Studies on chorion hardening inhibition and dechorionization in turbot embryos

Embryo dechorionization is a common practice used in certain fish species for different purposes. It facilitates techniques like microinjection, transfection or electroporation in embryos. Dechorionization is easily achieved in some fish species but is a more complex problem in species that have very thick chorions. In this study, we address this problem in turbot embryos, where chorion removal is practically unachievable post-chorion hardening. For this purpose, different solutions that lacked ions required for the hardening of this envelope or contained inhibitors of enzymes involved in the process were used during egg fertilization. The toxicity of the solutions was assessed, and their effect on embryo cleavage and on chorion structure was studied by light and scanning electron microscopy (SEM). The results demonstrated that embryos are very sensitive to these solutions and that first cellular cleavages are affected with most of them. This study also provides the first report on turbot chorion structure, analyzed by SEM. The chorion is a very thick envelope in this species, and its total removal was not observed with the employed treatments. Nevertheless, partial dechorionization was achieved when embryos were fertilized in some of the tested solutions and later treated with pronase (3 mg/ml).