Pattern of gonadotropin-releasing hormone (GnRH) secretion leading up to ovulation in the ewe: existence of a preovulatory GnRH surge

We have previously shown LH surges induced by physiological estradiol levels are invariably accompanied by robust and sustained GnRH surges in the ewe. Such an increase, however, has not been observed consistently during the preovulatory LH surge. In the present study, we examined GnRH secretion in Suffolk and Ile de France ewes during the preovulatory period using a method for pituitary portal blood collection which allows simultaneous portal and jugular blood samples to be taken at frequent intervals for up to 48 h. Ewes were sampled either during the mid-late luteal phase (n = 8) or follicular phase (n = 20). During the follicular phase, a robust increase in GnRH secretion occurred at the onset of the LH surge in 11 of 12 ewes sampled during the LH surge. The GnRH increase in most ewes was a massive surge, reaching values averaging 40-fold greater than baseline and extending well beyond the end of the preovulatory LH surge. In the single ewe not exhibiting a GnRH surge during the LH surge, postmortem inspection indicated blood was probably not sampled from the pituitary portal vessels. In the early follicular phase, GnRH-pulse frequency was greater than that observed in the luteal phase and, within the follicular phase, GnRH-pulse frequency increased further and amplitude decreased as the surge approached. These data demonstrate GnRH secretion leading up to ovulation in the ewe is dynamic, beginning with slow pulses during the luteal phase, progressing to higher frequency pulses during the follicular phase and invariably culminating in a robust surge of GnRH. The LH surge, however, ends despite continued elevation of GnRH.     

The central effect of beta-endorphin and naloxone on the expression of GnRH Gene and GnRH receptor (GnRH-R) gene in the hypothalamus, and on GnRH-R gene in the anterior pituitary gland in follicular phase ewes.

The effect of prolonged intermittent infusion of beta-endorphin or naloxone into the third cerebral ventricle in ewes during the follicular phase of the estrous cycle on the expression of GnRH gene and GnRH-R gene in the hypothalamus and GnRH-R gene in the anterior pituitary gland was examined by Real time-PCR. Activation of micro opioid receptors decreased GnRH mRNA levels in the hypothalamus and led to complex changes in GnRH-R mRNA: an increase of GnRH-R mRNA in the preoptic area, no change in the anterior hypothalamus and decrease in the ventromedial hypothalamus and stalk/median eminence. In beta-endorphin treated ewes the levels of GnRH-R mRNA in the anterior pituitary gland also decreased significantly. These complex changes in the levels of GnRH mRNA and GnRH-R mRNA were reflected in the decrease of LH secretion. Blockade of micro opioid receptors affected neither GnRH mRNA and GnRH-R mRNA nor LH levels secretion. These results indicate that beta-endorphin displays a suppressive effect on the expression of the GnRH gene in the hypothalamus and GnRH-R gene in the anterior pituitary gland, but affects GnRH-R gene expression in a specific manner in the various parts of hypothalamus; altogether these events lead to the decrease in GnRH/LH secretion.     

GnRH secretion throughout the ovine estrous cycle

In order to define the patterns of gonadotropin-releasing hormone (GnRH) secretion during the estrous cycle of the sheep we sampled hypophysial portal blood from conscious animals on day 1 of the cycle (n = 1), during the luteal phase (n = 8), during the follicular phase (n = 6) and during the preovulatory luteinizing hormone (LH) surge (n = 6). At the same time, we sampled jugular blood to measure plasma LH concentrations. During day 1 we noted regular GnRH pulses, whereas GnRH pulse amplitude and frequency were more variable in the luteal phase of the cycle. In the transition from the luteal phase to the follicular phase the GnRH pulse frequency increased and the amplitude decreased. Around the time of the LH surge we noted 3 types of secretory profiles for GnRH. In one sheep (type 1) there was a large GnRH pulse at the onset of the LH surge followed by very little activity during the surge. In two sheep (type 2) the GnRH profile did not change between the late follicular phase and the onset of the LH surge. In the remaining three sheep (type 3) there was a clear increase in the secretion of GnRH at the onset of the LH surge. With the exclusion of the type 1 sheep the GnRH pulse frequency was maximal (2 pulses/h) at the time of the LH surge; average portal GnRH levels were also maximal at this time.(ABSTRACT TRUNCATED AT 250 WORDS)     

