Histamine H 3 receptor binding sites in rat brain membranes: modulations by guanine nucleotides and divalent cations

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The binding of [ 3H](R)α-methylhistamine, a potent and specific agonist at histamine H 3 receptors, was investigated with membranes of rat cerebral cortex. In phosphate buffer the specific binding defined with thioperamide, an H 3 receptor antagonist, displayed characters of reversibility and saturability with a B max of ∼30 fmol/mg protein. The K D, derived from either dissociation/association rates or saturation kinetics at equilibrium, was ∼0.5 nM at 25°C. Competition studies indicated that the binding occurred with a stereoselectivity and pharmacological specificity similar to that of functional H 3 autoreceptors regulating histamine release in brain slices. However, whereas the potency of antagonists was closely similar in the two assay systems, that of agonists was ∼ 10-fold higher in the binding assay. Among antagonists burimamide was the only one to compete with a pseudo-Hill coefficient significantly different from unity (n H = 0.48 ± 0.03), indicating a possible heterogeneity among binding sites. The presence of 2.6 mM Ca 2+, in a modified Krebs-Ringer medium, promoted the conversion of a large fraction of sites into a low-affinity component with a K D of 16 nM. The presence of guanylnucleotides in the Krebs-Ringer medium with Ca 2+ abolished the binding to this low-affinity component whereas in a phosphate buffer only the K D was slightly increased. It is concluded that the H 3 receptor, like many other amine receptors, is coupled to its still unidentified effector system via a G-protein and regulated by Ca 2+. However, unlike the latter, the H 3 receptor is down-regulated by the divalent cation.