Evidence for parallel Na(+)-H+ and Na(+)-dependent Cl(-)-HCO3- exchangers in cultured bovine lens cells.
BCECF, a cell-entrapable dye with a pH-sensitive fluorescence spectrum, was used to identify transport mechanisms contributing to pH homeostasis of cultured bovine lens epithelial cells. Cells from a spontaneously established lineage were grown on glass coverslips that fit diagonally in a standard curvette and intracellular pH (pHi) was measured. Under perfusion with a CO2-HCO3(-)-free medium (pH 7.45), pHi was 7.19 +/- 0.21 (mean +/- S.D., n = 94 cell preparations). Cell acidifications (pHi to 6.65, n = 8) induced by the 'NH(4+)-loading' method were rapidly followed by a Na(+)-dependent, amiloride-inhibitable pHi recovery. Introduction of a CO2-HCO3(-)-rich medium (pH 7.45) resulted in a small acidification (0.18 +/- 0.04 U, n = 16; P < 0.002) due to rapid CO2 entry and an ensuing slow alkalinization to a pHi near the control CO2-HCO3(-)-free value. Subsequent removal of Cl- resulted in a further alkalinization of 0.18 +/- 0.02 U (n = 13; P < 0.001). This Cl- effect was completely inhibited by the absence of Na+, but was insensitive to amiloride, suggesting the presence of a Na(+)-dependent Cl(-)-HCO3- exchanger. Consistent with this posit, the reintroduction of Na+ to cells perfused in the absence of the cation with a HCO3(-)-containing, amiloride-complemented solution resulted in a gradual recovery from the acidic pHi induced by the baseline conditions (n = 6). The amiloride-insensitive, Na(+)- and HCO3(-)-dependent recovery was completely inhibited in cells pre-incubated with DIDS.(ABSTRACT TRUNCATED AT 250 WORDS)
Alvarez
L J
LJ
Department of Ophthalmology, Mount Sinai School of Medicine, New York, NY 10029.
Candia
O A
OA
Wolosin
J M
JM
D000818:Animals; D000837:Anion Exchange Resins / Q000378:metabolism*; D001639:Bicarbonates / Q000493:pharmacokinetics; D001692:Biological Transport; D002417:Cattle; D002478:Cells, Cultured; D002712:Chlorides / Q000493:pharmacokinetics; D004848:Epithelium; D005452:Fluoresceins / Q000378:metabolism*; D006859:Hydrogen / Q000493:pharmacokinetics; D006863:Hydrogen-Ion Concentration; D007475:Ion Exchange Resins / Q000378:metabolism*; D007908:Lens, Crystalline / Q000378:metabolism*; D012964:Sodium / Q000493:pharmacokinetics
D000837:Anion Exchange Resins; D001639:Bicarbonates; D002712:Chlorides; D005452:Fluoresceins; D007475:Ion Exchange Resins; D006859:Hydrogen; C043829:2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein; D012964:Sodium