A subset of gonadotropin-releasing hormone neurons in the ovine medial basal hypothalamus is activated during increased pulsatile luteinizing hormone secretion

GnRH neurons active in the preovulatory LH surge have been identified in several species using the early intermediate gene product, Fos, but the GnRH neurons active during episodic LH secretion remain unknown. In this study, we have used Fos and Fos-related antigens (FRA) to determine whether a subset of GnRH neurons is active when pulsatile LH secretion is acutely stimulated in sheep. In experiment 1, episodic LH secretion was stimulated in five of six ewes by injection of an opioid antagonist to luteal phase ewes. These five ewes had a 6-fold increase in the percentage of GnRH neurons in the medial basal hypothalamus (MBH) expressing Fos/FRA, compared with control ewes that had no LH pulses before death. Fos/FRA expression was not increased in GnRH neurons found in any other area. In experiment 2, episodic LH secretion was induced in rams by introduction of estrous ewes. This treatment increased Fos/FRA expression in MBH GnRH neurons approximately 10-fold compared with control rams. Again, this increase in Fos/FRA expression in GnRH neurons was limited to the MBH. This selective activation of MBH GnRH neurons could reflect the preferential inhibition of these perikarya by endogenous opioid peptides. It also raises the possibility that a subset of GnRH neurons in the MBH may be responsible for episodic GnRH secretion in sheep.     

Regulation of gonadotropin releasing hormone release by neuropeptide Y at the median eminence during the preovulatory period in ewes

The median eminence (ME) of the hypothalamus is known to be an important brain site where hypophysiotropic release might be regulated by excitatory and inhibitory signals impinging on their neuronal terminals. Since a role for neuropeptide Y (NPY) on preovulatory luteinizing hormone (LH) release has been suggested, we hypothesized that NPY might act at the ME to control preovulatory gonadotropin-releasing hormone (GnRH) release and thus the onset of the preovulatory surge of LH. To examine this possibility, we used the ewe as an animal model to determine: (a) immunocytochemical distribution of GnRH and NPY in the ewe ME; (b) changes in in vivo release of NPY and GnRH using ME push-pull cannula (PPC) perfusate samples, as well as in plasma LH, during the luteal, follicular and preovulatory phases of a synchronized estrous cycle, and (c) effects of ME perfusion of NPY or a Y1-NPY antagonist, or an NPY antiserum on in vivo release of ME-GnRH and plasma LH during a synchronized follicular phase. Immunolocalization reveals a dense plexus of beaded GnRH-containing neurites in the arcuate nucleus and in its vicinity, the pituitary stalk and the palisade. In contrast, a dense plexus of NPY-containing neurites occurs in the internal layer, with occasional fibers found in the intermediate and lateral external zone of the ME. In the area between the lateral internal and lateral external layers, both NPY and GnRH-containing processes were found, thus providing opportunities for synaptic and/or paracrine interactions between NPY- and GnRH-containing neurons. Hormonal analysis indicated that a synchronized preovulatory surge of LH is elicited within a 2-hour window by the sequential implantation and removal of silastic-encased estradiol (E2) or progesterone (P4) implants. In this paradigm, there was a parallel increase in ME release of both NPY and GnRH preceding the synchronized LH surge. The onset of this synchronized LH surge was advanced by ME perfusion of exogenous NPY and was both delayed and blunted by ME perfusion with the NPY antagonist (both were perfused through the PPC probe for 2 h, starting 2-3 h before the expected onset of the LH surge). In addition, NPY perfusion in the ME increases, while perfusion of the Y1-NPY antagonist or of the NPY antiserum decreases ME-PPC GnRH content and plasma levels of LH in early follicular ewes. Finally, perfusion of NPY antiserum during an ongoing LH surge disrupted LH release. These results suggest that interactions between NPY and GnRH neurons are important in controlling the timing, magnitude and maintenance of the preovulatory LH surge.     

