Coupling of M-2 muscarinic receptors to ERK MAP kinases and caldesmon phosphorylation in colonic smooth muscle

Coupling of M-2 and M-3 muscarinic receptors to activation of mitogen-activ ated protein (MAP) kinases and phosphorylation of caldesmon was studied in canine colonic smooth muscle strips in which M-3 receptors were selectively inactivated by N,N-dimethyl-4-piperidinyl diphenylacetate (4-DAMP) mustard (40 nM). ACh elicited activation of extracellular signal-regulated kinase (ERK) 1, ERK2, and p38 MAP kinases in control muscles and increased phospho rylation of caldesmon (Ser(789)), a putative downstream target of MAP kinas es. Alkylation of M-3 receptors with 4-DAMP had only a modest inhibitory ef fect on ERK activation, p38 MAP kinase activation, and caldesmon phosphoryl ation. Subsequent treatment with 1 mu M AF-DX 116 completely prevented acti vation of ERK and p38 MAP kinase and prevented caldesmon phosphorylation. C aldesmon phosphorylation was blocked by the MAP kinase/ERK kinase inhibitor PD-98509 but not by the p38 MAP kinase inhibitor SB-203580. These results indicate that colonic smooth muscle M-2 receptors are coupled to ERK and p3 8 MAP kinases. Activation of ERK, but not p38 MAP kinases, results in phosp horylation of caldesmon in vivo, which is a novel function for M-2 receptor activation in smooth muscle.