Pituitary Luteinizing Hormone Release Research Articles

A role for glial fibrillary acidic protein (GFAP)-expressing cells in the regulation of gonadotropin-releasing hormone (GnRH) but not arcuate kisspeptin neuron output in male mice

GnRH neurons are the final central neural output regulating fertility. Kisspeptin neurons in the hypothalamic arcuate nucleus (KNDy neurons) are considered the main regulator of GnRH output. GnRH and KNDy neurons are surrounded by astrocytes, which can modulate neuronal activity and communicate over distances. Prostaglandin E2 (PGE2), synthesized primarily by astrocytes, increases GnRH neuron activity and downstream pituitary release of luteinizing hormone (LH). We hypothesized that glial fibrillary acidic protein (GFAP)-expressing astrocytes play a role in regulating GnRH and/or KNDy neuron activity and LH release. We used adeno-associated viruses to target designer receptors exclusively activated by designer drugs (DREADDs) to GFAP-expressing cells to activate Gq- or Gi-mediated signaling. Activating Gq signaling in the preoptic area, near GnRH neurons, but not in the arcuate, increases LH release in vivo and GnRH firing in vitro via a mechanism in part dependent upon PGE2. These data suggest that astrocytes can activate GnRH/LH release in a manner independent of KNDy neurons.

Hypothalamic Astrocyte Development and Physiology for Neuroprogesterone Induction of the Luteinizing Hormone Surge.

Neural circuits in female rats sequentially exposed to estradiol and progesterone underlie so-called estrogen positive feedback that induce the surge release of pituitary luteinizing hormone (LH) leading to ovulation and luteinization of the corpus hemorrhagicum. It is now well-established that gonadotropin releasing hormone (GnRH) neurons express neither the reproductively critical estrogen receptor-α (ERα) nor classical progesterone receptor (PGR). Estradiol from developing ovarian follicles acts on ERα-expressing kisspeptin neurons in the rostral periventricular region of the third ventricle (RP3V) to induce PGR expression, and kisspeptin release. Circulating estradiol levels that induce positive feedback also induce neuroprogesterone (neuroP) synthesis in hypothalamic astrocytes. This local neuroP acts on kisspeptin neurons that express PGR to augment kisspeptin expression and release needed to stimulate GnRH release, triggering the LH surge. In vitro and in vivo studies demonstrate that neuroP signaling in kisspeptin neurons occurs through membrane PGR activation of Src family kinase (Src). This signaling cascade has been also implicated in PGR signaling in the arcuate nucleus of the hypothalamus, suggesting that Src may be a common mode of membrane PGR signaling. Sexual maturation requires that signaling between neuroP synthesizing astrocytes, kisspeptin and GnRH neurons be established. Prior to puberty, estradiol does not facilitate the synthesis of neuroP in hypothalamic astrocytes. During pubertal development, levels of membrane ERα increase in astrocytes coincident with an increase of PKA phosphorylation needed for neuroP synthesis. Currently, it is not clear whether these developmental changes occur in existing astrocytes or are due to a new population of astrocytes born during puberty. However, strong evidence suggests that it is the former. Blocking new cell addition during puberty attenuates the LH surge. Together these results demonstrate the importance of pubertal maturation involving hypothalamic astrocytes, estradiol-induced neuroP synthesis and membrane-initiated progesterone signaling for the CNS control of ovulation and reproduction.

Indução da ovulação e características ovarianas de marrãs pre-púberes tratadas com gonadotrofina coriônica humana ou kisspeptina

In view of its similar structure and function to luteinizing hormone (LH), human chorionic gonadotropin (hCG) has conventionally been used as the main hormone to induce estrus and ovulation in sows. Recently, it has been demonstrated that the neuropeptide kisspeptin is associated with pituitary LH release in different mammalian species, including swine. Thus, the aim of this study was to determine whether kisspeptin-10 combined with equine chorionic gonadotropin (eCG) could be an efficient method to induce ovulation in prepubertal gilts. Thirty 5-month-old gilts weighing 78.0 ± 8.4 kg were randomly divided into five groups of six animals each. Groups 1, 2 and 3 received 750 IU eCG, while groups 4 and 5 received saline solution. After 72 hours, the animals received 380 nmol kisspeptin-10 (group 2), 500 IU hCG (groups 3 and 4), or saline (groups 1 and 5). The treatments were applied by a single intramuscular injection. The animals were slaughtered 7 days later and ovulation was confirmed by the presence of corpora lutea in the recovered ovaries. Data were analyzed by the Kruskal-Wallis and Fisher exact tests. The number of gilts that ovulated was similar in groups 2 and 3. This number was higher in group 5 (P<0.01) and in groups 1 and 4 (P=0.08). The weight or size of the ovaries was not influenced by the treatments, although the number of ovulations was higher in groups 2 and 3 (P<0.01). These results suggest that kisspeptin-10 and hCG have similar biological activity to induce ovulation in prepubertal gilts.

