G protein-linked receptors labeled by [3H]histamine in guinea pig cerebral cortex. II. Mechanistic basis for multiple states of affinity [corrected].

Tritiated histamine labels multiple states of guanine nucleotide-binding protein-linked receptors in washed membranes from guinea pig cerebral cortex. The effects of guanylylimidodiphosphate identify the radioligand as an agonist, but the Hill coefficients can be as low as 0.78 for H2 and H3 agonists and 0.66 for antagonists. To examine the mechanistic basis of the binding patterns, the inhibitory behavior of 14 histaminergic ligands has been measured at 1.4 nM and 11 nM [3H]histamine. In such experiments, the radioligand is both a second independent variable and an internal control; it is therefore possible to differentiate among mechanistic schemes for binding at equilibrium. When the data are analyzed in terms of distinct and independent sites, the relative capacities and the inferred affinities of [3H]histamine differ significantly from ligand to ligand. Because the discrepancies persist for any degree of heterogeneity, the model can be rejected unequivocally. Several lines of evidence argue against the notion of a ligand-regulated equilibrium between two states of mutually independent sites. In particular, antagonists reveal a paradoxical arrangement in which the ligand appears to increase the relative number of sites in the state to which it binds more weakly; also, the Hill coefficients estimated at 1.4 nM [3H]histamine are 1.54 for the H2 agonist pyridylethylamine and 1.26 for the antagonist SK&F 93479 [Mol. Pharmacol. 43: 569-582 (1993)]. High values of nH suggest that one equivalent of bound ligand affects the affinity of the next, and a model based on cooperative interactions can predict most of the effects that are anomalous in the context of other schemes. H2 agonists can be distinguished empirically from antagonists on the basis of their inhibitory behavior at two concentrations of [3H]histamine. The trend includes two compounds that are H2 agonists but H3 antagonists, and the labeled sites therefore reveal an element of H2 specificity.