The Gonadotropin-Releasing Hormone Pulse Generator

Abstract The pulsatile release of GnRH and LH secretion is essential for fertility in all mammals. Pulses of LH occur approximately every hour in follicular-phase females and every 2 to 3 hours in luteal-phase females and males. Many studies over the last 50 years have sought to identify the nature and mechanism of the “GnRH pulse generator” responsible for pulsatile LH release. This review examines the characteristics of pulsatile hormone release and summarizes investigations that have led to our present understanding of the GnRH pulse generator. There is presently little compelling evidence for an intrinsic mechanism of pulse generation involving interactions between GnRH neuron cell bodies. Rather, data support the presence of an extrinsic pulse generator located within the arcuate nucleus, and attention has focused on the kisspeptin neurons and their projections to GnRH neuron dendrons concentrated around the median eminence. Sufficient evidence has been gathered in rodents to conclude that a subpopulation of arcuate kisspeptin neurons is, indeed, the GnRH pulse generator. Findings in other species are generally compatible with this view and suggest that arcuate/infundibular kisspeptin neurons represent the mammalian GnRH pulse generator. With hindsight, it is likely that past arcuate nucleus multiunit activity recordings have been from kisspeptin neurons. Despite advances in identifying the cells forming the pulse generator, almost nothing is known about their mechanisms of synchronicity and the afferent hormonal and transmitter modulation required to establish the normal patterns of LH pulsatility in mammals.

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Definition of the hypothalamic GnRH pulse generator in mice

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1713897114 ◽

2017 ◽

Vol 114 (47) ◽

pp. E10216-E10223 ◽

Cited By ~ 99

Author(s):

Jenny Clarkson ◽

Su Young Han ◽

Richard Piet ◽

Timothy McLennan ◽

Grace M. Kane ◽

...

Keyword(s):

Arcuate Nucleus ◽

Pulse Generator ◽

Hormone Secretion ◽

Pulse Generation ◽

Neuron Population ◽

Lh Pulsatility ◽

Freely Behaving ◽

Definition Of ◽

Lh Secretion

The pulsatile release of luteinizing hormone (LH) is critical for mammalian fertility. However, despite several decades of investigation, the identity of the neuronal network generating pulsatile reproductive hormone secretion remains unproven. We use here a variety of optogenetic approaches in freely behaving mice to evaluate the role of the arcuate nucleus kisspeptin (ARNKISS) neurons in LH pulse generation. Using GCaMP6 fiber photometry, we find that the ARNKISS neuron population exhibits brief (∼1 min) synchronized episodes of calcium activity occurring as frequently as every 9 min in gonadectomized mice. These ARNKISS population events were found to be near-perfectly correlated with pulsatile LH secretion. The selective optogenetic activation of ARNKISS neurons for 1 min generated pulses of LH in freely behaving mice, whereas inhibition with archaerhodopsin for 30 min suppressed LH pulsatility. Experiments aimed at resetting the activity of the ARNKISS neuron population with halorhodopsin were found to reset ongoing LH pulsatility. These observations indicate the ARNKISS neurons as the long-elusive hypothalamic pulse generator driving fertility.

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Kisspeptin, Neurokinin B, and Dynorphin Act in the Arcuate Nucleus to Control Activity of the GnRH Pulse Generator in Ewes

Endocrinology ◽

10.1210/en.2013-1331 ◽

2013 ◽

Vol 154 (11) ◽

pp. 4259-4269 ◽

Cited By ~ 101

Author(s):

Robert L. Goodman ◽

Stanley M. Hileman ◽

Casey C Nestor ◽

Katrina L. Porter ◽

John M. Connors ◽

...

Keyword(s):

