DOPAMINE D2-RECEPTOR MEDIATED SIGNAL TRANSDUCTION

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The overall goal of this project is to explore signal transduction mechanisms for dopamine acting through D2-receptors, in the belief that this may elucidate the role played by these receptors in regulating neuronal activity. Consequently, the present proposal takes two approaches - first, to establish a model cell system in which dopamine's role in modulating the intracellular second messengers, cAMP and [Ca2+]i can be delineated and secondly, to investigate the electrophysiological consequences of a biochemical perturbation of D2-receptor mediated signal transduction in striatum. The model system to be evaluated will be a neuroblastoma x Chinese hamster embryonic brain explant hybrid cell line (NCB-20), which expresses receptors for dopamine, serotonin, bradykinin, opiates and acetylcholine. The first group of experiments will determine whether D2-receptors inhibit adenylate cyclase in NCB-20 cells and whether these receptors also may perturb [Ca2+]i mobilization in these cells, by measuring D2-receptor effects on phosphatidylinositol metabolism and [Ca2+]i mobilization. In the second group of experiments, conditions will be optimized for the intrastriatal injection of pertussis toxin, so that a significant portion of the pertussis toxin substrates are ADP-ribosylated; a determination will be made on the effectiveness with which this treatment eliminates receptor-mediated inhibition of adenylate cyclase; subsequently, receptor-mediated alterations in electrophysiological properties will be examined in these animals compared to sham-injected controls.