Gonadotropin-releasing hormone differentially regulates expression of the genes for luteinizing hormone alpha and beta subunits in male rats.

Abstract

Gonadotropin-releasing hormone (GnRH) and gonadal steroids regulate synthesis and release of luteinizing hormone (LH). GnRH is secreted intermittently by the hypothalamus, producing pulsatile LH release, and a pulsatile GnRH stimulus is required to maintain LH secretion. We report the regulatory effects of GnRH Recently, cDNAs coding for LH a and /3 subunits were isolated (16,17), and we have used these cDNAs as hybridization probes to develop a sensitive assay for mRNAs using dot-blot hybridization of extracted cytoplasmic RNA from rat pituitaries.We employed this mRNA quantitation assay to study the effects of GnRH pulse injections on expression of the genes for LH a and p subunits in gonadectomized male rats that were given testosterone implants. In this animal model endogenous GnRH secretion is markedly reduced, as judged by infrequent serum LH pulses (18). Thus assessment of the effects of exogenous GnRH pulse injections, given to mimic the intermittent physiologic stimulus, is not unduly complicated by endogenous GnRH secretion. In addition, experiments are performed in the presence of stable physiologic concentrations of testosterone, which avoids the potential effects of fluctuating serum testosterone levels in intact animals (19). This model has been previously used to study the effects of testosterone in regulating GfiRH receptor and gonadotropin responses to a pulsatile GnRH stimulus (19). In the present study we examined the effects of different doses of GnRH per pulse injection on GnRH membrane receptors and LH subunit gene expression to assess their role in the regulation of gonadotropin secretion. MATERIALS AND METHODSMale Sprague-Dawley rats (250-300 g) were castrated and implanted with two 20-mm Silastic implants containing testosterone, to produce a serum testosterone concentration of 2.3 ± 0.12 ng/ml (mean ± SEM). Beginning immediately after recovery from anesthesia, GnRH pulse (10-250 ng per pulse) or control saline injections were given every 30 min for 48 For mRNA quantification pituitaries were individually homogenized in 10 mM Tris/0.5% Nonidet P-40/1 mM EI)TA, pH 7.4, and the homogenate was centrifuged (13,000x g for 5 mmin). DNA was measured (20) in the nuclear pellet, and total RNA was extracted from the cytosol supernatant by using a phenol/chloroform/isoamyl alcohol mixture (100:100:1, Abbreviations: GnRH, gonadotropin-releasing hormone; LH, luteinizing hormone; FSH, follicle-stimulating hormone. §To whom reprint requests should be addressed

A mechanism for the differential regulation of gonadotropin subunit gene expression by gonadotropin-releasing hormone.

“…GnRHR concentration in primary rat pituitary cells is 2-to 3-fold higher when the GnRH pulse frequency is every 30 min, compared to that at a frequency of every 2 h. This magnitude of change in GnRHR numbers is similar to that seen when 4 ,ug vs. 40 ,ug of GnRHR cDNA is transfected into GH3 cells, the concentrations at which the maximal differences in LH,3 and FSH3 gene expression were observed. Similar effects of varying GnRH pulse amplitudes have also been observed (9). We therefore speculate that varying GnRH pulse frequencies and amplitudes may regulate differentially LH and FSH subunit gene expression in vivo in rats by regulating pituitary GnRHR concentrations and hence determining the intracellular signaling pathways activated by GnRH.…”

“…Furthermore, the frequency and amplitude of GnRH pulses secreted by the hypothalamus, which vary during different phases of the menstrual or estrous cycle, regulate differentially . Similarly, the levels of gonadotropin subunit gene expression in the rat pituitary vary by 2-to 4-fold, depending on the GnRH pulse frequency and amplitude (8)(9)(10)(11) GnRHR activation include inositol phospholipid turnover, which leads to calcium mobilization and protein kinase C activation (19,20).…”

FOXO1 Transcription Factor Inhibits Luteinizing Hormone β Gene Expression in Pituitary Gonadotrope Cells

“…Our studies in L␤T2 cells also demonstrated that FOXO1 may regulate fertility through modulation of Lhb transcription, the rate-limiting step in the production of the mature hormone (8,9). Overexpression of wild-type and constitutively active FOXO1 reduced basal and GnRH-induced Lhb-luc expression, whereas overexpression of FOXO1-CA decreased endogenous GnRH-induced Lhb mRNA levels.…”

“…LH is a heterodimeric glycoprotein composed of an ␣ subunit and a unique ␤ subunit (7). Transcription of the ␤ subunit is the rate-limiting step for LH production and is essential for reproduction (8,9). Decreased Lhb mRNA levels result in infertility due to hypogonadotropic hypogonadism, whereas increased Lhb gene expression, such as in the LH␤CTP transgenic mouse model, result in infertility due to anovulation (1,10).…”

Pituitary transcription factor Prop-1 stimulates porcine pituitary glycoprotein hormone α subunit gene expression

“…Thus, aGSU gene is expressed within two different cell types in the pituitary and in different tissue in a cell-type specific as well as tissue-specific manner. Meanwhile, the expression of specific b-subunit genes, LHb and FSHb, are differently regulated (Papavasiliou et al 1986, Kato et al 1989. To clarify the specific regulatory mechanism of aGSU and specific b-subunit genes, several investigations have been conducted and several regulatory elements and transcription factors were reported (for review, see Brown & McNeilly 1999, Savage et al 2003, Jorgensen et al 2004.…”