A GTP-binding protein inhibits a gastric housekeeping chloride channel via intracellular production of superoxide.
A housekeeping basolateral Cl- channel of rabbit gastric parietal cells, the single channel conductance of which is about 0.3 picosiemens, is opened by prostaglandin E2 and closed by intracellular application of guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S). In the present patch clamp study, we found a novel GTP gamma S-dependent regulatory mechanism of the Cl- channel. GTP gamma S significantly decreased the open probability of the single Cl- channel without altering unit conductance. An intracellular application of superoxide dismutase (SOD; 100 units/ml) inhibited the GTP gamma S (50 microM)-induced closure of the Cl- channel. SOD plus catalase (100 units/ml) also inhibited the GTP gamma S-induced effect, while catalase alone did not inhibit it. In the absence of GTP gamma S, an intracellular application of hydrogen peroxide (H2O2; 30 microM) did not affect the Cl- channel current. Desferrioxamine (50 microM) which inhibits hydroxyl radical (.OH) production was without effect on the GTP gamma S-induced closure. These results suggest that the GTP gamma S-induced closure of the Cl- channel was due to intracellular production of superoxide (O2.-), but not due to .OH or H2O2. Furthermore, an artificial production of O2.- inside the cell by lumazine (50-100 microM) plus xanthine oxidase (0.5-1 milliunit/ml) in the absence of GTP gamma S also closed the channel. The lumazine/xanthine oxidase-induced closure of the channel was inhibited by SOD, but not by catalase or desferrioxamine. We conclude from these results that GTP-binding protein-coupled production of O2.- leads to closure of the Cl- channel in rabbit gastric parietal cells.
Sakai
H
H
Department of Pharmaceutical Physiology, Faculty of Pharmaceutical Sciences, Toyama Medical and Pharmaceutical University, Japan.
Takeguchi
N
N
D000818:Animals; D018118:Chloride Channels / Q000378:metabolism*; D015232:Dinoprostone / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism*; D005753:Gastric Mucosa / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D015640:Ion Channel Gating; D008297:Male; D011621:Pteridines / Q000494:pharmacology; D011817:Rabbits; D017382:Reactive Oxygen Species / Q000378:metabolism; D013482:Superoxide Dismutase / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism*
D018118:Chloride Channels; D011621:Pteridines; D017382:Reactive Oxygen Species; D013481:Superoxides; C008297:lumazine; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D013482:Superoxide Dismutase; D019204:GTP-Binding Proteins; D015232:Dinoprostone