In this report we have described several aspects of glial development in cultures containing dissociated DRG neurons and glial cells obtained from dissociated spinal cord at least a week prior to the onset of in vivo myelination. With time in culture the dissociated neurons and glia interact and become organized into 3-dimensional structures possessing many features characteristic of developing CNS in vivo. We have presented evidence that some of the glial cells proliferate and differentiate into galactocerebroside positive (GC+) cells and that some produce myelin sheaths. Thymidine was incorporated into precursors of GC+ cells, but not into cells which were already GC+. Nearly all the astrocytes in areas where neurons were present participated in the formation of large fascicles, and it was within such fascicles that most myelin was formed, suggesting a possible role for astrocytes in creating a favorable microenvironment for myelination. Both the formation of myelin sheaths and the morphological maturity of the oligodendrocytes within the fascicles indicated that oligodendrocyte differentiation proceeded practically to completion in these cultures. In conclusion we believe that the culture system herein described provides an excellent model for in vitro studies of CNS development, while retaining some of the advantages of dissociated cell cultures as well as the possibility of separating and re-uniting the various cell types of interest.