Cyclic AMP potentiates Ca2+-dependent exocytosis in pancreatic duct epithelial cells.

Exocytosis is evoked by intracellular signals, including Ca(2+) and protein kinases. We determined how such signals interact to promote exocytosis in exocrine pancreatic duct epithelial cells (PDECs). Exocytosis, detected using carbon-fiber microamperometry, was stimulated by [Ca(2+)](i) increases induced either through Ca(2+) influx using ionomycin or by activation of P2Y2 or protease-activated receptor 2 receptors. In each case, the exocytosis was strongly potentiated when cyclic AMP (cAMP) was elevated either by activating adenylyl cyclase with forskolin or by activating the endogenous vasoactive intestinal peptide receptor. This potentiation was completely inhibited by H-89 and partially blocked by Rp-8-Br-cAMPS, inhibitors of protein kinase A. Optical monitoring of fluorescently labeled secretory granules showed slow migration toward the plasma membrane during Ca(2+) elevations. Neither this Ca(2+)-dependent granule movement nor the number of granules found near the plasma membrane were detectably changed by raising cAMP, suggesting that cAMP potentiates Ca(2+)-dependent exocytosis at a later stage. A kinetic model was made of the exocytosis stimulated by UTP, trypsin, and Ca(2+) ionophores with and without cAMP increase. In the model, without a cAMP rise, receptor activation stimulates exocytosis both by Ca(2+) elevation and by the action of another messenger(s). With cAMP elevation the docking/priming step for secretory granules was accelerated, augmenting the releasable granule pool size, and the Ca(2+) sensitivity of the final fusion step was increased, augmenting the rate of exocytosis. Presumably both cAMP actions require cAMP-dependent phosphorylation of target proteins. cAMP-dependent potentiation of Ca(2+)-induced exocytosis has physiological implications for mucin secretion and, possibly, for membrane protein insertion in the pancreatic duct. In addition, mechanisms underlying this potentiation of slow exocytosis may also exist in other cell systems.