Selvaggio A M, Schwartz J H, Bengele H H, Gordon F D, Alexander E A
Thorndike Memorial Laboratory, Boston City Hospital, Massachusetts.
Am J Physiol. 1988 Mar;254(3 Pt 2):F391-400. doi: 10.1152/ajprenal.1988.254.3.F391.
Inner medullary collecting duct cells were isolated from rat papillae and grown to confluence on cover slips. H+ secretion was estimated by intracellular pH (pHi) changes measured with the fluorescent probe 2,7-biscarboxyethyl-5(6)-carboxyfluorescein. In buffered NaCl, pHi was 7.14 +/- 0.04 (n = 78). After acidification about 40% of monolayers exhibited Na+-independent alkalinization. In 5 mM glucose, cell alkalinization occurred at a rate of 47 +/- 4 nM H+/min. However, cell alkalinization did not occur in the presence of 2-deoxy-D-glucose (5-15 mM), iodoacetate (5 mM), or KCN (5 mM). All monolayers tested exhibited amiloride-inhibitable Na+-dependent cell alkalinization that appeared to be a first-order kinetic process; Km [Na+] was approximately 52 mM and Vmax was approximately 250 nM [H+]/min. At a constant extracellular [Na+] (110 mM), Na+-dependent H+ efflux was a first-order function of pHi; Km for intracellular [H+] = 321 nM and Vmax = 182 nM H+/min. The data are consistent with the presence of a primary active H+ pump and a secondary active Na+ exchanger. The metabolic energy for the active H+ pump can be provided by glycolysis and oxidative phosphorylation.
从大鼠乳头分离出髓质集合管细胞，并使其在盖玻片上生长至汇合。通过用荧光探针2,7 - 双（羧乙基）-5（6）-羧基荧光素测量细胞内pH值（pHi）变化来估计H⁺分泌。在缓冲的NaCl中，pHi为7.14±0.04（n = 78）。酸化后，约40%的单层细胞表现出不依赖Na⁺的碱化。在5 mM葡萄糖存在下，细胞碱化以47±4 nM H⁺/分钟的速率发生。然而，在2 - 脱氧 - D - 葡萄糖（5 - 15 mM）、碘乙酸盐（5 mM）或KCN（5 mM）存在下，细胞碱化未发生。所有测试的单层细胞均表现出氨氯地平可抑制的依赖Na⁺的细胞碱化，这似乎是一个一级动力学过程；Km[Na⁺]约为52 mM，Vmax约为250 nM[H⁺]/分钟。在恒定的细胞外[Na⁺]（110 mM）下，依赖Na⁺的H⁺外流是pHi的一级函数；细胞内[H⁺]的Km = 321 nM，Vmax = 182 nM H⁺/分钟。这些数据与存在一个原发性活性H⁺泵和一个继发性活性Na⁺交换体一致。活性H⁺泵的代谢能量可由糖酵解和氧化磷酸化提供。
