimulated cAMP levels. This effect seems to be selective for this adenylyl cyclase isoform because the D3 receptor does not inhibit adenylyl cyclase types I or VI and only weakly stimulates adenylyl cyclase type II. In contrast, the D2 receptor inhibits cAMP accumulation in 293 cells in the absence of cotransfected adenylyl cyclases and stimulates adenylyl cyclase type II to a greater extent than the D3 receptor. The inhibition of adenylyl cyclase type V by the D3 receptor is sensitive to pertussis toxin, suggesting the involvement of G proteins of the Gi family. Guanosine-5'-O-(3-thio)triphosphate binding studies indicate that the D3 receptor weakly activates all three Gialpha subunits, whereas the D2 receptor activates these G proteins to a substantially greater extent. However, despite its relative inability to promote G protein activation, the D3 receptor is capable of substantial and consistent inhibition of adenylyl cyclase type V. The robust second messenger coupling of the D3 receptor in a heterologous system with defined components provides a system for further studies of the function of this receptor and should facilitate the development and characterization of new D3 receptor ligands.

Authors
Robinson, SW; Caron, MG
MLA Citation
Robinson, SW, and Caron, MG. "Selective inhibition of adenylyl cyclase type V by the dopamine D3 receptor." Mol Pharmacol 52.3 (September 1997): 508-514.
PMID
9281614
Source
pubmed
Published In
Molecular pharmacology
Volume
52
Issue
3
Publish Date
1997
Start Page
508
End Page
514

Dopamine transporter is required for in vivo MPTP neurotoxicity: evidence from mice lacking the transporter.

The neurotoxic effect of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) was tested on mice lacking the dopamine (DA) transporter (DAT-/- mice). Striatal tissue DA content and glial fibrillary acidic protein (GFAP) mRNA expression were assessed as markers of MPTP neurotoxicity. MPTP (30 mg/kg, s.c., b.i.d.) produced an 87% decrease in tissue DA levels and a 29-fold increase in the level of GFAP mRNA in the striatum of wild-type animals 48 h after administration. Conversely, there were no significant changes in either parameter in DAT-/- mice. Heterozygotes demonstrated partial sensitivity to MPTP administration as shown by an intermediate value (48%) of tissue DA loss. Direct intrastriatal infusion of the active metabolite of MPTP, 1-methyl-4-phenylpyridinium (MPP+; 10 mM), via a microdialysis probe produced a massive efflux of DA in wild-type mice (>320-fold). In the DAT-/- mice the same treatment produced a much smaller increase in extracellular DA (sixfold), which is likely secondary to tissue damage due to the implantation of the dialysis probe. These observations show that the DAT is a mandatory component for expression of MPTP toxicity in vivo.

Authors
Gainetdinov, RR; Fumagalli, F; Jones, SR; Caron, MG
MLA Citation
Gainetdinov, RR, Fumagalli, F, Jones, SR, and Caron, MG. "Dopamine transporter is required for in vivo MPTP neurotoxicity: evidence from mice lacking the transporter." J Neurochem 69.3 (September 1997): 1322-1325.
PMID
9282960
Source
pubmed
Published In
Journal of Neurochemistry
Volume
69
Issue
3
Publish Date
1997
Start Page
1322
End Page
1325

Anterior pituitary hypoplasia and dwarfism in mice lacking the dopamine transporter.

Deletion of the dopamine transporter (DAT) results in increased dopaminergic tone, anterior pituitary hypoplasia, dwarfism, and an inability to lactate. DAT elimination alters the spatial distribution and dramatically reduces the numbers of lactotrophs and somatotrophs in the pituitary. Despite having normal circulating levels of growth hormone and prolactin in blood, hypoplastic glands from DAT-/- mice fail to respond to secretagog stimulation. The effects of DAT deletion on pituitary function result from elevated DA levels that down-regulate the lactotroph D2 DA receptors and depress hypothalamic growth hormone-releasing hormone content. These results reveal an unexpected and important role or DA in the control of developmental events in the pituitary gland and assign a critical role for hypothalamic DA reuptake in regulating these events.

Authors
Bossé, R; Fumagalli, F; Jaber, M; Giros, B; Gainetdinov, RR; Wetsel, WC; Missale, C; Caron, MG
MLA Citation
Bossé, R, Fumagalli, F, Jaber, M, Giros, B, Gainetdinov, RR, Wetsel, WC, Missale, C, and Caron, MG. "Anterior pituitary hypoplasia and dwarfism in mice lacking the dopamine transporter." Neuron 19.1 (July 1997): 127-138.
PMID
9247269
Source
pubmed
Published In
Neuron
Volume
19
Issue
1
Publish Date
1997
Start Page
127
End Page
138

HIV-1-induced cell fusion is mediated by multiple regions within both the viral envelope and the CCR-5 co-receptor.

