Innervation of GnRH Neuron Distal Projections and Activation by Kisspeptin in a New GnRH-Cre Rat Model

Endocrinology ◽  
2020 ◽  
Vol 162 (1) ◽  
Author(s):  
Siew Hoong Yip ◽  
Pauline Campos ◽  
Xinhuai Liu ◽  
Robert Porteous ◽  
Allan E Herbison
Keyword(s):  
Median Eminence ◽  
Pulse Generator ◽  
Ectopic Expression ◽  
Brain Slices ◽  
Gnrh Neuron ◽  
External Zone ◽  
Pulsatile Gnrh ◽  
Gnrh Neurons ◽  
Gnrh Release ◽  
Presynaptic Nerve Terminals

Abstract The neural mechanisms generating pulsatile GnRH release from the median eminence (ME) remain unclear. Studies undertaken in the mouse demonstrate that GnRH neurons extend projections to the ME that have properties of both dendrites and axons, termed “dendrons,” and that the kisspeptin neuron pulse generator targets these distal dendrons to drive pulsatile GnRH secretion. It presently remains unknown whether the GnRH neuron dendron exists in other species. We report here the generation of a knock-in Gnrh1-Ires-Cre rat line with near-perfect targeting of Cre recombinase to the GnRH neuronal phenotype. More than 90% of adult male and female GnRH neurons express Cre with no ectopic expression. Adeno-associated viruses were used in adult female Gnrh1-Ires-Cre rats to target mCherry or GCAMP6 to rostral preoptic area GnRH neurons. The mCherry tracer revealed the known unipolar and bipolar morphology of GnRH neurons and their principal projection pathways to the external zone of the ME. Synaptophysin-labeling of presynaptic nerve terminals revealed that GnRH neuron distal projections received numerous close appositions as they passed through the arcuate nucleus and into the median eminence. Confocal GCaMP6 imaging in acute horizontal brain slices demonstrated that GnRH neuron distal projections lateral to the median eminence were activated by kisspeptin. These studies indicate the presence of a dendron-like arrangement in the rat with GnRH neuron distal projections receiving synaptic input and responding to kisspeptin.

Indirect suppression of pulsatile LH secretion by CRH neurons in the female mouse

Endocrinology ◽  
2020 ◽  
Author(s):  
Siew Hoong Yip ◽  
Xinhuai Liu ◽  
Sabine Hessler ◽  
Isaiah Cheong ◽  
Robert Porteous ◽  
...  
Keyword(s):  
Female Mouse ◽  
Pulse Generator ◽  
Acute Stress ◽  
Brain Slices ◽  
Gnrh Neuron ◽  
Ovariectomized Mice ◽  
Electrophysiological Recordings ◽  
Gnrh Neurons ◽  
Gnrh Release ◽  
Lh Secretion

Abstract Acute stress is a potent suppressor of pulsatile LH secretion but the mechanisms through which corticotrophin-releasing hormone (CRH) neurons inhibit GnRH release remain unclear. The activation of paraventricular nucleus (PVN) CRH neurons with Cre-dependent hM3Dq in Crh-Cre female mice resulted in the robust suppression of pulsatile LH secretion. Channelrhodopsin (ChR2)-assisted circuit mapping revealed that PVN CRH neuron projections existed around kisspeptin neurons in the arcuate nucleus (ARN) although many more fibers made close appositions with GnRH neuron distal dendrons in the ventral ARN. Acutely-prepared brain slice electrophysiology experiments in GnRH-GFP mice showed a dose-dependent (30 and 300 nM CRH) activation of firing in ~20% of GnRH neurons in both intact diestrous and ovariectomized mice with inhibitory effects being uncommon (<8%). Confocal GCaMP6 imaging of GnRH neuron distal dendrons in acute para-horizontal brain slices from GnRH-Cre mice injected with Cre-dependent GCaMP6s AAVs demonstrated no effects of 30-300 nM CRH on GnRH neuron dendron calcium concentrations. Electrophysiological recordings of ARN kisspeptin neurons in Crh-Cre,Kiss1-GFP mice revealed no effects of 30 -300 nM CRH on basal or neurokinin B-stimulated firing rate. Similarly, the optogenetic activation (2-20 Hz) of CRH nerve terminals in the ARN of Crh-Cre,Kiss1-GFP mice injected with Cre-dependent ChR2 had no effect on kisspeptin neuron firing. Together, these studies demonstrate that PVN CRH neurons potently suppress LH pulsatility but do not exert direct inhibitory control over GnRH neurons, at their cell body or dendron, or the ARN kisspeptin neuron pulse generator in the female mouse.

