Sphingosine-1-phosphate induces a Ca2+ signal in primary rat astrocytes and a Ca2+ signal and shape changes in C6 rat glioma cells

Abstract

Treatment of rat glioma C6 cells with the beta-adrenergic agonist L-isoproterenol leads to a rise in cAMP level and a subsequent change in cell morphology from an epithelial to an astrocytic type of appearance. This morphological response is reverted by the addition of sphingosine-1-phosphate (S1P) with an EC50 of 10 nM. In rat glioma C6 cells loaded with the Ca2+ indicator Fura-2, S1P evoked Ca2+ release from internal stores and Ca2+ influx from the external medium. Half-maximal stimulation of the Ca2+ increase was 10-20 nM. A similar Ca2+ signal was observed in primary rat astrocytes loaded with the Ca2+ indicator fluo-3. Pretreatment of the C6 cells with PMA (162 nM) prevented both the S1P-induced Ca2+ increase and the morphological reversion. Ca2+ ions therefore seem essential for the morphological reversion by S1P. Pretreatment of the cells with the Clostridium botulinum C3 exoenzyme did not affect the reversion of the morphological response by S1P, indicating that the small GTP-binding protein Rho is not involved in the S1P-induced reversion.