In vivo modulation of follicle-stimulating hormone release and beta subunit gene expression by activin A and the GnRH agonist buserelin in female rats.
ABSTRACT The effects of separate and simultaneous recombinant bovine (rb) activin A and buserelin administration on the FSH release and pituitary FSH beta subunit gene expression in vivo were examined in ovariectomised, estradiol pretreated rats. The animals received a single injection of either rb activin A (50 ng), buserelin (1 micro g) or activin/buserelin (50 ng+1 micro g/0.1 ml PBS) into the jugular vein and were killed 30 min, 1, 3 and 5h later. Activin A stimulated FSH release and effect appeared 1h after injection (168% increase of controls) reaching a maximum at 3h (437% of controls). Activin A and buserelin exerted their effects with a distinct time courses: activin's stimulation was not so rapid when compared with buserelin. The simultaneous administration of rb activin A and buserelin amplified FSH release (118, 309, 1006 and 779% of controls). The low dose of activin A was sufficient to elevate FSH beta mRNA level as early as 3 and 5h after administration (170 and 140%, respectively). Activin plus buserelin stimulation resulted in a higher (340 and 360% of controls) FSH beta gene expression than after their separate administration. These results suggest that activin and buserelin may act independently and synergistically in the regulation of FSH release and beta subunit mRNA level.
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ABSTRACT: Both activin and GnRH can independently stimulate expression of the FSHbeta subunit gene. In this study, we used the gonadotrope-derived LbetaT2 cell line to investigate the potential interaction between activin and GnRH in regulating the transcriptional activity of the rat FSHbeta gene promoter. Activin A and GnRH synergistically enhanced rat FSHbeta transcriptional activity. Overexpression of SMAD3 (mediator of decapentaplegic-related protein 3), but not of SMAD2, increased transcriptional activation of the rat (r) FSHbeta gene promoter, which was further enhanced by the combined overexpression of SMAD3 and 4 (3+4). The stimulatory effects of SMAD3 overexpression were localized to -472/-256 of the rFSHbeta gene promoter, and activin- and GnRH-responsive proteins were shown to bind to region -284/-252. Sequence analysis identified a consensus palindromic SMAD-binding site at -266/-259 of the rFSHbeta gene promoter. Mutation of two bases located in the center of this palindrome effectively abrogated SMAD4 binding, markedly reduced SMAD3 and 3+4 stimulation of the rFSHbeta gene promoter, and significantly decreased the synergistic enhancement of promoter activity by both activin A and GnRH, and SMAD3 and GnRH. Blockade of the MAPK-signaling pathway did not significantly affect the response to combined stimulation with activin and GnRH. In contrast, interference with SMAD3 signaling caused a significant reduction in activin and GnRH synergy. The results indicate that SMAD3 plays an important role in the synergistic effects of activin and GnRH and demonstrate that this synergy is mediated by a palindromic cis-element located at -266/-259 of the rFSHbeta gene promoter.Molecular Endocrinology 02/2005; 19(1):237-54. · 4.75 Impact Factor
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ABSTRACT: Secretion of LH and FSH from the anterior pituitary is regulated primarily by hypothalamic GnRH and ovarian steroid hormones. More recent evidence indicates regulatory roles for certain members of the transforming growth factor beta (TGFbeta) superfamily including inhibin and activin. The aim of this study was to identify expression of mRNAs encoding key receptors and ligands of the inhibin/activin system in the hen pituitary gland and to monitor their expression throughout the 24-25-h ovulatory cycle. Hens maintained on long days (16 h light/8 h dark) were killed 20, 12, 6 and 2 h before predicted ovulation of a midsequence egg (n = 8 per group). Anterior pituitary glands were removed, RNA extracted and cDNA synthesized. Plasma concentrations of LH, FSH, progesterone and inhibin A were measured. Real-time quantitative PCR was used to quantify pituitary expression of mRNAs encoding betaglycan, activin receptor (ActR) subtypes (type I, IIA), GnRH receptor (GnRH-R), LH beta subunit, FSH beta subunit and GAPDH. Levels of mRNA for inhibin/activin betaA and betaB subunits, inhibin alpha subunit, follistatin and ActRIIB mRNA in pituitary were undetectable by quantitative PCR (<2 amol/reaction). Significant changes in expression (P<0.05) of ActRIIA and betaglycan mRNA were found, both peaking 6 h before ovulation just prior to the preovulatory LH surge and reaching a nadir 2 h before ovulation, just after the LH surge. There were no significant changes in expression of ActRI mRNA throughout the cycle although values were correlated with mRNA levels for both ActRIIA (r = 0.77; P<0.001) and beta-glycan (r = 0.45; P<0.01). Expression of GnRH-R mRNA was lowest 20 h before ovulation and highest (P<0.05) 6 h before ovulation; values were weakly correlated with betaglycan (r = 0.33; P = 0.06) and ActRIIA (r = 0.34; P = 0.06) mRNA levels. Expression of mRNAs encoding LH beta and FSH beta subunit were both lowest (P<0.05) after the LH surge, 2 h before ovulation. These results are consistent with an endocrine, but not a local intrapituitary, role of inhibin-related proteins in modulating gonadotroph function during the ovulatory cycle of the hen, potentially through interaction with betaglycan and ActRIIA. In contrast to mammals, intrapituitary expression of inhibin/activin subunits and follistatin appears to be extremely low or absent in the domestic fowl.Journal of Endocrinology 10/2005; 186(3):447-55. · 4.06 Impact Factor