Mechanism of chloride uptake in rabbit corneal epithelium

The mechanism of chloride uptake at the basal membrane (stromal side) of rabbit corneal epithelium was examined by observing the effects of ion transport inhibitors and ion concentrations on the stimulated epithelial short-circuit current (I(sc)). Loop diuretics inhibited the theophylline-stimulated peak and sustained I(sc). Treatment with 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS, 0.2 mM) and/or 5-(N,N-dimethyl)amiloride (0.1 mM) as well as the potent anion exchange inhibitor, 5c(+)[(2,3,9,9a-tetrahydro-1H-fluoren-7-yl)oxy]acetic acid (0.01 mM), had no significant effect on I(sc). These results are consistent with Cl^- uptake by a Na⁺-Cl^- or Na⁺-K⁺-2Cl^- cotransport mechanism rather than Cl^--HCO₃^--OH^- exchange coupled to Na⁺-H⁺ exchange. Incubation in low [Na⁺] or [Cl^-] before stimulation with forskolin (0.1 mM) reduced both peak and sustained I(sc), and saturation kinetics were exhibited. Hill coefficients for [Na⁺] and [Cl^-] were 0.99 and 1.04, respectively, for peak I(sc) and 0.66 and 1.18, respectively, for sustained I(sc). Apparent ion affinities for Na⁺ and Cl^- were 13.5 and 18 mM, respectively, for peak I(sc) and 15 and 22 mM, respectively, for sustained I(sc). These results favor Cl^- uptake by a 1 Na⁺:1 Cl^- cotransport mechanism for the rabbit corneal epithelium, but involvement of K⁺ in this process has not been eliminated.