The long-term goal of this project is the characterization of neurotransmitter receptor-mediated information transduction, and its regulation, across neuronal membranes. The primary, but not exclusive, model systems under investigation are those for dopamine (DA) receptors. In order to characterize DA and related receptors at the biochemical and molecular levels and study their regulation, there are two major interrelated lines of research which are ongoing: 1) investigation of the cell biology, function and regulation of the receptors at the protein level; and 2) the molecular cloning of the receptor genes and investigation of gene structure and regulation in normal and pathophysiologic states. 1. Cell Biology and Regulation of DA Receptors. Characterization of the functional and regulatory properties of D1 and D2 DA receptors on various neuroblastoma and cDNA-transfected cell lines were continued. The D1 receptors were shown to undergo an agonist-induced form of desensitization which is partially cAMP-mediated and involves both functional uncoupling and down-regulation of the receptors. D2 receptors were also shown to undergo desensitization in response to agonist treatment but, in contrast to D1 receptors, demonstrate an up-regulation response. A variety of anti-peptide antibody probes for DA receptors were developed and used to map the expression of the D1, D2 and D3 receptor proteins at the cellular level in the CNS. 2. Molecular Cloning of DA and Other Receptors. A cDNA encoding a second D1 receptor (designated D1B) linked to adenylyl cyclase in rat kidney was identified and cloned. The distribution of the D1A and D1B receptors were mapped in the kidney. Work continued on the cloning of a third """"""""D1 like"""""""" receptor which apparently is linked to the stimulation of phosphatidyinositol turnover and calcium mobilization. The genes for the D1A, D1B, and D2 receptors were isolated. Transgenic """"""""knock-out"""""""" experiments for several of the dopamine receptor subtypes were initiated. Two completely novel serotonin receptors were cloned and expressed. Several other cDNA clones encoding putative """"""""orphan"""""""" G protein-linked neurotransmitter receptors were identified.
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