Although the human hCCR-5 chemokine receptor can serve as a co-receptor for both M-tropic (ADA and BaL) and dual-tropic (89.6) strains of human immunodeficiency virus type 1 (HIV-1), the closely related mouse mCCR-5 homolog is inactive. We used chimeric hCCR-5-mCCR-5 receptor molecules to examine the functional importance of the three extracellular domains of hCCR-5 that differ in sequence from their mCCR-5 equivalents. While this analysis revealed that all three of these extracellular domains could participate in the functional interaction with HIV-1 envelope, clear differences were observed when different HIV-1 strains were analyzed. Thus, while the ADA HIV-1 isolate could effectively utilize chimeric human-mouse CCR-5 chimeras containing any single human extracellular domain, the BaL isolate required any two human extracellular sequences while the 89.6 isolate would only interact effectively with chimeras containing all three human extracellular sequences. Further analysis using hybrid HIV-1 envelope proteins showed that the difference in co-receptor specificity displayed by the ADA and BaL isolates was due partly to a single amino acid change in the V3 loop, although this interaction was clearly also modulated by other envelope domains. Overall, these data indicate that the interaction between HIV-1 envelope and CCR-5 is not only complex but also subject to marked, HIV-1 isolate-dependent variation.

Authors
Bieniasz, PD; Fridell, RA; Aramori, I; Ferguson, SS; Caron, MG; Cullen, BR
MLA Citation
Bieniasz, PD, Fridell, RA, Aramori, I, Ferguson, SS, Caron, MG, and Cullen, BR. "HIV-1-induced cell fusion is mediated by multiple regions within both the viral envelope and the CCR-5 co-receptor." EMBO J 16.10 (May 15, 1997): 2599-2609.
PMID
9184207
Source
pubmed
Published In
EMBO Journal
Volume
16
Issue
10
Publish Date
1997
Start Page
2599
End Page
2609
DOI
10.1093/emboj/16.10.2599

Synergistic regulation of beta2-adrenergic receptor sequestration: intracellular complement of beta-adrenergic receptor kinase and beta-arrestin determine kinetics of internalization.

Two of the common mechanisms regulating G protein-coupled receptor (GPCR) signal transduction are phosphorylation and sequestration (internalization). Agonist-mediated receptor phosphorylation by the beta-adrenergic receptor kinase (betaARK) facilitates subsequent interaction with an arrestin protein, resulting in receptor desensitization. Studies of the beta2-adrenergic receptor (beta2AR) receptor in human embryonic kidney (HEK) 293 cells indicate that betaARK and arrestin proteins (beta-arrestins) also regulate sequestration. Consistent with this notion, we show in HEK 293 cells that reduction in or removal of the ability of the beta2AR to be phosphorylated by betaARK or to interact normally with beta-arrestin substantially reduces agonist-mediated sequestration. To evaluate betaARK and beta-arrestin regulation of beta2AR sequestration, we examined the relationship between betaARK and/or beta-arrestin expression and beta2AR sequestration in a variety of cultured cells, including HEK 293, COS 7, CHO, A431, and CHW. COS cells had both the lowest levels of endogenous beta-arrestin expression and beta2AR sequestration, whereas HEK 293 had the highest. Overexpression of beta-arrestin, but not betaARK, in COS cells increased the extent of wild-type beta2AR sequestration to levels observed in HEK 293 cells. However, a betaARK phosphorylation-impaired beta2AR mutant (Y326A) required the simultaneous overexpression of both betaARK and beta-arrestin for this to occur. Among all cell lines, sequestration correlated best with the product of betaARK and beta-arrestin expression. Moreover, an agonist-mediated translocation of wild-type beta2AR and endogenous beta-arrestin 2 to endocytic vesicles prepared from CHO fibroblasts was observed. These data suggest not only that the complement of cellular betaARK and arrestin proteins synergistically regulate beta2AR sequestration but also that beta-arrestins directly regulate beta2AR trafficking as well as desensitization.

Authors
Ménard, L; Ferguson, SS; Zhang, J; Lin, FT; Lefkowitz, RJ; Caron, MG; Barak, LS
MLA Citation
Ménard, L, Ferguson, SS, Zhang, J, Lin, FT, Lefkowitz, RJ, Caron, MG, and Barak, LS. "Synergistic regulation of beta2-adrenergic receptor sequestration: intracellular complement of beta-adrenergic receptor kinase and beta-arrestin determine kinetics of internalization." Mol Pharmacol 51.5 (May 1997): 800-808.
PMID
9145918
Source
pubmed
Published In
Molecular pharmacology
Volume
51
Issue
5
Publish Date
1997
Start Page
800
End Page
808

Internal trafficking and surface mobility of a functionally intact beta2-adrenergic receptor-green fluorescent protein conjugate.

