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

We have shown previously that the G protein-coupled production of superoxide anion (O-2(-)) 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-1beta (IL-1beta) on the Cl- channel were investigated. In the whole-cell patch-clamp recording, IL-1beta (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-1beta is 0.37 pS. The IL-1beta (1 ng ml(-1))-induced decrease of the Cl- current was abolished by anti-IL-1beta antibody (2 mug ml(-1)), recombinant IL-1 receptor antagonist (500 ng ml(-1)), GDPbetaS (500 muM) and superoxide dismutase (100 units ml(-1)), a scavenger of O-2(-). 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-1beta (0.3-10 ng ml(-1)) stimulated the production of oxygen radicals. Y-27632 (1-10 muM), a specific Rhokinase inhibitor, and fluvastatin (10 muM), an indirect inhibitor for Rho proteins, significantly inhibited the IL-1beta-induced effects on the channel activity and production of oxygen radicals. IL-1beta (0.3-10 ng ml(-1)) activated Rho in the parietal cells. These results indicate that IL-1beta 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 O-2(-).