CHARACTERIZATION OF 2 DISTINCT CHLORIDE CHANNELS IN CULTURED DOG PANCREATIC DUCT EPITHELIAL-CELLS

Cl- secretion by pancreatic duct epithelial cells (PDEC) regulates cel lular HCO3- secretion, an important component of the exocrine pancreas . In cystic fibrosis, for example, impaired function of the cystic fib rosis transmembrane conductance regulator (CFTR) Cl- channel results i n decreased pancreatic secretion and secondary pancreatic insufficienc y. Studies of ion transport by PDEC have been hindered by the lack of a practical in vitro model. We have successfully cultured nontransform ed dog PDEC on Vitrogen-coated permeable membranes overlying a feeder layer of myofibroblasts and report the characterization of Cl- channel s in these cells. Cl- conductance, assessed through efflux of I-125- f rom PDEC, was stimulated by agents acting via adenosine 3',5'-cyclic m onophosphate (cAMP) or cytosolic Ca2+. The Cl- conductances activated by cAMP and Ca2+ were distinct, since they were differentially inhibit ed by 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid and, to a lesse r extent, by 5-nitro-2-(3-phenylpropylamino)benzoic acid and diphenyla mine-2 carboxylate. Patch-clamp studies confirmed the presence of Cl- channels activated by cAMP and Ca2+ with differential inhibition by 4, 4'-diisothiocyanostilbene-2,2'-disulfonic acid. The presence of CFTR C l- channels in PDEC was confirmed by immunoblotting. These cultured PD EC are an optimal model for studies of pancreatic duct secretion.