Cytosolic-free Ca2 and cell killing in hepatoma lclc7 cells exposed to chemical anoxia

Exposure of cultured hepatoma lclc7 cells to KCN and iodoacetate, to produce chemical anoxia, caused a rapid and sustained increasein cytosolic-free Ca" con- centration, which was associated with depletion of in- tracellular ATP and glutathione. These changes oc- curred before the loss of cell viability and were accompanied by the appearance of plasma membrane blebs. Pretreatment of the cells with the Ca" chelators Quin 2 or BAPTA markedly delayed both the onset of Although some studies support this hypothesis (6-8), others have questioned the involvement of Ca" in anoxic cell injury (9, 10). A recent study has reported that exposure of cultured hepatocytes to chemical anoxia caused by KCN and iodoacetate resulted in surface blebbing and cell death without measurable changes in cytosolic Ca" (11). This has stimulated a debate regard- ing whether or not an elevation of cytosolic Ca" is in- volved in cell killing in toxic and anoxic injury. The present study was designed to investigate the role of Ca" in the toxicity of KCN and iodoacetate in cultured hepatoma lclc7 cells. The hepatoma cells, which have retained most of the properties of normal liver cells (12, 13), were found to be a suitable model to measure changes in cytosolic Ca" with the use of the fluorescent indicator Fura 2.2 Exposure of hepatoma lclc7 cells to KCN and iodoacetate caused a rapid and sustained increase in cytosolic Ca", accompanied by a depletion of intracellular ATP and GSH, which oc- curred before the appearance of surface blebs and the loss of cell viability. Cytotoxicity was markedly delayed when KCN- and iodoacetate-treated cells were pre- loaded with the intracellular Ca" chelators Quin 2 or BAPTA to buffer changes in cytosolic Ca". Our results suggest that a sustained elevation of cytosolic-free Ca" is directly involved in cell killing by chemical anoxia.