Protein kinase C mediates spontaneous tone in the cat lower esophageal sphincter

The intracellular pathways responsible for maintenance of tone in the lower esophageal sphincter (LES) are not well understood. We show that the protein kinase C (PKC) antagonists (1-(5-isoquinolinesulphonyl)-2- methylpiperazine dihydrochloride) and calphostin C reduce spontaneous resting tone in LES muscle strips, whereas the calmodulin antagonist N-(6-aminohexyl- 5-chloro-1-naphthalenesulfonamide hydrochloride) has no effect, which suggests that LES tone is maintained by a PKC-mediated mechanism. In addition, U73122, an inhibitor of phosphatidylinositol-4,5-bisphosphate (PIP₂)-specific phosphotipase C, and D609, an inhibitor of phosphatidycholine-specific phospholipase C, reduced diacylglycerol formation and LES tone in a concentration-dependent manner. Finally, diacylglycerol levels and PKC activity were reduced during relaxation of the LES induced by the inhibitory neurotransmitter vasoactive intestinal peptide. These data suggest that resting LES tone is associated with elevated diacylglycerol levels and PKC activity, which are reduced during relaxation. Diacylglycerol is derived from at least two different sources. Hydrolysis of PIP₂ by PIP₂- specific phospholipase C produces equimolar amounts of inositol 1,4,5- triphosphate and diacylglycerol, which may interact synergistically to activate PKC and develop tone. Furthermore, PKC-mediated contraction may be augmented by additional diacylglycerol production arising from the hydrolysis of phosphatidylcholine by phosphatidylcholine-specific phospholipase C.