The glutathione reductase inhibitor cartmustine induces an influx of Ca2+ in PC12 cells

We studied the effects of carmustine (1,3-bis(2-chloroethyl)-1-nitrosourea) on the intracellular Ca2+ concentration ([Ca2+](i)) in PC12 cells using fura-2 fluorescence imaging. Carmustine (100 muM) caused a delayed increase in [Ca2+](i) that developed within similar to3 h. This effect was enhanced in cells that were pretreated with an inhibitor of glutathione (GSH) synthesis, buthionine sulfoximine (BSO, 200 muM, 24 h), and was suppressed in cells that were treated with an antioxidant deferoxamine (50 muM). The carmustine-induced increase in [Ca2+](i) was absolutely dependent on the presence of extracellular Ca2+ and could be inhibited by dihydropyridine blockers of L-type voltage-gated Ca2+ channels (nimodipine or nitrendipine, 10 muM). The increase in [Ca2+](i) was also suppressed in Cl--free solution and in the presence of the Cl- channel blockers, indanyloxyacetic acid 94 (IAA-94, 100 muM) and 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB, 100 muM). The inhibition was complete when the blockers were applied simultaneously with carmustine and was partial when the blockers were applied after the initial increase in [Ca2+](i). We conclude that carmustine induces an influx of extracellular Ca2+ through L-type Ca2+ channels and that this effect is mediated by oxidative stress that results from the depletion of GSH following the inhibition by carmustine of glutathione reductase. (C) 2004 Elsevier B.V. All rights reserved.