Radiotelemetric monitoring of hypothalamic gonadotropin-releasing hormone pulse generator activity throughout the menstrual cycle of the rhesus monkey

Continuous monitoring of the electrophysiological manifestations of GnRH pulse generator activity was achieved by radiotelemetry throughout the menstrual cycles of unrestrained rhesus monkeys. The characteristic increases in hypothalamic multiunit activity (MUA volleys) associated with each LH pulse measured in the peripheral circulation were of lower frequency during the luteal phase than in the follicular phase of the cycle. Multiunit activity volley frequency increased as functional luteolysis progressed and achieved maxima of approximately one volley per hour within the first few days of the follicular phase. Unexpectedly, a dramatic decline in pulse generator frequency was observed coincidentally with the initiation of the preovulatory LH surge. Evidence is presented to support the conclusion that this deceleration of pulse generator activity is the consequence of the preovulatory rise in plasma estrogen concentration. As reported in women, a significant reduction in GnRH pulse generator frequency was observed at night during the follicular phase, but not during the luteal phase, of the menstrual cycle.     

Kisspeptin is essential for the full preovulatory LH surge and stimulates GnRH release from the isolated ovine median eminence

Kisspeptins are the product of the Kiss1 gene and potently stimulate GnRH secretion. In sheep, Kiss1 mRNA-expressing cells are found in the arcuate nucleus (ARC) and dorsal-lateral preoptic area and both appear to mediate the positive feedback effect of estradiol to generate the preovulatory GnRH/LH surge. To determine the role of kisspeptin in transmitting estrogen-positive feedback in the hypothalamus, we administered the kisspeptin antagonist p-271 to ewes subjected to an estradiol benzoate-induced LH surge. Kisspeptin antagonist treatment significantly attenuated these LH surges. We further examined the response to kisspeptin treatment prior to the LH surge. Kisspeptin significantly stimulated GnRH secretion into the hypophysial portal system, but the response to kisspeptin was similar in luteal and late-follicular phase ewes. Kiss1r mRNA expression in GnRH neurons was also similar across the estrous cycle. To examine alternative pathways for kisspeptin stimulation of GnRH neurons, we examined the origin of kisspeptin neuronal fibers in the external zone of the median eminence (ME) using neuronal tracing and immunohistochemical techniques. ARC populations of kisspeptin neurons project fibers to the ME. Finally, we showed kisspeptin stimulates GnRH release from ovine ME-cultured explants. This suggests direct kisspeptin to GnRH terminal-to-terminal communication within the ME. Overall, these data indicate an essential role for kisspeptin in receiving stimulatory estrogen signals and generating the full positive feedback GnRH/LH surge. Kisspeptin neurons of the ARC project to the external zone of the ME and kisspeptin acts upon the GnRH fibers at this level.     

Hypothalamic dysfunction

A pulsatile GnRH stimulus is required to maintain gonadotropin synthesis and secretion. The frequency and amplitude of GnRH pulses determine gonadotropin subunit gene expression and secretion of pituitary LH and FSH. Rapid frequency (more than 1 pulse per h) GnRH pulses favor LH while slower frequencies favor FSH secretion. During ovulatory cycles, an increase in GnRH frequency during the follicular phase favors LH synthesis prior to the LH surge, while following ovulation, luteal steroids slow GnRH pulses to favor FSH synthesis. Thus, a changing frequency of GnRH stimulation of the gonadotrope is one of the mechanisms involved in differential gonadotropin secretion during ovulatory cycles. In hypothalamic amenorrhea a majority of women exhibit a persistent slow frequency of LH (GnRH) pulses, which reflects excess hypothalamic opioid tone and can be temporarily reversed by opioid antagonists. At the other end of the spectrum, in polycystic ovarian syndrome, LH (GnRH) pulses are persistently rapid and favor LH synthesis, hyperandrogenism and impaired follicular maturation. Administration of progesterone can slow GnRH pulse secretion, favor FSH secretion and induce follicular maturation. Thus, the ability to change the pattern of GnRH secretion is an important factor in the maintenance of cyclic ovulation, and loss of this function leads to anovulation and amenorrhea.     

