Critical Role of Phospholipase C gamma 1 in the Generation of H sub(2)O sub(2)-evoked [Ca super(2+)] sub(i) Oscillations in Cultured Rat Cortical Astrocytes

• Published in: The Journal of biological chemistry 2006-05, Vol.281 (19), p.13057-13067
• Main Authors: Hong, Jeong Hee, Moon, Seok Jun, Byun, Hae Mi, Kim, Min Seuk, Jo, Hae, Bae, Yun Soo, Lee, Syng-Ill, Bootman, Martin D, Roderick, HLlewelyn, Shin, Dong Min, Seo, Jeong Taeg
• Format: Article
• Language: English
• ISSN: 0021-9258; 1083-351X

Reactive oxygen species, such as the superoxide anion, H sub(2)O sub(2), and the hydroxyl radical, have been considered as cytotoxic by-products of cellular metabolism. However, recent studies have provided evidence that H sub(2)O sub(2) serves as a signaling molecule modulating various physiological functions. Here we investigated the effect of H sub(2)O sub(2) on the regulation of intracellular Ca super(2+) signaling in rat cortical astrocytes. H sub(2)O sub(2) triggered the generation of oscillations of intracellular Ca super(2+) concentration ([Ca super(2+)] sub(i)) in a concentration-dependent manner over the range 10-100 mu M. The H sub(2)O sub(2)-induced [Ca super(2+)] sub(i) oscillations persisted in the absence of extracellular Ca super(2+) and were prevented by depletion of intracellular Ca super(2+) stores with thapsigargin. The H sub(2)O sub(2)-induced [Ca super(2+)] sub(i) oscillations were not inhibited by pretreatment with ryanodine but were prevented by 2-aminoethoxydiphenyl borate and caffeine, known antagonists of inositol 1,4,5-trisphosphate receptors. H sub(2)O sub(2) activated phospholipase C (PLC) gamma 1 in a dose-dependent manner, and U73122, an inhibitor of PLC, completely abolished the H sub(2)O sub(2)-induced [Ca super(2+)] sub(i) oscillations. In addition, RNA interference against PLC gamma 1 and the expression of the inositol 1,4,5-trisphosphate-sequestering "sponge" prevented the generation of [Ca super(2+)] sub(i) oscillations. H sub(2)O sub(2)-induced [Ca super(2+)] sub(i) oscillations and PLC gamma 1 phosphorylation were inhibited by pretreatment with dithiothreitol, a sulfhydryl-reducing agent. Finally, epidermal growth factor induced H sub(2)O sub(2) production, PLC gamma 1 activation, and [Ca super(2+)] sub(i) increases, which were attenuated by N-acetylcysteine and diphenyleneiodonium and by the overexpression of peroxiredoxin type II. Therefore, we conclude that low concentrations of exogenously applied H sub(2)O sub(2) generate [Ca super(2+)] sub(i) oscillations by activating PLC gamma 1 through sulfhydryl oxidation-dependent mechanisms. Furthermore, we show that this mechanism underlies the modulatory effect of endogenously produced H sub(2)O sub(2) on epidermal growth factor-induced Ca super(2+) signaling in rat cortical astrocytes.