Clozapine interaction with the M₂ and M₄ subtypes of muscarinic receptors

Available evidence indicates that the antipsychotic drug clozapine acts as a partial agonist at the muscarinic M₄ and as an antagonist at the M₂ receptors. We wondered whether there is indeed a fundamental difference between its action on these two receptor subtypes, and whether it interacts with their classical or allosteric binding sites. In experiments on Chinese hamster ovary cells stably expressing the M₂ or M₄ receptors, clozapine inhibited the binding of the specific muscarinic ligand [³H]N-methylscopolamine to either receptor subtype. The affinity of the high-affinity sites for clozapine was diminished by GTP in the way expected for agonists on both the M₂ and the M₄ receptor subtypes. Arunlakshana-Schild plots of data obtained in saturation binding experiments with [³H]N-methylscopolamine at different concentrations of clozapine were linear with a slope of unity. Clozapine did not alter the time course of [³H]N-methylscopolamine dissociation from muscarinic M₂ or M₄ receptors. It inhibited the synthesis of cyclic AMP in cells expressing the M₄ receptor subtype, but did not measurably inhibit the synthesis of cyclic AMP in cells expressing the M₂ receptor subtype. We conclude that clozapine has a high affinity for muscarinic M₂ and M₄ receptor subtypes, that it associates with the classical and not with the allosteric binding site, and that it acts as a partial agonist on both the M₂ and the M₄ receptor subtype. Copyright (C) 1999 Elsevier Science B.V.