Inhibition of small-conductance Cl- channels by the interleukin-1beta-stimulated production of superoxide in rabbit gastric parietal cells

Abstract

We have shown previously that the G protein-coupled production of superoxide anion (O2−) leads to closure of small-conductance Cl− channels (0.3–0.4 pS) in the basolateral membrane of rabbit parietal cells. In the present study, effects of interleukin-1β (IL-1β) on the Cl− channel were investigated. In the whole-cell patch-clamp recording, IL-1β (0.3–10 ng ml−1) inhibited the whole-cell Cl− current recorded from a parietal cell within isolated rabbit gastric glands. Variance noise analysis of the whole-cell Cl− current showed that the single channel conductance of the Cl− channel that is sensitive to IL-1β is 0.37 pS. The IL-1β (1 ng ml−1)-induced decrease of the Cl− current was abolished by anti-IL-1β antibody (2 μg ml−1), recombinant IL-1 receptor antagonist (500 ng ml−1), GDPβS (500 μM) and superoxide dismutase (100 units ml−1), a scavenger of O2−. Northern blot analysis showed that the mRNA of the IL-1 receptor was selectively expressed in rabbit gastric parietal cells. In the dihydrofluorescein diacetate-loaded single parietal cells in gastric glands, IL-1β (0.3–10 ng ml−1) stimulated the production of oxygen radicals. Y-27632 (1–10 μM), a specific Rho-kinase inhibitor, and fluvastatin (10 μM), an indirect inhibitor for Rho proteins, significantly inhibited the IL-1β-induced effects on the channel activity and production of oxygen radicals. IL-1β (0.3–10 ng ml−1) activated Rho in the parietal cells. These results indicate that IL-1β binds to the IL-1 receptor of gastric parietal cells and inhibits the small-conductance Cl− channel via the G protein-mediated Rho/Rho-kinase-dependent production of O2−.