Studies on the isolation and propagation of multipotent neural precursors as neurospheres suggest their potential use in the reconstitution of neurons and oligodendrocytes in neurodegenerative diseases. To ensure that an adequate number of functionally relevant cells are present after transplantation, in vitro manipulation of cell fate before transplantation may be necessary to control the terminal phenotype of these cells. Using growth factors known to have a role in oligodendrocyte development such as sonic hedgehog, platelet-derived growth factor (PDGF), and basic fibroblast growth factor (FGF-2), we have tried to increase the number of oligodendroglia derived from E18 cortical neurospheres. We have shown that although all of the growth factor combinations induce the formation of oligodendroglia, they do so in varying proportions, with PDGF favouring the formation of oligodendrocyte progenitor cells and sonic hedgehog favouring the formation of mature oligodendrocytes. To further enhance the generation of oligodendroglia we exposed neurospheres to B104-cell conditioned medium (B104 CM). Long-term growth of the neurospheres in this B104 CM increased markedly the number of cells committed to the oligodendrocyte lineage, specifically oligodendrocyte progenitor cells. These were then referred to as oligospheres. Our results suggest that the oligosphere culture system may provide a valuable source of cells for the reconstitution of oligodendrocytes in neurologic disorders. © 2007 Wiley-Liss, Inc.