Localization and Quantification of Binding Sites for Follicle-Stimulating Hormone, Luteinizing Hormone, Growth Hormone, and Insulin-Like Growth Factor I in Sheep Ovarian Follicles1

Abstract

In sheep, growth and development of ovarian follicles beyond 2 mm in diameter is acutely dependent on gonadotropin support. As a consequence, following hypophysectomy (HPX) or hypothalamic-pituitary stalk disconnection (HPD), growth of follicles beyond 2 mm is arrested and all follicles > 2 mm undergo atresia. Although administration of exogenous gonadotropins stimulates follicular growth and ovulation in HPD ewes, follicles in HPX ewes remain unresponsive unless growth hormone (GH) is also given. To determine whether the difference in follicular sensitivity to gonadotropins after HPD (gonadotropin sensitive) or HPX (gonadotropin insensitive) is related to the distribution and quantity of binding sites for FSH, LH, and/or insulin-like growth factor I (IGF-I), binding sites for these hormones were localized and quantified using topical autoradiography in healthy follicles from control (pituitary-intact), HPD, and HPX ewes. In addition, in situ hybridization was performed to localize mRNA for GH and FSH receptors. Irrespective of treatment, binding of FSH and mRNA for FSH receptor were greatest (p < 0.05) in the membrana granulosa; LH binding was greatest (p < 0.05) in the theca interna; and IGF-I binding was greatest (p < 0.05) in the theca externa. Although the relative number of binding sites for LH did not differ among treatments, those for FSH and IGF-I were lower (p < 0.05) in HPD and HPX ewes compared to controls. Attempts to quantify binding sites for GH were unsuccessful due to high nonspecific binding. However, mRNA for GH receptor was most abundant (p < 0.05) in the membrana granulosa and oocytes of small antral and preantral follicles. Compared to levels in controls and HPD ewes, the level of GH receptor mRNA was lower (p < 0.05) in follicles obtained from HPX ewes. On the basis of these data, failure of small antral follicles in HPX ewes to respond to exogenous gonadotropins is not due to a reduction in receptors for FSH, LH, or IGF-I. The observed reduction of mRNA for GH receptor in the membrana granulosa of follicles from HPX ewes provides evidence that GH may play an important role in early stages of folliculogenesis and that it is involved in the maintenance of sensitivity to gonadotropins.

Effect of exogenous insulin and fasting on growth hormone receptor and IGF-I expression in the pre-ovulatory follicle of ewes

“…Regarding gene expression, the present results indicate that there is GHR mRNA expression in the granulosa cells at higher levels than theca cells of the pre-ovulatory follicle, in agreement with observations from Eckery et al (1997). In addition, expression of IGF-I mRNA was detected in the theca interna and granulosa cells of the pre-ovulatory follicle as reported elsewhere (Leeuwenberg et al, 1995;Khalid et al, 2000), although, according to Spicer et al (1995), only 5% of the ovine pre-ovulatory follicles expressed IGF-I mRNA and no follicular expression of IGF-I mRNA was detectable by in situ hybridization (Perks et al, 1995).…”

“…However, GHR is also found in other tissues, including the ovary, where it is expressed in the corpus luteum (Juengel et al, 1997) and follicle (theca and granulosa cells) (Eckery et al, 1997) of ewes.…”

The secretory patterns of growth hormone in pregnant and hysterectomized ewes

“…The failure of antral follicles in hypophysectomized ewes to grow in response to exogenous gonadotrophins (13) is attributed to a reduction of mRNA for GH receptor in the membrana granulosa of follicles (39). Cattle with a GH receptor deficiency fail to maintain follicular development during the oestrous cycle (40).…”

Effects of luteinizing hormone and follicle stimulating hormone on the developmental competence of porcine preantral follicle oocytes grown in vitro

“…LH receptors are expressed on theca cells from preantral and antral follicles, and on granulosa cells from large antral follicles [15][16][17][18]. LH may play multiple roles throughout follicular development, but most studies have focused on the action in late-stage follicles and during the periovulatory period.…”