Ion dependence of cytotoxicity of carmustine against PC12 cells

Cytotoxicity is a major complication of carmustine (1,3-bis(2-chloroethyl)-1-nitrosourea, BCNU) therapy for treatment of brain tumors and lymphomas. Using the lactate dehydrogenase (LDH) cell death assay in PC12 cells, we studied the role in this phenomenon of transmembrane ion fluxes that could be activated following inhibition by carmustine of glutathione reductase. The cytotoxic effect of carmustine developed during 4-6 h, with the EC50 of 27 microM. It depended on the extracellular Ca2+ concentration and substantially decreased upon Ca2+ removal. An almost complete suppression of toxicity was achieved when, additionally, monovalent cations were also replaced with impermeant organic cations. A similar loss of toxicity occurred in the presence of Ca2+ when extracellular Cl- was replaced with impermeable gluconate. Various blockers of cation and Cl- channels, as well as antioxidants also protected cells from carmustine. We conclude that carmustine toxicity against PC12 cells requires an influx of Ca2+ ions, supposedly through redox-sensitive cation channels.