The Inhibitory Effects of RFamide-Related Peptide 3 on Luteinizing Hormone Release Involves an Estradiol-Dependent Manner in Prepubertal but Not in Adult Female Mice.

The mammalian gonadotropin-inhibitory hormone (GnIH) ortholog, RFamide-related peptide (RFRP), is considered to act on gonadotropin-releasing hormone (GnRH) neurons and the pituitary to inhibit gonadotropin synthesis and release. However, there is little evidence documenting whether RFamide-related peptide 3 (RFRP-3) plays a primary role in inhibition of the hypothalamo-pituitary-gonadal (HPG) axis prior to the onset of puberty. The present study aimed to understand the functional significance of the neuropeptide on pubertal development. The developmental changes in reproductive-related gene expression at the mRNA level were investigated in the hypothalamus of female mice. The results indicated that RFRP-3 may be an endogenous inhibitory factor for the activation of the HPG axis prior to the onset of puberty. In addition, centrally administered RFRP-3 significantly suppressed plasma luteinizing hormone (LH) levels in prepubertal female mice. Surprisingly, centrally administered RFRP-3 had no effects on plasma LH levels in ovariectomized (OVX) prepubescent female mice. In contrast, RFRP-3 also inhibited plasma LH levels in OVX prepubescent female mice that were treated with 17beta-estradiol replacement. Our study also examined the effects of RFRP-3 on plasma LH release in adult female mice that were ovariectomized at dioestrus, with or without estradiol (E2). Our results showed that the inhibitory effects of RFRP-3 were independent of E2 status. Quantitative real-time PCR and immunohistochemistry analyses showed that RFRP-3 inhibited GnRH expression at both the mRNA and protein levels in the hypothalamus. These data demonstrated that RFRP-3 could effectively suppress pituitary LH release, via the inhibition of GnRH transcription and translation in prepubescent female mice, which is associated with estrogen signaling pathway and developmental stages.

From Precocious Puberty to Infertility: Metabolic Control of the Reproductive Function

From Precocious Puberty to Infertility: Metabolic Control of the Reproductive Function

Comparative Study of Sex Steroid Levels of Persian Sturgeon, Acipenser persicus Males in Responding Negative and Positive to LHRH-A2 Hormone

Gonadotropin-releasing hormone agonists (GnRHa or LHRHa) have been used extensively in order to stimulate the release of pituitary luteinizing hormone (LH) required to induce sexual maturation (e.g., spermiation). In this research, blood serum testosterone (T), 11-ketotestosterone (11-K) and progesterone (P4) levels were measured in Persian sturgeon, Asipenser persicus (Borodin, 1897), males during propagation season. The type and dose of hormone administration for artificial propagation were LH-RH-A2 and 5 μg kg-1, respectively. In males which responded by spermiating, serum steroids levels (T, 11-K and P4) were higher than in non spermiating males. Following hormonal stimulation, levels of all three steroids increased significantly in spermiting males 14h after LHRH- A2 injection. T levels increased slightly in non spermiting males and other steroids did not change during the spermiation process. The rise of T levels in both males show that this steroid as a dominant and key androgen related with maturation in Persian sturgeon.

GnRH dose reduction decreases pituitary LH release and ovulatory response but does not affect corpus luteum (CL) development and function in llamas

