Ca(2+)-activated C1- channels in a human biliary cell line: regulation by Ca2+/calmodulin-dependent protein kinase

Biliary epithelial cells contribute to bile formation through absorption and secretion of fluid and electrolytes. Recent studies indicate that membrane Cl- permeability is regulated in part by increases in intracellular Ca2+ concentration. The purpose of these studies was to evaluate the effects of intracellular Ca2+ on channel activity, using the human Mz-ChA-1 cholangiocarcinoma cell line as a model, and to assess the possible roles of Ca(2+)-dependent kinases in channel regulation. Exposure to ionomycin (1 microM) activated ion channels in the cell-attached configuration in 63 of 74 attempts, increasing open probability (NPo) from 0 to 0.26 +/- 0.15 (n = 17). Multiple channels were present in each patch, and the effects of ionomycin were reversed by subsequent addition of ethylene glycol-bis (beta-aminoethyl ether)-N,N,N',N'- tetraacetic acid (2 mM) to the bath. With Cl(-)-containing solutions, channels had a slope conductance of 14 +/- 4 pS (n = 11), and the mean open time was estimated to be 5.3 +/- 1.8 ms. These channels were anion selective, and currents were carried by efflux of Cl- at the resting potential. Exposure to the Ca2+/calmodulin-dependent protein kinase II (CaMKII) antagonist calmidazolium (100 microM) decreased NPo in ionomycin-stimulated cells to 0.02 +/- 0.06 (n = 19). The protein kinase C antagonist chelerythrine (50 microM) was without effect. In parallel studies in subconfluent cell monolayers, CaMKII antagonists were also potent inhibitors of ionomycin-stimulated 125I efflux. These findings indicate that Ca(2+)-dependent increases in membrane Cl- permeability are related in part to opening of 14.pS anion channels through a mechanism that depends on both Ca2+ and CaMKII. These channels represent a potential target for pharmacological modulation of biliary cell transport and function.