Early embryonic development in pikeperch (Sander lucioperca) related to micromanipulation

Recently, transplantation of germ cells has attracted attention as a potential technique for efficient reproduction of fish. One of the well-proven techniques to deliver donor germ cells into a recipient is the transplantation of primordial germ cells (PGCs) during the blastula stage. Nevertheless, the application of such techniques so far has been limited to model fish species such as zebrafish, due to the lack of information about early development in many fish species. We propose that pikeperch (Sander lucioperca) can be a useful model species for establishing this technique in the order Perciformes, which includes commercially and ecologically important marine species. In this study, we described the important events, namely, embryonic staging, yolk syncytial layer (YSL) formation, and midblastula transition (MBT) during the blastula stage in pikeperch to obtain basic information about early embryonic development. The chorion was softened by treating with 0.2% trypsin and 0.4% urea in Ringer's solution so as to remove it easily by forceps. Although the first cleavage occurred at about 2.5 h post fertilization, blastomeres divided approximately every one hour after this at 15 degrees C. The YSL was formed after the breakdown of marginal cells during the 512- to 1k-cell stage. Cell division analysis by 4'-6-diaminido-2-phenylindole (DAPI) staining revealed that transition from synchronous to asynchronous division occurred after the 10th cleavage (1k-cell stage). Our results indicate that zygotic gene expression (MBT) starts after this stage. Next, we performed blastodisc isolation assay to find the competent stage for embryonic manipulation. Embryos were manipulated by using a microneedle every hour from the 512-cell to the sphere stage, and then developmental rates were evaluated at the hatching stage. The highest survival rate was obtained when we performed this manipulation at the 1k-cell stage. These results clearly showed that the MBT is the best stage for transplantation of PGCs or any cells in pikeperch.