Arcuate Nucleus ◽

Pulse Generator ◽

Pulse Frequency ◽

Afferent Input ◽

Pulse Generation ◽

Orphanin Fq ◽

Neurokinin B ◽

Control Activity ◽

Kndy Neurons ◽

Lh Secretion

Recent work has led to the hypothesis that kisspeptin/neurokinin B/dynorphin (KNDy) neurons in the arcuate nucleus play a key role in GnRH pulse generation, with kisspeptin driving GnRH release and neurokinin B (NKB) and dynorphin acting as start and stop signals, respectively. In this study, we tested this hypothesis by determining the actions, if any, of four neurotransmitters found in KNDy neurons (kisspeptin, NKB, dynorphin, and glutamate) on episodic LH secretion using local administration of agonists and antagonists to receptors for these transmitters in ovariectomized ewes. We also obtained evidence that GnRH-containing afferents contact KNDy neurons, so we tested the role of two components of these afferents: GnRH and orphanin-FQ. Microimplants of a Kiss1r antagonist briefly inhibited LH pulses and microinjections of 2 nmol of this antagonist produced a modest transitory decrease in LH pulse frequency. An antagonist to the NKB receptor also decreased LH pulse frequency, whereas NKB and an antagonist to the receptor for dynorphin both increased pulse frequency. In contrast, antagonists to GnRH receptors, orphanin-FQ receptors, and the N-methyl-D-aspartate glutamate receptor had no effect on episodic LH secretion. We thus conclude that the KNDy neuropeptides act in the arcuate nucleus to control episodic GnRH secretion in the ewe, but afferent input from GnRH neurons to this area does not. These data support the proposed roles for NKB and dynorphin within the KNDy neural network and raise the possibility that kisspeptin contributes to the control of GnRH pulse frequency in addition to its established role as an output signal from KNDy neurons that drives GnRH pulses.

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Androgenic and oestrogenic steroid participation in feedback control of luteinizing hormone secretion in male sheep

Acta Endocrinologica ◽

10.1530/acta.0.1020499 ◽

1983 ◽

Vol 102 (4) ◽

pp. 499-504 ◽

Cited By ~ 15

Author(s):

M. J. D'Occhio ◽

B. D. Schanbacher ◽

J. E. Kinder

Keyword(s):

Luteinizing Hormone ◽

Pulse Generator ◽

Hormone Secretion ◽

Pulse Interval ◽

Serum Concentrations ◽

Luteinizing Hormone Secretion ◽

Lh Release ◽

Lh Secretion ◽

Additional Feedback ◽

Male Sheep

Abstract. The acute castrate ram (wether) was used as an experimental model to investigate the site(s) of feedback on luteinizing hormone (LH) by testosterone, dihydrotestosterone and oestradiol. At the time of castration, wethers were implanted subdermally with Silastic capsules containing either crystalline testosterone (three 30 cm capsules), dihydrotestosterone (five 30 cm capsules) or oestradiol (one 6.5 cm capsule). Blood samples were taken at 10 min intervals for 6 h 2 weeks after implantation to determine serum steroid concentrations and to characterize the patterns of LH secretion. Pituitary LH response to exogenous LRH (5 ng/kg body weight) were also determined at the same time. The steroid implants produced serum concentrations of the respective hormones which were either one-third (testosterone) or two-to-four times (dihydrotestosterone, oestradiol) the levels measured in rams at the time of castration. Non-implanted wethers showed rhythmic pulses of LH (pulse interval 40–60 min) and had elevated LH levels (16.1 ± 1.6 ng/ml; mean ± se) 2 weeks after castration. All three steroids suppressed pulsatile LH release and reduced mean LH levels (to below 3 ng/ml) and pituitary LH responses to LRH. Inhibition of pulsatile LH secretion by all three steroids indicated that testosterone as well as its androgenic and oestrogenic metabolites can inhibit the LRH pulse generator in the hypothalamus. Additional feedback on the pituitary was indicated by the dampened LH responses to exogenous LRH.

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Effect of androgen on Kiss1 expression and luteinizing hormone release in female rats

Journal of Endocrinology ◽

10.1530/joe-16-0568 ◽

2017 ◽

Vol 233 (3) ◽

pp. 281-292 ◽

Cited By ~ 8

Author(s):

Kinuyo Iwata ◽

Yuyu Kunimura ◽

Keisuke Matsumoto ◽

Hitoshi Ozawa

Keyword(s):

