The Expression Of RFRP-3 In Lamb Tissues And Its Role Between Nutrition And Reproduction

RFamide-related peptide-3 (RFRP-3) is the mammalian ortholog of gonadotropin-inhibiting hormone (GnIH). RFRP-3 was reported to have many effects on reproduction, feeding, stress and behavior. As RFRP-3 have an dual effect on regular reproduction and feeding, RFRP-3 was thought to regular the reproduction and feeding. The genetic structure, distribution and physiological function of RFRP-3 have been studied in many species, the distribution on female sheep has never been reported. Thus, we chose 3-month old prepubertal ewe lambs as the main research objects in the present studies. First we tested the RFRP-3 expression in sheep tissues by RT-PCR and qRT-PCR. Immuohistochemical localization of RFRP-3 in the ovaries was also conducted to find the direct effect of RFRP-3 on ovarian activity. Eighteen prepubertal ewe lambs were treated with low and high level of food restriction to detect the effect of food restriction on ovarian development. To understand the effect of food restriction on RFRP-3 expression in hypothalamic-piturary-ovary(HPO) axis, we detected the mRNA level of RFRP-3, GnRHR, FSHR and LHR in the HPO axis using qRT-PCR. This paper will present a new approach that RFRP-3 may invovled in regular energy balance and reproduction.1. mRNA expression of RFRP-3 in different tissues of prepubertal ewesmRNA expression of RFRP-3 was detected by quantitative PCR in various tissues of juvenile sheep showed that:mRNA expression of RFRP-3 in the medulla oblongata, eyes, ovaries, spinal cord, uterus,and hypothalamus were relatively higher. While in the tissues of heart, liver, spleen, lung and kidney the mRNA expression of RFRP-3 is relatively lower.2. Immunohistochemical localization of RFRP-3 in the ovaries of prepubertal ewesRFRP-3 immunoreactivity showed distinct cell specific distribution. RFRP-3 immunoreactivity was observed in the oocytes of follicles. In the granulosa cells of primordial and primary follicles, RFRP-3 generally showed no immunoreactivity. Whereas in the secondary follicles presented no to mild RFRP-3 immunoreactivity in the granulosa cells and theca cells. In the granulosa and theca cells of tertiary follicles, RFRP-3 immunoreactivity was more abundant than that in the secondary follicles. RFRP-3 immunoreactivity was observed weakly in the interstitial cells and ovarian theca.3. Effect of food restriction on ovarian development of prepubertal ewesWe test the effect of high and low level of food restriction on ovarian development. Both the high and low level of food restriction significantly reduced the volume of paired ovaries, the weight of paired ovaries and the body weight of prepubertal ewes compared to that in the C group. The "paired ovarian weight/body weight" ratio in the HR group was significantly lower than that of the C group. The data indicate that food restriction suppresses ovarian development in prepubertal ewes.Food restriction did not affect the population of primordial and primary follicles in prepubertal ewes. However, a significant decrease in the population of secondary and tertiary follicles was observed in the HR group compared to the C group. Though food restriction tended to increase the rate of follicular atresia, no significant difference was observed. The follicular recruitment rate in the HR group was lower than that of the C group. These results indicate that food restriction suppresses follicular growth and recruitment.4. Effect of food restriction on mRNA expression of RFRP-3, GnRHR, FSHR and LHR in the hypothalamic-pituitary-ovarian (HPO) axisWe investigated the mRNA expression of RFRP-3, GnRHR, FSHR, and LHR in the hypothalamic-pituitary-ovarian (HPO) axis by quantitative PCR. The mRNA expression of RFRP-3 was significantly increased in the HPO axis of prepubertal ewes in the HR and LR groups compared to that in the C group. In contrast, both high and low level of food restriction decreased the mRNA level of GnRHR, FSHR and LHR in the HPO axis of prepubertal ewes.