Bidet M, Merot J, Tauc M, Poujeol P
Department of Biologie, CEN Saclay, Gif-sur-Yvette, France.
Biochim Biophys Acta. 1988 Feb 18;938(2):257-69. doi: 10.1016/0005-2736(88)90164-2.
Isolated proximal cells were prepared from rabbit kidney cortex by mechanical dissociation. The intracytoplasmic pH (pHi) was measured in HCO3(-)-free media (external pH (pHe), 7.3) using the fluorescent dye 2,7-biscarboxyethyl-5,6-carboxyfluorescein (BCECF). Cells were acid-loaded by the nigericin technique. Addition of 70 mM Na+ to the cells caused a rapid pHi recovery, which was blocked by 0.5 mM amiloride. When the cells were exposed to 5 mM sodium butyrate in the presence of 1 mM amiloride, the H+ efflux was significantly increased and followed Michaelis-Menten kinetics. Increasing pHe from 6.4 to 7.6 at a constant pHi of 6.4 enhanced the butyrate activation of the H+ efflux. Increasing pHi from 6.5 to 7.2 at a constant pHe of 7.2 reduced the butyrate effect. 22Na uptake experiments in the presence of 1 mM amiloride showed that 1.5 mM butyrate increased the Na+ flux in the proximal cells (pHi 7.10). The efficiency of monocarboxylic anions in promoting a pHi recovery increased with the length of their straight chain (acetate less than propionate less than butyrate less than valerate). The data show that when the Na+/H+ antiporter is blocked, the proximal cells can regulate their pHi by a Na+-coupled absorption of butyrate followed by non-ionic diffusion of butyric acid out of the cell and probably also by OH- influx by means of the OH-/anion exchanger.
通过机械分离法从兔肾皮质制备出分离的近端细胞。使用荧光染料2,7 - 双（羧乙基）- 5,6 - 羧基荧光素（BCECF）在无HCO3(-)的培养基（外部pH（pHe）为7.3）中测量细胞内pH（pHi）。采用尼日利亚菌素技术使细胞酸负荷。向细胞中添加70 mM Na+会导致pHi快速恢复，这被0.5 mM氨氯吡咪阻断。当细胞在1 mM氨氯吡咪存在下暴露于5 mM丁酸钠时，H+外流显著增加，并遵循米氏动力学。在pHi恒定为6.4时，将pHe从6.4提高到7.6会增强丁酸盐对H+外流的激活作用。在pHe恒定为7.2时，将pHi从6.5提高到7.2会降低丁酸盐的作用。在1 mM氨氯吡咪存在下进行的22Na摄取实验表明，1.5 mM丁酸盐增加了近端细胞中的Na+通量（pHi 7.10）。一元羧酸阴离子促进pHi恢复的效率随着其直链长度的增加而增加（乙酸盐＜丙酸盐＜丁酸盐＜戊酸盐）。数据表明，当Na+/H+反向转运体被阻断时，近端细胞可以通过与Na+偶联的丁酸盐吸收来调节其pHi，随后丁酸通过非离子扩散排出细胞，并且可能还通过OH - /阴离子交换器的OH - 内流来调节。
