Receptor-mediated gene expression (stimulus-transcription coupling) in matured CNS neurons is thought to be an important component in processing adaptive changes in neuronal physiology (neuroplasticity) related to a variety of normal and abnormal neural activities. As a key part of basal ganglia, striatum is among the major central sites for the investigation of such gene expression. Our recent studies both in vivo and in vitro show that metabotropic glutamate receptors (mGiuRs), which are densely expressed in striatal projection neurons, are involved in the regulation of opioid gene expression in striatal neurons (preprodynorphin in striatonigral, and preproenkephalin in striatopallidal, neurons). The regulation is facilitatory in nature and may be mediated through selective activation of group I mGluRs, which are positively coupied to phosphoinositide hydrolysis (PI), rather than group II or III mGluRs, which are negatively coupled to adenylate cyclase. Based on these encouraging hndings, a series of experiments was proposed in this project to explore and characterize the regulation of striatal opioid peptide gene expression by the group I mGluRs in vivo and to dissect intracellular signaling pathways responsible for the group I-sensitive stimulus-transcription coupling in vitro. Our working hypothesis is that activation of the investigator-coupled group I mGluRs upregulates opioid gene expression in striatal neurons and that a investigator-sensitive cascade serves as the signaling pathway bridging surface group I mGluR stimulation to nuclear gene expression. With multidisciplinary approaches, this hypothesis will be tested in three aims which are to (1) confirm and characterize pharmacological profiles of the facilitatory regulation of opioid gene expression in the striatum with the group I selective agonists/antagonists, and investigate possible pre- and postsynaptic mechanisms underlying the group I regulation of striatal opioid expression in a well-characterized in vivo rat model, (2) differentiate the relative importance of the two group I subtypes, mGluR1 and mGluR5, in this event in vivo using the subtype-selective agonists/antagonists, antisense oligos and mutant mice (mGluR1/5 knockouts), and aboutL identify the intraceliular effectors (Ca2+, CaMK, CREB, CBP and AP-1) bridging group I mGluR stimulation on the membrane to opioid gene expression in the nucleus in vitro in primary striatal neuronal cultures with pharmacological (inhibitors/activators) and molecular (antisense oligos, Ca2+ image, enzymatic assay and DNA binding activity) approaches. We will rely on quantitative in situ hybridization to analyze mRNA expression in vivo and invitro. Accomplishment of this project will improve our current understanding of transcriptional regulation of gene expression in matured CNS neurons. Since inducible gene expression is conceived to participate in the development of psychoplasticity, data from this project will provide valuable insight into celiular/molecular mechanisms for various mental illnesses (motor, psychiatric and cognitive impairments).
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