Molecular Mechanisms of Calcium-sensing Receptor-mediated Calcium Signaling in the Modulation of Epithelial Ion Transport and Bicarbonate Secretion

Background: Calcium-sensing receptor (CaSR) plays a critical role in the regulation of epithelial ion transport. Results: CaSR activators induce Ca2+ signaling and duodenal bicarbonate secretion (DBS). Conclusion: CaSR triggers Ca2+-dependent DBS, likely through receptor-operated channels, intermediate conductance Ca2+-activated K+ channels, and the cystic fibrosis transmembrane conductance regulator. Significance: Dietary CaSR activators may modulate the physiological process of DBS that is critical for duodenal mucosal protection. Epithelial ion transport is mainly under the control of intracellular cAMP and Ca2+ signaling. Although the molecular mechanisms of cAMP-induced epithelial ion secretion are well defined, those induced by Ca2+ signaling remain poorly understood. Because calcium-sensing receptor (CaSR) activation results in an increase in cytosolic Ca2+ ([Ca2+](cyt)) but a decrease in cAMP levels, it is a suitable receptor for elucidating the mechanisms of [Ca2+](cyt)-mediated epithelial ion transport and duodenal bicarbonate secretion (DBS). CaSR proteins have been detected in mouse duodenal mucosae and human intestinal epithelial cells. Spermine and Gd3+, two CaSR activators, markedly stimulated DBS without altering duodenal short circuit currents in wild-type mice but did not affect DBS and duodenal short circuit currents in cystic fibrosis transmembrane conductance regulator (CFTR) knockout mice. Clotrimazole, a selective blocker of intermediate conductance Ca2+-activated K+ channels but not chromanol 293B, a selective blocker of cAMP-activated K+ channels (KCNQ1), significantly inhibited CaSR activator-induced DBS, which was similar in wild-type and KCNQ1 knockout mice. HCO3- fluxes across epithelial cells were activated by a CFTR activator, but blocked by a CFTR inhibitor. CaSR activators induced HCO3- fluxes, which were inhibited by a receptor-operated channel (ROC) blocker. Moreover, CaSR activators dose-dependently raised cellular [Ca2+](cyt), which was abolished in Ca2+-free solutions and inhibited markedly by selective CaSR antagonist calhex 231, and ROC blocker in both animal and human intestinal epithelial cells. Taken together, CaSR activation triggers Ca2+-dependent DBS, likely through the ROC, intermediate conductance Ca2+-activated K+ channels, and CFTR channels. This study not only reveals that [Ca2+](cyt) signaling is critical to modulate DBS but also provides novel insights into the molecular mechanisms of CaSR-mediated Ca2+-induced DBS.