Gonadotrophin releasing hormone (GnRH) is commonly used in llamas to induce ovulation; however, the consequence of reduced doses of GnRH on luteinizing hormone (LH) release, ovulatory response, and subsequent corpus luteum (CL) development and function have apparently not been investigated. Hence, we examined the effect of gradual reduction of gonadorelin acetate (GnRH) dosage on pituitary LH release, ovulatory response, CL development, and plasma progesterone concentrations in llamas. Non-pregnant, non-lactating adult llamas were examined once daily by transrectal ultrasonography, and those with a follicle ≥8 mm in diameter that had grown for three consecutive days were randomly assigned to receive 50 (GnRH50, n = 23), 25 (GnRH25, n = 29), 12.5 (GnRH12.5, n = 29), or 6.25 μg (GnRH6.25, n = 29) of GnRH, or 0.5 mL of PBS (Control group, n = 16) im. In a subset (7 or 8 animals/group), intense blood sampling was done to measure LH concentrations. All females were examined by ultrasonography every 12 h from treatment (Day 0) to Day 2 to determinate ovulation, and thereafter on alternate days until Day 16 to evaluate CL development (9–13 animals/group). Also, blood samples for progesterone determination were taken (9 or 10 animals/group) on alternate days from Days 0–16. Ovulatory response (%) was highest (P < 0.05) in the GnRH50 (82.6), intermediate in the GnRH25 (72.3) and GnRH12.5 (75.9) groups, and lowest in the GnRH6.25 group (48.3). No ovulations were detected in the Control group. Mean peak LH concentrations (ng/mL) were highest (P < 0.05) for GnRH50 (6.2), intermediate for GnRH25 (4.4) and GnRH12.5 (2.9), and lowest for GnRH6.25 (2.2) groups. In addition, based on regression analysis, llamas with an LH peak <4 ng/mL were less likely to ovulate. Llamas given 50 μg of GnRH released more (P < 0.05) pituitary LH and had an LH surge of longer duration than those given 25, 12.5, or 6.25 μg. However, in those that ovulated, neither GnRH treatment nor treatment by time interaction affected (P > 0.05) CL diameter or plasma progesterone concentrations. In summary, reducing the dose of GnRH gradually decreased the magnitude of the preovulatory LH surge and ovulatory response; however, subsequent CL development and plasma progesterone concentrations were not affected.

Components of the porcine anterior pituitary insulin-like growth factor system throughout the estrous cycle

Components of the circulating and anterior pituitary insulin-like growth factor (IGF) system vary in response to steroids in pigs. However, whether serum and anterior pituitary concentrations of the IGF system vary throughout the estrous cycle has not been determined. To further examine this relationship, estrus was synchronized in 40 gilts of similar age and weight (180 d; 120 kg) by feeding 15 mg altrenogest for 15 d to synchronize estrus. Gilts were checked twice daily for expression of estrus beginning 3 d after the end of altrenogest treatment and continuing for 7 d. The first day each gilt exhibited estrus was designated as day 1 of the estrous cycle. Blood samples were obtained by jugular venipuncture on days 1, 4, 7, 10, 13, 16, 19, and 22 of the estrous cycle. On days 7, 13, 19, and 22 of the estrous cycle 10 pigs were killed and anterior pituitary glands (AP) were collected. Serum concentrations of IGF-I and AP concentrations of IGF-I were determined by radioimmunoassay. Relative amounts of AP IGF binding protein (IGFBP) were determined by western ligand blot analysis. Relative expression of AP IGF-I, IGF-I receptor (IGF-I-R), gonadotropin-releasing hormone receptor (GnRHR), and luteinizing hormone (LH)-β subunit were determined by real-time reverse transcription polymerase chain reaction. Serum concentrations of IGF-I fluctuated throughout the estrous cycle. Mean serum concentrations of IGF-I decreased ( P < 0.02) from day 1 through day 10, increased ( P < 0.02) on days 13 through 16, and then decreased ( P < 0.02) from days 19 through 22. Mean AP concentrations of IGF-I were greater ( P < 0.03) on day 19 than on all other days, whereas no difference was detected ( P > 0.05) in mean AP concentrations of IGF-I on days 7, 13, and 22. Mean relative amounts of AP IGFBP-2 and -5 were each greater ( P < 0.02) in gilts on day 19 than on all other days, whereas no difference was detected ( P > 0.05) in mean relative amounts of AP IGFBP-2 and -5 among pigs on days 7, 13, and 22 of the estrous cycle. Relative expression AP IGF-I was greater ( P < 0.05) on days 13, 19, and 22 than on day 7 of the estrous cycle. Similarly, the relative expression of AP IGF-IR was increased ( P < 0.05) in gilts on days 13, 19, and 22 compared with day 7. The relative expression of GnRHR was greater ( P < 0.05) on days 13 and 22 of the estrous cycle than on day 7. The relative expression of LHβ subunit was greater ( P < 0.05) on day 19 of the estrous cycle than on days 7, 13, and 22. Anterior pituitary release of LH throughout the porcine estrous cycle may be modulated by changes in the intrapituitary IGF system.

