Sphingosine-1-phosphate induces apoptosis of cultured hippocampal neurons that requires protein phosphatases and activator protein-1 complexes

In this study, we report that mobilization of internal Ca2+ by sphingosine- 1-phosphate, a metabolite of ceramide, induces apoptosis in cultured hippoc ampal neurons. This sphingosine 1-phosphate-induced apoptosis is dependent upon the activation of protein phosphatases, possibly calcineurin and phosp hatase 2A (or a related phosphatase). In addition, pretreatment of neurons with double-stranded oligonucleotides containing the metallothionein phorbo l-12-myristate-13-acetate response element sequence as transcription factor decoys suppressed apoptosis. In contrast, double-stranded oligonucleotides containing either the c-jun or SV40 phorbol 12-myristate-13-acetate respon se element sequences were ineffective. Electrophoretic mobility shift assay s and supershift assays revealed that c-Fos-containing activator protein-1 complexes preferentially bound the metallothionein phorbol-12-myristate-13- acetate response element sequence-containing oligonucleotides. Furthermore, antisense oligonucleotides to c-fos and c-jun were also protective. The apoptotic death of hippocampal neurons has been hypothesized to contrib ute to the cognitive impairments observed following insults to the brain. W hile increases in intracellular calcium are thought to be key mediators of neuronal apoptosis, the biochemical cascade(s) activated as a result of inc reased Ca2+ which mediates apoptosis of hippocampal neurons is (are) not we ll understood. The findings presented in this study suggest that mobilizati on of internal calcium via prolonged exposure of sphingosine-1-phosphate in duces apoptosis of hippocampal neurons in culture. Sustained increases in i ntracellular calcium activate a phosphatase cascade that includes calcineur in and a phosphatase 2A-like phosphatase, and leads to the expression of ge nes containing metallothionein phorbol-12-myristate-13-acetate response ele ment (TGAGTCA)-type enhancer sequences. The expression of genes containing TGAGTCA-type enhancer sequences appears to be essential for sphingosine-1-p hosphate-induced apoptosis of hippocampal neurons. (C) 1999 IBRO. Published by Elsevier Science Ltd.