Folliculogenesis and ovarian expression of mRNA encoding aromatase in anoestrous sheep after 5 days of glucose or glucosamine infusion or supplementary lupin feeding

Abstract

Improved nutrition increases ovulation rate in sheep and there is evidence that intra-ovarian pathways mediate responses to nutrition. An experiment was conducted to examine the effect of dietary energy on folliculogenesis. Anoestrous Merino ewes were fed a diet of wheat straw alone (control, n = 5), or wheat straw supplemented with lupins (500 g day(-1), n = 5). Other ewes were fed wheat straw and infused with glucose (50 mmol h(-1), n = 5) or with glucosamine (3.5 mmol h(-1), n = 5). Intravaginal progestagen sponges were inserted for 12 days, and nutritional treatments were started 5 days before sponge removal. At sponge removal, the ewes were injected with a regimen of GnRH pulses (500 ng every 4 h from 0 to 12 h; 250 ng every 2 h from 14 to 24 h; and 200 ng every 1 h from 25 to 36 h) to simulate normal follicular development. Thirty-six hours after sponge removal, the animals were killed and the ovaries were collected and stored at -80 degrees C. The ovaries were sectioned serially every 10 microm. Every 20th section was stained (to estimate number and diameter of follicles) and every 17-19th section was probed by in situ hybridization for P(450) aromatase. Data were analysed using ANOVA and chi-squared tests. There was an effect of treatment (P < 0.05) on the number of follicles 2-3, 3-4 and 6-7 mm in diameter. Aromatase-positive follicles (1.6-7.9 mm) were detected in 31 follicles from 15 ewes across all four groups. In ten animals, the largest follicle was aromatase-positive. The diameters of aromatase-positive follicles were larger (P = 0.004) in lupin fed compared with glucose-infused ewes (4.9 +/- 0.5, 3.6 +/- 0.7, 5.3 +/- 0.5 and 4.2 +/- 0.5 mm for control, glucose-infused, lupin-fed and glucosamine-infused groups, respectively). Treatment did not affect the plasma concentration of FSH when compared with controls, indicating that the energy supplements were modifying recruited (2-3 mm and 3-4 mm) and selected follicles (> 6 mm) directly. In conclusion, dietary energy can directly stimulate folliculogenesis in recruited and selected follicles, and this effect may be mediated by changes in systemic leptin concentrations and the hexosamine energy-sensing pathway in the follicle.