Differentiation of embryonic chick brain cells in monolayer and reaggregate cultures: A potential model in vitro for neurotoxicity.

In order to develop a model for potential neurotoxicity and teratogenicity, chick brain cells (embryonic day 7) were mechanically dissociated and cultured for up to several months. Differentiation of nerve and glial cells (monolayers in petri dishes and reaggregates in suspension cultures) were monitored with monoclonal antibodies against 68 kDa neurofilament protein (anti-NF) and glial fibrillary acidic protein (anti-GFAP). Anti-NF stains neurons in vitro as they differentiate morphologically; at day 1 many nerve processes already are feebly stained. In monolayer cultures extensive neural networks develop at day 6, which are strongly stained by anti-NF. At day 8 staining fades, days before deterioration becomes visible. In reaggregates a stable differentiation of nerve cells could be observed by intensive anti-NF staining for as long as 3 wk in culture; 5-wk-old cultures still showed substantial staining. The differentiation of GFAP-positive cells takes 3 days in vitro in monolayers as well as in reaggregate cultures. Small groups of marked cells continually increase in number; after 2 wk in culture they are present throughout the reaggregates. The differentiation of astrocytes as measured by anti-GFAP immunostaining is considerably faster in vitro than in vivo. It is suggested that the expression of NF in nerve cells could be used as a sensitive cytotoxicity endpoint, whereas GFAP expression could serve as an endpoint for monitoring differentiation of neural tissue.