This proposal seeks to develop and characterize a functional assay for measuring GABA responses in primary spinal cord cultured neurons. GABA is known to produce its responses by increased conductance to C1- ions. A detailed study to characterize GABA-mediated 36C1-influx in cultured neurons and the effect of various modulators, like barbiturates, benzodiazepines and pyrazolopyridines on GABA-induced 36C1-influx will be examined. These classes of drugs possess a similar pharmacological profile and are known to facilitate GABAergic transmission. Further, we will grow the cultures in the presence of GABA agonists and selected modulators to determine desensitization or supersensitivity. Group selective modification studies will be aimed to determine which of the GABAA receptors is functionally relevant. All the studies utilizing GABA functional assay will be compared with the published electrophysiological data and the published binding data and, finally, with our own binding data in the intact cultures. The overall objective is to study the function and binding in the same preparation and using physiological conditions. These studies are aimed at understanding GABAergic transmission and the possible mechanisms by which diverse classes of drugs modulate CNS function mediated by the inhibitory neurotransmitter GABA.
