Hydrogen peroxide potentiates the EDHF phenomenon by promoting endothelial Ca2+ mobilization

Objective— The purpose of this study was to test the hypothesis that H2O2 contributes to the EDHF phenomenon by mobilizing endothelial Ca2+ stores. Methods and Results— Myograph studies with rabbit iliac arteries demonstrated that EDHF-type relaxations evoked by the SERCA inhibitor cyclopiazonic acid (CPA) required activation of KCa channels and were potentiated by exogenous H2O2 and the thiol oxidant thimerosal. Preincubation with a submaximal concentration of CPA unmasked an ability of exogenous H2O2 to stimulate an EDHF-type response that was sensitive to KCa channel blockade. Imaging of cytosolic and endoplasmic reticulum [Ca2+] in rabbit aortic valve endothelial cells with Fura-2 and Mag-fluo-4 demonstrated that H2O2 and thimerosal, which sensitizes the InsP3 receptor, both enhanced CPA-evoked Ca2+ release from stores, and that the potentiating effect of H2O2 was suppressed by the cell-permeant thiol reductant glutathione monoethylester. CPA-evoked relaxations were attenuated by exogenous catalase and potentiated by the catalase inhibitor 3-aminotriazole, and were abolished by the connexin-mimetic peptide 43Gap26, which interrupts intercellular communication via gap junctions constructed from connexin 43. Conclusions— H2O2 can enhance EDHF-type relaxations by potentiating Ca2+ release from endothelial stores, probably via redox modification of the InsP3 receptor, leading to the opening of hyperpolarizing endothelial KCa channels and an electrotonically-mediated relaxant response