The mouse neural plate as starting material for studying neuronal differentiation in vitro

Tissue from the mouse neural plate and neural tube was studied, by light and electron microscopy, as starting material for tissue culture. In vivo, up to embryonic day 9 (E 9, stage Th 14; Theiler 1972) all neuroepithelial cells of the neural plate were mitotically active. As judged from their light microscopic or ultrastructural appearance, they could hardly be distinguished from one another or from neuroepithelial cells of more mature embryos. The earliest few immature neurons in the mesencephalic anlage were discernible on day 9 1/2 (stage Th 15) in the prospective intermediate layer of the neural tube, concomitantly with the development of processes containing neurotubules and vesicles which were oriented in parallel to the basal lamina. For tissue culture, explants of the mesencephalic anlage of embryonic days 8 (Th 12/13), 9 1/2 (Th 15), and 11 (Th 18) were kept in vitro and their development was compared with each other and with the corresponding developmental stage in vivo in the initial phase of culture (e.g., E 8, day of explanation, kept in vitro for 2 days, E 10 in vivo being the stage for comparison). The study demonstrated that further in vitro development proceeded in an accelerated manner, independent of the developmental stage of the embryo from which the tissue was explanted. In vitro, proliferation of the explanted neuronal progenitor cells stopped in all explants within 24 h of culture as revealed by autoradiographic and electron microscopic techniques. Cytoplasmic transformation was observed corresponding to that found in vivo, but always greatly accelerated. Earliest axons had formed after 24 h in vitro; synapses with clear vesicles and dense core vesicles were observed after at least 3 days in culture in all explants regardless of age at the time of explantation (E 8 or E 11). The present ultrastructural results indicate that prospective neurons within the neuroepithelium of the neural plate and early neural tube were immediately able to develop into neurons without the complete sequence of mitotic events normally occurring under in vivo conditions.