Arsenis G
Endocrine Section, Veterans Administration Medical Center, Bay Pines, Florida 33504, USA.
Endocrinology. 1995 May;136(5):1920-7. doi: 10.1210/endo.136.5.7720639.
The activity of the Na+/H+ exchanger was examined by acidifying the intracellular pH (pHi) with Na+ propionate (NaP) and monitoring the recovery in the absence of HCO3- in rat adipocytes. Acidification of pHi, monitored with 2',7'-bis-(2-carboxyethyl)-5(6)-carboxyfluorescein, decreased the resting pHi from 7.25 +/- 0.01 to 6.70 +/- 0.01. A spontaneous pHi recovery to 6.90 +/- 0.02 was inhibited by 300 microM amiloride. This effect was Na+ specific, as recovery did not occur in cells exposed to K+ propionate (KP). The addition of NaCl (30 mM) to KP induced pHi alkalinization. Acidification of pHi increased 22Na+ transport from 0.60 +/- 0.12 nmol/10(5) cells.min at resting pHi to 2.893 +/- 0.129 (P < 0.001) and 7.984 +/- 0.312 (P < 0.001) in the first and tenth minutes, respectively. Amiloride inhibited this 5- and 14-fold stimulation by 85% (P < 0.001). Insulin in the presence of 100 microM ouabain stimulated Na+ influx by more than 15% (P < 0.01). Ethylisopropylamiloride (10 microM) inhibited the effect of insulin by 85% (P < 0.001). Intracellular Na+, measured with a Na(+)-specific electrode, increased by 10-fold in acid-loaded cells compared to that in Na(+)-depleted cells (10.750 +/- 0.479 vs. 1.045 +/- 0.100 mM; P < 0.001). Amiloride decreased NaP-stimulated intracellular Na+ by 82% (P < 0.001). To our knowledge, this is the first report showing the presence of an insulin-responsive and amiloride-sensitive Na+/H+ exchanger that regulates pHi by a Na(+)-specific and pHi-dependent mechanism in rat adipocytes.
通过用丙酸钠（NaP）酸化大鼠脂肪细胞的细胞内pH值（pHi）并在无HCO3-的情况下监测其恢复情况，来检测Na+/H+交换体的活性。用2',7'-双（2-羧乙基）-5（6）-羧基荧光素监测pHi的酸化情况，结果显示静息pHi从7.25±0.01降至6.70±0.01。300微摩尔的氨氯吡咪抑制了pHi自发恢复至6.90±0.02。这种作用具有Na+特异性，因为在暴露于丙酸钾（KP）的细胞中未出现恢复情况。向KP中添加氯化钠（30 mM）会诱导pHi碱化。pHi的酸化使22Na+转运在静息pHi时从0.60±0.12纳摩尔/10^5个细胞·分钟分别增加到第一分钟时的2.893±0.129（P<0.001）和第十分钟时的7.984±0.312（P<0.001）。氨氯吡咪抑制了这种5倍和14倍的刺激，抑制率达85%（P<0.001）。在存在100微摩尔哇巴因的情况下，胰岛素刺激Na+内流超过15%（P<0.01）。乙基异丙基氨氯吡咪（10微摩尔）抑制胰岛素作用的抑制率为85%（P<0.001）。用Na+特异性电极测量，与Na+耗尽的细胞相比，酸负荷细胞中的细胞内Na+增加了10倍（10.750±0.479对1.045±0.100毫摩尔；P<0.001）。氨氯吡咪使NaP刺激的细胞内Na+降低了82%（P<0.001）。据我们所知，这是第一份表明在大鼠脂肪细胞中存在一种胰岛素反应性且对氨氯吡咪敏感的Na+/H+交换体的报告，该交换体通过一种Na+特异性且依赖pHi的机制调节pHi。
