犬肾Madin-Darby细胞中不依赖钠离子的氯离子-碳酸氢根离子交换。在细胞内pH调节中的作用。

Na+-independent Cl(-)-HCO-3- exchange in Madin-Darby canine kidney cells. Role in intracellular pH regulation.

作者信息

Kurtz I, Golchini K

出版信息

J Biol Chem. 1987 Apr 5;262(10):4516-20.

Abstract

The role of plasma membrane Cl(-)-HCO-3-exchange in regulating intracellular pH (pHi) was examined in Madin-Darby canine kidney cell monolayers. In cells bathed in 25 mM HCO-3, pH 7.4, steady state pHi was 7.10 +/- 0.03 (n = 14) measured with the fluorescent pH probe 2',7'-biscarboxyethyl-5,6-carboxyfluorescein. Following acute alkaline loading, pHi recovered exponentially in approximately 4 min. The recovery rate was significantly decreased by Cl- or HCO-3 removal and in the presence of 50 microM 4,4'-diisothiocyano-2,2'-disulfonic stilbene (DIDS). Na+ removal or 10(-3) M amiloride did not inhibit the pHi recovery rate after an acute alkaline load. Following acute intracellular acidification, the pHi recovery rate was significantly inhibited by 10(-3) M amiloride but was not altered by Cl- removal or 50 microM DIDS. At an extracellular pH (pHo) of 7.4, pHi remained unchanged when the cells were bathed in either Cl- free media, HCO-3 free media, or in the presence of 50 microM DIDS. As pHo was increased to 8.0, steady state pHi was significantly greater than control in Cl(-)-free media and in the presence of 50 microM DIDS. It is concluded that Madin-Darby canine kidney cells possess a Na+-independent Cl(-)-HCO-3 exchanger with a Km for external Cl- of approximately 6 mM. The exchanger plays an important role in pHi regulation following an elevation of pHi above approximately 7.1. Recovery of pHi following intracellular acidification is mediated by the Na+/H+ antiporter and not the anion exchanger.

摘要

在Madin-Darby犬肾细胞单层中研究了质膜Cl⁻-HCO₃⁻交换在调节细胞内pH（pHi）中的作用。在用25 mM HCO₃、pH 7.4的溶液孵育的细胞中，用荧光pH探针2',7'-双羧乙基-5,6-羧基荧光素测量的稳态pHi为7.10±0.03（n = 14）。急性碱负荷后，pHi在约4分钟内呈指数恢复。去除Cl⁻或HCO₃以及存在50 μM 4,4'-二异硫氰酸-2,2'-二磺酸芪（DIDS）时，恢复速率显著降低。去除Na⁺或10⁻³ M氨氯吡脒并不抑制急性碱负荷后的pHi恢复速率。急性细胞内酸化后，pHi恢复速率受到10⁻³ M氨氯吡脒的显著抑制，但不受去除Cl⁻或50 μM DIDS的影响。在细胞外pH（pHo）为7.4时，当细胞在无Cl⁻培养基、无HCO₃培养基或存在50 μM DIDS的情况下孵育时，pHi保持不变。当pHo增加到8.0时，在无Cl⁻培养基和存在50 μM DIDS的情况下，稳态pHi显著高于对照。得出结论，Madin-Darby犬肾细胞具有一种不依赖Na⁺的Cl⁻-HCO₃交换体，其对细胞外Cl⁻的Km约为6 mM。该交换体在pHi升高至约7.1以上后的pHi调节中起重要作用。细胞内酸化后pHi的恢复是由Na⁺/H⁺反向转运体而非阴离子交换体介导的。