The beta2-adrenergic receptor (beta2AR) is prototypic of the large family of G protein-coupled receptors (GPCRs) whose desensitization and resensitization are regulated by intracellular kinases, arrestin proteins, phosphatases, and ill-defined components of the cellular endocytic machinery. The study of beta2AR signal transduction and behavior in living cells is technically difficult because of the relatively low cellular expression of the receptor and a lack of useful biological reagents. Availability of a functional beta2AR tagged with the highly sensitive Green Fluorescent Protein (GFP) could allow measurements of the various properties of the beta2AR. We demonstrate that a fully functional beta2AR/GFP can be engineered. In mammalian cells, beta2AR/S65T/GFP demonstrates strong, diffuse plasma membrane fluorescence when observed with 480 nm excitation. The fluorescent receptor binds agonist and antagonist, stimulates adenylyl cyclase, undergoes phosphorylation, and is internalized in a manner indistinguishable from wild-type receptor. We then show that its internal trafficking and surface mobility can be determined by measuring only the endogenous fluorescence of the conjugate. beta2AR/S65T/GFP was found to be localized on endosomal membranes in living cells within minutes of agonist treatment, and within 15 min it is observed in more complicated structures formed from fusion of multiple endosomes. Finally, its free diffusion (diffusion coefficient, 4.0-12 x 10(-9) cm2/sec) was assessed on living cells using photobleaching recovery measurements. This approach and the fidelity of the biochemical properties of the beta2AR/S65T/GFP demonstrate that real-time optical measurements of beta2AR (as well as other GPCR) interactions and dynamics on living cells are feasible.

Authors
Barak, LS; Ferguson, SS; Zhang, J; Martenson, C; Meyer, T; Caron, MG
MLA Citation
Barak, LS, Ferguson, SS, Zhang, J, Martenson, C, Meyer, T, and Caron, MG. "Internal trafficking and surface mobility of a functionally intact beta2-adrenergic receptor-green fluorescent protein conjugate." Mol Pharmacol 51.2 (February 1997): 177-184.
PMID
9203621
Source
pubmed
Published In
Molecular pharmacology
Volume
51
Issue
2
Publish Date
1997
Start Page
177
End Page
184

Cocaine and amphetamine elicit differential effects in rats with a unilateral injection of dopamine transporter antisense oligodeoxynucleotides.

We have developed an antisense oligodeoxynucleotide to the dopamine transporter and used it to discriminate the behavioral properties of amphetamine and cocaine. In SK-N-MC cells permanently transfected with the dopamine transporter complementary DNA, treatment with 5 mM antisense oligodeoxynucleotide reduced dopamine uptake by 25% when compared to sense control. Unilateral intranigral administration of dopamine transporter antisense (50 microM) twice daily in freely moving rats for 2.5 days was sufficient to reduce dopamine transporter messenger RNA by 70% as measured by in situ hybridization, but not protein levels as measured by [3H]mazindol binding. However, intranigral treatment via implanted osmotic minipump over a period of seven days produced reductions in both dopamine transporter messenger RNA and protein levels (32%) at a dose of 500 pmol/day. These results indicate a longer half-life for the dopamine transporter than expected. Potassium chloride depolarization of ipsilateral striatal slices showed a greater than 200% increase in dopamine overflow on the antisense-treated side compared to the control side. Since imbalance of dopamine tone is known to induce rotational activity, we tested this behavioral paradigm in rats treated with various oligodeoxynucleotides at different doses and time-points. We have found that antisense-treated animals did not rotate spontaneously under any experimental conditions. Using various psychostimulants that target the dopamine transporter and increase dopamine levels, we found that the antisense-treated animals consistently rotated contralaterally in response to amphetamine (2 mg/kg), but not to cocaine (10 mg/kg) or nomifensine (10 mg/kg). These results bring in vivo evidence for a different mode of action of amphetamine and cocaine on the dopamine transporter and lend direct support to the view that amphetamine acts as a dopamine releaser, whereas cocaine acts by blocking dopamine transport.

Authors
Silvia, CP; Jaber, M; King, GR; Ellinwood, EH; Caron, MG
MLA Citation
Silvia, CP, Jaber, M, King, GR, Ellinwood, EH, and Caron, MG. "Cocaine and amphetamine elicit differential effects in rats with a unilateral injection of dopamine transporter antisense oligodeoxynucleotides." Neuroscience 76.3 (February 1997): 737-747.
PMID
9135047
Source
pubmed
Published In
Neuroscience
Volume
76
Issue
3
Publish Date
1997
Start Page
737
End Page
747

Molecular mechanisms of G protein-coupled receptor signaling: role of G protein-coupled receptor kinases and arrestins in receptor desensitization and resensitization.

Dynamic regulation of G protein-coupled receptor signaling demands a coordinated balance between mechanisms leading to the generation, turning off and re-establishment of agonist-mediated signals. G protein-coupled receptor kinases (GRKs) and arrestin proteins not only mediate agonist-dependent G protein-coupled receptor desensitization, but also initiate the internalization (sequestration) of activated receptors, a process leading to receptor resensitization. Studies on the specificity of beta-arrestin functions reveal a multiplicity of G protein-coupled receptor endocytic pathways and suggest that beta-arrestins might serve as adaptors specifically targeting receptors for dynamin-dependent clathrin-mediated endocytosis. Moreover, inactivation of the GRK2 gene in mice has lead to the discovery of an unexpected role of GRK2 in cardiac development, further emphasizing the pleiotropic function of GRKs and arrestins.

