AN IMMORTALIZED MOUSE NEUROEPITHELIAL CELL-LINE WITH NEURONAL AND GLIAL PHENOTYPES

Evidence from retroviral marking techniques and immortalized cell line s indicates that multipotential stem cells exist in many areas of the developing central nervous system. However, the factors that influence the commitment of these stem cells into distinct neuronal or glial li neages are not known. We have created an immortalized hypothalamic cel l line derived from embryonic day 14 hypothalamic cells with a replica tion-defective retroviral construct containing a temperature-sensitive allele (tsA58) of the large T antigen of the simian virus 40. The clo nality of this cell line, which we have named V1, was established by s ingle cell cloning and by Southern blot analysis. V1 cells exhibit two different morphologies: the vast majority of cells are flat and stell ate, and a smaller number are phase-bright round cells with processes. V1 cells express nestin and neural-cell adhesion molecule, typical of proliferating neuroepithelial cells. They also express glial fibrilla ry acidic protein and S100 as well as the low molecular weight neurofi lament protein. In addition, the phase-bright, process-bearing V1 cell s stain intensely for many typical neuronal proteins, such as low, med ium and high molecular weight neurofilament proteins, tau protein, mic rotubule-associated protein-2, and neuron-specific enolase. The phase- bright cells also have condensed chromatin and display mitotic spindle s, indicating that they are in mitosis. When V1 cells are transferred from the permissive temperature (33 degrees C) to the restrictive temp erature (39 degrees C), there is a decrease in expression of NF-L and an increase in expression of NF-H and glial fibrillary acidic protein in the flat V1 cells. The enhanced expression of neuronal antigens in mitotically active V1 cells is novel and may represent a more general property of the differentiation process. We suggest that V1 cells aris e from a mixed neural/glial neuroepithelial progenitor cell that expre sses both neuronal- and glial-specific proteins in the developing hypo thalamus.