Effect of a GnRH agonist on the FSH receptors in prepubertal ewes

The present study was aimed at investigating the effects of the GnRH agonist, alarelin, on the expression of the follicle-stimulating hormone receptor (FSHR) in the pituitary gland. Also to evaluate the presence and immunolocalization of FSHR in the ovary, as well as to confirm the efficacy on the development of the follicles in the ovaries of ewes. Twenty-eight prepubertal ewes (5-6 months of age) were randomly assigned to four experimental groups (EG, n = 7 per group). The animals in experimental group EG-I, EG-II and EG-III were subcutaneously injected with 200 fig, 300 mu g or 400 mu g alarelin antigens twice (day 0 and 14), respectively. Animals in the control group (CG) were twice subcutaneously injected with 2.0 mL of a solvent (day 0 and 14). Samples of the pituitary gland and ovaries were collected aseptically on day 70 following treatment. Blood samples were taken from the jugular vein on day 0, 7, 14, 21, 28,35, 50, 60 and 70 after the first alarelin antigen injection and ELISA used to measure the serum concentration of the GnRH antibody and FSH. Fluorescence quantitative RT-PCR was implemented to detect the gene expression of FSHR mRNA in the pituitary. Immunohistochemistry was performed to localize the FSHR in the ovary. GnRH antibody concentrations in the EG-I, EG-II and EG-III treatment groups increased gradually and were higher than that of the CG (P < 0.05) or control from day 14 to day 60. Pituitary FSHR mRNA levels were significantly reduced (P < 0.05) 1.38, 7.33 and 10.11 times in the EG-I, EG-II and EG-III groups, respectively. Immunohistochemistry identified that the immunostained cells of the FSHR were present in the ewes' ovaries, which predominantly concentrated in the cytomembrane, cytoplasm and nuclei of the follicle cells. The oocytes had different immunostaining intensities at different developing stages. The gray values of microscopy images in EG-I, EG-II and EG-III groups increased, when compared to the CG. Ovaries in alarelin treated ewes of groups EG-II and EG-III were significantly higher (P < 0.05), when compared to ewes in group EG-I and CG. The follicle vertical diameter (FVD), follicle transverse diameter (FTD), follicle-wall thickness (FWT), follicle externatheca thickness (FET) and follicle internatheca thickness (FIT) in the alarelin immunity ewes were greater than those in the CG. The serum FSH concentrations of the treated ewes remained higher than that in the CG (P < 0.05). In conclusion, active immunity with alarelin stimulated the production of the GnRH antibody, inhibited the expression of FSHR mRNA in the pituitary gland, improved the distribution of FSHR in the ovary, increased the FSH secretion and thereby promoted the development of the ovaries and follicles in the ewes. This has important potential for developing a novel technique in the superovulation and regulation of reproductive functions in ewes. (C) 2012 Elsevier B.V. All rights reserved.