Zhou X, Nakamura S, Xia S L, Wingo C S
Division of Nephrology, Hypertension and Transplantation, Department of Medicine, College of Medicine, University of Florida, 32608-1197, USA.
Am J Physiol Renal Physiol. 2001 Aug;281(2):F366-73. doi: 10.1152/ajprenal.2001.281.2.F366.
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 PCO(2) 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% CO(2) in CCD, and this effect was persistent for the entire 10% CO(2) period, whereas 10 microM SCH-28080 in the perfusate totally abolished the stimulation of Rb reabsorption by 10% CO(2). After stimulation of Rb reabsorption by 10% CO(2), subsequent addition of 0.1 mM methazolamide, an inhibitor of carbonic anhydrase, failed to affect Rb reabsorption. However, simultaneous exposure to 10% CO(2) and methazolamide prevented the stimulation of Rb reabsorption. Treatment with the intracellular calcium chelator MAPTAM (0.5 microM) inhibited the stimulation of Rb reabsorption by 10% CO(2). Similar inhibition was also observed in the presence of either a calmodulin inhibitor, W-7 (0.5 microM), 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% CO(2) 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.
皮质集合管（CCD）中的顶端H⁺-K⁺-ATP酶在尿液酸化和钾重吸收中起重要作用。我们之前的研究表明，H⁺-K⁺-ATP酶在一定程度上介导了兔CCD中的铷重吸收（周X和温戈CS。《美国生理学杂志：肾脏流体电解质生理学》263：F1134 - F1141，1992）。这些实验的目的是使用来自低钾饮食兔子的体外微灌流CCD来研究：1）PCO₂急性升高以及由此推测的细胞内酸中毒是否刺激钾吸收通量；2）这种刺激是否依赖于功能性H⁺-K⁺-ATP酶的存在。在CCD中暴露于10% CO₂后，铷重吸收显著增加，并且在整个10% CO₂时间段内这种作用持续存在，而灌流液中10 μM的SCH - 28080完全消除了10% CO₂对铷重吸收的刺激。在10% CO₂刺激铷重吸收后，随后添加0.1 mM的碳酸酐酶抑制剂甲醋唑胺，并未影响铷重吸收。然而，同时暴露于10% CO₂和甲醋唑胺可防止铷重吸收的刺激。用细胞内钙螯合剂MAPTAM（0.5 μM）处理可抑制10% CO₂对铷重吸收的刺激。在存在钙调蛋白抑制剂W - 7（0.5 μM）或微管蛋白聚合抑制剂秋水仙碱（0.5 mM）的情况下也观察到类似的抑制作用。在时间对照研究中，灌流时间对铷重吸收没有显著影响。我们得出以下结论：1）暴露于10% CO₂时铷重吸收的刺激依赖于功能性H⁺-K⁺-ATP酶的存在，并且似乎部分通过该酶通过胞吐作用插入顶端质膜来调节；2）H⁺-K⁺-ATP酶的插入需要细胞内pH的变化并且需要细胞内钙浓度的基础水平；3）H⁺-K⁺-ATP酶的插入通过微管依赖过程发生。
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