Many modern models of receptor-G protein function assume that there is a direct relationship between high-affinity agonist binding and efficacy. The validity of this assumption has been recently questioned for the serotonin 5-HT2, receptor. We examined the intrinsic activities of various ligands in activating phosphoinositide hydrolysis and measured their respective binding affinities to the high-and low-affinity states of the 5-HTzC (VNV isoform) and 5-HT2, receptors. Ligand binding affinities for the high-affinity state of the receptors were determined using 1 -(4-[1251]iodo-2,5-dimethoxyphenyl)-2-aminopropane, whereas rH]mesulergine and N-rH]-methylspiperone were used, in the presence of excess guanine nucleotide [guanosine 5'-0-(3-thiotriphosphate)], to define binding to the low-affinity state of the 5-HTzc and 5-HT2, receptors, respectively. Antagonists labeled the high-and low-affinity states of each receptor with comparable affinities. Previously identified inverse agonists of the 5-HT2, receptor behaved as silent antagonists in our systems even when the receptor was overexpressed at a relatively high density. In contrast, the ability of agonists to bind differentially to the high-and low-affinity states of the 5-HT2, and 5-HT2, receptors was highly correlated (r2 = 0.86 and 0.96, respectively) with their intrinsic activities. These data suggest that high-affinity agonist states can account for agonist efficacy at human 5-HTz, or 5-HTz, receptors without the need for considering additional transition or active states of the receptor-ligand complex. The procedure described herein may expedite drug discovery efforts by predicting intrinsic activities of ligands solely from ligand binding assays.