scholarly journals Vasoactive Intestinal Polypeptide Can Excite Gonadotropin-Releasing Hormone Neurons in a Manner Dependent on Estradiol and Gated by Time of Day

Endocrinology ◽  
2008 ◽  
Vol 149 (6) ◽  
pp. 3130-3136 ◽  
Author(s):  
Catherine A. Christian ◽  
Suzanne M. Moenter
Keyword(s):  
Vasoactive Intestinal Polypeptide ◽  
Brain Slices ◽  
Time Of Day ◽  
Gnrh Neuron ◽  
Suprachiasmatic Nuclei ◽  
Neuron Firing ◽  
Neuron Response ◽  
Gnrh Neurons ◽  
Gnrh Release ◽  
Intestinal Polypeptide

A surge of GnRH release signals the LH surge that triggers ovulation. The GnRH surge is dependent on a switch in estradiol feedback from negative to positive and, in rodents, a daily neural signal, likely from the suprachiasmatic nuclei. Vasoactive intestinal polypeptide (VIP) may be involved in suprachiasmatic nuclei-GnRH neuron communication. Here we assessed the effects of acute VIP (5 min treatment) on GnRH neuron function using targeted extracellular recordings of firing activity of GnRH neurons in brain slices. We examined the effect of VIP on firing rate at different times of day using an established ovariectomized, estradiol-treated (OVX+E) mouse model that exhibits daily LH surges timed to the late afternoon. Cells from OVX animals (no estradiol) did not respond to VIP, regardless of time of day. With estradiol, the effect of VIP on GnRH neurons was dependent on the time of recording. During negative feedback, OVX+E cells did not respond. VIP increased firing in cells recorded during surge onset, but this excitatory response was reduced at surge peak. Acute treatment of OVX+E cells during surge peak with a VIP receptor antagonist decreased GnRH neuron firing. This suggests endogenous VIP may both increase GnRH neuron firing during the surge and occlude response to exogenous VIP. These data provide functional evidence for VIP effects on GnRH neurons and indicate that both estradiol and time of day gate the GnRH neuron response to this peptide. VIP may provide an excitatory signal from the circadian clock that helps time the GnRH surge.

scholarly journals Kisspeptin Is Essential for the Full Preovulatory LH Surge and Stimulates GnRH Release from the Isolated Ovine Median Eminence

Endocrinology ◽  
2011 ◽  
Vol 152 (3) ◽  
pp. 1001-1012 ◽  
Author(s):  
Jeremy T. Smith ◽  
Qun Li ◽  
Kai Sing Yap ◽  
Muhammad Shahab ◽  
Antonia K. Roseweir ◽  
...  
Keyword(s):  
Median Eminence ◽  
Positive Feedback ◽  
Estradiol Benzoate ◽  
Portal System ◽  
Neuronal Tracing ◽  
Lh Surge ◽  
External Zone ◽  
Gnrh Secretion ◽  
Gnrh Neurons ◽  
Gnrh Release

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.

scholarly journals Both Estrogen and Androgen Modify the Response to Activation of Neurokinin-3 and κ-Opioid Receptors in Arcuate Kisspeptin Neurons From Male Mice

Endocrinology ◽  
2015 ◽  
Vol 157 (2) ◽  
pp. 752-763 ◽  
Author(s):  
Kristen A. Ruka ◽  
Laura L. Burger ◽  
Suzanne M. Moenter
Keyword(s):  
Estrogen Receptor ◽  
Firing Pattern ◽  
Pulse Generator ◽  
Brain Slices ◽  
Gonadal Steroids ◽  
Gnrh Neuron ◽  
Male Mice ◽  
Neurokinin B ◽  
Neuron Firing ◽  
Kndy Neurons

