Immunochemical Studies of the Muscarinic Acetylcholine Receptor

Abstract

Muscerinic receptors have been purified from calf forebrain plasma cell membranes by affinity chromatography on a dexetimide-agarose gel. SDS-PAGE analysis showed a single 70kDa band. Monoclonal antibodies have been prepared against these affinity purified 70kDa protein(s). One antibody, M-35, Immunoprecipitated up to 80% of digitonin-solubilized muscarinic receptors. M-35 had agonist-like effects on guinea-pig myometrium: it increased the intracellular cyclic GMP content, decreased prostaglandin-induced cyclic AMP accumulation and caused muscle contractions. The two first affects were inhibited by atropine. M-35 was used to visualize muscarinic receptors at the surface of human fibroblastic cells. In the particular cell line used, the receptors have a low affinty for pirenzepine, were negatively coupled to adenylate cyclase and mediated increase in the phosphatidyl-insitol breakdown.