Dual Somatic Recordings from Gonadotropin-Releasing Hormone (GnRH) Neurons Identified by Green Fluorescent Protein (GFP) in Hypothalamic Slices

Gonadotropin-Releasing Hormone (GnRH) is a small neuropeptide that regulates pituitary release of luteinizing hormone (LH) and follicle-stimulating hormone (FSH). These gonadotropins are essential for the regulation of reproductive function. The GnRH-containing neurons are distributed diffusely throughout the hypothalamus and project to the median eminence where they release GnRH from their axon terminals into the hypophysiotropic portal system (1). In the portal capillaries, GnRH travels to the anterior pituitary gland to stimulate release of gonadotropins into systemic circulation. GnRH release is not continuous but rather occurs in episodic pulses. It is well established that the intermittent manner of GnRH release is essential for reproduction (2, 3).Coordination of activity of multiple GnRH neurons probably underlies GnRH pulses. Total peptide content in GnRH neurons is approximately 1.0 pg/cell (4), of which 30% likely comprises the releasable pool. Levels of GnRH during a pulse (5, 6), suggest multiple GnRH neurons are probably involved in neurosecretion. Likewise, single unit activity extracted from hypothalamic multi-unit recordings during LH release indicates changes in activity of multiple neurons (7). The electrodes with recorded activity during LH pulses are associated with either GnRH somata or fibers (8). Therefore, at least some of this activity arises from GnRH neurons.The mechanisms that result in synchronized firing in hypothalamic GnRH neurons are unknown. Elucidating the mechanisms that coordinate firing in GnRH neurons is a complex problem. First, the GnRH neurons are relatively few in number. In rodents, there are 800-2500 GnRH neurons. It is not clear that all GnRH neurons are involved in episodic GnRH release. Moreover, GnRH neurons are diffusely distributed (1). This has complicated our understanding of coordination of firing and has made many technical approaches intractable. We have optimized loose cell-attached recordings in current-clamp mode for the direct detection of action potentials and developed a recording approach that allows for simultaneous recordings from pairs of GnRH neurons.

Dual Somatic Recordings from Gonadotropin-Releasing Hormone (GnRH) Neurons Identified by Green Fluorescent Protein (GFP) in Hypothalamic Slices

Gonadotropin-Releasing Hormone (GnRH) is a small neuropeptide that regulates pituitary release of luteinizing hormone (LH) and follicle-stimulating hormone (FSH). These gonadotropins are essential for the regulation of reproductive function. The GnRH-containing neurons are distributed diffusely throughout the hypothalamus and project to the median eminence where they release GnRH from their axon terminals into the hypophysiotropic portal system (1). In the portal capillaries, GnRH travels to the anterior pituitary gland to stimulate release of gonadotropins into systemic circulation. GnRH release is not continuous but rather occurs in episodic pulses. It is well established that the intermittent manner of GnRH release is essential for reproduction (2, 3). Coordination of activity of multiple GnRH neurons probably underlies GnRH pulses. Total peptide content in GnRH neurons is approximately 1.0 pg/cell (4), of which 30% likely comprises the releasable pool. Levels of GnRH during a pulse (5, 6), suggest multiple GnRH neurons are probably involved in neurosecretion. Likewise, single unit activity extracted from hypothalamic multi-unit recordings during LH release indicates changes in activity of multiple neurons (7). The electrodes with recorded activity during LH pulses are associated with either GnRH somata or fibers (8). Therefore, at least some of this activity arises from GnRH neurons. The mechanisms that result in synchronized firing in hypothalamic GnRH neurons are unknown. Elucidating the mechanisms that coordinate firing in GnRH neurons is a complex problem. First, the GnRH neurons are relatively few in number. In rodents, there are 800-2500 GnRH neurons. It is not clear that all GnRH neurons are involved in episodic GnRH release. Moreover, GnRH neurons are diffusely distributed (1). This has complicated our understanding of coordination of firing and has made many technical approaches intractable. We have optimized loose cell-attached recordings in current-clamp mode for the direct detection of action potentials and developed a recording approach that allows for simultaneous recordings from pairs of GnRH neurons.

