G protein coupling to M1 and M3 muscarinic receptors in sublingual glands

Rat sublingual gland M-1 and M-3 muscarinic receptors each directly activate exocrine secretion. To investigate the functional role of coreceptor expression, we determined receptor-G protein coupling. Although membrane proteins of 40 and 41 kDa are ADP-ribosylated by pertussis toxin (PTX), and 44 kDa proteins by cholera toxin (CTX), both carbachol-stimulated high-affinity GTPase activity and the GTP-induced shift in agonist binding are insensitive to CTX or PTX. Carbachol enhances photoaffinity labeling ([alpha P-32] GTP-azidoaniline) of only 42-kDa proteins that are subsequently tractable to immunoprecipitation by antibodies specific for G alpha (q) or G alpha (11) but not G alpha (12) or G alpha (13). Carbachol-stimulated photoaffinity labeling as well as phosphatidylinositol 4,5-bisphosphate (PIP2) hydrolysis is reduced 55% and 60%, respectively, by M-1 receptor blockade with m1-toxin. G alpha (q/11)-specific antibody blocks carbachol-stimulated PIP2 hydrolysis. We also provide estimates of the molar ratios of receptors to G alpha (q) and G alpha (11). Although simultaneous activation of M-1 and M-3 receptors is required for a maximal response, both receptor subtypes are coupled to G alpha (q) and G alpha (11) to stimulate exocrine secretion via redundant mechanisms.