Authors
Zhang, J; Ferguson, SS; Barak, LS; Aber, MJ; Giros, B; Lefkowitz, RJ; Caron, MG
MLA Citation
Zhang, J, Ferguson, SS, Barak, LS, Aber, MJ, Giros, B, Lefkowitz, RJ, and Caron, MG. "Molecular mechanisms of G protein-coupled receptor signaling: role of G protein-coupled receptor kinases and arrestins in receptor desensitization and resensitization." Receptors Channels 5.3-4 (1997): 193-199. (Review)
PMID
9606723
Source
pubmed
Published In
Receptors & channels
Volume
5
Issue
3-4
Publish Date
1997
Start Page
193
End Page
199

Molecular mechanism of desensitization of the chemokine receptor CCR-5: Receptor signaling and internalization are dissociable from its role as an HIV-1 co-receptor

The chemokine receptor, CCR-5, a G protein-coupled receptor (GPCR) which mediates chemotactic responses of certain leukocytes, has been shown to serve as the primary co-receptor for macrophagetropic human immunodeficiency virus type 1 (HIV-1). Here we describe functional coupling of CCR-5 to inhibition of forskolin-stimulated cAMP formation via a pertussis toxin-sensitive G(i) protein mechanism in transfected HEK 293 cells. In response to chemokines, CCR-5 was desensitized, phosphorylated and sequestered like a prototypic GPCR only following overexpression of G protein-coupled receptor kinases (GRKs) and β-arrestins in HEK 293 cells. The lack of CCR-5 desensitization in HEK 293 cells in the absence of GRK overexpression suggests that differences in cellular complements of GRK and/or β-arrestin proteins could represent an important mechanism determining cellular responsiveness. When tested, the activity of CCR-5 as an HIV-1 co-receptor was dependent neither upon its ability to signal nor its ability to be desensitized and internalized following agonist stimulation. Thus, while chemokine-promoted cellular signaling, phosphorylation and internalization of CCR-5 may play an important role in regulation of chemotactic responses in leukocytes, these functions are dissociable from its HIV-1 co-receptor function.

Authors
Aramori, I; Zhang, J; Ferguson, SSG; Bieniasz, PD; Cullen, BR; Caron, MG
MLA Citation
Aramori, I, Zhang, J, Ferguson, SSG, Bieniasz, PD, Cullen, BR, and Caron, MG. "Molecular mechanism of desensitization of the chemokine receptor CCR-5: Receptor signaling and internalization are dissociable from its role as an HIV-1 co-receptor." EMBO Journal 16.15 (1997): 4606-4616.
PMID
9303305
Source
scival
Published In
EMBO Journal
Volume
16
Issue
15
Publish Date
1997
Start Page
4606
End Page
4616
DOI
10.1093/emboj/16.15.4606

Pleiotropic role for GRKs and β-arrestins in receptor regulation

G protein-coupled receptor kinases and arrestin proteins are well-characterized mediators of agonist-dependent G protein-coupled receptor desensitization. These proteins are now shown to play a dual role in receptor regulation by mediating both receptor uncoupling and sequestration, a process important for receptor resensitization. β-Arrestins bound to phosphorylated β2-adrenergic and angiotensin II type 1A receptors act as intracellular trafficking molecules specifically targeting these receptors for dynamin-dependent clathrin-coated vesicle-mediated sequestration.

Authors
Ferguson, SSG; Zhang, J; Barak, LS; Caron, MG
MLA Citation
Ferguson, SSG, Zhang, J, Barak, LS, and Caron, MG. "Pleiotropic role for GRKs and β-arrestins in receptor regulation." News in Physiological Sciences 12.4 (1997): 145-151.
Source
scival
Published In
News in physiological sciences : an international journal of physiology produced jointly by the International Union of Physiological Sciences and the American Physiological Society
Volume
12
Issue
4
Publish Date
1997
Start Page
145
End Page
151

A β-arrestin/green fluorescent protein biosensor for detecting G protein-coupled receptor activation

G protein-coupled receptors (GPCR) represent the single most important drug targets for medical therapy, and information from genome sequencing and genomic data bases has substantially accelerated their discovery. The lack of a systematic approach either to identify the function of a new GPCR or to associate it with a cognate ligand has added to the growing number of orphan receptors. In this work we provide a novel approach to this problem using a β-arrestin2/green fluorescent protein conjugate (βarr2-GFP). It provides a real-time and single cell based assay to monitor GPCR activation and GPCR-G protein-coupled receptor kinase or GPCR-arrestin interactions. Confocal microscopy demonstrates the translocation of βarr2-GFP to more than 15 different ligand-activated GPCRs. These data clearly support the common hypothesis that the β-arrestin binding of an activated receptor is a convergent step of GPCR signaling, increase by 5-fold the number of GPCRs known to interact with β-arrestins, demonstrate that the cytosol is the predominant reservoir of biologically active β-arrestins, and provide the first direct demonstration of the critical importance of G protein-coupled receptor kinase phosphorylation to the biological regulation of β-arrestin activity and GPCR signal transduction in living cells. The use of βarr2-GFP as a biosensor to recognize the activation of pharmacologically distinct GPCRs should accelerate the identification of orphan receptors and permit the optical study of their signal transduction biology intractable to ordinary biochemical methods.

