Native rat hippocampal 5-HT1A receptors show constitutive activity

Previous studies have shown that human 5-hydroxytryptamine (5-HT)(1A) receptors stably expressed in transfected cell lines show constitutive G-protein activity, as revealed by the inhibitory effect of inverse agonists, such as spiperone, on basal guanosine- 5'-O-(3-[S-35]thio)-triphosphate ([S-35]GTP gamma S) binding. In the present study, we evaluated the constitutive activity of native rat 5-HT 1A receptors in hippocampal membranes. Using anti-G alpha(o)-antibody capture coupled to scintillation proximity assay under low sodium (30 mM) conditions, we observed high basal [S-35]GTP gamma S binding to G alpha(o) subunits (defined as 100%). Under these conditions, 5-HT and the prototypic selective 5-HT1A agonist (+) 8-hydroxy-2-(di-n-propylamino) tetralin [(+)-8-OH-DPAT] both stimulated [S-35]GTP gamma S binding to G alpha(o) to a similar extent, raising binding to approximately 130% of basal with pEC(50) values of 7.91 and 7.87, respectively. The 5-HT1A-selective neutral antagonist [O-methyl-3H]-N-(2-(4-(2-methoxyphenyl)-pheynl)-1-piperazinyl)ethyl)-N-(2-pyridinyl)cyclohexanecarboxamide trihydrochloride (WAY100,635) could block these effects in a competitive manner with pK(b) values (5-HT, 9.57; (+)-8-OHDPAT, 9.52) that are consistent with its pK(i) value at r5-HT1A receptors (9.33). In this native receptor system, spiperone and methiothepin reduced basal [S-35] GTP gamma S binding to G alpha(o) in a concentration-dependent manner to 90% of basal with pIC(50) values of 7.37 and 7.98, respectively. The inhibition of basal [S-35]GTP gamma S binding induced by maximally effective concentrations of spiperone (10 mu M) or methiothepin (1 mu M) was antagonized by WAY100,635 in a concentration-dependent manner (pK(b), 9.52 and 8.87, respectively), thus indicating that this inverse agonism was mediated by 5-HT1A receptors. These data provide the first demonstration that native rat serotonin 5-HT1A receptors can exhibit constitutive activity in vitro.