While the molecular locus of antidepressant and anti-bipolar drug action has not yet been established, it has become increasingly likely that the targets of such drugs lie distal to neuro- transmitter receptors. These targets may involve G protein-mediated signal transduction systems such as adenylyl cyclase and phospholipase C. For this study, both rats and cultured cells will be treated chronically with a varity of antidepressants (amitriptyline, iprindole, fluoxetine, phenylzine and chlorpromazine as a control). Previous studies showed an increased association between a G protein (Gs) and adenylyl cyclase. Cultured Cg glioma cells treated with antidepressant drugs also showed increased coupling between Gs and adenylyl cyclase. This allowed the conclustion that the """"""""presynaptic"""""""" component is not required for antidepressant effects. These studies also showed a dissociation between receptor desensitization and increased Gs/adenylyl cyclase coupling. Although increased Gs-adenylyl cyclase coupling consistently results from antidepressant treatment, neither Gs nor adenylyl cyclase appears to be the actual target for antidepressant action. It is hypothesized that the effect of antidepressant treatment on increasing the coupling between Gs and adenylyl cyclase may be due to a perturbation of the membrane which alters the relationship between GTs and the cytoskeleton (specifically tubulin). This will be investigated by subjecting the membrane to differential extraction with detergents as well as to an electron microscopic examination of the effect of antidepressant treatment on the synaptic distribution of Gs and tubulin as well as their co-localization. Functional G protein will be compared to the absolute content with the photoaffinity GTP analog, azidoanilido GTP (AAGTP). Subtype specificity of adenylyl cyclase relative to antidepressant treatment will also be examined in transfected cells. A """"""""dominant negative"""""""" chimera of Gi alpha and tranducin, which blocks normal tubulin-Gs interaction has been developed. Expression of this construct, concommitant with antidepressant treatment will help to verify the importance of tubulin-Gs interaction for the observed antidepressant response. It is hoped that successful completion of these studies will allow a greater realization of how a number of diverse drugs all exert antidepressant effects. Such knowledge should assist in the design of more efficient antidepressant drug therapy and may lead to a true understanding of the molecular basis of mood.
