Search PubMed⌕ Search

Biomedical subjects

M G Caron

Publications and source records attributed to M G Caron.

At least 55 records · Page 3Linked to original sources
PubMed2000

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.

Analgesia↗
PubMed2000

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.

Animals↗
PubMed2000

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.

Animals↗
PubMed1999

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.

Analgesia↗
PubMed1999

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.

Amino Acid Sequence↗
PubMed1999

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.

Animals↗
PubMed1999

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.

Arrestins↗
PubMed1999

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.

Animals↗
PubMed1999

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.

Animals↗
PubMed1999

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.

Aggression↗
PubMed1999

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.

Acoustic Stimulation↗
PubMed1999

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.

Arrestins↗
PubMed1999

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.

Animals↗
PubMed1999

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.

Adaptor Protein Complex alpha Subunits↗
PubMed1999

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.

Arrestins↗
PubMed1999

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.

Adrenergic beta-Agonists↗
PubMed1999

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.

Animals↗