Authors
Barak, LS; Ferguson, SSG; Zhang, J; Caron, MG
MLA Citation
Barak, LS, Ferguson, SSG, Zhang, J, and Caron, MG. "A β-arrestin/green fluorescent protein biosensor for detecting G protein-coupled receptor activation." Journal of Biological Chemistry 272.44 (1997): 27497-27500.
Source
scival
Published In
The Journal of biological chemistry
Volume
272
Issue
44
Publish Date
1997
Start Page
27497
End Page
27500
DOI
10.1074/jbc.272.44.27497

Images in neuroscience. Molecular biology, II. A dopamine transporter mouse knockout.

Authors
Caron, MG
MLA Citation
Caron, MG. "Images in neuroscience. Molecular biology, II. A dopamine transporter mouse knockout." Am J Psychiatry 153.12 (December 1996): 1515-.
PMID
8942444
Source
pubmed
Published In
American Journal of Psychiatry
Volume
153
Issue
12
Publish Date
1996
Start Page
1515
DOI
10.1176/ajp.153.12.1515

Essential role of beta-adrenergic receptor kinase 1 in cardiac development and function.

The beta-adrenergic receptor kinase 1 (beta ARK1) is a member of the G protein-coupled receptor kinase (GRK) family that mediates the agonist-dependent phosphorylation and desensitization of G protein-coupled receptors. We have cloned and disrupted the beta ARK1 gene in mice by homologous recombination. No homozygote beta ARK1-/- embryos survive beyond gestational day 15.5. Prior to gestational day 15.5, beta ARK1-/- embryos display pronounced hypoplasia of the ventricular myocardium essentially identical to the "thin myocardium syndrome" observed upon gene inactivation of several transcription factors (RXR alpha, N-myc, TEF-1, WT-1). Lethality in beta ARK1-/- embryos is likely due to heart failure as they exhibit a > 70% decrease in cardiac ejection fraction determined by direct in utero intravital microscopy. These results along with the virtual absence of endogenous GRK activity in beta ARK1-/- embryos demonstrate that beta ARK1 appears to be the predominant GRK in early embryogenesis and that it plays a fundamental role in cardiac development.

Authors
Jaber, M; Koch, WJ; Rockman, H; Smith, B; Bond, RA; Sulik, KK; Ross, J; Lefkowitz, RJ; Caron, MG; Giros, B
MLA Citation
Jaber, M, Koch, WJ, Rockman, H, Smith, B, Bond, RA, Sulik, KK, Ross, J, Lefkowitz, RJ, Caron, MG, and Giros, B. "Essential role of beta-adrenergic receptor kinase 1 in cardiac development and function." Proc Natl Acad Sci U S A 93.23 (November 12, 1996): 12974-12979.
Website
http://hdl.handle.net/10161/7833
PMID
8917529
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
93
Issue
23
Publish Date
1996
Start Page
12974
End Page
12979

Silencing of the constitutive activity of the dopamine D1B receptor. Reciprocal mutations between D1 receptor subtypes delineate residues underlying activation properties.

Recently, we have shown that the dopamine D1B/D5 receptor displays binding and coupling properties that are reminiscent of those of the constitutively activated G protein-coupled receptors when compared with the related D1A/D1 receptor subtype (Tiberi, M., and Caron, M. G. (1994) J. Biol. Chem. 269, 27925-27931). The carboxyl-terminal region of the third cytoplasmic loop of several G protein-coupled receptors has been demonstrated to be important for the regulation of the equilibrium between inactive and active receptor conformations. In this cytoplasmic region, the primary structure of dopamine D1A and D1B receptors differs by only two residues: Phe264/Arg266 are present in D1A receptor compared with Ile288/Lys290 in the D1B receptor. To investigate whether these structural differences could account for the distinct binding and coupling properties of these dopamine receptor subtypes, we swapped the variant residues located in the carboxyl-terminal region by site-directed mutagenesis. The exchange of the D1A receptor residue Phe264 by the D1B receptor counterpart isoleucine led to a D1A receptor mutant exhibiting D1B-like constitutive properties. In contrast, substitution of D1B receptor Ile288 by the D1A receptor counterpart phenylalanine resulted in a loss of constitutive activation of the D1B receptor with binding and coupling properties similar to the D1A receptor. The Arg/Lys substitution had no effect on the function of either receptor. These results demonstrate that the carboxyl-terminal region, and in particular residue Ile288, is a major determinant of the constitutive activity of the dopamine D1B receptor. Moreover, these results establish that not only can agonist-independent activity of a receptor be induced, but when given the appropriate mutation, it can be reversed or silenced.

Authors
Charpentier, S; Jarvie, KR; Severynse, DM; Caron, MG; Tiberi, M
MLA Citation
Charpentier, S, Jarvie, KR, Severynse, DM, Caron, MG, and Tiberi, M. "Silencing of the constitutive activity of the dopamine D1B receptor. Reciprocal mutations between D1 receptor subtypes delineate residues underlying activation properties." J Biol Chem 271.45 (November 8, 1996): 28071-28076.
PMID
8910419
Source
pubmed
Published In
The Journal of biological chemistry
Volume
271
Issue
45
Publish Date
1996
Start Page
28071
End Page
28076

G-protein-coupled receptor kinases and arrestins: regulators of G-protein-coupled receptor sequestration.