Luteinizing Hormone ◽

Arcuate Nucleus ◽

Female Rats ◽

Hormone Release ◽

Lh Surge ◽

Continuous Release ◽

Ovulatory Dysfunction ◽

Lh Release ◽

Gonadotrophin Releasing Hormone ◽

Lh Secretion

Hyperandrogenic women have various grades of ovulatory dysfunction, which lead to infertility. The purpose of this study was to determine whether chronic exposure to androgen affects the expression of kisspeptin (ovulation and follicle development regulator) or release of luteinizing hormone (LH) in female rats. Weaned females were subcutaneously implanted with 90-day continuous-release pellets of 5α-dihydrotestosterone (DHT) and studied after 10 weeks of age. Number of Kiss1-expressing cells in both the anteroventral periventricular nucleus (AVPV) and arcuate nucleus (ARC) was significantly decreased in ovary-intact DHT rats. Further, an estradiol-induced LH surge was not detected in DHT rats, even though significant differences were not observed between DHT and non-DHT rats with regard to number of AVPV Kiss1-expressing cells or gonadotrophin-releasing hormone (GnRH)-immunoreactive (ir) cells in the presence of high estradiol. Kiss1-expressing and neurokinin B-ir cells were significantly decreased in the ARC of ovariectomized (OVX) DHT rats compared with OVX non-DHT rats; pulsatile LH secretion was also suppressed in these animals. Central injection of kisspeptin-10 or intravenous injection of a GnRH agonist did not affect the LH release in DHT rats. Notably, ARC Kiss1-expressing cells expressed androgen receptors (ARs) in female rats, whereas only a few Kiss1-expressing cells expressed ARs in the AVPV. Collectively, our results suggest excessive androgen suppresses LH surge and pulsatile LH secretion by inhibiting kisspeptin expression in the ARC and disruption at the pituitary level, whereas AVPV kisspeptin neurons appear to be directly unaffected by androgen. Hence, hyperandrogenemia may adversely affect ARC kisspeptin neurons, resulting in anovulation and menstrual irregularities.

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KNDy Neurons Modulate the Magnitude of the Steroid-Induced Luteinizing Hormone Surges in Ovariectomized Rats

Endocrinology ◽

10.1210/en.2015-1070 ◽

2015 ◽

Vol 156 (11) ◽

pp. 4200-4213 ◽

Cited By ~ 23

Author(s):

Cleyde V. Helena ◽

Natalia Toporikova ◽

Bruna Kalil ◽

Andrea M. Stathopoulos ◽

Veronika V. Pogrebna ◽

...

Keyword(s):

Negative Feedback ◽

Arcuate Nucleus ◽

Ovariectomized Rats ◽

Neurokinin B ◽

Lh Surge ◽

Neuronal Populations ◽

Lh Release ◽

Positive And Negative Feedback ◽

Kndy Neurons ◽

Lh Secretion

Kisspeptin is the most potent stimulator of LH release. There are two kisspeptin neuronal populations in the rodent brain: in the anteroventral periventricular nucleus (AVPV) and in the arcuate nucleus. The arcuate neurons coexpress kisspeptin, neurokinin B, and dynorphin and are called KNDy neurons. Because estradiol increases kisspeptin expression in the AVPV whereas it inhibits KNDy neurons, AVPV and KNDy neurons have been postulated to mediate the positive and negative feedback effects of estradiol on LH secretion, respectively. Yet the role of KNDy neurons during the positive feedback is not clear. In this study, ovariectomized rats were microinjected bilaterally into the arcuate nucleus with a saporin-conjugated neurokinin B receptor agonist for targeted ablation of approximately 70% of KNDy neurons. In oil-treated animals, ablation of KNDy neurons impaired the rise in LH after ovariectomy and kisspeptin content in both populations. In estradiol-treated animals, KNDy ablation did not influence the negative feedback of steroids during the morning. Surprisingly, KNDy ablation increased the steroid-induced LH surges, accompanied by an increase of kisspeptin content in the AVPV. This increase seems to be due to lack of dynorphin input from KNDy neurons to the AVPV as the following: 1) microinjections of a dynorphin antagonist into the AVPV significantly increased the LH surge in estradiol-treated rats, similar to KNDy ablation, and 2) intra-AVPV microinjections of dynorphin in KNDy-ablated rats restored LH surge levels. Our results suggest that KNDy neurons provide inhibition to AVPV kisspeptin neurons through dynorphin and thus regulate the amplitude of the steroid-induced LH surges.