Abstract Gonadal steroids regulate the pattern of GnRH secretion. Arcuate kisspeptin (kisspeptin, neurokinin B, and dynorphin [KNDy]) neurons may convey steroid feedback to GnRH neurons. KNDy neurons increase action potential firing upon the activation of neurokinin B receptors (neurokinin-3 receptor [NK3R]) and decrease firing upon the activation of dynorphin receptors (κ-opioid receptor [KOR]). In KNDy neurons from intact vs castrated male mice, NK3R-mediated stimulation is attenuated and KOR-mediated inhibition enhanced, suggesting gonadal secretions are involved. Estradiol suppresses spontaneous GnRH neuron firing in male mice, but the mediators of the effects on firing in KNDy neurons are unknown. We hypothesized the same gonadal steroids affecting GnRH firing pattern would regulate KNDy neuron response to NK3R and KOR agonists. To test this possibility, extracellular recordings were made from KNDy neurons in brain slices from intact, untreated castrated or castrated adult male mice treated in vivo with steroid receptor agonists. As observed previously, the stimulation of KNDy neurons by the NK3R agonist senktide was attenuated in intact vs castrated mice and suppression by dynorphin was enhanced. In contrast to observations of steroid effects on the GnRH neuron firing pattern, both estradiol and DHT suppressed senktide-induced KNDy neuron firing and enhanced the inhibition caused by dynorphin. An estrogen receptor-α agonist but not an estrogen receptor-β agonist mimicked the effects of estradiol on NK3R activation. These observations suggest the steroid modulation of responses to activation of NK3R and KOR as mechanisms for negative feedback in KNDy neurons and support the contribution of these neurons to steroid-sensitive elements of a GnRH pulse generator.

scholarly journals Autocrine Regulation of Gonadotropin-Releasing Hormone Secretion in Cultured Hypothalamic Neurons

Endocrinology ◽  
1999 ◽  
Vol 140 (3) ◽  
pp. 1423-1431 ◽  
Author(s):  
Lazar Z. Krsmanovic ◽  
Antonio J. Martinez-Fuentes ◽  
Krishan K. Arora ◽  
Nadia Mores ◽  
Carlos E. Navarro ◽  
...  
Keyword(s):  
Hormone Secretion ◽  
Receptor Activation ◽  
Gnrh Receptor ◽  
Hypothalamic Neurons ◽  
Gnrh Receptors ◽  
Pulsatile Gnrh ◽  
Gnrh Neurons ◽  
Gnrh Release ◽  
Hypothalamic Cells

Abstract Episodic hormone secretion is a characteristic feature of the hypothalamo-pituitary-gonadal system, in which the profile of gonadotropin release from pituitary gonadotrophs reflects the pulsatile secretory activity of GnRH-producing neurons in the hypothalamus. Pulsatile release of GnRH is also evident in vitro during perifusion of immortalized GnRH neurons (GT1–7 cells) and cultured fetal hypothalamic cells, which continue to produce bioactive GnRH for up to 2 months. Such cultures, as well as hypothalamic tissue from adult rats, express GnRH receptors as evidenced by the presence of high-affinity GnRH binding sites and GnRH receptor transcripts. Furthermore, individual GnRH neurons coexpress GnRH and GnRH receptors as revealed by double immunostaining of hypothalamic cultures. In static cultures of hypothalamic neurons and GT1–7 cells, treatment with the GnRH receptor antagonist, [d-pGlu1, d-Phe2, d-Trp3,6]GnRH caused a prominent increase in GnRH release. In perifused hypothalamic cells and GT1–7 cells, treatment with the GnRH receptor agonist, des-Gly10-[d-Ala6]GnRH N-ethylamide, reduced the frequency and increased the amplitude of pulsatile GnRH release, as previously observed in GT1–7 cells. In contrast, exposure to the GnRH antagonist analogs abolished pulsatile secretion and caused a sustained and progressive increase in GnRH release. These findings have demonstrated that GnRH receptors are expressed in hypothalamic GnRH neurons, and that receptor activation is required for pulsatile GnRH release in vitro. The effects of GnRH agonist and antagonist analogs on neuropeptide release are consistent with the operation of an ultrashort-loop autocrine feedback mechanism that exerts both positive and negative actions that are necessary for the integrated control of GnRH secretion from the hypothalamus.

