Abstract :
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. Twentyeight
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 g, 300 g or 400 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.
