Effect of reduced water intake on ovarian reserve, leptin immunoexpression and impact of leptin on the in vitro culture of sheep secondary follicles

Abstract

This study evaluated the effect of reduced water intake on survival, apoptosis and immunoexpression of leptin in sheep preantral follicles, activation of primordial follicles, serum levels of leptin, estradiol (E2) and progesterone (P4), and in vitro maturation (IVM) of oocytes antral follicles, as well evaluated the effects of leptin on in vitro culture of secondary follicles isolated these animals. Ewes (n = 32) were divided into four groups: water ad libitum (Control – 100%), 80%; 60% and 40% of ad libitum intake. Blood was collected to determine, leptin, E2 and P4, before and after experiment. After the slaughter, ovarian cortex was used to histological and immunohistochemistry analysis and oocytes IVM. Moreover, isolated secondary follicles were cultured in vitro for 12 days in control medium (α-MEM+) or α-MEM+ with 10 or 25 ng/mL leptin. The reduction of water intake caused a linear decreasing effect on the percentages of normal preantral follicles, especially of primordial (P < 0.05), increased the apoptosis (P < 0.05) and decreased leptin expression in preantral follicles. The treatment with 60% of water intake showed greater total growth rate of isolated secondary follicles cultured with 25 ng/L leptin (P < 0.05), compared to those cultured in α-MEM+ . In conclusion, reduced water intake impaired the number of normal sheep preantral follicles, especially of primordial follicles, increased apoptosis and decreased leptin expression in preantral follicles. Moreover, secondary follicles from of ewes that receive 60% water intake increased follicular growth after in vitro culture with 25 ng/mL leptin.