Acute stress may induce ovulation in women

BackgroundThis study aims to gather information either supporting or rejecting the hypothesis that acute stress may induce ovulation in women. The formulation of this hypothesis is based on 2 facts: 1) estrogen-primed postmenopausal or ovariectomized women display an adrenal-progesterone-induced ovulatory-like luteinizing hormone (LH) surge in response to exogenous adrenocorticotropic hormone (ACTH) administration; and 2) women display multiple follicular waves during an interovulatory interval, and likely during pregnancy and lactation. Thus, acute stress may induce ovulation in women displaying appropriate serum levels of estradiol and one or more follicles large enough to respond to a non-midcycle LH surge.MethodsA literature search using the PubMed database was performed to identify articles up to January 2010 focusing mainly on women as well as on rats and rhesus monkeys as animal models of interaction between the hypothalamic-pituitary-adrenal (HPA) and hypothalamic-pituitary-gonadal (HPG) axes.ResultsWhereas the HPA axis exhibits positive responses in practically all phases of the ovarian cycle, acute-stress-induced release of LH is found under relatively high plasma levels of estradiol. However, there are studies suggesting that several types of acute stress may exert different effects on pituitary LH release and the steroid environment may modulate in a different way (inhibiting or stimulating) the pattern of response of the HPG axis elicited by acute stressors.ConclusionWomen may be induced to ovulate at any point of the menstrual cycle or even during periods of amenorrhea associated with pregnancy and lactation if exposed to an appropriate acute stressor under a right estradiol environment.

The efficiency of progesterone vaginal gel versus intramuscular progesterone for luteal phase supplementation in gonadotropin-releasing hormone antagonist cycles: a prospective clinical trial

We compared the efficiency of progesterone vaginal gel (PVG) with intramuscular progesterone (IMP) supplementation for luteal phase support after in vitro fertilization and embryo transfer in gonadotropin-releasing hormone (GnRH) antagonist cycles. The treatment outcomes were similar for PVG and IMP for luteal support in GnRH-antagonist protocols. With its ease of use, high tolerability by patients, and fewer side effects, PVG can be a successful alternative to IMP.

Testicular function and prolactin responsiveness to TRH and cimetidine after renal transplantation.

The hypothalamic-pituitary-testicular axis and the regulation of prolactin secretion were investigated in eleven male renal transplant recipients. Mean serum levels of testosterone and estrone were normal, whereas those of androstenedione and estradiol were low. Mean basal luteinizing hormone (LH) levels were slightly elevated, but the peak responses to 50 micrograms i.v. gonadotropin-releasing hormone (GnRH) were not dissimilar from controls. Both basal and GnRH-stimulated follicle-stimulating hormone (FSH) levels were elevated (p less than 0.02-0.05) and also positively correlated with the time spent on hemodialysis (p less than 0.005-0.002). Basal prolactin (PRL) levels were normal, in all subjects. Nine out of 11 patients had a normal PRL response to Thyrotropin-releasing Hormone (TRH). However only six out of 11 had a normal response to 200 mg i.v. Cimetidine (Cim). Three subjects normally responding to TRH failed to respond to Cim. Uremic primary hypogonadism is not fully reversed by renal transplantation: a slight defect in the pituitary LH release may persist and the impairment of the tubular testicular function is left unchanged. While uremic hyperprolactinemia is corrected, the responsiveness to PRL-stimulating agents, particularly Cim, is not restored to normal, reflecting a derangement at the pituitary as well as the hypothalamic level.

Spawning performance and plasma levels of GnRHa and sex steroids in cultured female Senegalese sole ( Solea senegalensis) treated with different GnRHa-delivery systems

The aquaculture of Senegalese sole ( Solea senegalensis) is limited by the poor spawning performance of the F1 generation cultured broodstock. The present study compared the efficiency of two sustained-release delivery systems loaded with gonadotropin-releasing hormone agonist (GnRHa) and a single GnRHa injection on the stimulation of oocyte maturation (OM), sex steroid hormone secretion and spawning. Analysis with an ELISA showed that the administered GnRHa was no longer detectable in the plasma 3, 7 and 14 d after administration via a single injection, microspheres and an implant, respectively. All GnRHa treatments induced a transient elevation of testosterone (T) and 17β estradiol (E 2) plasma levels after 1–3 d. Only three spawns were obtained from controls on days 35, 41 and 42 of the study and ovarian biopsies showed abundant fully vitellogenic oocytes, but little OM. Fish given a GnRHa injection spawned sporadically with the first spawn at 20 days post-treatment (dpt). In contrast, GnRHa implants and microspheres stimulated OM and daily spawning starting at 4 and 5 dpt, respectively, and lasting for a period of 28 d. Total fecundity in the broodstock treated with GnRHa-delivery systems was 15–22 times greater than controls and 3–5 times greater than GnRHa-injected fish. Although egg quality appeared to be high, no fertilization was obtained. The results suggest that GnRHa-delivery systems are highly efficient on stimulating OM and spawning in Senegalese sole, presumably through the long-term elevation of pituitary luteinizing hormone release, which in turn stimulated the appropriate changes in ovarian steroidogenesis. However, the effectiveness of the treatment was negated by possible dysfunctions of the male breeders or of the reproductive behavior of the broodstock.

