Anion secretion evoked by ATP in rabbit cornea and its suppression in dry eye model

Abstract

To explore the regulation of anion secretion across rabbit corneal epithelia by extracellular ATP and to investigate the mechanism of benzalkonium chloride (BAC) induced dry eye model involved in this secretion, short circuit current and patch clamp were used. Changes in cytosolic Ca2+ concentration were measured using Fluo3. The cornea responded to apical application of ATP in short circuit current (Isc). Ion substitution experiments showed the rise in Isc due to Cl‐ and HCO3‐ secretion. Pretreatment with Cl‐ channel blockers, such as DIDS, NPPB and DPC attenuated the response. Moreover, the Isc was inhibited by U73122 and BAPTA/AM, indicating the requirement of PLC and intracellular calcium (Ca2+i) in the anion secretion. In whole‐cell patch clamp study, ATP stimulated a Ca2+‐activate Cl‐ channel and this current showed an outward rectifying I‐V relationship. In BAC‐pretreated epithelial cell, the whole‐cell current was suppressed. Dynamic changes of Ca2+i demonstrated the elevations of Ca2+i was smaller in BAC‐treated epithelia compared to control cells when they were challenged by ATP and Thapsigagin. These findings suggested ATP could induce anion secretion, and this secretion was inhibited in BAC‐evoked rabbit dry eye model. Abnormality of intracellular Ca2+ and Ca2+ activated Cl‐ channel may take charge of inhibitory anion secretion in the rabbit corneal epithelial cells of dry eye model.