Much evidence implicates the involvement of the serotonin 5-HT1A receptor in anxiety and depression. 5-HT1A receptor agonists display anxiolytic/antidepressant effects, and 5-HT1A receptor function is altered after repeated treatment with anxiolytic and antidepressant drugs. While much data has accumulated about 5- HT1A receptor signal transduction in cells genetically engineered to express the receptor, much less is known about the receptor in its native environment. The potential importance of regional differences in 5-HT1A receptor function is underscored by our previous studies demonstrating regional variations in receptor/effector coupling efficiency and recent evidence that chronic treatment with anxiolytic/antidepressant drugs elicits regionally specific 5-HT1A receptor adaptation. This project therefore proposes to gain a better understanding of 5-HT1A receptor signal transduction, especially with regard to regional differences in receptor/G protein coupling, G protein specificity, and 5-HT1A receptor adaptation after repeated treatment with various classes of anxiolytic/antidepressant drugs. The following Specific Aims are proposed: 1) To determine if previously described regional differences in receptor/effector coupling efficiency are demonstrable at the level of receptor/G protein coupling. Dose-response curves for agonist-stimulated binding of [35S]GTPgammaS (in membranes and brain sections) before and after partial irreversible receptor blockade will be generated to test the hypothesis that 5-HT1A receptor activation of G protein coupling will exhibit greater efficiency at somatodendritic dorsal raphe (DR) autoreceptors than at postsynaptic receptors in hippo-campus. Coupling efficiency at postsynaptic receptors in the lateral septum (LS) and medial prefrontal cortex (mPFC) will also be determined; 2) To determine if 5-HT1A receptors couple preferentially to specific G proteins in different brain regions. Selective knockdown of pertussis toxin (PTX)-sensitive G protein alpha subunits by intracerebroventricular or intra-tissue infusion of antisense oligodeoxynucleotides (AS ODNs) will test the hypothesis that 5- HT1A receptor coupling to G proteins will be region- and agonist- specific. We predict that in the DR, 5-HT1A receptors will couple preferentially to Galphai3 to mediate agonist-induced inhibition of 5-HT synthesis; in the hippocampus, 5-HT1A receptors will couple preferentially to Galphai2 to mediate inhibition of forskolin-stimulated adenylyl cyclase. We will also test the hypothesis that some agonists may preferentially couple to different Galphai/Galphao subunits to mediate these effects; 3) To determine if repeated treatment with different classes of anxiolytic/antidepressant drugs results in regionally- specific 5-HT1A receptor adaptation and changes in receptor/G protein coupling. Agonist inhibition of 5-HT synthesis in forebrain regions will be assessed to test the hypothesis that repeated treatment (21 days) with ipsapirone, paroxetine or clorgyline (but not imipramine) will desensitize somatodendritic 5-HT1A autoreceptors and reduce agonist-stimulated binding of [35S]GTPgammaS in the DR. We will also test the hypothesis that in the hippocampus, agonist-stimulated binding of [35S]GTPgammaS will be increased by repeated treatment with imipramine, decreased by clorgyline, but remain unchanged after ipsapirone or paroxetine.