The dorsomedial suprachiasmatic nucleus times circadian expression of Kiss1 and the luteinizing hormone surge

Ovulation in mammals is gated by a master circadian clock in the suprachiasmatic nucleus (SCN). GnRH neurons represent the converging pathway through which the brain triggers ovulation, but precisely how the SCN times GnRH neurons is unknown. We tested the hypothesis that neurons expressing kisspeptin, a neuropeptide coded by the Kiss1 gene and necessary for the activation of GnRH cells during ovulation, represent a relay station for circadian information that times ovulation. We first show that the circadian increase of Kiss1 expression, as well as the activation of GnRH cells, relies on intact ipsilateral neural input from the SCN. Second, by desynchronizing the dorsomedial (dm) and ventrolateral (vl) subregions of the SCN, we show that a clock residing in the dmSCN acts independently of the light-dark cycle, and the vlSCN, to time Kiss1 expression in the anteroventral periventricular nucleus of the hypothalamus and that this rhythm is always in phase with the LH surge. In addition, we show that although the timing of the LH surge is governed by the dmSCN, its amplitude likely depends on the phase coherence between the vlSCN and dmSCN. Our results suggest that whereas dmSCN neuronal oscillators are sufficient to time the LH surge through input to kisspeptin cells in the anteroventral periventricular nucleus of the hypothalamus, the phase coherence among dmSCN, vlSCN, and extra-SCN oscillators is critical for shaping it. They also suggest that female reproductive disorders associated with nocturnal shift work could emerge from the desynchronization between subregional oscillators within the master circadian clock.     

Role of the progesterone receptor in restrained glutamic acid decarboxylase gene expression in the hypothalamus during the preovulatory luteinizing hormone surge

Previous work by our laboratory demonstrated that activation of the progesterone receptor through exogenous administration of progesterone suppressed glutamic acid decarboxylase-67 (GAD(67)) mRNA in the hypothalamus of the estrogen-primed ovariectomized rat. Since GAD(67) is the major synthetic enzyme for the inhibitory transmitter, gamma-aminobutyric acid, the finding raised the possibility that the endogenous activation of the progesterone receptor may act to restrain GAD(67) expression during the natural preovulatory gonadotropin surge during proestrus in the rat, thereby allowing GnRH secretion and the resultant LH surge. To test this hypothesis, the progesterone receptor antagonist, RU486, was administered to regularly cycling proestrous rats and the effect on GAD(67) and GAD(65) mRNA levels in the preoptic area (POA) and medial basal hypothalamus (MBH) was examined. Serum luteinizing hormone (LH) levels were also examined in order to identify correlations between changes in POA and MBH GAD levels and production of the LH surge. GAD(67) mRNA levels in the POA were increased in the cycling rat during proestrus at 18.00 h at the peak and just preceding the termination of the LH surge. There was no change in GAD(67) mRNA levels in the MBH, and GAD(65) expression was also unchanged during proestrus in the POA and MBH. Treatment with the antiprogestin RU486 resulted in an increase in GAD(67) mRNA levels at 12.00 and 14.00 h in the POA, and in the MBH at 14.00, 16.00, and 18.00 h during proestrus, effects which preceded and correlated with the attenuated LH surge in RU486-treated rats at 18.00 h. GAD(65) mRNA levels were also elevated by RU486 at 14.00 and 16.00 h in the POA, and at 14.00 h in the MBH during proestrus. These findings suggest that the progesterone receptor plays a role in restraining GAD expression in the hypothalamus during proestrus, and that this effect may be important for the production of the GnRH and LH surge.     

