Vol. 61, Issue 3, 486-494, March 2002

Distinct Effects of Different Calcium-Mobilizing Agents on Cell Death in NG108-15 Neuroblastoma X Glioma Cells

Ting-Yu Chin, Hsiou-Min Hwang, and Sheau-Huei Chueh

Department of Biochemistry, National Defense Medical Center, Taipei, Taiwan, Republic of China

The effects of different calcium-mobilizing agents on cell death were characterized in NG108-15 neuroblastoma x glioma hybrid cells. Carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP) increased the cytosolic Ca²⁺ concentration ([Ca²⁺]_i) and caused cell death. Thapsigargin (TG) not only increased the [Ca²⁺]_i and caused cell death but also induced neurite outgrowth via activation of phospholipase A₂ and cytochrome P450 epoxygenase. In contrast, bradykinin increased the [Ca²⁺]_i, but had no effect on cell morphology or cell death. Cell death occurred by two different mechanisms, one of which was caspase-3-dependent and the other caspase-3-independent. Caspase-3 activation was Ca²⁺-dependent, whereas neurite outgrowth was Ca²⁺-independent. TG- or FCCP-induced caspase-3 activation occurred at the same time, but the cell death induced by TG was delayed. TG treatment did not enhance the generation of nitric oxide or cAMP or secretion of glial-derived neurotrophic factor or neurotrophin-3, but activated sphingosine kinase. Furthermore, inhibition of sphingosine kinase accelerated TG-induced cell death, and exogenous sphingosine 1-phosphate (S1P) protected cells from FCCP-induced cell death by about 60%. These results indicate that, in these cells, depletion of intracellular nonmitochondrial or mitochondrial Ca²⁺ stores causes cell death, that TG activates phospholipase A₂ and sphingosine kinase, and that arachidonic acid induces neurite outgrowth, whereas S1P delays cell death.