Increase in daily LH secretion in response to short-term calorie restriction in obese women with PCOS.

We hypothesized that short-term calorie restriction would blunt luteinizing hormone (LH) hypersecretion in obese women with polycystic ovary syndrome (PCOS) and thereby ameliorate the anovulatory endocrine milieu. To test this hypothesis, 15 obese patients with PCOS and nine age- and body mass index-matched healthy women underwent 24-h blood sampling to quantitate plasma LH, leptin, and insulin levels. PCOS subjects were prescribed a very low caloric liquid diet (4.2 MJ/day) for 7 days and were then resampled. Basal and pulsatile LH secretion was threefold higher in PCOS subjects, but plasma insulin and leptin levels were not different in the calorie-replete state. Contrary to expectation, calorie restriction enhanced basal and pulsatile LH secretion even further. As expected, plasma glucose, insulin, and leptin concentrations decreased by 18, 75, and 50%, respectively. Serum total testosterone concentration fell by 23%, whereas serum estrone, estradiol, sex hormone-binding globulin (SHBG), and androstenedione concentrations remained unchanged. Enhanced LH secretion in the presence of normal metabolic and hormonal adaptations to calorie restriction points to anomalous feedback control of pituitary LH release in PCOS.

GnRH and Gonadotropin Release is Decreased in Chronic Nitric Oxide Deficiency

Nitric oxide synthetase (NOS), the conversion enzyme for nitric oxide (NO) is localized in the anterior pituitary of female rats, particularly in gonadotrophs and folliculo-stellate cells, suggesting that NO regulates the release of luteinizing hormone (LH) and follicle stimulating hormone (FSH) from the anterior pituitary. The focus of this study was to determine the effect of chronic NO deficiency on the subsequent pituitary release of LH and FSH in vitro and the hypothalamic immunoexpression of GnRH in vivo. NO deficiency was induced by adding the NOS inhibitor, N-nitro-L-arginine (L-NNA, 0.6 g/L) to the drinking water of female Wistar rats. After 8 weeks, the animals were euthanized, the pituitaries were removed, and they were incubated in vitro. Pituitaries were perfused for 4 hr in the presence of pulsatile gonadotropin release hormone (GnRH, 500 ng/pulse) every 30 min. S-Nitroso-L-acetyl penicillamine (SNAP, an NO donor, 0.1 mM) or L-nitro-argine methyl ester (L-NAME, a NOS inhibitor, 0.1 mM) was added to the media and perfusate samples were collected at 10-min intervals. LH and FSH levels in the perfusate were measured by double antibody radioimmunoassays. Pituitaries from the NO-deficient rats had a significantly smaller GnRH-stimulated release of LH and FSH compared with proestrous control rats. The addition of S-NAP to the perfusate resulted in decreased LH and FSH secretion in the control group, but increased LH secretion in the NO-deficient group. The addition of L-NAME to the perfusate suppressed LH secretion from control pituitaries, but not in pituitaries from NO-deficient animals. Immunohistochemistry of brain slices demonstrated that NO-deficient rats had a large qualitative decrease of GnRH in the median eminence compared with their controls. This decrease was particularly evident in the external capillary plexus of the median eminence. We concluded that chronic NO deficiency is associated with a decreased GnRH in neurosecretory terminals in the external capillary layer of the median eminence, accompanied by a decrease in LH and FSH release from the pituitaries.

Phytoestrogens and Gonadotropin-Releasing Hormone Pulse Generator Activity and Pituitary Luteinizing Hormone Release in the Rat

Phytoestrogens and Gonadotropin-Releasing Hormone Pulse Generator Activity and Pituitary Luteinizing Hormone Release in the Rat

Phytoestrogens and gonadotropin-releasing hormone pulse generator activity and pituitary luteinizing hormone release in the rat.

