Inhibitory effects of oxidants on n-type K+ channels in T lymphocytes and Xenopus oocytes

Reactive oxygen species (ROS) appear to be involved in Fas-induced programmed cell death, We have previously demonstrated a tyrosine-kinase-dependent inhibition of the n-type K+ channels (K-n) by Fas stimulation. Thus, the effect of hydrogen peroxide (H2O2) on the function of K-n was examined using the patch-clamp technique. Incubation of Jurkat human T lymphocytes with 100 mu M H2O2 resulted in a 46 +/- 5% inhibition of the macroscopic whole-cell current, Experiments performed at the single-channel level using the cell-attached configuration revealed that the probability of the channel being open diminished upon incubation in H2O2. The effect was not dependent on src-like kinases, since H2O2 did not trigger tyrosine phosphorylation of the K-n channel protein and herbimycin A did not prevent channel inhibition. K(v)1.3 channels underly the K-n of T lymphocytes and were expressed Xenopus oocytes and subjected to electrophysiological analysis by the two-electrode voltage-clamp technique. Application of 1 mM H2O2 and 500 mu M t-BOOH (tert, butylhydroperoxide) resulted in a marked inhibition of the K+ current within 20 min. Both the membrane-permeable thiol-group oxidizing agent DTNP [2,2'-dithiobis-(5-nitropyridine)] and the membrane-impermeable DTNB [5,5'-Dithiobis-(2-nitrobenzoic acid)] (50 mu M) inhibited K(v)1.3 channels, suggesting that extracellular domains of K(v)1.3 are affected. These results point to a direct modulation of K-n by various oxidative agents.