In the mammalian brain, thyroid hormones regulate myelination. Their actions are mediated by interactions with nuclear receptors that function as ligand-regulated transcription factors. Two genes, alpha and beta, encode different isoforms, of which only the beta and alpha1 isoforms are authentic nuclear triiodothyronine (T3)-receptors (NT3R). In agreement with the important role of T3 on myelination and oligodendrocyte generation, the presence of NT3Rs has been reported in oligodendrocytes and their precursors. We and others have shown that both progenitors and oligodendrocytes in vitro express the alpha1 and alpha2 isoforms, but the expression of the beta1 isoform is confined to differentiated oligodendrocytes, suggesting that they have different functions. To establish if this is the case during development in vivo, we have studied NT3R isoform expression in glial cells isolated by density gradient centrifugation from rat brains of various ages. We report the presence of the alpha1 NT3R and its variant alpha2, but not that of the beta1 isoform, in newborn rat glial progenitors. The pattern of expression of beta1, both at the level of mRNA and protein, parallels the increase in the number of oligodendrocytes. We found a significant change in the kinetic parameters of [125I]-T3 binding to NT3Rs in these cells during the first month of life, consisting of an increase in the binding capacity that peaks with myelination, and a significative decrease in Kd that coincides with the switch from the alpha to the beta1 isoform. Thus, the expression of NT3R isoforms in the rat oligodendrocyte lineage changes radically from the alpha to the beta1 isoform during the period when oligodendrocytes differentiate from progenitors.