In non-excitable cells, thiol-oxidizing agents have been shown to evoke oscillations in cytosolic free Ca(2+) concentration (Ca(2+)) by increasing the sensitivity of the inositol 1,4,5-trisphosphate (IP(3)) receptor (IP(3)R) to IP(3). Although thiol modification of the IP(3)R is implicated in this response, the molecular nature of the modification(s) responsible for changes in channel activity is still not well understood. Diamide is a chemical oxidant that selectively converts reduced glutathione (GSH) to its disulfide (GSSG) and promotes the formation of protein–glutathione (P-SSG) mixed disulfide, i.e. glutathionylation. In the present study, we examined the effect of diamide, and the model oxidant hydrogen peroxide (H(2)O(2)), on oscillations in Ca(2+) in fura-2-loaded bovine (BAECs) and human (HAECs) aortic endo-thelial cells using time-lapse fluorescence video microscopy. In the absence of extracellular Ca(2+), acute treatment with either diamide or H(2)O(2) increased the number of BAECs exhibiting asynchronous Ca(2+) oscillations, whereas HAECs were unexpectedly resistant. Diamide pretreatment increased the sensitivity of HAECs to histamine-stimulated Ca(2+) oscillations and BAECs to bradykinin-stimulated Ca(2+) oscillations. Moreover, in both HAECs and BAECs, diamide dramatically increased both the rate and magnitude of the thapsigargin-induced Ca(2+) transient suggesting that Ca(2+)-induced Ca(2+) release (CICR) via the IP(3)R is enhanced by glutathionylation. Similar to diamide, H(2)O(2) increased the sensitivity of HAECs to both histamine and thapsigargin. Lastly, biochemical studies showed that glutathionylation of native IP(3)R(1) is increased in cells challenged with H(2)O(2). Collectively our results reveal that thiol-oxidizing agents primarily increase the sensitivity of the IP(3)R to Ca(2+), i.e. enhanced CICR, and suggest that glutathionylation may represent a fundamental mechanism for regulating IP(3)R activity during physiological redox signalling and during pathologicalical oxidative stress.