Comparative studies on early neural-type differentiation between isolated blastomeres of protochordate embryos and teratocarcinoma cell lines derived from mouse primitive ectoderm.

如先前报道，原脊索动物Halocynthia胚胎在发育早期（1- 4细胞阶段）处于卵裂阻滞状态，并进一步培养至正常孵化期，大卵裂球的膜只表达表皮型兴奋性，而在8细胞阶段后卵裂阻滞时，一些卵裂球表现出神经型兴奋性。这种差异可能是由于卵细胞质因子的进一步分离和/或8细胞期后细胞间的相互作用造成的。在本项目中，从膜兴奋性和免疫反应性判断，在8细胞胚胎中，单个前动物卵裂球（包括假定的神经区域）在分离、卵裂阻止和溶液培养时，完全分化为表皮型。然而，当同一卵裂球与单个前植物卵裂球（包括推定的脊索区）接触时，它显示出Na尖峰，其特征是Na和…更多的K通道，并且未显示表皮抗原的表达，这表明“神经诱导”导致单个细胞与单个相邻细胞相互作用。动物胚前裂球与植物胚前裂球诱导相互作用所需的接触在接触失效前后有一个关键时期，这个时期在对照胚胎的64细胞期和原肠胚中期之间。此外，我们还寻找了一些能模拟动物前卵裂球诱导作用的物质，发现一种蛋白水解酶能产生与细胞-细胞接触诱导的钠峰完全相同的钠峰。最后研究哺乳动物胚胎系统，并与原脊索动物诱导系统进行比较，从5天小鼠胚胎的原始外胚层提取克隆畸胎癌细胞系，化学诱导其干细胞分化为神经元。神经分化早期以Na通道、t型Ca通道和160 kD神经丝蛋白在无神经突延伸的圆形细胞内表达为特征。少

As previously reported, when the Halocynthia embryo, a protochordate, is cleavage-arrested at early developmental stages, 1- to 4-cell, and cultured further until the normal hatching time, the membrane of the large blastomeres expresses exclusively the epidermal-type excitability, while when cleavage-arrested after 8-cell stage, some of blastomeres show neural-type excitability. The difference can be caused by further segregation of egg cytoplasmic factors and/or cell-cell interaction after 8-cell stage. In the present project, a single anterior-animal blastomere which includes the presumptive neural region in the 8-cell embryo, when dissociated, cleavage-arrested and cultured in solution, differentiated exclusively to epidermal type judging from membrane excitability and immunoreactivity. However, when the same blastomere was cultured in contact with a single anterior-vegetal blastomere which includes the presumptive notochordal region, it displayed Na spikes, characterized by Na and … More K channels, and showed no expression of the epidermal antigen, suggesting "neural induction" resulted in a single cell during the interaction with a single neighboring cell. The contact necessary for this inductive interaction between an anterior-animal blastomere and an anterior-vegetal blastomere had a critical period before and after which the contact became ineffective and the period was between 64-cell stage and mid-gastrula stage of the control embryo. Further, some agents which mimic the inductive effects upon the anterior-animal blastomere were looked for and a proteolytic enzyme was found to develop Na spikes exactly identical with those induced by the cell-cell contact. Finally studying the mammalian embryonic system in comparison with the protochordate inductive system, the stem cells of cloned teratocarcinoma cell line derived from the primitive ectoderm of the 5 day mouse embryo was chemically induced to differentiate into neurons. The early stage of neural differentiation was characterized by expression of Na channels, T-type Ca channels, and 160 kD neurofilament protein in a round cell without neurite extension. Less