title: Estradiol differentially modulates the exocytotic proteins SNAP-25 and munc-18 in pituitary gonadotrophs.
abstract: Long-term treatment with estradiol increases LH secretion from female gonadotrophs. The mechanisms are not fully clarified yet. Our previous data indicated that sexual steroids might affect late steps in GnRH signal transduction such as exocytosis. The secretion of hormones from neuroendocrine cells requires the merger of secretory vesicles with the plasma membrane. This regulated exocytosis is mediated by specific proteins, which are present in the pituitary gland. Here, we examined whether two of these crucial exocytotic proteins, SNAP-25 and munc-18, are affected by estradiol in female gonadotrophs. Female rat anterior pituitary cells and alphaT3-1 cells, derived from a murine immortalized gonadotroph cell line, were treated with 100 pM estradiol for 48 h. LH secretion of anterior pituitary cells, additionally stimulated with eight consecutive pulses of 1 nM GnRH for 15 min at an interval of 1 h, was determined by RIA. Gene expression was measured by quantitative RT-PCR and protein expression by immunoblotting. Additionally, quantitative RT-PCR was performed in single rat gonadotrophs to ascribe effects exclusively to intact gonadotrophs. Pulsatile GnRH enhanced the mRNA expression of SNAP-25 and munc-18 in accordance with the LH secretory response with the greatest increase at the third pulse of GnRH. Estradiol treatment further increased GnRH-induced LH secretion at all GnRH pulses. SNAP-25 gene expression was significantly decreased at the fifth GnRH pulse and unaffected at basal after 48 h of estradiol treatment. In contrast, munc-18 mRNA levels were not significantly affected by estradiol at different GnRH-pulses in mixed anterior pituitary cells, whereas munc-18 gene expression was significantly increased at basal. In alphaT3-1 cells and single gonadotrophs, long-term estradiol treatment significantly reduced SNAP-25 protein and gene expression. In contrast, the protein and gene expression of munc-18 was significantly enhanced in both alphaT3-1 cells and single gonadotrophs. In conclusion, munc-18 and SNAP-25 were oppositionally influenced by estradiol. The results suggest that estradiol modulates the expression of exocytotic proteins in gonadotrophs and thus affects LH secretion.
authors.affiliation: Department of Obstetrics and Gynaecology, University of Luebeck, Luebeck, Germany. jmweiss1@hotmail.com
authors.forename: Jürgen M
authors.initials: JM
authors.lastname: Weiss
authors.forename: Heike
authors.initials: H
authors.lastname: Hüller
authors.forename: Stephan
authors.initials: S
authors.lastname: Polack
authors.forename: Michael
authors.initials: M
authors.lastname: Friedrich
authors.forename: Klaus
authors.initials: K
authors.lastname: Diedrich
authors.forename: Oliver
authors.initials: O
authors.lastname: Treeck
authors.forename: Georg
authors.initials: G
authors.lastname: Pfeiler
authors.forename: Olaf
authors.initials: O
authors.lastname: Ortmann
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D004958:Estradiol / Q000502:physiology*; D005089:Exocytosis / Q000502:physiology*; D005260:Female; D005786:Gene Expression Regulation / Q000502:physiology; D052681:Gonadotrophs / Q000378:metabolism*; D007987:Gonadotropin-Releasing Hormone / Q000502:physiology; D051938:Munc18 Proteins / Q000235:genetics / Q000378:metabolism*; D010902:Pituitary Gland / Q000166:cytology / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D050825:Synaptosomal-Associated Protein 25 / Q000235:genetics / Q000378:metabolism*
chemical_list: D051938:Munc18 Proteins; C494210:Snap25 protein, rat; D050825:Synaptosomal-Associated Protein 25; D007987:Gonadotropin-Releasing Hormone; D004958:Estradiol