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Division of Nephrology, Hypertension and Transplantation, Department of Medicine, College of Medicine, University of Florida, and Nephrology Section, Veterans Affairs Medical Center, Gainesville, Florida 32608-1197
Apical H-K-ATPase in the cortical collecting duct (CCD) plays an important role in urinary acidification and K reabsorption. Our previous studies demonstrated that an H-K-ATPase mediates, in part, Rb reabsorption in rabbit CCD (Zhou X and Wingo CS. Am J Physiol Renal Fluid Electrolyte Physiol 263: F1134-F1141, 1992). The purpose of these experiments was to examine using in vitro microperfused CCD from K-restricted rabbits 1) whether an acute increase in PCO2 and, presumably, intracellular acidosis stimulate K absorptive flux; and 2) whether this stimulation was dependent on the presence of a functional H-K-ATPase. Rb reabsorption was significantly increased after exposure to 10% CO2 in CCD, and this effect was persistent for the entire 10% CO2 period, whereas 10 µM SCH-28080 in the perfusate totally abolished the stimulation of Rb reabsorption by 10% CO2. After stimulation of Rb reabsorption by 10% CO2, subsequent addition of 0.1 mM methazolamide, an inhibitor of carbonic anhydrase, failed to affect Rb reabsorption. However, simultaneous exposure to 10% CO2 and methazolamide prevented the stimulation of Rb reabsorption. Treatment with the intracellular calcium chelator MAPTAM (0.5 µM) inhibited the stimulation of Rb reabsorption by 10% CO2. Similar inhibition was also observed in the presence of either a calmodulin inhibitor, W-7 (0.5 µM), or colchicine (0.5 mM), an inhibitor of tubulin polymerization. In time control studies, the perfusion time did not significantly affect Rb reabsorption. We conclude the following: 1) stimulation of Rb reabsorption on exposure to 10% CO2 is dependent on the presence of a functional H-K-ATPase and appears to be regulated in part by the insertion of this enzyme into the apical plasma membrane by exocytosis; 2) insertion of H-K-ATPase requires changes in intracellular pH and needs a basal level of intracellular calcium concentration; and 3) H-K-ATPase insertion occurs by a microtubule-dependent process.
exocytosis; intracellular pH; calcium; calmodulin; microtubules; cortical collecting duct
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