Influence of different concentrations of LH and FSH on in vitro caprine primordial ovarian follicle development

Abstract

The roles of gonadotrophins in the regulation of primordial ovarian follicular development remains unclear. The aims of the present study were to investigate the effects of LH alone or in combination with FSH on the survival, activation and growth of in vitro caprine primordial follicles, using histology and transmission electron microscopy (TEM). To this end, samples of the caprine ovarian cortex were cultured for 1 or 7 days (39 °C), in an atmosphere of 5% CO 2, in minimum essential medium (MEM—control medium) supplemented with different concentrations of LH (0, 1, 5, 10, 50 or 100 ng/ml—Experiment 1). In Experiment 2, the control medium was supplemented with FSH (50 ng/ml) and the different concentrations of LH (0, 1, 5, 10 or 50 ng/ml also used). Small samples of non-cultured ovarian tissue, as well as those cultured for 1 or 7 days in a specific medium were processed for classical histological evaluation and TEM to evaluate the follicular integrity and to calculate the percentage of normal follicles. Additionally, the effects of FSH on the oocyte and follicle diameters of the cultured follicles were evaluated. Results showed that after 7 days of culture, the presence of LH (10, 50 or 100 ng/ml) in the culture media significantly increases the percentage of developing follicles ( P < 0.05). In addition, fragments cultured in media supplemented with FSH alone or FSH plus LH had a higher percentage of developing follicles after 1 and 7 days of culture, compared to the control ( P < 0.05). At the end of culture period, 1 and 5 ng/ml LH increased the follicular diameter, while the addition of 1, 5 or 10 ng/ml LH increased the oocyte diameter ( P < 0.05). A combination of FSH with 1 and 5 ng/ml LH increased the follicular diameter. With regard to follicle survival, low doses of LH (1 ng/ml) maintained follicle viability—similar to day 0 and in the control medium after 7 days of culture. However, higher concentrations of LH (5, 10, 50 and 100 ng/ml) induced atresia in the goat follicles. Culture of the ovarian cortex for 7 days in medium supplemented with FSH alone or FSH plus 1 ng/ml LH kept the ultrastructural characteristics of follicles similar to that of day 0. In conclusion, it can be said that, the addition of low concentrations of LH (1 ng/ml) combined with or without FSH maintained the goat follicular ultrastructural integrity, but LH in doses higher than 5 ng/ml induced atresia in in vitro goat preantral follicles.