Phytoestrogens can produce inhibitory effects on gonadotropin secretion in both animals and humans. The aims of this study were 2-fold: 1) to determine in vivo whether genistein and coumestrol act on the GnRH pulse generator to suppress hypothalamic multiunit electrical activity volleys and associated LH pulses and/or on the pituitary to suppress the LH response to GnRH; and 2) to examine the effect of these phytoestrogens on GnRH-induced pituitary LH release in vitro and to determine whether estrogen receptors are involved. Wistar rats were ovariectomized and chronically implanted with recording electrodes and/or indwelling cardiac catheters, and blood samples were taken every 5 min for 7--11 h. Intravenous infusion of coumestrol (1.6-mg bolus followed by 2.4 mg/h for 8.5 h) resulted in a profound inhibition of pulsatile LH secretion, a 50% reduction in the frequency of hypothalamic multiunit electrical activity volleys, and a complete suppression of the LH response to exogenous GnRH. In contrast, both genistein (1.6-mg bolus followed by 2.4 mg/h for 8.5 h) and vehicle were without effect on pulsatile LH secretion. Coumestrol (10(-5) M; over 2 or 4 h) suppressed GnRH-induced pituitary LH release in vitro, an effect blocked by the antiestrogen ICI 182,780. It is concluded that coumestrol acts centrally to reduce the frequency of the hypothalamic GnRH pulse generator. In addition, the inhibitory effects of coumestrol on LH pulses occur at the level of the pituitary by reducing responsiveness to GnRH via an estrogen receptor-mediated process.

Evidence that Stimulation of Two Modalities of Pituitary Luteinizing Hormone Release in Ovarian Steroid-Primed Ovariectomized Rats May Involve Neuropeptide Y Y1 and Y4 Receptors

A large body of evidence indicates that neuropeptide Y (NPY) is involved in stimulation of basal and cyclic release of hypothalamic LHRH and pituitary LH. To identify the NPY receptor subtypes that mediate the excitatory effects of NPY in these two modalities of LH release, we studied the effects of 1229U91, a selective Y1 receptor antagonist and Y4 receptor agonist, in two experimental paradigms that reproduce the two modalities of LH secretion in steroid-primed ovariectomized (OVX) rats. Rats were ovariectomized and implanted with a permanent cannula into the lateral cerebroventricle. In the first experiment, rats received estradiol benzoate (EB, 30 μg/rat) on day 5, followed 2 days later with progesterone (2 mg/rat) at 1000 h to induce an afternoon LH surge. 1229U91 (30 μg/3 μl) or vehicle (control) was injected intracerebroventricularly into these rats either once at 1300 h or twice (15 μg/injection) at 1100 and 1200 h. Blood samples were collected before progesterone injection at 1000 h and at hourly intervals from 1300–1800 h via an intrajugular cannula implanted on the previous day. In control rats, serum LH levels rose significantly at 1400 h, and these high levels were maintained until 1700 h. After two injections of 1229U91, LH levels displayed a tendency to rise at 1300–1400 h, as in controls, but thereafter, decreased rapidly below the control range. In the second experiment, the acute effect of 1229U91 on LH release was evaluated in OVX rats pretreated with EB alone. Saline alone or saline containing 1, 3, 10, or 30 μg 1229U91 was injected intracerebroventricularly at 1000 h, and the effects on LH release were analyzed at 10, 20, 30, and 60 min. 1229U91 elicited a dose-dependent stimulation of LH release, with maximal response (950% of basal levels) occurring at 10 min after the 30-μg dose; elevated levels were maintained for 1 h. Because 1229U91 is a potent Y4 agonist with some affinity for Y5 receptors, these results raised the possibility that activation of Y4/Y5 receptors by 1229U91 may augment LH release. Therefore, we examined the effects of icv administration of rat pancreatic polypeptide, a Y4-selective agonist, and[ D-Trp32]-NPY, a Y5 agonist on LH release in EB-primed rats. Rat pancreatic polypeptide (0.5–2 μg/rat) stimulated LH release in a dose-related manner, and peak levels (280% of basal levels) were seen at 10–20 min; the response evoked by a higher dose (10 μg) was smaller than that induced by 0.5 or 2 μg.[ D-Trp32]-NPY was relatively less effective, because only the highest (10-μg) dose elicited a modest stimulation (244% of basal levels). These results demonstrate that 1229U91, a Y1 antagonist and Y4 agonist, evokes two types of responses; it suppresses the protracted ovarian steroid-induced LH surge, and acutely, it also stimulates LH. These results imply that normally two different types of NPY receptors may mediate the stimulation of LH release. Because 1229U91 is a Y1 receptor antagonist, inhibition of the steroid-induced LH surge by 1229U91 suggests that Y1 receptors may mediate the cyclic release of LH. On the other hand, acute stimulation of LH by 1229U91 implies that the Y4 agonist-like activity of 1229U91 may mediate the basal release of LH and that either NPY or a yet-to-be-identified endogenous Y4 receptor agonist may activate Y4 receptors in the hypothalamus to stimulate LH release.

