REDUCTION OF WATER PERMEABILITY BY ANISOTONIC SOLUTIONS IN FROG CORNEAL EPITHELIUM

PURPOSE. To study the effects of bathing solution osmolarity and Cl- s ecretagogues on the diffusional water permeability (P-dw) of the isola ted frog corneal epithelium. METHODS. Isolated frog corneas, with the endothelium scraped off, were mounted as a partition between Ussing-ty pe hemichambers. Unidirectional diffusional water fluxes (J(dw)) were measured by adding (H2O)-H-3 to one hemichamber and sampling from the other. Electrical parameters were measured simultaneously. J(dw) was d etermined in control isosmotic conditions and after either changes in osmolarity of the bathing solutions or the additions of amphotericin B , epinephrine, Ca2+ ionophore, and other agents, RESULTS. Apical addit ion of 0.5 mM HgCl2 elicited an 11-fold increase in paracellular condu ctance and inhibited J(dw) by 36%, suggesting that J(dw) was predomina ntly transcellular and that there was a negligible contribution of the paracellular pathway. Pretreatment of corneas with 2-mercaptoethanol prevented the effects of Hg2+ an the paracellular conductance and J(dw ). A hypotonic medium on the basolateral side reversibly reduced J(dw) proportionately to the reduction in osmolarity, with 40 mOsm exerting a 29% decrease. Results from an Arrhenius plot suggest that water cha nnels closed under this condition. Apical hypertonicity (350 mOsm) red uced J(dw) by approximately 12%. Basolateral hypertonicity (450 mOsm), after permeabilization of the apical membrane with amphotericin B, re duced J(dw) by 15%. Epinephrine was the only Cl- secretagogue that red uced J(dw), on average by 12%. This effect, which was also observed wi th amphotericin B-treated corneas, was not mediated by classical beta- receptors based on the results obtained with isoproterenol and propran olol. CONCLUSIONS. Changes in basolateral osmolarity or the presence o f an apical hypertonic solution decreased the diffusional water permea bility (P-dw) of the corneal epithelium. Epinephrine also decreased P- dw, and this effect was localized to the basolateral membrane. The sim ilarities, of a sequence motif found in potassium channels and beta-ad renergic receptor kinases that are regulated by the beta gamma subunit of G proteins with that found in aquaporins 2 and 5, could explain th e link with epinephrine. Regardless of the mechanism, these results in dicate that corneal epithelial water permeability; can be regulated, p resumably to protect cell volume from changes in solution osmolarity.