Intact Cell Binding for In Vitro Prediction of Sedative and Non-sedative Histamine H₁–Receptor Antagonists Based on Receptor Internalization

Abstract

We evaluated changes in the binding properties of sedative and non-sedative histamine H₁–receptor antagonists induced by internalization of H₁ receptors in intact human U373 MG astrocytoma cells. Internalization of H₁ receptors was induced without their degradation by treatment with 0.1 mM histamine for 30 min at 37°C, and then the intact cell binding assay was performed at 4°C. The binding properties of [³H]mepyramine, a cell-penetrating radioligand for H₁ receptors, were not changed by histamine pretreatment. Displacement curves for sedative H₁-receptor antagonists (diphenhydramine, chlorpheniramine, promethazine, ketotifen, azelastine and oxatomide) against [³H]mepyramine binding were not changed by histamine pretreatment. In contrast, the displacement curves for non-sedative H₁-receptor antagonists (mequitazine, bepotastine, olopatadine, epinastine, carebastine, desloratadine and fexofenadine) were changed by histamine pretreatment: two types of changes, i.e. a rightward shift in the monophasic curve or an increase in the proportion of the low affinity component of the biphasic curve, were prevented under hypertonic conditions, in which clathrin-mediated receptor internalization is known to be inhibited. Thus, internalization-mediated changes in the binding properties of H₁-receptor antagonists were well correlated with their sedative and non-sedative behaviors, which might confirm their permeability through the biomembrane and possibly the blood brain barrier.