Chloride Channels Regulate HIT Cell Volume but Cannot Fully Account for Swelling-Induced Insulin Secretion

Chloride Channels Regulate HIT Cell Volume but Cannot Fully Account for Swelling-Induced Insulin Secretion Tracie A. Kinard 1 , Paulette B. Goforth 1 , Qing Tao 1 , Mary E. Abood 1 4 , Jeanette Teague 5 and Leslie S. Satin 1 2 3 1 Departments of Pharmacology and Toxicology 2 Department of Physiology 3 Medicine (Endocrinology), Medical College of Virginia, Virginia Commonwealth University, Richmond, Virginia 4 Forbes Norris ALS Research Center, California Pacific Medical Center, San Francisco, California 5 Division of Endocrinology and Metabolism and Nutrition Department of Medicine, School of Medicine, University of Washington, Seattle, Washington Abstract Insulin-secreting pancreatic islet β-cells possess anion-permeable Cl − channels (I Cl,islet ) that are swelling-activated, but the role of these channels in the cells is unclear. The Cl − channel blockers 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid (DIDS) and niflumic acid were evaluated for their ability to inhibit I Cl,islet in clonal β-cells (HIT cells). Both drugs blocked the channel, but the blockade due to niflumic acid was less voltage-dependent than the blockade due to DIDS. HIT cell volume initially increased in hypotonic solution and was followed by a regulatory volume decrease (RVD). The addition of niflumic acid and, to a lesser extent, DIDS to the hypotonic solution potentiated swelling and blocked the RVD. In isotonic solution, niflumic acid produced swelling, suggesting that islet Cl − channels are activated under basal conditions. The channel blockers glyburide, gadolinium, or tetraethylammonium-Cl did not alter hypotonic-induced swelling or volume regulation. The Na/K/2Cl transport blocker furosemide produced cell shrinkage in isotonic solution and blocked cell swelling normally induced by hypotonic solution. Perifused HIT cells secreted insulin when challenged with hypotonic solutions. However, this could not be completely attributed to I Cl,islet -mediated depolarization, because secretion persisted even when Cl − channels were fully blocked. To test whether blocker-resistant secretion occurred via a distal pathway, distal secretion was isolated using 50 mmol/l potassium and diazoxide. Under these conditions, glucose-dependent secretion was blunted, but hypotonically induced secretion persisted, even with Cl − channel blockers present. These results suggest that β-cell swelling stimulates insulin secretion primarily via a distal I Cl,islet -independent mechanism, as has been prop