Characterization of Na+/H+ exchange in a rabbit corneal epithelial cell line (SIRC).
Continuous intracellular pH (pHi) measurements were performed in SIRC rabbit corneal epithelial cells using the pH-sensitive absorbance of intracellularly trapped 5(and 6)-carboxy-4',5'-dimethylfluorescein. Steady-state pHi in nominally bicarbonate free Ringer's solution averaged 6.87 +/- 0.02 (mean +/- S.E., n = 53). After intracellular acidification induced by the NH4Cl-prepulse technique, there was a sodium-dependent pHi recovery towards the normal steady-state pHi. The initial pHi recovery rate was a saturable function of extracellular sodium concentration with an apparent Km for external sodium of about 25 mM and a Vmax of about 0.28 pH units/min. Virtually no pHi recovery was observed in the absence of extracellular sodium. Sodium removal during steady state acidified the cells by 0.36 +/- 0.05 pH units (mean +/- S.E., n = 13) within 5 min. There was a dose-dependent inhibition of pHi recovery after NH4Cl prepulse by amiloride with an IC50 of about 15 microM. Amiloride in a concentration of 1 mM almost completely abolished pHi recovery. Amiloride (1 mM) applied during steady state induced an intracellular acidification of 0.2 +/- 0.03 pH units (mean +/- S.E., n = 7) within 5 min. These findings suggest that a Na+/H+ exchange is present in SIRC rabbit corneal epithelial cells. Na+/H+ exchange seems to be the major process involved in pHi recovery in SIRC cells after an intracellular acid load. Na+/H+ exchange also plays a role in the maintenance of steady-state pHi.
Korbmacher
C
C
Institut für Klinische Physiologie, Freie Universität Berlin, Germany.
Helbig
H
H
Förster
C
C
Wiederholt
M
M
D000584:Amiloride / Q000494:pharmacology; D000643:Ammonium Chloride / Q000378:metabolism; D000818:Animals; D001693:Biological Transport, Active; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D003315:Cornea / Q000378:metabolism*; D004848:Epithelium / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D011817:Rabbits; D012964:Sodium / Q000378:metabolism*; D017923:Sodium-Hydrogen Exchangers
D002352:Carrier Proteins; D017923:Sodium-Hydrogen Exchangers; D000643:Ammonium Chloride; D000584:Amiloride; D012964:Sodium