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The neurobiological mechanism underlying hypothalamic GnRH pulse generation: the role of kisspeptin neurons in the arcuate nucleus

F1000Research ◽

10.12688/f1000research.18356.2 ◽

2020 ◽

Vol 8 ◽

pp. 982 ◽

Cited By ~ 1

Author(s):

Tony M. Plant

Keyword(s):

Arcuate Nucleus ◽

Pulse Generator ◽

Hormone Secretion ◽

Pulse Generation ◽

Luteinizing Hormone Secretion ◽

Gnrh Release ◽

Exciting Time ◽

Kndy Neurons ◽

Hypophysial Portal

This review recounts the origins and development of the concept of the hypothalamic gonadotropin-releasing hormone (GnRH) pulse generator. It starts in the late 1960s when striking rhythmic episodes of luteinizing hormone secretion, as reflected by circulating concentrations of this gonadotropin, were first observed in monkeys and ends in the present day. It is currently an exciting time witnessing the application, primarily to the mouse, of contemporary neurobiological approaches to delineate the mechanisms whereby Kiss1/NKB/Dyn (KNDy) neurons in the arcuate nucleus of the hypothalamus generate and time the pulsatile output of kisspeptin from their terminals in the median eminence that in turn dictates intermittent GnRH release and entry of this decapeptide into the primary plexus of the hypophysial portal circulation. The review concludes with an examination of questions that remain to be addressed.

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The neurobiological mechanism underlying hypothalamic GnRH pulse generation: the role of kisspeptin neurons in the arcuate nucleus

F1000Research ◽

10.12688/f1000research.18356.1 ◽

2019 ◽

Vol 8 ◽

pp. 982 ◽

Cited By ~ 12

Author(s):

Tony M. Plant

Keyword(s):

Arcuate Nucleus ◽

Pulse Generator ◽

Hormone Secretion ◽

Pulse Generation ◽

Luteinizing Hormone Secretion ◽

Gnrh Release ◽

Exciting Time ◽

Kndy Neurons ◽

Hypophysial Portal

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Deficiency of Arcuate Nucleus Kisspeptin Results in Post-Pubertal Central Hypogonadism

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00088.2021 ◽

2021 ◽

Author(s):

Nimisha Nandankar ◽

Ariel L. Negron ◽

Andrew Wolfe ◽

Jon E Levine ◽

Sally Radovick

Keyword(s):

Feedback Control ◽

Arcuate Nucleus ◽

Critical Role ◽

Knock Out ◽

Protein Levels ◽

Reproductive Axis ◽

Lh Pulsatility ◽

Estrous Cyclicity ◽

Lh Release ◽

Kndy Neurons

Kisspeptin (encoded by Kiss1), a neuropeptide critically involved in neuroendocrine regulation of reproduction, is primarily synthesized in two hypothalamic nuclei: the anteroventral periventricular nucleus (AVPV) and arcuate nucleus (ARC). AVPV kisspeptin is thought to regulate the estrogen-induced positive feedback control of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH), and the pre-ovulatory LH surge in females. In contrast, ARC kisspeptin neurons, which largely co-express neurokinin B and dynorphin A (collectively named KNDy neurons), are thought to mediate estrogen-induced negative feedback control of GnRH/LH and be the major regulators of pulsatile GnRH/LH release. However, definitive data to delineate the specific roles of AVPV versus ARC kisspeptin neurons in the control of GnRH/LH release is lacking. Therefore, we generated a novel mouse model targeting deletion of Kiss1 to the ARC nucleus (Pdyn-Cre/Kiss1fl/fl KO) to determine the functional differences between ARC and AVPV kisspeptin neurons on the reproductive axis. The efficacy of the knock-out was confirmed at both the mRNA and protein levels. Adult female Pdyn-Cre/Kiss1fl/fl KO mice exhibited persistent diestrus and significantly fewer LH pulses when compared to controls, resulting in arrested folliculogenesis, hypogonadism, and infertility. Pdyn-Cre/Kiss1fl/fl KO males also exhibited disrupted LH pulsatility, hypogonadism, and variable, defective spermatogenesis and subfertility. The timing of pubertal onset in males and females was equivalent to controls. These findings add to the current body of evidence for the critical role of kisspeptin in ARC KNDy neurons in GnRH/LH pulsatility in both sexes, while directly establishing ARC kisspeptin's role in regulating estrous cyclicity in female mice, and gametogenesis in both sexes, and culminating in disrupted fertility. The Pdyn-Cre/Kiss1fl/fl KO mice present a novel mammalian model of post-pubertal central hypogonadism.

