Prochloraz-induced oocyte maturation in rainbow trout (Oncorhynchus mykiss), a molecular and functional analysis

Abstract

In the present study, we aimed at characterizing the effect of prochloraz, an imidazole fungicide, on the oocyte meiotic maturation process in a freshwater teleost species, the rainbow trout (Oncorhynchus mykiss). Full-grown post-vitellogenic ovarian follicles were incubated in vitro with prochloraz, Luteinizing Hormone (LH), or a combination of prochloraz and LH. The occurrence of oocyte maturation was assessed by monitoring germinal vesicle breakdown (GVBD) after 62-h in vitro incubation. Experiments were repeated in presence of actinomycin D, cycloheximide, or trilostane. The effect of prochloraz on the production of 17,20β-dihydroxy-4-pregnen-3-one (17,20βP), the natural maturation-inducing steroid, was quantified by radioimmunoassay. In addition, the effect of prochloraz on ovarian expression of 12 genes was monitored by real-time PCR. Prochloraz (10(-5)M) administered alone was able to induce 100% GVBD in the most responsive females. The occurrence of GVBD observed after prochloraz stimulation of follicles originating from various females was similar and highly correlated with the occurrence of GVBD observed after stimulation with low LH concentration. In addition, oocyte maturation induced by LH or prochloraz was totally inhibited by actinomycin D, cycloheximide, and trilostane. Similarly to LH, prochloraz was able to trigger 17,20βP production by the ovarian follicle. Finally, prochloraz induced the overexpression of genes participating in 17,20βP production, intercellular communication, and paracrine control of preovulatory follicular differentiation such as igf, igf2, connexin 43, and 20β hydroxysteroid dehydrogenase (hsbd20). Together, our results demonstrate that prochloraz administered alone is able to trigger oocyte maturation through the induction of specific genes, some of them being also triggered by LH. Finally, our results clearly indicate that the effects of prochloraz and LH on oocyte maturation are synergistic.