Mechanisms and sites of action of endothelins 1 and 2 on the opossum internal anal sphincter smooth muscle tone in vitro

Endothelins, localized in the enteric nervous system, may play important ro les in the morphogenesis of the gastrointestinal (GI) tract and in the regu lation of GI motility. However, the role of endothelins in the GI sphincter s, including the internal anal sphincter (IAS) have not been examined. We e xamined the actions of endothelins on the basal tone of the opossum IAS cir cular smooth muscle strips before and after different neurohumoral antagoni sts or inhibitors. Endothelins 1 and 2 produced a concentration-dependent b iphasic effect on the basal tone of the IAS, an initial brief fall followed by a sustained rise. The fall in the IAS smooth muscle tone was not modifi ed by atropine, guanethidine, or tetrodotoxin but was significantly attenua ted by the nitric oxide synthase inhibitor L-NNA, the specific neuronal nit ric oxide synthase inhibitor, 1-(2-trifluoromethylphenyl)imidazole, the N-t ype neuronal Ca++-channel blocker omega-conotoxin GVIA, and by the calmodul in antagonist W-13. Endothelin-induced contraction of the IAS, on the other hand, was not affected by any of the neurohumoral antagonists but was sign ificantly inhibited by the selective protein kinase C inhibitor H-7 or the calmodulin inhibitor W-13. The combination of H-7 and W-13 had no additive effect in attenuating the contractile action of endothelin 1. There was cle ar evidence of a cross-tachyphylaxis to the actions of endothelin 1 and end othelin 2. We conclude that the endothelins exert important neuromodulatory effects on the basal tone of the IAS. The contractile action occurs direct ly at the smooth muscle and the relaxant action by the activation of neuron al nitric oxide synthase at the nerve terminals. The contraction and relaxa tion of the smooth muscle caused by endothelins 1 and 2 may involve distinc t receptors that are similar for both endothelins, The excitatory actions o f endothelin 1 involve both the protein kinase C and the Ca++-calmodulin pa thways that may lie in series.