Mellergård P, Ouyang Y B, Siesjö B K
Laboratory for Experimental Brain Research, Lund University Hospital, Sweden.
Exp Brain Res. 1993;95(3):371-80. doi: 10.1007/BF00227129.
We studied the regulation of intracellular pH (pHi) and the mechanisms of pHi regulation in cultured rat astrocytes using microspectrofluorometry and the pH-sensitive fluorophore 2',7'-bis(carboxyethyl-)-5,6-carboxyfluorescein. Control pHi was 7.00 +/- 0.02 in HCO3(-)-containing solutions at an extracellular pH of 7.35. Addition of 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS) or amiloride decreased pHi, as did removal of extracellular Na+, while removal of extracellular Cl- was followed by an increase in pHi. Following exposure to an acid transient induced by increasing the CO2 content from 5 to 15%, pHi rapidly returned to base line, with an average initial rate of recovery of 0.10 pH units min-1 (corresponding to a mean acid extrusion rate of 6.3 +/- 0.36 mmolo l-1 min-1). Regulation of pHi was impaired when either amiloride or DIDS was added or Cl- was removed. This inhibition was enhanced when both DIDS and amiloride were present, and pHi regulation was completely blocked in the absence of extracellular Na+. The rapid regulation of pHi normally seen following a transient alkalinisation was not inhibited by amiloride or removal of Na+, but was partially inhibited by DIDS and by the absence of extracellular Cl-. The results are compatible with the presence of at least three different pHi-regulating mechanisms: a Na+/H+ antiporter, a Na(+)-dependent HCO3-/Cl- exchanger (both regulating pHi during a transient acidification), and a passive Cl-/HCO3- exchanger (regulating pHi during transient alkalinisation). The results fail to provide firm evidence of the presence of an electrogenic Na+/HCO3- symporter.
我们使用显微分光荧光测定法和pH敏感荧光团2',7'-双（羧乙基）-5,6-羧基荧光素，研究了培养的大鼠星形胶质细胞内pH（pHi）的调节及其调节机制。在细胞外pH为7.35的含HCO3(-)溶液中，对照pHi为7.00±0.02。添加4,4'-二异硫氰酸根合芪-2,2'-二磺酸（DIDS）或氨氯吡咪会降低pHi，去除细胞外Na+也会如此，而去除细胞外Cl-后pHi会升高。在将CO2含量从5%增加到15%诱导的酸性瞬变暴露后，pHi迅速恢复到基线，平均初始恢复速率为0.10 pH单位/分钟（对应于平均酸排出速率为6.3±0.36 mmol·l-1·分钟-1）。当添加氨氯吡咪或DIDS或去除Cl-时，pHi的调节受损。当同时存在DIDS和氨氯吡咪时，这种抑制作用增强，并且在没有细胞外Na+的情况下pHi调节完全被阻断。短暂碱化后通常可见的pHi快速调节不受氨氯吡咪或去除Na+的抑制，但部分受DIDS和没有细胞外Cl-的抑制。结果与至少三种不同的pHi调节机制的存在相符：Na+/H+反向转运体、Na(+)依赖性HCO3-/Cl-交换体（两者在短暂酸化期间调节pHi）和被动Cl-/HCO3-交换体（在短暂碱化期间调节pHi）。结果未能提供确凿证据证明存在电中性Na+/HCO3-同向转运体。
