Amiloride-sensitive Na+/H+ antiporter in basolateral membrane of hamster ascending thin limb of Henle's loop

I. Fujiwara, Y. Kondo, Y. Igarashi, C. N. Inoue, N. Takahashi, K. Tada, K. Abe

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4 Citations (Scopus)

Abstract

The mechanisms of intracellular pH (pH(i)) regulation were investigated in the in vitro microperfused hamster ascending thin limb (ATL) of Henle's loop with the fluorescent pH indicator, 2',7'-bis(carboxyethyl)-5(6)- carboxyfluorescein. pH(i) of ATL cells was 7.05 ± 0.02 (n = 30) when microperfused with a CO2/HCO3/--buffered solution. In HEPES-buffered solution, pH(i) was 7.10 ± 0.02 (n = 16), which was significantly higher than the value in CO2/HCO3/--buffered solution (P < 0.05, n = 16). In HEPES-buffered solution, elimination of Na+ and addition of 1 mM amiloride to basolateral solution decreased the pH(i) by 0.12 ± 0.03 (n = 6) and 0.11 ± 0.02 (n = 5) at 1 min, respectively. The same manipulations in the luminal solution had no effect on pH(i). One millimolar of N-ethylmaleimide (NEM) added to either side of ATL caused no significant change in pH(i). Elimination of K+ on either side of ATL did not alter pH(i). After adding 20 mM NH4Cl to basolateral solution, pH(i) instantaneously increased from 7.17 ± 0.01 to 7.51 ± 0.03 (n = 3), and then returned to steady-state level of 7.21 ± 0.05 (n = 15) in 3 min. Removal of NH4Cl from basolateral solution then caused a rapid fall in pH(i) to 6.31 ± 0.05 (n = 15), followed by spontaneous recovery at a rate of 0.43 ± 0.06 unit/min (n = 15). In presence of 1 mm amiloride in basolateral solution, NH4Cl removal caused a fall in pH(i) to 6.10 ± 0.05 (n = 6), followed by a recovery at a rate of 0.14 ± 0.03 unit/min (n = 6), significantly smaller than that in absence of amiloride. In absence of Na+ in basolateral solution, NH4Cl removal caused a fall in the pH(i) to 6.22 ± 0.06 (n = 5), followed by a recovery at a rate of 0.05 ± 0.02 unit/min (n = 5), which is also significantly smaller than that in presence of Na+. In CO2/HCO3/--buffered solution, 1 mM NEM added either to lumen or to bath did not change pH(i), whereas 1 mM amiloride added to bath significantly acidified the ATL cells. We conclude that there is an amiloride-sensitive Na+/H+ antiporter in the basolateral membrane of the ATL, and that this transporter is the main regulatory mechanism of pH(i) in ATL cells. The results also show that other pH regulatory mechanisms, such as an NEM-sensitive proton pump or H+-K+-adenosinetriphosphatase, may not play a role in pH(i) regulation from acidification in the ATL.

Original languageEnglish
Pages (from-to)F410-F415
JournalAmerican Journal of Physiology - Renal Fluid and Electrolyte Physiology
Volume268
Issue number3 37-3
DOIs
Publication statusPublished - 1995

Keywords

  • 2',7'- bis(carboxyethyl)-5(6)-carboxyfluorescein
  • amiloride
  • intracellular pH
  • renal medulla
  • sodium/hydrogen antiport

ASJC Scopus subject areas

  • Physiology

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