HCO₃^- transport in the toad lens epithelium is mediated by an electronegative Na⁺-dependent symport

The pH-sensitive cell-entrapable dye 2',7'-bis(carboxyethyl)-5,6-carboxyfluorescein (BCECF) was used to continuously monitor epithelial intracellular pH (pH(i)) of intact toad lenses, enabling a description of a HCO₃^- transport mechanism that contributes to pH(i) homeostasis of this organ. In physiological medium, pH 7.40, the steady-state pH(i) was 7.48 ± 0.03 (SE; n = 93). Induction of cell depolarization by either elevation of [K⁺] to 50 mM, addition of 0.2 mM quinidine, a K⁺-channel blocker, or addition of 0.1 mM Li⁺ ionophore that equalizes Na⁺ and K⁺ permeabilities elicited pH(i) increases (ΔpH(i) = 0.18 ± 0.02; P < 0.0005; n = 13, for K⁺). These increases could be blocked or reverted by DIDS and were not affected by amiloride. Removal of Na⁺ induced an amiloride-insensitive acidification. pH(i) recovery seen upon Na⁺ reintroduction in the presence of amiloride was inhibited by DIDS. Despite the effects of DIDS on induced pH(i) changes, the agent did not affect control pH(i). Elevation of medium HCO₃^- (pH to 7.7) produced a pH(i) increase followed by a spontaneous reversal. This increase was both DIDS and Na⁺ sensitive. pH(i) was not affected in any condition by removal (or addition) of Cl^-, unless the lens was pretreated with the artificial Cl^--HCO₃^- exchanger tributyltin. Collectively, these results suggest that the primary mechanism for HCO₃^- movement across the lens epithelial membrane is an electronegative Na⁺ cotransporter and that this system is near equilibrium under normal physiological conditions.