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 receptors. In order to characterize dopamine 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 cDNAs/genes and investigation of receptor structure and regulation in normal and pathophysiological states. (1) Cell Biology and Regulation of Dopamine Receptors. Characterization of the functional and regulatory properties of D-1 and D-2 dopamine receptors in various cDNA-transfected cell lines was continued. The role of cAMP in agonist-induced regulation of D-1 and D-2 receptors is being investigated using cell lines deficient in the cAMP-dependent protein kinase as well as using site-directed mutagenesis techniques to alter potential phosphorylation sites in the receptor proteins. Chimeric D- 1/D-2 receptors have been constructed to further investigate regulatory mechanisms of these receptors. D-2, D-3 and D-4 receptors were transfected into the mesencephalic cell line, MES 23.5. D-2 and D-3 receptors were found to increase K+ currents whereas D-4 receptors decreased this response. Agonist-induced desensitization of the 5-HT-6 serotonin receptor was characterized. Interestingly, this receptor exhibited a desensitization response in the absence of down-regulation. (2) Molecular Biology of Dopamine Receptors. Work continued on cloning of a third """"""""D-1 like"""""""" receptor which apparently is linked to the stimulation of phosphatidylinositol turnover and mobilization of calcium. Transgenic """"""""knock-out"""""""" mice lacking a functional D-1 receptor were further characterized and transgenic mice lacking the D-3 receptors were produced. Other transgenic mice lacking the D-5 receptors are in production. Chimeric D-2/D-3 and D-2/D-4 dopamine receptors were constructed and expressed for characterization. These results indicated that multiple domains in the D-2 receptor are important for coupling to adenylyl cyclase and that transmembrane region III is an important structural determinant for D-4 pharmacology. The D-4 receptor pharmacology was further found not to be predictive of atypical antipsychotic efficacy.
Showing the most recent 10 out of 32 publications
