Sonic hedgehog facilitates dopamine differentiation in the presence of a mesencephalic glial cell line

The aim of this study was to establish a cellular system to investigate the requirement for cell surface and diffusible molecules in the differentiati on of fetal mesencephalic cells toward the dopamine lineage. Toward this en d, we immortalized rat embryonic day 14 (E14) mesencephalon with a regulata ble retroviral vector encoding v-myc. The stably transduced cells were pool ed and designated as VME14 cells. VME14 cells proliferated rapidly, stopped proliferating, extended processes, and expressed GFAP after suppression of the v-myc expression with tetracycline, suggesting that VME14 cells differ entiated into glial cells. Dissociated cells derived from the E11 rat mesen cephalon gave rise to only a small number of tyrosine hydroxylase (TH)-posi tive neurons. However, when grown on a monolayer of the differentiated VME1 4 cells, a significantly higher number of cells differentiated into TH-posi tive neurons. VME14 cells were transduced with the secreted N-terminal clea vage product of the Sonic hedgehog gene (SHH-N), an inducer of mesencephali c dopaminergic neurons. This monoclonal, SHH-N-overexpressing cell line fur ther enhanced dopaminergic differentiation of E11 rat mesencephalon cells. Thus, SHH-N and signals derived from fetal mesencephalic glia act cooperati vely to facilitate dopaminergic differentiation. These fetal mesencephalon- derived cell lines will provide tools for the study of signals involved in dopaminergic differentiation.