Changes in oxytocin receptor in bovine preovulatory follicles between the gonadotropin surge and ovulation

In cattle, production of oxytocin by granulosa cells of preovulatory follicles is induced by the LH/FSH surge and intrafollicular oxytocin increases dramatically toward the end of the interval between the surge and ovulation. We reported previously that oxytocin modulates steroid production by both theca and granulosa cells obtained from bovine preovulatory follicles, implying actions of oxytocin on both cell types of preovulatory follicles. The objective of the present study was to examine the temporal expression of oxytocin receptor mRNA and protein in both theca and granulosa cells of bovine periovulatory follicles. To induce luteal regression and initiate a follicular phase, heifers were injected with prostaglandin F₂ on Day 6 or 7 of the estrous cycle and 36 h later, a GnRH analogue was administered to induce the LH/FSH surge. The periovulatory follicle was isolated at 0, 3.5, 12, or 24 h after GnRH injection. A significant increase in the levels of mRNA for oxytocin was detected in granulosa, but not theca, cells of periovulatory follicles at 12 and 24 h after GnRH injection, relative to time 0. In contrast, the levels of oxytocin receptor mRNA and specific binding sites for oxytocin in granulosa cells had decreased significantly at 12 and 24 h post-GnRH. In theca cells, the levels of oxytocin receptor mRNA were significantly lower at 12 and 24 h compared with values at 3.5 h, but specific binding of oxytocin to thecal cell membranes was not different at any time point. Immunopositive staining for oxytocin receptor was localized to both the theca and granulosa cell layer of periovulatory follicles at all four times of follicle isolation. These results suggest the direct action of oxytocin on both theca and granulosa cells of bovine periovulatory follicles through binding to its receptor, supporting the hypothesis that follicular oxytocin plays an important role(s) in the regulation of the final stage of follicular development. Down-regulation of oxytocin receptor mRNA and oxytocin binding may serve to temporally limit the actions of oxytocin on the preovulatory follicle.     

Neurotensin gene expression increases during proestrus in the rostral medial preoptic nucleus: potential for direct communication with gonadotropin-releasing hormone neurons.

Neurotensin (NT)-containing neurons in the rostral portion of the medial preoptic nucleus (rMPN) of the brain may play a key role in regulating the pattern of secretion of GnRH, thereby influencing the reproductive cycle in females. The major goals of this study were to determine whether NT messenger RNA (mRNA) levels in the rMPN exhibit a unique pattern of expression in temporal association with the preovulatory LH surge and to assess whether NT neurons may communicate directly with GnRH neurons. We analyzed NT gene expression in rats using in situ hybridization over the day of proestrus and compared this with diestrous day 1. We also determined whether the high-affinity NT receptor (NT1) is expressed in GnRH neurons using dual-label in situ hybridization and whether this expression varies over the estrous cycle. We found that NT mRNA levels in the rMPN increase significantly on the day of proestrus, rising before the LH surge. No such change was detected on diestrous day 1, when the LH surge does not occur. Furthermore, we observed that a significant number of GnRH neurons coexpress NT1 mRNA and that the number of GnRH neurons expressing NT1 mRNA peaks on proestrus. Together with previous findings, our results suggest that increased expression of NT in the rMPN may directly stimulate GnRH neurons on proestrus, contributing to the LH surge. In addition, our results suggest that responsiveness of GnRH neurons to NT stimulation is enhanced on proestrus due to increased expression of NT receptors within GnRH neurons.     

Effects of cycle stage on regionalised galanin, galanin receptors 1-3, GNRH and GNRH receptor mRNA expression in the ovine hypothalamus

The neurotransmitter galanin has been implicated in the steroidogenic regulation of reproduction based on work mainly conducted in rodents. This study investigated the temporal changes in the expression of galanin and its three receptor isoforms and GNRH and GNRHR mRNA in specific hypothalamic nuclei known to be involved in the regulation of reproductive cyclicity, namely the medial pre-optic area (mPOA), the rostral mPOA/organum vasculosum of the lamina terminalis, the paraventricular nucleus and the arcuate nucleus using an ovine model. Following synchronisation of their oestrous cycles, tissues were collected from ewes at five time points: the early follicular, mid follicular (MF) and late follicular phases and the early luteal and mid luteal phases. The results indicated significant differences in regional expression of most of the genes studied, with galanin mRNA expression being highest during the MF phase at the start of the GNRH/LH surge and the expression of the three galanin receptor (GalR) isoforms and GNRH and its receptor highest during the luteal phase. These findings are consistent with a role for galanin in the positive feedback effects of oestradiol (E(2)) on GNRH secretion and a role for progesterone induced changes in the pattern of expression of GalRs in the regulation of the timing of E(2)'s positive feedback through increased sensitivity of galanin-sensitive systems to secreted galanin.     

