The AA dimeric form of platelet-derived growth factor (PDGF-AA) is implicated in the differentiation of cells of the oligodendrocyte lineage, which express PDGF receptors of the alpha subunit type (PDGF-alphaR). In the present study, we show that a single injection of PDGF-AA into the cerebrospinal fluid of neonatal rats delays oligodendrocyte differentiation and interrupts the progress of myelination in the anterior medullary velum (AMV), a white matter tract roofing the IVth ventricle of the brain. PDGF-AA or saline was injected intrathecally in postnatal day (P) 7 rats, and the AMV was subsequently removed and immunolabelled with the oligodendrocyte-specific antibody Rip, at P9, P12, and P21, corresponding to postinjection days (PID) 2, 5, and 14. At P9 (PID2), myelination was retarded in PDGF-AA-treated rats as opposed to saline-treated controls but progressed rapidly after P12 (PID5). Quantification supported the qualitative observations that PDGF-AA mediated an acute decrease in the number of Rip+ oligodendrocytes at P9-12, which largely recovered by P21, suggesting that PDGF-AA may have delayed recruitment of myelinating oligodendrocytes. However, the definitive number of Rip+ oligodendrocytes in the AMV was not increased, suggesting that its action as a promoter of early oligodendrocyte survival may not ultimately affect the definitive number of myelinating oliogdendrocytes in vivo. We discuss the possibilities that excess PDGF-AA may have acted on early oligodendrocytes (precursors or preoligodendrocytes) to either (1) delay their differentiation by maintaining them in the cell cycle or (2) accelerate their differentiation, which may result in premature cell death in the absence of synchronised survival signals. This study supports a role for PDGF-AA in the timing of oligodendrocyte differentiation in vivo, as has been shown in vitro.