Sphingosine-1-phosphate (S1P) and its receptors are important in nervous system development. Reliable in vitro human model systems are needed to further define specific roles for S1P signaling in neural development. We have recently reported that human embryonic stem cell-derived neuroepithelial progenitor cells (hES-NEP) express functional S1P receptors. These cells can be further differentiated to a neuronal cell type, and therefore represent a good model system to study the role of S1P signaling in human neural development. The following sections describe in detail the culture of hES-NEP cells and two assays to measure S1P signaling in these cells.