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Modulation of pulsatile GnRH dynamics across the ovarian cycle: the role of excitability within the arcuate kisspeptin network

10.1101/2021.03.22.436428 ◽

2021 ◽

Author(s):

Margaritis Voliotis ◽

Xiao Feng Li ◽

Ross De Burgh ◽

Geffen Lass ◽

Deyana Ivanova ◽

...

Keyword(s):

Arcuate Nucleus ◽

Pulse Generator ◽

Pulse Generation ◽

Ovarian Cycle ◽

Gonadal Steroids ◽

Pulsatile Gnrh ◽

Gnrh Release ◽

Kndy Neurons ◽

Stimulation Of

AbstractPulsatile GnRH release is essential for normal reproductive function. Kisspeptin secreting neurons found in the arcuate nucleus, known as KNDy neurons for co-expressing neurokinin B, and dynorphin, drive pulsatile GnRH release. Furthermore, gonadal steroids regulate GnRH pulsatile dynamics across the ovarian cycle by altering KNDy neurons’ signalling properties. However, the precise mechanism of regulation remains mostly unknown. Here we investigate these mechanisms using a combination of mathematical and in-vivo approaches. We find that optogenetic stimulation of KNDy neurons stimulates pulsatile GnRH/LH secretion in estrous mice but inhibits it in diestrous mice. Our mathematical modelling suggests that this differential effect is due to well-orchestrated changes in neuropeptide signalling and the excitability of the KNDy population controlled via glutamate signalling. Guided by model predictions, we show that blocking glutamate signalling in the arcuate nucleus in diestrous animals inhibits LH pulses, and that optic stimulation of the KNDy population mitigates this inhibition. In estrous mice, disruption of glutamate signalling inhibits pulses generated via sustained low-frequency optic stimulation of the KNDy population, supporting the idea that the level of network excitability is critical for pulse generation. Our results reconcile previous puzzling findings regarding the estradiol-dependent effect that several neuromodulators have on the GnRH pulse generator dynamics. Therefore, we anticipate our model to be a cornerstone for a more quantitative understanding of the pathways via which gonadal steroids regulate GnRH secretion dynamics. Finally, our results could inform useful repurposing of drugs targeting the glutamate system in reproductive therapy.

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Localization of kisspeptin, NKB, and NK3R in the hypothalamus of gilts treated with the progestin altrenogest

Biology of Reproduction ◽

10.1093/biolre/ioab103 ◽

2021 ◽

Author(s):

Ashley N Lindo ◽

Jennifer F Thorson ◽

Michelle N Bedenbaugh ◽

Richard B McCosh ◽

Justin A Lopez ◽

...

Keyword(s):

Arcuate Nucleus ◽

Preoptic Area ◽

Pulse Frequency ◽

Peptide Expression ◽

Hypothalamic Tissue ◽

Lh Release ◽

Neurokinin 3 Receptor ◽

Almost All ◽

Lh Secretion ◽

The Brain

Abstract Mechanisms in the brain controlling secretion of gonadotropin hormones in pigs, particularly luteinizing hormone (LH), are poorly understood. Kisspeptin is a potent LH stimulant that is essential for fertility in many species, including pigs. Neurokinin B (NKB) acting through neurokinin 3 receptor (NK3R) is involved in kisspeptin-stimulated LH release, but organization of NKB and NK3R within the porcine hypothalamus is unknown. Hypothalamic tissue from ovariectomized (OVX) gilts was used to determine the distribution of immunoreactive kisspeptin, NKB, and NK3R cells in the arcuate nucleus (ARC). Almost all kisspeptin neurons coexpressed NKB in the porcine ARC. Immunostaining for NK3R was distributed throughout the preoptic area (POA) and in several hypothalamic areas including the periventricular and retrochiasmatic areas but was not detected within the ARC. There was no colocalization of NK3R with gonadotropin-releasing hormone (GnRH), but NK3R-positive fibers in the POA were in close apposition to GnRH neurons. Treating OVX gilts with the progestin altrenogest decreased LH pulse frequency and reduced mean circulating concentrations of LH compared with OVX control gilts (P < 0.01), but the number of kisspeptin and NKB cells in the ARC did not differ between treatments. The neuroanatomical arrangement of kisspeptin, NKB, and NK3R within the porcine hypothalamus confirm they are positioned to stimulate GnRH and LH secretion in gilts, though differences with other species exist. Altrenogest suppression of LH secretion in the OVX gilt does not appear to involve decreased peptide expression of kisspeptin or NKB.

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