Differentiation in vitro of mouse embryos to the stage of early somite

Abstract

Mouse blastocysts continuously differentiate in vitro to the early somite stage with reconstituted rat tail collagen as the substrate for the attachment. In order for this to occur, it appears that two differentiation barriers must be overcome. The first, the formation of egg cylinders from the inner cell mass, can be overcome by incubating embryos in heat-inactivated fetal calf serum. The second, the formation of the early somite from the presomite stage, can be overcome by replacing fetal calf serum with human cord serum. Mouse blastocysts were initially incubated with calf serum in Eagle's minimum essential medium. After shedding the zona pellucida, the denuded blastocysts lay flat on the surface of the collagen. Soon thereafter, trophoblastic cells invaded the underlying collagen leaving the rounded inner cell mass protruding from the surface of the collagen. By replacing calf serum in the medium with fetal calf serum the inner cell mass differentiated into endoderm and ectoderm to form an egg cylinder. The egg cylinder rapidly became elongated and formed extraembryonic and embryonic regions. However, the embryonic region shrank from this point on in the fetal calf serum, and the resulting yolk sac formation did not contain the embryo proper. When fetal calf serum was replaced with human cord serum at the end of the egg cylinder stage (equivalent to embryos of about 7.5 days gestation) neural tissue, cardiac chambers, and somites were formed.