Evidence that stimulation of two modalities of pituitary luteinizing hormone release in ovarian steroid-primed ovariectomized rats may involve neuropeptide Y Y1 and Y4 receptors.

A large body of evidence indicates that neuropeptide Y (NPY) is involved in stimulation of basal and cyclic release of hypothalamic LHRH and pituitary LH. To identify the NPY receptor subtypes that mediate the excitatory effects of NPY in these two modalities of LH release, we studied the effects of 1229U91, a selective Y1 receptor antagonist and Y4 receptor agonist, in two experimental paradigms that reproduce the two modalities of LH secretion in steroid-primed ovariectomized (OVX) rats. Rats were ovariectomized and implanted with a permanent cannula into the lateral cerebroventricle. In the first experiment, rats received estradiol benzoate (EB, 30 microg/rat) on day 5, followed 2 days later with progesterone (2 mg/rat) at 1000 h to induce an afternoon LH surge. 1229U91 (30 microg/3 microl) or vehicle (control) was injected intracerebroventricularly into these rats either once at 1300 h or twice (15 microg/injection) at 1100 and 1200 h. Blood samples were collected before progesterone injection at 1000 h and at hourly intervals from 1300 -1800 h via an intrajugular cannula implanted on the previous day. In control rats, serum LH levels rose significantly at 1400 h, and these high levels were maintained until 1700 h. After two injections of 1229U91, LH levels displayed a tendency to rise at 1300-1400 h, as in controls, but thereafter, decreased rapidly below the control range. In the second experiment, the acute effect of 1229U91 on LH release was evaluated in OVX rats pretreated with EB alone. Saline alone or saline containing 1, 3, 10, or 30 microg 1229U91 was injected intracerebroventricularly at 1000 h, and the effects on LH release were analyzed at 10, 20, 30, and 60 min. 1229U91 elicited a dose-dependent stimulation of LH release, with maximal response (950% of basal levels) occurring at 10 min after the 30-microg dose; elevated levels were maintained for 1 h. Because 1229U91 is a potent Y4 agonist with some affinity for Y5 receptors, these results raised the possibility that activation of Y4/Y5 receptors by 1229U91 may augment LH release. Therefore, we examined the effects of icv administration of rat pancreatic polypeptide, a Y4-selective agonist, and [D-Trp32]-NPY, a Y5 agonist on LH release in EB-primed rats. Rat pancreatic polypeptide (0.5-2 microg/rat) stimulated LH release in a dose-related manner, and peak levels (280% of basal levels) were seen at 10-20 min; the response evoked by a higher dose (10 microg) was smaller than that induced by 0.5 or 2 microg. [D-Trp32]-NPY was relatively less effective, because only the highest (10-microg) dose elicited a modest stimulation (244% of basal levels). These results demonstrate that 1229U91, a Y1 antagonist and Y4 agonist, evokes two types of responses; it suppresses the protracted ovarian steroid-induced LH surge, and acutely, it also stimulates LH. These results imply that normally two different types of NPY receptors may mediate the stimulation of LH release. Because 1229U91 is a Y1 receptor antagonist, inhibition of the steroid-induced LH surge by 1229U91 suggests that Y1 receptors may mediate the cyclic release of LH. On the other hand, acute stimulation of LH by 1229U91 implies that the Y4 agonist-like activity of 1229U91 may mediate the basal release of LH and that either NPY or a yet-to-be-identified endogenous Y4 receptor agonist may activate Y4 receptors in the hypothalamus to stimulate LH release.