Co-localisation of gonadotrophins and granins in gonadotrophs at different stages of the oestrous cycle in sheep

Abstract

Associations between granins (secretogranin II (SgII) and chromogranin A and B (CgA and CgB)) and gonadotrophins (LH and FSH) have been reported in rodents and they may interact to facilitate differential storage and secretion of LH and FSH. This study investigated the relationship between granins and gonadotrophins in sheep at different stages of the oestrous cycle. Thirty-four cycling ewes had their oestrous cycles synchronised, and were divided into late luteal (LL; n=5) and early (EF; n=4), mid (n=3) and late (LF; n=11) follicular stages, and 24-53 h (n=5), 80-100 h (n=3) and 120-144 h (n=3) after the preovulatory LH surge (PS). LH mRNA levels were low in LF ewes (when plasma levels and pulse frequency of LH were high) but had increased by 80-100 h PS. In contrast, FSH mRNA levels decreased during the follicular phase and plasma FSH concentrations followed a similar pattern, to peak at 24-53 h PS due to low plasma oestradiol levels. While -gonadotrophin subunit ( -GSU), SgII and CgA mRNA levels did not change, CgB mRNA levels were elevated in EF ewes and had declined in ewes around the surge. Four distinctly sized mRNA transcripts (1·3, 2·0, 2·8 and 3·2 kb) were observed for CgA mRNA, while a double band was observed for LH mRNA that was subsequently reduced to a single band after 3 -poly(A) tail truncation. The long and short LH transcripts were prevalent in follicular and luteal ewes respectively. Numbers of LH +ve /FSH -ve granules stored within gonadotrophs were not different in LL and LF ewes (even though proportions of LH +ve granules were higher in LF ewes), but were reduced at 24-53 h PS. The majority of LH +ve granules also contained SgII, although few CgA +ve granules were found. Granule partitioning was evident whereby FSH and CgA were located near the periphery, and LH and SgII throughout the matrix. In conclusion, increases in both storage of LH +ve granules and secretion of LH in LF ewes despite constant LH mRNA levels was facilitated, at least in part, by improved LH mRNA transcript stability. Fewer LH +ve /FSH -ve granules were in storage after the PS, which was mirrored by a reduction in LH pulsatile release. Surprisingly, in view of results in rodents indicating significant changes, SgII and CgA mRNA levels did not change over the oestrous cycle in sheep. Conversely, CgB mRNA levels decreased around the time of PS. These novel results illustrate major differences in granin-gonadotrophin interactions between sheep and rodents.

Caloric restriction: Impact upon pituitary function and reproduction

“…Additionally, it has been demonstrated that the episodic secretion of FSH continues when GnRH input is blocked by GnRH antagonist treatment (Padmanabhan et al, 2003). Since LH and FSH are present within the same gonadotrope cell (Taragnat et al, 1998;Crawford and McNeilly, 2002), this asynchrony between the release of FSH and LH has been proposed to be due to the fact that LH and FSH are released by separate mechanisms from the gonadotrope cell (McNeilly, 1988;Farnworth, 1995;McNeilly et al, 2003), and while GnRH is known to be crucial for the pulsatile release of LH, basal secretion of LH and the majority of FSH secretion occurs independently of signals arising from the GnRH receptor to the release mechanisms. Gonadal hormones can decrease gonadotropin release both by decreasing GnRH release from the hypothalamus and by affecting the ability of GnRH to stimulate gonadotropin secretion from the pituitary itself.…”

GnRHa active immunity regulates expression of LHR protein and development of uteri in ewes

“…GnRH analogue acts on the gonadotrophin cells to influence the differential synthesis, storage and secretion of the luteinising hormone (LH) and follicle-stimulating hormone (FSH) (Crawford & McNeilly 2002;Schneider et al 2006). The administration of GnRH or its analogues (GnRH-A) has been shown to increase the release of LH and FSH in the pituitary gland, and therefore improves the pregnancy rate by 12Á15% in sheep (Schneider et al 2006).…”

Secretoneurin stimulates the production and release of luteinizing hormone in mouse LβT2 gonadotropin cells

“…The latter granule contained exclusively the immunoreactive signals of SgII and LH (41). In addition to the pituitaries of cow, rat and mouse, SgII was found in the LH-positive granules from female sheep gonadotrophs (6). The above immunocytochemical studies and other work on in vivo LH release (7) reveal an important association of SgII-related products and LH in mammalian gonadotrophs.…”

“…Prior to each static incubation experiment, the cells were plated in 24-well culture plates at a density of ϳ2.5 ϫ 10 5 cells/well or six-well culture plates at a density of ϳ2.0 ϫ 10 6 cells/well and cultured in the 5% CO2 air at 37°C for 48 h. At the beginning of experimentation, the medium was replaced with DMEM containing either 10 nM mGnRH-A or various doses of SN (1-100 nM). After the individual static treatments of 3,6, and/or 12 h, depending on the experiment, media were collected and stored at Ϫ20°C for radioimmunoassay of LH. Moreover, cells were removed from the plate bottom and kept at Ϫ80°C for RNA or protein extraction.…”