Generation of neurospheres from embryonic stem cells for investigating the temporal specification of neural stem/progenitor cells

This culture system, developed for the induction and selective amplification of neural stem/progenitor cells (NSPCs) from embryonic stem (ES) cells, allows the systematic characterization of their temporal specification. The use of ES cells in this system enables us to obtain a large number of NSPCs with early temporal identities and to control their regional identity by exposing them to extrinsic signals, recapitulating in vivo central nervous system (CNS) development. In this system, primary neurospheres derived from embryoid bodies (EBs) differentiate exclusively into neurons, and gliogenesis is activated in subsequent generations of neurospheres. The culture and differentiation assay take approximately 1 month to complete.