ACELL Mar. 45/3

Alexander, EdwardA., Dennis Brown, Theodora Shih, Mary McKee, and John H. Schwartz. Effect of acidification on the location of H1-ATPase in cultured inner medullary collecting duct cells. Am. J. Physiol. 276 (Cell Physiol. 45): C758–C763, 1999.—In previous studies, our laboratory has utilized a cell line derived from the rat inner medullary collecting duct (IMCD) as a model system for mammalian renal epithelial cell acid secretion. We have provided evidence, from a physiological perspective, that acute cellular acidification stimulates apical exocytosis and elicits a rapid increase in proton secretion that is mediated by an H1ATPase. The purpose of these experiments was to examine the effect of acute cellular acidification on the distribution of the vacuolar H1-ATPase in IMCD cells in vitro. We utilized the 31-kDa subunit of the H1-ATPase as a marker of the complete enzyme. The distribution of this subunit of the H1-ATPase was evaluated by immunohistochemical techniques (confocal and electron microscopy), and we found that there is a redistribution of these pumps from vesicles to the apical membrane. Immunoblot evaluation of isolated apical membrane revealed a 237 6 34% (P , 0.05, n 5 9) increase in the 31-kDa subunit present in the membrane fraction 20 min after the induction of cellular acidification. Thus our results demonstrate the presence of this pump subunit in the IMCD cell line in vitro and that cell acidification regulates the shuttling of cytosolic vesicles containing the 31-kDa subunit into the apical membrane.