In vitro maturation of African catfish (Clarias gariepinus, Burchell, 1822) oocytes results in viable larvae

In this study, we describe a protocol for in vitro maturation and ovulation of African catfish (Clarias gariepinus) postvitellogenic ovarian follicles, resulting in fertilizable and developmentally competent eggs. Such culture systems enable detailed studies of fish oocyte development and present a useful tool in assisted reproduction of fish. The maturation-inducing effects of 17 alpha,20 beta-dihydroxy-4-pregnen-3-one (DHP), human chorionic gonadotropin (hCG) and recombinant human insulin-like growth factor 1 (rhIGF-1) were evaluated based on their ability to promote ooplasm translucency and germinal vesicle breakdown (GVBD), which indicates meiosis resumption. Among the tested groups, only follicles exposed to DHP underwent GVBD in a time-dependent manner, with the highest rates (>80%) observed within 10 to 12 h of incubation. Furthermore, adding rhIGF-1 or hCG prior to or during incubations with DHP did not influence the GVBD outcome. This suggested that the oocytes acquired maturational competence prior to isolation, as high concentrations of DHP (1 mu g/ml) were alone sufficient to induce complete nuclear and cytoplasmic maturation; however, only 4 +/- 3% of these oocytes ovulated in culture. To promote ovulation, we incorporated prostaglandins (PGs) as an additional incubation step following DHP-induced maturation. Exposure to both prostaglandin F-2 alpha (PGF(2 alpha)) and prostaglandin E2 (PGE(2)) led to increased rupture of the follicular layer, most notably in groups with 5 mu g/ml PGF(2 alpha) (71 +/- 8%). In addition to identifying optimal hormonal stimulation, effects of culture media pH in combination with media supplementation-in the form of bovine serum albumin (BSA) and fetal bovine serum (FBS)-were considered. Compared to results in media with pH of 7.5, significantly less follicles matured and ovulated in a more alkaline environment (pH 8.5), regardless of supplement type and concentration. Although the presence of FBS (5-20%) slightly improved ovulation rates, this benefit was not reflected in the fertilization outcomes. In vitro matured and ovulated oocytes obtained in non-supplemented media (pH 7.5) maintained their developmental competence and were successfully fertilized. The hatching rate was 39%, after which the survival rate of larvae was 8% at 72 h post fertilization (hpf).