Cyclic AMP regulates PDGF-stimulated signal transduction and differentiation of an immortalized optic-nerve-derived cell line

To facilitate the study of the molecular events underlying the development of optic-nerve-derived oligodendrocytes and their growth-factor-related sig nal transduction events, we immortalized perinatal rat optic nerve cells wi th a temperature-sensitive simian virus 40 large T-antigen, carrying the ts A58 and U19 mutations, via a retrovirus vector. The line, tsU19-9, was sele cted on the basis of the expression of the neural precursor marker nestin, At the permissive temperature, 33 degrees C, tsU19-9 cells had a flat epith elial morphology, In contrast, following exposure to platelet-derived growt h factor (PDGF), a factor important in the lineage progression of oligodend rocytes, or in the presence of dibutyryl cyclic AMP at 39 degrees C (the no n-permissive temperature), the cells underwent morphological and antigenic differentiation to cells characteristic of the oligodendrocyte lineage. We used this cell line to investigate the binding characteristics of PDGF and related signalling cascades. Competition binding, phosphoinositide hydrolys is and intracellular Ca2+ mobilization assays all demonstrated that the thr ee different isoforms of PDGF (AA, AB and BB) bound to and acted on the cel l line. Overnight exposure to forskolin, a treatment that initiated morphol ogical and phenotypic progression into an oligodendrocyte lineage, decrease d PDGF-BB-induced intracellular Ca2+ mobilization and inhibited basal and P DGF-stimulated [H-3]thymidine incorporation. Our results demonstrate that t sU19-9 may serve as a resource to study early optic-nerve oligodendrocyte d evelopment.