Regulation of pituitary gonadotropin-releasing hormone receptors by pulsatile gonadotropin-releasing hormone injections in male rats. Modulation by testosterone.

Garcia, Alexandra N.; Schiff, Melissa A.; Marshall, John C.

doi:10.1172/jci111510

Cited by 29 publications

(10 citation statements)

References 52 publications

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“…The maximal values were similar to receptor concentrations observed in male rats that received saline pulses for 48 hr after castration (600 ± 50 fmol/mg of protein, mean ± SEM). Higher doses of GnRH per pulse reduced GnRH receptors; a similar biphasic pattern of receptor regulation has been observed after GnRH pulses (19) and after continuous infusions of GnRH in intact or castrated male rats (25). a subunit and LH P subunit mRNA concentrations are shown in Fig.…”

Section: Resultssupporting

confidence: 67%

“…Beginning immediately after recovery from anesthesia, GnRH pulse (10-250 ng per pulse) or control saline injections were given every 30 min for 48 hr through a carotid cannula (19 …”

Section: Methodsmentioning

confidence: 99%

“…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).…”

mentioning

confidence: 99%

“…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.…”

mentioning

confidence: 99%

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

Papavasiliou¹,

Zmeili²,

Khoury³

et al. 1986

Proc. Natl. Acad. Sci. U.S.A.

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132

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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

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Section: Resultssupporting

confidence: 67%

“…Beginning immediately after recovery from anesthesia, GnRH pulse (10-250 ng per pulse) or control saline injections were given every 30 min for 48 hr through a carotid cannula (19 …”

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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

Papavasiliou¹,

Zmeili²,

Khoury³

et al. 1986

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

132

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“…The dissociation constant (Kd) of GnRH binding was reported to be independent of experimental conditions and of the endocrine status of the animal (1-5). However, the number of receptors seems to depend upon several endocrine parameters, such as pulsatile secretion of endogenous GnRH, estrous cycle, pregnancy, lactation or castration (3,(6)(7)(8)(9)(10)(11)(12)(13).…”

mentioning

confidence: 99%

Temperature and Protein Kinase C Modulation of Gonadotropin‐Releasing Hormone Receptors in Pituitary Cells from Intact or Castrated Male Rats

Leblanc

I’Heritier

Mounier

et al. 1990

J Neuroendocrinology

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Unite de recherches sur la dynamique des ensembles neuroendocriniens, U 159 INSERM, 2ter rue d'AIesia, 75014 Paris, France.t Pharmacologie Neuro-lmmuno-Endocrinienne (CNRS UA 11 13), lnstitut Pasteur, 28 rue du Dr Roux, 75015 Paris, France.Key words: gonadotropin-releasing hormone receptors, protein kinase C, pituitary cell culture, castration. AbstractBinding constants of ['251]Des-Gly,0-(D-Ala,)-gonadotropin-releasing hormone-ethylamide (GnRHa) to dispersed pituitary cells were evaluated in a 4-day culture. Cells were sampled either from intact or from castrated male rats and binding was measured at various temperatures before or after treatment with phorbol-12-myristate-13-acetate (PMA) or l-5-(isoquinolinyl-sulfc.xyl) 2-methylpiperazine (H7), which activate or inhibit, respectively, protein kinase C (PKC). In cells from intact rats incubated with increasing concentrations of ligand at 21 "C for 25 rnin, the Scatchard plot was not linear and calculation of the Hill coefficient (NH) was indicative of positive cooperativity (NH= 1.26_+0.02). Such non-linearity was not observed when cells were incubated at 0.5 "C for 3 h. In that condition the maximal number of binding sites measured at equilibrium (Bmax) increased (15.1 t0.05 versus 9.3k0.5 fmoles x mg-' proteins at 21 "C). Two control experiments permitted us to rule out the possibility that lower B , , , at 21 "C might reflect internalization: 1) Cells were first incubated with the ligand at 21 "C for 25 min and subsequently for 3 additional hours at 0.5 "C. Preincubation did not affect the B , , , obtained at 0.5 "C; 2) when the radioligand bound to the cell surface was washed out with an acidic buffer, only 13% of the specific radioactivity was retained irrespective of the ligand concentration applied, a much lower value than the 40% binding difference observed between 05°C and 21°C. When the cells were incubated with PMA, the Scatchard plot was linearized and the B, , , recorded at 21 "C increased by 50% over control cells (13k0.7 fmolesxrng-' proteins). Conversely, inhibition of PKC by H7, a preferential PKC inhibitor, was ineffective. In contrast, cells sampled from castrates exhibited linear and comparable Scatchard plots at either 0.5 or 21 "C, with B , , , values of 14.4k0.3 and 15k0.34 fmoles x mg-' proteins, respectively. PKC activation did not affect binding in that model, but H7 decreased the number of sites (B,,,= 10.7t0.9) and induced appearance of positive cooperativity (NH= 1.36k0.07). Taken together, these experiments reveal a pool of GnRH receptors in the pituitary of intact rats that recognizes the ligand in a phosphorylation-dependent manner. These binding sites also became evident when biological properties of the membrane were modified by temperature or cell homogenization. After castration, PKC activation was no longer a prerequisite for recruitment of the total population of receptors whereas protein kinase inhibition resulted in a reduction of maximal binding. Finally, our observations demonstrate that GnRH binding can exhibit positiv...

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Inhibins, Activins, Follistatin, and GnRH: Regulators of Gonadotropin Subunit Gene Expression

Marshall¹,

Dalkin²,

Haisenleder³

et al. 1997

Inhibin, Activin and Follistatin

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Regulation of pituitary gonadotropin-releasing hormone receptors by pulsatile gonadotropin-releasing hormone injections in male rats. Modulation by testosterone.

Cited by 29 publications

References 52 publications

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

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

Temperature and Protein Kinase C Modulation of Gonadotropin‐Releasing Hormone Receptors in Pituitary Cells from Intact or Castrated Male Rats

Inhibins, Activins, Follistatin, and GnRH: Regulators of Gonadotropin Subunit Gene Expression

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