Authors
Ferguson, SS; Zhang, J; Barak, LS; Caron, MG
MLA Citation
Ferguson, SS, Zhang, J, Barak, LS, and Caron, MG. "G-protein-coupled receptor kinases and arrestins: regulators of G-protein-coupled receptor sequestration." Biochem Soc Trans 24.4 (November 1996): 953-959. (Review)
PMID
8968491
Source
pubmed
Published In
Biochemical Society transactions
Volume
24
Issue
4
Publish Date
1996
Start Page
953
End Page
959

Images in neuroscience. A mouse knockout.

Authors
Caron, MG
MLA Citation
Caron, MG. "Images in neuroscience. A mouse knockout." Am J Psychiatry 153.11 (November 1996): 1387-.
PMID
8890669
Source
pubmed
Published In
American Journal of Psychiatry
Volume
153
Issue
11
Publish Date
1996
Start Page
1387
DOI
10.1176/ajp.153.11.1387

G-protein-coupled receptor regulation: role of G-protein-coupled receptor kinases and arrestins.

G-protein-coupled receptors (GPCRs) represent a large family of proteins that transduce extracellular signals to the interior of cells. Signalling through these receptors rapidly desensitized primarily as the consequence of receptor phosphorylation, but receptor sequestration and downregulation can also contribute to this process. Two families of serine/threonine kinases, second messenger dependent protein kinases and receptor-specific G-protein-coupled receptor kinases (GRKs), phosphorylate GPCRs and thereby contribute to receptor desensitization. Receptor-specific phosphorylation of GPCRs promotes the binding of cytosolic proteins referred to as arrestins, which function to further uncouple GPCRs from their heterotrimeric G-proteins. To date, the GRK protein family consists of six members, which can be further classified into subgroups according to sequence homology and functional similarities. The arrestin protein family also comprises six members, which are subgrouped on the basis of sequence homology and tissue distribution. While the molecular mechanisms contributing to GPCR desensitization are fairly well characterized, little is known about the mechanism(s) by which GPCR responsiveness is reestablished, other than that receptor sequestration (internalization) might be involved. The goal of the present review is to overview current understanding of the regulation of GPCR responsiveness. In particular, we will review new evidence suggesting a pleiotropic role for GRKs and arrestins in the regulation of GPCR responsiveness. GRK-mediated phosphorylation and arrestin binding are not only involved in the functional uncoupling of GPCRs but they are also intimately involved in promoting GPCR sequestration and as such likely play an important role in mediating the subsequent resensitization of GPCRs.

Authors
Ferguson, SS; Barak, LS; Zhang, J; Caron, MG
MLA Citation
Ferguson, SS, Barak, LS, Zhang, J, and Caron, MG. "G-protein-coupled receptor regulation: role of G-protein-coupled receptor kinases and arrestins." Can J Physiol Pharmacol 74.10 (October 1996): 1095-1110. (Review)
PMID
9022829
Source
pubmed
Published In
Canadian Journal of Physiology and Pharmacology
Volume
74
Issue
10
Publish Date
1996
Start Page
1095
End Page
1110

Dynamin and beta-arrestin reveal distinct mechanisms for G protein-coupled receptor internalization.

The process of agonist-promoted internalization (sequestration) of G protein-coupled receptors (GPCRs) is intimately linked to the regulation of GPCR responsiveness. Following agonist-mediated desensitization, sequestration of GPCR is presumably associated with the dephosphorylation and recycling of functional receptors. However, the exact mechanisms responsible for GPCR sequestration, even for the prototypic beta2-adrenergic receptor (beta2AR), have remained controversial. We demonstrate here that dynamin, a GTPase that regulates the formation and internalization of clathrin-coated vesicles, is essential for the agonist-promoted sequestration of the beta2AR, suggesting that the beta2AR internalizes via the clathrin-coated vesicle-mediated endocytic pathway. In contrast, internalization of the angiotensin II type 1A receptor (AT1AR), another typical GPCR, does not require dynamin. In addition, the AT1AR internalizes independent of the function of beta-arrestin, a critical component for beta2AR cellular trafficking, but additional AT1ARs are mobilized to the dynamin-dependent pathway upon overexpression of beta-arrestin. These findings demonstrate that GPCRs can utilize distinct endocytic pathways, distinguishable by dynamin and beta-arrestin, and that beta-arrestins function as adaptor proteins specifically targeting GPCRs for dynamin-dependent endocytosis via clathrin-coated vesicles.

Authors
Zhang, J; Ferguson, SS; Barak, LS; Ménard, L; Caron, MG
MLA Citation
Zhang, J, Ferguson, SS, Barak, LS, Ménard, L, and Caron, MG. "Dynamin and beta-arrestin reveal distinct mechanisms for G protein-coupled receptor internalization." J Biol Chem 271.31 (August 2, 1996): 18302-18305.
PMID
8702465
Source
pubmed
Published In
The Journal of biological chemistry
Volume
271
Issue
31
Publish Date
1996
Start Page
18302
End Page
18305

Chimeric D2/D3 dopamine receptors efficiently inhibit adenylyl cyclase in HEK 293 cells.

