Inactivation of the beta-adrenergic receptor in skeletal muscle by dithiols.

The effect of sulfhydryl compounds on binding of the beta-adrenergic antagonist (-)-[3H]dihydroalprenolol [(-)-[3H]DHA] to a microsomal fraction from rabbit skeletal muscle was examined. Inhibition of binding by a variety of adrenergic agonists and antagonists and the effects of these agents on adenylate cyclase were consistent with the beta-adrenergic receptor in this tissue being of the beta 2-subtype. Binding of (-)-[3H]DHA was reduced by incubating the membranes with dithiols such as dithiothreitol (DTT), 1,3-dimercapto-2-propanol and 1,4-dimercaptobutane; monothiols were much less potent. DTT-induced decline in (-)-[3H]DHA binding resulted primarily from a decrease in receptor number. Inactivation was partially reversed by the oxidant H2O2. Binding sites could be locked in the inactivated state by incubating DTT-treated membranes with the alkylating agent iodoacetamide. Both beta-adrenergic agonists and antagonists protected against inactivation. Adenylate cyclase activity in the membranes was increased by DTT. The enzyme was rapidly inactivated by H2O2, and this could be partially reversed by DTT. It is concluded that the beta-adrenergic receptor of skeletal muscle contains an essential disulfide moiety which can be inactivated by reducing dithiols. Adenylate cyclase, on the other hand, contains at least one essential sulfhydryl which is preserved by dithiols.