Involvement of Sphingosine 1-phosphate in Nerve Growth Factor-mediated Neuronal Survival and Differentiation

Sphingolipid metabolites, such as ceramide and sphingosine-1-phosphate (SPP), are emerging as a new class of second messengers involved in cellular proliferation, differentiation, and apoptosis. Nerve growth factor (NGF), a neurotrophic factor for pheochromocytoma PC12 cells, induced a biphasic increase in the activity of sphingosine kinase, the enzyme that catalyzes the formation of SPP. This activation was blocked by K252a, an inhibitor of tyrosine kinase A (trkA). A rapid 1.7-fold increase was followed by a marked prolonged increase reaching a maximum of fourfold to fivefold stimulation with a concomitant increase in SPP levels and a corresponding decrease in endogenous sphingosine levels. Levels of ceramide, the precursor of sphingosine, were only slightly decreased by NGF in serum-containing medium. However, NGF decreased the elevation of ceramide induced by serum withdrawal. Treatment of PC12 cells with SPP did not induce neurite outgrowth or neurofilament expression, yet it enhanced neurofilament expression elicited by suboptimal doses of NGF. Moreover, SPP also protected PC12 cells from apoptosis induced by serum withdrawal. To further substantiate a role for SPP in the cytoprotective actions of NGF, we found that N, N-dimethylsphingosine, a competitive inhibitor of sphingosine kinase, also induced apoptosis and interfered with the survival effect of NGF. These effects were counteracted by exogenous SPP. Moreover, other structurally related compounds, such as dihydrosphingosine 1-phosphate and lysophosphatidic acid, had no significant protective effects. Our results suggest that activation of sphingosine kinase and subsequent formation of SPP may play an important role in the differentiation and survival effects induced by NGF.