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 c ellular proliferation, differentiation, and apoptosis. Nerve growth fa ctor (NGF), a neurotrophic factor for pheochromocytoma PC12 cells, ind uced a biphasic increase in the activity of sphingosine kinase, the en zyme that catalyzes the formation of SPP. This activation was blocked by K252a, an inhibitor of tyrosine kinase A (trkA). A rapid 1.7-fold i ncrease was followed by a marked prolonged increase reaching a maximum of fourfold to fivefold stimulation with a concomitant increase in SP P 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 t he elevation of ceramide induced by serum withdrawal. Treatment of PC1 2 cells with SPP did not induce neurite outgrowth or neurofilament exp ression, yet it enhanced neurofilament expression elicited by suboptim al doses of NGF. Moreover, SPP also protected PC12 cells from apoptosi s induced by serum withdrawal. To further substantiate a role for SPP in the cytoprotective actions of NGF, we found that N,N-dimethylsphing osine, a competitive inhibitor of sphingosine kinase, also induced apo ptosis and interfered with the survival effect of NGF. These effects w ere counteracted by exogenous SPP. Moreover, other structurally relate d compounds, such as dihydrosphingosine I-phosphate and lysophosphatid ic acid, had no significant protective effects. Our results suggest th at activation of sphingosine kinase and subsequent formation of SPP ma y play an important role in the differentiation and survival effects i nduced by NGF.