Monitoring of cell viability in suspensions of embryonic CNS tissue and its use as a criterion for intracerebral graft survival.

Neuronal cell suspensions, prepared by trypsination and mechanical disruption from embryonic CNS tissue, are currently used for intracerebral neuronal grafting to deep brain sites. In the present study the viability of suspended neurons from different brain regions has been monitored with a fluorescent vital stain, and studied as a function of time after dissociation and age of the donor rat embryos. Subsequently, the validity of the in vitro viability rates as a criterion for the in vivo survivability of each individual suspension was tested for suspensions prepared from the developing mesencephalon, rich in dopamine-containing neurons. The results indicate that mechanically dissociated embryonic CNS neurons remain viable for several hours in a simple glucose-saline solution at room temperature. The in vitro viability scores declined faster in suspensions prepared from mesencephalon than in those prepared from telencephalon (striatum and basal forebrain), and they declined faster in suspensions prepared from older embryos. The fetal cells were sensitive to the mechanical trauma caused by excessive pipetting, and tissue from older embryos seemed generally more vulnerable in the trypsination-dissociation procedure. The grafting experiments showed a good correlation between the in vitro cell viability counts and in vivo neuronal survival after grafting, indicating that the vital stain, at least under certain conditions, can be used as a simple and practical routine test to check and standardize cell suspensions to be used in intracerebral grafting experiments.