Several laboratories have shown that cyclic AMP (cAMP) plays an important role in inducing oligodendrocyte differentiation and myelin synthesis. Our previous results have shown that oligodendrocytes contain a nuclear protein that binds to the DNA sequence TGACGTCA or cAMP response element (CRE) known to be involved in the transcriptional regulation of cAMP-responsive genes. In this report the oligodendroglial CRE-binding protein was further identified by using two different antibodies which specifically recognize the CRE-binding protein known as CREB. In DNA-shift assays CREB-1 (X-12) antibody interacted with the CRE-protein complexes resulting in further retardation (“super shift”) of the mobility of the bands in the gels. Immunoprecipitation of oligodendroglial nuclear extracts with CREB(240) antibody prior to the DNA binding assays resulted in a lack of formation of CRE-protein complexes. In addition immunoreaction with CREB(240) antibody identified the CRE-binding species as a 45 kDa phosphoprotein. Immunocytochemical staining with CREB(240) antibody in oligodendrocytes from 10-, 14-, and 18-day-old and adult rats indicated that this protein in expressed before the appearance of myelin basic protein (MBP) which was used as a marker of myelin synthesis. Collectively, these observations support our previous results and indicate that the oligodendroglial CRE-binding protein species is highly homologous to the CREB protein. The developmental expression of this CREB protein supports the idea of a possible role during the early stages of oligodendrocyte differentiation preceding the peak of myelin synthesis in rat CNS. © 1994 Wiley-Liss, Inc.