Relationship between a <f>HCO3−</f>-permeable conductance and a CLCA protein from rat pancreatic zymogen granules

<f>Ca2+</f>-induced enzyme secretion in the exocrine pancreas is not completely understood. We have proposed that <f>Ca2+</f>-induced enzyme secretion in the exocrine pancreas involves activation of ion conductances in the membrane of zymogen granules (ZG). Here we have identified a <f>Ca2+</f>-activated anion conductance in rat pancreatic ZG membranes (ZGM). <f>Ca2+</f> (2.5–50 <f>μ</f>M) increased the conductance for <f>I−</f>, <f>NO3−</f>, <f>Br−</f>, or <f>HCO3−</f>, but not for <f>Cl−</f>, as determined by the rate of valinomycin-induced osmotic lysis of ZG suspended in isotonic <f>K+</f>-salts. 4,<f>4′</f>-Diisothiocyanatodihydrostilbene-2,<f>2′</f>-disulfonate (100 <f>μ</f>M) or 25 <f>μ</f>M dithiothreitol strongly inhibited <f>Ca2+</f>-dependent lysis. The permeability sequence, <f>Ca2+</f> dependence, and inhibitor sensitivity of ZG anion conductance are reminiscent of a family of epithelial <f>Ca2+</f>-activated anion channels (CLCA). CLCA expression was confirmed by RT-PCR with rat pancreatic mRNA and mouse CLCA1 primers. A PCR product (580 bp) exhibited 81%, 77%, and 57% amino acid similarity to the three mouse isoforms mCLCA-1, -2, and -3 (mgob-5), respectively. Antibodies against bovine tracheal CLCA1 showed CLCA expression in ZGM by immunoblotting, immunoperoxidase light microscopy, and immunogold labeling. These findings suggest that a CLCA-related protein could account for the <f>Ca2+</f>-activated <f>HCO3−</f> conductance of rat pancreatic ZGM and contribute to hormone-stimulated enzyme secretion. [Copyright &y& Elsevier]