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 cuvette and intracellular pH (pHi) was measured. Under perfusion with a CO2HCO3--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 'NH4+-loading' method were rapidly followed by a Na+-dependent, amiloride-inhibitable pHi recovery. Introduction of a CO2HCO3--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 CO2HCO3--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. Two independent manipulations known to depolarize cell potential failed to elicit a pHi change indicating the absence of an electrogenic acid-base pathway, although the presence of an electroneutral Na+HCO3--cotransporter could not be excluded. It is concluded that these cells possess a highly efficient Na+H+ exchanger and a low Na+-dependent Cl-HCO3- exchange activity since in cells pre-acidified by the 'NH4+-loading' method, virtually all of the acid extruding capacity could be accounted for by the action of the former. © 1992.