SUSCEPTIBILITY OF CAFFEINE-INDUCED AND INS(1,4,5)P-3-INDUCED CONTRACTIONS TO OXIDANTS IN PERMEABILIZED VASCULAR SMOOTH-MUSCLE

Two principal pathways of Ca2+ release from the sarcoplasmic reticulum of excitable and non-excitable cells have been described: one pathway dependent on the second messenger D-myo-inositol 1,4,5-trisphosphate (Ins(1,4,5)P-3), and a second pathway sensitive to Ca2+ and regulated by caffeine and ryanodine. It was found that the Ca2+-pump activity of vascular smooth muscle sarcoplasmic reticulum is inhibited by superox ide anion radicals (O-2(.-)); however, the effects of reactive oxygen intermediates on sarcoplasmic reticulum Ca2+ release in vascular muscl e cells are not well defined. The purpose of the present study was to evaluate the effects of reactive oxygen intermediates generated from t he hypoxanthine/xanthine oxidase reaction system on contractions induc ed by caffeine, Ins(1,4,5)P, and norepinephrine in staphylococcal oc-t oxin-permeabilized rabbit mesenteric arteries. This system generates O -2(.-), H2O2, and hydroxyl radicals. We wished to identify which class of reactive oxygen intermediates is responsible for the associated lo ss of vascular smooth muscle contractile function. Caffeine and Ins(1, 4,5)P, produced a transient contraction when the sarcoplasmic reticulu m of the permeabilized preparations was preloaded with pCa 7.0 solutio n for 5 min before washing with 0.5 mM EGTA solution; norepinephrine a lso produced a transient contraction. Exposure of the preparations to hypoxanthine/xanthine oxidase (for 30 min) attenuated caffeine-induced contraction, but was without effect on Ins(1,4,5)P-3-induced contract ion. The observed effect of hypoxanthine/xanthine oxidase exposure was superoxide dismutase-inhibitable, suggesting O-2(.-) involvement. Hyp oxanthine/xanthine oxidase also inhibited norepinephrine-induced contr action. The effect of hypoxanthine/xanthine oxidase on norepinephrine contraction was protected by catalase, but not by superoxide dismutase and dimethyl sulfoxide; exogenously added H2O2 mimicked the effect of hypoxanthine/xanthine oxidase exposure. H2O2, added exogenously, was without effect on Ins(1,4,5)P-3-induced contraction. It is suggested t hat the pathway of Ca2+ release from the sarcoplasmic reticulum depend ent on Ins(1,4,5)P-3 is insensitive to O-2(.-). Instead, caffeine-indu ced Ca2+ release mechanisms may be susceptible to O-2(.-) and H2O2, ra ther than O-2(.-) and hydroxyl radicals, may be the active agent in th e norepinephrine-induced contraction. Our results are also consistent with the view that the attenuation by H2O2 of the norepinephrine-induc ed contraction may be linked to the receptor-associated pathway of Ins (1,4,5)P-3 formation, but not to degradation processes of Ins(1,4,5)P- 3.