Despite a high degree of sequence homology, the dopamine D2 and D3 receptors have substantially different second messenger coupling properties. We have used chimeric D2/D3 receptors to investigate the contribution of the intracellular loops to the signaling properties of these receptors. In HEK 293 cells, D2 receptors inhibit prostaglandin E1-stimulated cyclic AMP levels by >90%, whereas D3 receptors inhibit cyclic AMP accumulation by only 20%. In chimeras that have the second or third intracellular loop, or both loops simultaneously, switched between the D2 and D3 receptors, the maximal inhibition of adenylyl cyclase is 60-90%. In addition, the potency of quinpirole to inhibit adenylyl cyclase activity at some of the chimeras is altered compared with the wild-type receptors. It appears that the intracellular loops of the D3 receptor are capable of interacting with G proteins, as when these loops are expressed in the D2 receptor, the chimeras inhibit adenylyl cyclase similarly to the wild-type D2 receptor. Our data suggest that the overall conformation of the D3 receptor may be such that it interacts with G proteins only weakly, but when the intracellular loops are expressed in another context or the D3 receptor structure is altered by the introduction of D2 receptor sequence, this constraint may be lifted.

Authors
Robinson, SW; Caron, MG
MLA Citation
Robinson, SW, and Caron, MG. "Chimeric D2/D3 dopamine receptors efficiently inhibit adenylyl cyclase in HEK 293 cells." J Neurochem 67.1 (July 1996): 212-219.
PMID
8666994
Source
pubmed
Published In
Journal of Neurochemistry
Volume
67
Issue
1
Publish Date
1996
Start Page
212
End Page
219

Ontogeny of the striatal neurons expressing the D1 dopamine receptor in humans.

We studied D1 dopamine receptor (D1R) gene expression in the human striatum during ontogeny by in situ hybridization, immunohistochemistry, and D1R ligand autoradiography. D1R mRNA, protein, and binding sites ([3H]SCH 23390) were detected in the striatum from week 12 of fetal life. At this time, D1R mRNA was predominant in the striosomal neurons; D1R immunoreactivity (D1R-IR) and D1R binding sites displayed a pattern similar to D1R mRNA. D1R-IR was essentially present in striosomal cell bodies and neuropil, whereas only a few cell bodies were detected in the matrix. From week 20 of fetal life, D1R gene expression developed in the matrix neurons as well, thus leading to an even D1R mRNA expression throughout striosomes and matrix compartments at birth. Comparative analysis of the expression of D1R and dynorphin mRNA show the same developmental patchy pattern up to week 26. Indeed, neurons expressing the D1R gene contain dynorphin mRNA; in contrast, they do not express the preproenkephalin A gene. At birth, the pattern of D1R mRNA expression level was sharply different from that of dynorphin (DYN) gene expression. High DYN mRNA expression was restricted to the striosomes, whereas high D1R mRNA expression was present in the whole striatum. These results demonstrate that, during human ontogeny, functional D1 receptors are expressed as early as week 12 in the striatum, developing initially in the striosomal neurons containing high dynorphin mRNA content. Toward the end of fetal life, there is a dissociation between D1R and DYN expression levels, suggesting that neuroanatomical or neurochemical modifications occur at this period, which may contribute to the regulation of the tone of the striatal D1R and DYN gene with topological specificity.

Authors
Brana, C; Caille, I; Pellevoisin, C; Charron, G; Aubert, I; Caron, MG; Carles, D; Vital, C; Bloch, B
MLA Citation
Brana, C, Caille, I, Pellevoisin, C, Charron, G, Aubert, I, Caron, MG, Carles, D, Vital, C, and Bloch, B. "Ontogeny of the striatal neurons expressing the D1 dopamine receptor in humans." J Comp Neurol 370.1 (June 17, 1996): 23-34.
PMID
8797154
Source
pubmed
Published In
The Journal of Comparative Neurology
Volume
370
Issue
1
Publish Date
1996
Start Page
23
End Page
34
DOI
10.1002/(SICI)1096-9861(19960617)370:1<23::AID-CNE3>3.0.CO;2-N

Catecholamine transporters and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine neurotoxicity: studies comparing the cloned human noradrenaline and human dopamine transporter.

