Ca2+ is mobilized by hydroxyl radical but not by superoxide in RTH-149 cells:: The oxidative switching-on of Ca2+ signaling

Differential effects of superoxide and hydroxyl radical on intracellular calcium were investigated in trout hepatoma cells (RTH-149). [Ca2+](i) variations were recorded using confocal imaging, fluo-3 loading, and exposure to various mixtures consisting of hypoxanthine/xanthine oxidase (HX/XO), and of sub-stimulatory concentrations of H2O2 and Cu2+. No [Ca2+](i) variation was found with HX/XO, a slight [Ca2+](i) rise with a mixture of Cu2+ and HX/XO, a sustained rise with Cu2+ and H2O2, and the highest rise with Cu2+, H2O2 and HX/XO. Fluorimetric assay using dihydrorhodamine 123 revealed a correlation between the oxidizing power of a mixture and its effect on [Ca2+](i). The [Ca2+](i) rise induced by Cu2+, H2O2 and HX/XO, was partially reduced in Ca2+ free medium or in the presence of SOD, converted into Ca2+ transient by verapamil, and almost abolished by the PLC inhibitor U73122 or in the presence of the hydroxyl radical quencher TEMPOL. Data indicate that Ca2+ is mobilized by hydroxyl radical but not by superoxide. The mechanism consists of PLC activation causing intracellular Ca2+ release, while Ca2+ entry potentiates Ca2+ release thus leading to sustained [Ca2+](i) rise. A role of hydroxyl radicals in the oxidative switching-on of Ca2+ signaling is discussed. (c) 2005 Elsevier Ltd. All rights reserved.