Dynamics of steroid regulation of GnRH secretion during the oestrus cycle of the ewe.

The oestrus cycle of the ewe is characterised by a long luteal phase followed by a short follicular phase and these periods are related to the production by the ovary of two major steroids: progesterone and oestrogen. Progesterone exerts a strong inhibitory effect on GnRH secretion during the luteal phase by a mechanism which is still unknown. Using an oestrogen-free ovine model and the portal blood collection technique we have obtained new insights into this mechanism. While progesterone removal induces a rapid increase in GnRH pulse frequency, progesterone reinsertion inhibits GnRH release even faster: less than 50 minutes. This action of progesterone is specific to the gonadotrophic axis and is mediated through an action on the nuclear receptor. Interestingly, this rapid mechanism is also strongly dependent of prior exposure to both progesterone and oestradiol. During the follicular phase, the rise in circulating oestradiol induces a robust preovulatory GnRH surge. In the ewe, this positive feedback effect is mainly exerted by an action of oestradiol on the mediobasal hypothalamus. Finally, we have also obtained evidence that progesterone priming is important for the full expression of the positive feedback action of oestradiol on GnRH secretion. In summary, progesterone and oestradiol sequentially exert opposite feedback effects on GnRH secretion during the oestrus cycle of the ewe but there is also clear evidence that the systems affected by these steroids are intimately linked.     

Oxytocin/neurophysin-I messenger ribonucleic acid in bovine granulosa cells increases after the luteinizing hormone (LH) surge and is stimulated by LH in vitro.

Bovine granulosa cells express the oxytocin/neurophysin-I (OT/NP-I) gene and secrete OT in vitro. We have shown previously that bovine granulosa cells isolated from the preovulatory follicle after the LH surge secrete 20 times more OT over 5 days in culture than granulosa cells obtained before the surge. LH or FSH stimulates OT secretion in vitro by granulosa cells isolated before the LH surge. We also observed that granulosa cells of preovulatory follicles isolated before the LH surge respond to OT with an increase in progesterone secretion, suggesting that OT may be involved in regulating the follicular/luteal phase shift, or ovulation, in an autocrine fashion. The objective of this study was to determine whether the increase in OT secretion from granulosa cells after the LH surge is regulated at the level of mRNA accumulation, peptide synthesis, and/or peptide secretion. Bovine preovulatory follicles were obtained during the early follicular phase (approximately 36 h before the LH surge), during the midfollicular phase (approximately 12 h before the LH surge), or during the late follicular phase (after the LH surge). Total RNA isolated from granulosa cells and theca interna at the time of cell isolation or after culture with or without LH was subjected to Northern analysis for OT/NP-I mRNA and quantified by densitometry. OT/NP-I mRNA was not detectable or was barely detectable in granulosa cells collected during the early or midfollicular phase (n = 6 and n = 4 follicles, respectively), but a strong hybridization signal was obtained from RNA isolated after the LH surge (n = 5 follicles; P < 0.01). In contrast, OT/NP-I mRNA was not detectable in theca interna before or after the LH surge. Although OT/NP-I mRNA was not detectable in granulosa cells isolated 24 h after prostaglandin F2 alpha injection, after 24 h in culture, a weak OT/NP-I mRNA hybridization signal was observed in RNA from granulosa cells in LH-containing cultures. After 72 h in culture, granulosa cells cultured in control, as well as in LH-containing medium, exhibited a strong signal for OT/NP-I mRNA, but granulosa cells treated with LH exhibited a stronger OT/NP-I hybridization signal than control cultures (P < 0.01). Theca interna did not yield any OT/NP-I hybridization signal initially, and none was induced in culture.(ABSTRACT TRUNCATED AT 400 WORDS)     

Gonadotropin-releasing hormone gene expression during the rat estrous cycle: effects of pentobarbital and ovarian steroids.