The uptake and cytotoxicity of 1-methyl-4-phenylpyridinium (MPP+), the toxic metabolite of the parkinsonism inducing agent 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), were studied in COS-7 cells transiently transfected with the cloned human noradrenaline and dopamine transporters and in permanently transfected SK-N-MC neuroblastoma cells. MPP+ had a 10- to 20-fold lower K(m) value for the noradrenaline than for the dopamine transporter. In dopamine transporter expressing cells, the maximal transport rate (Vmax) of MPP+, dopamine and noradrenaline was the same, but in noradrenaline transporter expressing cells the Vmax of MPP+ and dopamine was only one-half of the Vmax of noradrenaline. The turnover numbers (Vmax of uptake/maximal binding sites of binding) were 5 times higher for the dopamine transporter (as measured with [3H]dopamine and [3H]-2 beta-carbomethoxy-3 beta-(4-fluorophenyl) tropane than for the noradrenaline transporter (as measured with [3H]noradrenaline and [3H]nisoxetine). In SK-N-MC cells with similar Vmax values for both catecholamines, noradrenaline transporter expressing cells were killed by lower concentrations of MPP+ in the medium than dopamine transporter expressing cells. Desipramine blocked the toxicity of MPP+ toward the noradrenaline transporter, but not the dopamine transporter expressing cells. We conclude that the toxic effect of MPTP at the striatal dopamine system in the MPTP primate model of Parkinson's disease is not correlated with the affinity profile of MPP+ for catecholamine transporters, but rather with the higher turnover number of MPP+ at the dopamine transporter. In contradistinction, the toxicity of MPTP at the noradrenaline neurons in the primate cerebral cortex (Pifl et al., 1991) may involve the higher affinity of MPP+ for the noradrenaline transporter.

Authors
Pifl, C; Hornykiewicz, O; Giros, B; Caron, MG
MLA Citation
Pifl, C, Hornykiewicz, O, Giros, B, and Caron, MG. "Catecholamine transporters and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine neurotoxicity: studies comparing the cloned human noradrenaline and human dopamine transporter." J Pharmacol Exp Ther 277.3 (June 1996): 1437-1443.
PMID
8667208
Source
pubmed
Published In
The Journal of pharmacology and experimental therapeutics
Volume
277
Issue
3
Publish Date
1996
Start Page
1437
End Page
1443

Expression of mRNA coding for the serotonin transporter in aged vs. young rat brain: differential effects of glucocorticoids.

Serotonin transporter mRNA expression in midbrain of young and aged rats was measured after long-term infusion of dexamethasone (0.01 and 0.05 mg/kg/day). Aging alone had no effect. Dexamethasone significantly decreased expression in both young and old rats but the effect was greater in the aged group. Adrenocortical dysregulation is common in elderly depression; our results suggest that glucocorticoids interact with aging to exacerbate abnormalities of serotonergic function, contributing to reduced antidepressant effectiveness.

Authors
Fumagalli, F; Jones, SR; Caron, MG; Seidler, FJ; Slotkin, TA
MLA Citation
Fumagalli, F, Jones, SR, Caron, MG, Seidler, FJ, and Slotkin, TA. "Expression of mRNA coding for the serotonin transporter in aged vs. young rat brain: differential effects of glucocorticoids." Brain Res 719.1-2 (May 6, 1996): 225-228.
PMID
8782886
Source
pubmed
Published In
Brain Research
Volume
719
Issue
1-2
Publish Date
1996
Start Page
225
End Page
228

Members of the G protein-coupled receptor kinase family that phosphorylate the beta2-adrenergic receptor facilitate sequestration.

We recently reported that a beta2-adrenergic receptor (beta2AR) mutant, Y326A, defective in its ability to sequester in response to agonist stimulation was a poor substrate for G protein-coupled receptor kinase (GRK)-mediated phosphorylation; however, its ability to be phosphorylated and sequestered could be restored by overexpressing GRK2 [Ferguson et al. (1995) J. Biol. Chem. 270, 24782]. In the present report, we tested the ability of each of the known GRKs (GRK1-6) to phosphorylate and rescue the sequestration of the Y326A mutant in HEK-293 cells. We demonstrate that in addition to GRK2, GRK3-6 can phosphorylate the Y326A mutant and rescue its sequestration; however, GRK1 was totally ineffective in rescuing either the phosphorylation or the sequestration of the mutant receptor. We found that the agonist-dependent rescue of Y326A mutant phosphorylation by GRK2, -3, and -5 was associated with the agonist-dependent rescue of sequestration. In contrast, overexpression of GRK4 and -6 led mainly to agonist-independent phosphorylation of the Y326A mutant accompanied by increased basal receptor sequestration. Our results demonstrate that phosphorylation per se, but not the interaction with a specific GRK, is required to facilitate beta2AR sequestration.

Authors
Ménard, L; Ferguson, SS; Barak, LS; Bertrand, L; Premont, RT; Colapietro, AM; Lefkowitz, RJ; Caron, MG
MLA Citation
Ménard, L, Ferguson, SS, Barak, LS, Bertrand, L, Premont, RT, Colapietro, AM, Lefkowitz, RJ, and Caron, MG. "Members of the G protein-coupled receptor kinase family that phosphorylate the beta2-adrenergic receptor facilitate sequestration." Biochemistry 35.13 (April 2, 1996): 4155-4160.
PMID
8672451
Source
pubmed
Published In
Biochemistry
Volume
35
Issue
13
Publish Date
1996
Start Page
4155
End Page
4160
DOI
10.1021/bi952961+

Differential regulation of dopamine D1A receptor responsiveness by various G protein-coupled receptor kinases.