Maturation of cerebellar granule cells is delayed in cultures derived from ethanol-treated chick embryos: survival and proliferation studies.

Previous studies from this laboratory have shown that ethanol administration to chick embryos during embryonic days 1-3, a critical period of neuroembryogenesis, differentially affects primordial CNS structures. In this study, chick embryos were treated in ovo with ethanol (10 mg/50 microliter/day) at E1 to E3. At 14 days of embryonic age cerebellar (E14CE) granule cell cultures were prepared from both control and ethanol-treated embryos. Growth patterns were evaluated morphologically and the neuronal nature of these cultures was evaluated immunocytochemically. E14CE granule cell cultures exhibited neurofilament immunoreactivity demonstrating the neuronal-nature of these cultures. In addition E14CE granule cultures contained numerous glutamatergic neurons as assessed by positive glutamate immunoreactivity and also some GABAergic neurons as assessed by positive GABA immunoreactivity. Cultures derived from both control and ethanol-treated embryos were labeled with 3H-thymidine and assessed for effects on survival and proliferation in culture. Cultures derived from ethanol-treated embryos showed a higher rate of proliferation and survival during the first 3 days in culture as compared to those derived from controls. However, after 3 days in culture, survival was lower in the cultures from ethanol-treated embryos as compared to those derived from controls. We interpret these findings to mean that (a) ethanol arrested cerebellar granule cell development at an immature state; (b) immature neurons have a higher survival capacity than differentiated neurons; and (c) ethanol accelerates normal neuronal cell death as previously reported.