Inhibition of ACh-stimulated exocytosis by NSAIDs in guinea pig antral mucous cells: autocrine regulation of mucin secretion by PGE2

Abstract

The effects of indomethacin (IDM) and aspirin (ASA) on ACh (10 μM) -stimulated exocytotic events were studied in guinea pig antral mucous cells by using video optical microscopy. IDM or ASA, which inhibits cyclooxygenase (COX), decreased the frequency of ACh-stimulated exocytotic events by 30% or 60%, respectively. The extent of inhibition induced by ASA (60%) decreased by 30% when IDM or arachidonic acid (AA, the substrate of COX) was added. IDM, unlike ASA, appears to induce the accumulation of AA, which enhances the frequency of ACh-stimulated exocytotic events in ASA-treated cells. ONO-8713 (100 μM; an inhibitor of the EP1–EP4 prostaglandin receptors) and N-[2-(( p-bromocinnamyl)amino)ethyl]-5-isoquinolinesulfonamide, HCl (H-89, 20 μM; an inhibitor of PKA) also decreased the frequency of ACh-stimulated exocytotic events by 60%. However, the supplementation of PGE2 (1 μM) prevented the IDM-induced decrease in the frequency of ACh-stimulated exocytotic events. SC-560 (an inhibitor of COX-1) decreased the frequency of ACh-stimulated exocytotic events by 30%, but NS-398 (an inhibitor of COX-2) did not. Moreover, IDM decreased the frequency of exocytotic events stimulated by ionomycin, suggesting that COX-1 activity is stimulated by an increase in intracellular Ca2+ concentration ([Ca2+]i). ACh and ionomycin increased PGE2 release in antral mucosal cells. In conclusion, in ACh-stimulated antral mucous cells, an increase in [Ca2+]i activates Ca2+-regulated exocytotic events and PGE2 release mediated by COX-1. The released PGE2 induces the accumulation of cAMP, which enhances the Ca2+-regulated exocytosis. The autocrine mechanism mediated by PGE2 maintains the high-level mucin release from antral mucous cells during ACh stimulation.