Although hypothalamic GnRH release is known to be modulated by neural and hormonal factors, the relationship between altered GnRH secretion and GnRH synthesis remains unclear. In an attempt to address this question, we examined GnRH gene expression in the rat hypothalamus using in situ hybridization histochemistry. An 25S-labeled antisense RNA probe was used to identify neurons expressing GnRH mRNA in an area that included the diagonal band of Broca, the organum vasculosum of the lamina terminalis, and the preoptic area. The number of GnRH mRNA-expressing cells was determined at various times during the rat estrous cycle. During proestrus, the number of GnRH mRNA-expressing cells decreased somewhat at 1400-1600 h, increased significantly at 1800 h (the time of the LH surge), then gradually returned to basal levels at 2200 h. Expression did not change substantially at other times during the estrous cycle. To understand this close temporal relationship between the LH surge and increased GnRH mRNA levels, we examined GnRH gene expression in proestrous animals in which the LH surge was blocked with pentobarbital. Pentobarbital treatment blocked the increase in the number of GnRH mRNA-expressing cells normally observed at 1800 h in saline-treated controls, suggesting that the increase in GnRH gene expression is closely coupled to secretion of GnRH from the hypothalamus. Finally, we addressed the question of whether ovarian steroids have direct effects on GnRH gene expression by examining GnRH mRNA levels in ovariectomized steroid-treated rats at a time before (1100 h) and a time after (1800 h) hypothalamic GnRH hypersecretion. At 1100 h, no significant changes were observed, but at 1800 h, estrogen-treated rats showed a significant increase in both the number of GnRH mRNA-expressing cells and serum LH levels. This suggests that estrogen influences GnRH gene expression indirectly, perhaps by altering hypothalamic GnRH release. Our results in each of these models suggest that GnRH mRNA levels increase in response to GnRH hypersecretion at the time of the LH surge.     

Inhibitory influence of the nuclei of the posterior hypothalamus on the pro-oestrous surge of LH

The effect of transecting caudal afferents to the medial basal hypothalamus on the pro-oestrous surge of LH was studied in cyclic female rats. Rats with transverse cuts placed just in front of the mammillary bodies and caudal to the ventromedial hypothalamic nucleus showed an earlier time of onset of pro-oestrous surge of LH. Conversely, rats with transverse cuts placed 2 mm more caudally or with cuts along the lateral edges of the hypothalamus showed no altered release of LH. Advanced release of LH occurred also in rats in which the ventral premammillary nuclei or the posterior hypothalamic nuclei were bilaterally destroyed but not in those sham operated or with lesions in the dorsal premammillary nuclei. The number of ova ovulated was higher in rats bearing lesions of any of these nuclei but enhanced LH release was seen only in animals with lesions of the posterior hypothalamic nuclei. Electrochemical stimulation (anodic d.c., 100 microA, 15 s) applied at noon of pro-oestrus to the ventral premammillary nucleus, dorsal premammillary nucleus or posterior hypothalamic nucleus prevented ovulation and the preovulatory discharge of LH. It is concluded that inputs from nuclei of the posterior hypothalamus are inhibitory for LH release and could participate in determining the timing and magnitude of the pro-oestrous surge of the hormone.     

Control of the preovulatory luteinizing hormone surge by gonadotropin-releasing hormone antagonists Prospects for clinical application.

The preovulatory LH surge of the primate menstrual cycle represents a number of positive influences, a major component of which is a direct action of estradiol on the anterior pituitary lobe. Whether the LH surge also requires a corresponding burst of GnRH release from the hypothalamus has been debated. After many years of investigation, there is now conclusive evidence that a midcycle GnRH surge does occur in the primate. This is supported by studies in women with normal ovulatory cycles that demonstrate that blockade of the GnRH receptor by potent GnRH antagonists administered within 1-2 days of the expected midcycle can delay the LH surge. The ability to prevent the positive feedback effects of estradiol by GnRH antagonists is being employed for the controlled induction of follicular development and ovulation in the treatment of infertility and in in vitro fertilization programs.