We have been studying oligodendrocyte generation in vitro to obtain insights into how the timely generation of these cells might be regulated. Our studies suggest the existence of timing mechanisms quite different from those of existing models, wherein it is proposed that timely oligodendrocyte generation is associated with synchronous and symmetric differentiation controlled by cell-intrinsic biological clocks. Our results are most consistent with the hypothesis that the propensity of a clone of dividing oligodendrocyte type-2 astrocyte (O-2A) progenitors initially to generate at least one oligodendrocyte may be regulated by cell-intrinsic mechanisms, but that cell-extrinsic signals regulate the extent of further oligodendrocyte generation. In cultures of embryonic rat cortex grown in the presence of platelet-derived growth factor (PDGF), oligodendrocytes appeared in a timely manner in the absence of clonal differentiation. In contrast with previous suggestions, the presence or absence of thyroid hormone (T3) did not alter the probability of individual clones of O-2A progenitors generating at least one oligodendrocyte in vitro at a time equal to the rat's day of birth. Instead, T3 increased the proportion of oligodendrocytes generated within clones. For postnatally derived progenitor cells, the initial appearance of oligodendrocytes also was followed by further asymmetric generation of these cells, with the ratio of progenitors to oligodendrocytes within clones being regulated by environmental signals. T3 and ciliary neurotrophic factor increased oligodendrocyte generation, while neurotrophin-3 (NT-3) suppressed oligodendrocyte generation. Also in contrast to previous reports, NT-3 was not required for the promotion of extensive division of O-2A progenitor cells by PDGF.