Field emission scanning electron microscopy and transmission electron microscopy studies of the chorion, plasma membrane and syncytial layers of the gastrula-stage embryo of the zebrafish Brachydanio rerio: a consideration of the structural and functional relationships with respect to cryoprotectant penetration

The structure of the chorion and plasma membranes of gastrula-stage zebrafi sh Brachydanio rerio embryos were studied using field emission scanning ele ctron microscopy (FE-SEM) and transmission electron microscopy (TEM). These studies confirm the outer chorion membrane complex to be 1.5-2.5 mu m in t hickness and to consist of three layers, electron-dense outer and innermost layers (0.2-0.3 and 1.0-1.6 mu m in thickness respectively) separated by a n electron-lucent middle layer (0.3-0.6 mu m in thickness). The middle and inner layers are pierced by pore canals. A granular to farinaceous nature o f the thin outer surface of the outer layer of the chorion has been reveale d for the first time. The study provides original TEM images of the plasma membrane structures of gastrula-stage embryos, and FE-SEM and TEM images sh owing the plasma membrane to have three morpohologically distinct regions, being prominently ridged and folded at the surface of the blastoderm, smoot h over the syncytial layer at the vegetal pole and with an intermediate reg ion between the animal and vegetal pole where folding develops in advance o f the expanding blastodermal disc of cells. FE-SEM and TEM studies reveal d etails of the syncytial layer (1-4 mu m thick) beneath the smooth plasma me mbrane at the vegetal pole, containing cytoplasmic organelles and small yol k globules. The significance of the structural detail shown in these studie s is considered in the light of the difficulties experienced in cryopreserv ation of the embryo resulting from the inability of achieving cryoprotectan t penetration of the yolk mass.