Role of Cu/Zn-SOD in the Synthesis of Endothelium-Derived Hyperpolarizing Factor during Reactive Hyperemia in Mouse Mesenteric Microvessels in Vivo

Abstract

Background: We have recently identified that endothelium-derived hydrogen peroxide (H2O2) is an endothelium-derived hyperpolarlizing factor (EDHF) both in vitro and in vivo and that endothelial Cu/Zn-superoxide dismutase (SOD) plays an important role as an EDHF synthase in vitro. In this study, we examined the possible role of Cu/Zn-SOD in the synthesis of EDHF/ H2O2 during reactive hyperemia in mouse mesenteric microcirculation in vivo. Methods: Mesenteric arterioles in wild-type and Cu/Zn-SOD-/- mice (n=6 each, 20–60 µm in diameter) were continuously observed by a newly developed pencil-typed intravital microscope during acetylcholine (ACh)-induced vasodilatation and reactive hyperemia (RH, reperfusion after 20 sec of mesenteric artery occlusion) under cyclooxygenase blockade (indomethacin, 5×10−5 M, topically administration, TA) under the following 3 conditions; control, NO synthase inhibitor (L-NMMA, 10−4 M, TA) and LNMMA+catalase (PEG-catalase 10,000 U/kg, a specific decomposer of H2O2, intra-arterial administration). Results: In mesenteric arterioles of wild-type mice (n=6), endothelium-dependent relaxations to acetylcholine and vasodilatation during RH were totally resistant to L-NMMA, but were markedly inhibited by L-NMMA+catalase. In mesenteric arterioles of Cu/Zn-SOD-/- mice (n=6), L-NMMA markedly inhibited the relaxations to ACh and vasodilatation during RH. The responses were not inhibited by the addition of catalase. By contrast, endothelium-independent relaxations to sodium nitroprusside (10−4 M, TA), a direct NO donor, were comparable between the two strains in the presence of LNMMA+catalase. Conclusions: Endothelial Cu/Zn-SOD plays an important role as an EDHF synthase during reactive hyperemia in mouse mesenteric microcirculation in vivo.