Sphingosine-1-phosphate, a putative second messenger, mobilizes calcium from internal stores via an inositol trisphosphate-independent pathway.

Sphingosine-1-phosphate, a metabolite of sphingolipids which has previously been shown to stimulate DNA synthesis and cell division in quiescent cultures of Swiss 3T3 fibroblasts (Zhang, H., Desai, N. N., Olivera, A., Seki, T., Brooker, G., and Spiegel, S. (1991) J. Cell Biol. 114, 155-167), induced a transient increase in intracellular free calcium independent of extracellular calcium. The increase in calcium was completely abolished when intracellular calcium pools were depleted with thapsigargin, an inhibitor of the endoplasmic reticulum Ca(2+)-ATPase. The dose-response for calcium release induced by sphingosine-1-phosphate correlated closely with the concentration required for stimulation of DNA synthesis. The magnitude of the calcium response decreased with successive challenges, although sphingosine-1-phosphate did not attenuate the responses to either bradykinin or ionomycin. Conversely, prior stimulation of the cells with bradykinin had no effect on the sphingosine-1-phosphate-induced calcium signal. Although sphingosine-1-phosphate increased inositol (1,4,5)-trisphosphate levels, complete inhibition of inositol phosphate formation by pretreatment with 12-O-tetradecanoylphorbol-13-acetate did not block sphingosine-1-phosphate-mediated calcium responses. Moreover, in permeabilized cells, heparin, an inositol (1,4,5)-trisphosphate antagonist, blocked Ca2+ release induced by inositol (1,4,5)-trisphosphate, but did not significantly alter the Ca2+ release induced by sphingosine-1-phosphate. Sphingosine-1-phosphate did not stimulate the release of arachidonic acid, another signaling molecule known to elevate [Ca2+]i without inositol lipid turnover or calcium influx. Our data suggest that sphingosine-1-phosphate mobilizes Ca2+ from internal stores primarily through a mechanism independent of inositol lipid hydrolysis and arachidonic acid release and that sphingolipid metabolism may be important in calcium homeostasis.