AGONIST AND INVERSE AGONIST ACTIVITY AT THE DOPAMINE D-3 RECEPTOR MEASURED BY GUANOSINE 5'-[GAMMA-THIO]TRIPHOSPHATE-[S-35] BINDING

In this study, the ligand-receptor-G protein interactions of the dopam ine D-3 receptor expressed in Chinese hamster ovary cells were investi gated using guanosine 5'-[gamma-thio]triphosphate-[S-35] (S-35]GTP gam ma S) and receptor binding experiments. Dopamine stimulated the [S-35] GTP gamma S binding in a guanine nucleotide, magnesium and sodium-depe ndent manner. Dopamine and quinpirole produced maximal stimulation of the [S-35]GTP gamma S binding whereas (+)-7-OH-DPAT and (-)-3-PPP were partial agonists. Interestingly, several compounds previously classif ied as D-2 receptor antagonists behaved as inverse agonists at the D-3 receptor, i.e., they inhibited the basal [S-35]GTP gamma S binding in a dose dependent fashion. Haloperidol, (+)-UH-232, (+)-AJ-76 and racl opride were full inverse agonists but clozapine was a partial inverse agonist. Pertussis toxin treatment abolished the D-3 receptor-mediated agonist as well as inverse agonist responses, indicating the involvem ent of G(i)/G(o) proteins in both processes. According to the ternary complex model, agonists should bind with higher affinity to the G prot ein coupled receptor (RG) and thereby shift the equilibrium from free receptor (R) toward RG, which produces a biological response. However, an inverse agonist should bind with higher affinity to R than to RG a nd thereby inhibit the basal activity of the cell. We found that the h igh affinity agonist binding site (RG) was abolished by pertussis toxi n treatment of the cells. However, the inverse agonists bound with the same affinity to untreated and pertussis toxin treated D-3 receptor m embranes. Thus, we found no evidence for the hypothesis that inverse a gonists would shift the equilibrium from RG toward R by binding with h igher affinity to R than to RG.