Suppression of Apoptosis by Gonadotropin, 17β-Estradiol, and Epidermal Growth Factor in Rainbow Trout Preovulatory Ovarian Follicles

Abstract

In this study we present the first evidence for the occurrence of apoptotic cell death in ovarian follicles from teleost fish. Preovulatory ovarian follicles from mature hatchery-raised rainbow trout (Oncorhynchus mykiss) were collected and either immediately frozen in liquid nitrogen or incubated in serum-free medium at 18° for 24 hr. The extent of ovarian apoptotic DNA fragmentation was determined using 3′-end labeling of DNA with [32P]dideoxy-ATP, size fractionation by agarose gel electrophoresis, and quantification of low-molecular-weight (<15 kb) DNA using autoradiography and liquid scintillation counting. The extent of apoptotic DNA fragmentation was eightfold greater in immediately frozen preovulatory follicles than in previtellogenic ovarian follicles collected from immature rainbow trout (P<0.05), suggesting differences in the degree of apoptosis at different stages of follicular development. In preovulatory trout follicles, the extent of apoptotic DNA fragmentation was fivefold greater in follicles incubated for 24 hr. Treatment of incubated preovulatory follicles with either par tially purified salmon gonadotropin SG-G100 (1 μg/ml) or epidermal growth factor (EGF; 100 ng/ml) suppressed apoptotic DNA fragmentation by 31 and 41%, respectively, in comparison to untreated incubated follicles (P<0.01). Treatment of incubated follicles with 17β-estradiol (1–100 ng/ml) caused a concentration-dependent suppression of apoptotic DNA fragmentation (P<0.05). These results suggest that apoptosis is involved in teleost ovarian development and that several of the hormonal factors acting as follicle survival factors in mammalian and avian ovaries may play a similar role in teleost ovarian follicles.