scholarly journals Kisspeptin Increases γ-Aminobutyric Acidergic and Glutamatergic Transmission Directly to Gonadotropin-Releasing Hormone Neurons in an Estradiol-Dependent Manner

Endocrinology ◽  
2010 ◽  
Vol 151 (1) ◽  
pp. 291-300 ◽  
Author(s):  
Justyna Pielecka-Fortuna ◽  
Suzanne M. Moenter
Keyword(s):  
Negative Feedback ◽  
Positive Feedback ◽  
Brain Slices ◽  
Cell Voltage ◽  
Dependent Manner ◽  
Glutamatergic Transmission ◽  
Neuron Response ◽  
Gnrh Neurons ◽  
Gnrh Release ◽  
In Cells

Abstract GnRH neurons are the final central pathway controlling fertility. Kisspeptin potently activates GnRH release via G protein-coupled receptor 54 (GPR54). GnRH neurons express GPR54, and kisspeptin can act directly; however, GPR54 is broadly expressed, suggesting indirect actions are possible. Transsynaptic mechanisms are involved in estradiol-induced potentiation of GnRH neuron response to kisspeptin. To investigate these mechanisms, separate whole-cell voltage-clamp recordings were performed of γ-aminobutyric acid (GABA)-ergic and glutamatergic transmission to GnRH neurons in brain slices before and during kisspeptin treatment. To determine whether estradiol alters the effect of kisspeptin on synaptic transmission, mice were ovariectomized and either left with no further treatment (OVX) or treated with estradiol implants (OVX+E). Cells were first studied in the morning when estradiol exerts negative feedback. Kisspeptin increased frequency and amplitude of GABAergic postsynaptic currents (PSCs) in GnRH neurons from OVX+E mice. Blocking action potentials eliminated the effect on frequency, indicating presynaptic actions. Amplitude changes were due to postsynaptic actions. Kisspeptin also increased frequency of glutamatergic excitatory PSCs in cells from OVX+E animals. Kisspeptin did not affect either GABAergic or glutamatergic transmission to GnRH neurons in cells from OVX mice, indicating effects on transmission are estradiol dependent. In contrast to stimulatory effects on GABAergic PSC frequency during negative feedback, kisspeptin had no effect during positive feedback. These data suggest estradiol enables kisspeptin-mediated increases in GABA and glutamate transmission to GnRH neurons. Furthermore, the occlusion of the response during positive feedback implies one consequence of estradiol positive feedback is an increase in transmission to GnRH neurons mediated by endogenous kisspeptin.

Can obestatin modulate the GnRH neurons activity?

2016 ◽  
Vol 62 (5) ◽  
pp. 49-50 ◽  
Author(s):  
Michal Szlis ◽  
Jolanta Polkowska ◽  
Anna Wójcik-Gładysz
Keyword(s):  
Gene Expression ◽  
Central Nervous System ◽  
Nervous System ◽  
Pulse Generator ◽  
Receptor Gene ◽  
Secretory Activity ◽  
Gnrh Neurons ◽  
Gnrh Release ◽  
The Central Nervous System ◽  
Anorexigenic Peptide

Obestatin, an anorexigenic peptide acting at the central nervous system and on the periperial level, can co-create neuroendocrine network, which modulate the gonadotrophic axis activity. The aim of this study was to investigate the role of intracerebroventricular obestatin infusion on the activity of the gonadoliberine (GnRH) neurons activity.The experiment was performed on peripubertal Polish Merino sheep (n=24). Animals were divided into 2 groups: control (Ringer-Lock solution infusions; n=12) and experimental (obestatin infusion, 25μl/120μl/h; n=12). Infusions were performed over three consecutive days; blood samples were collected on day 0 and day 3. After the experiment, the animals were slaughtered, and the chosen brain tissue was preserved for IHC and Real Time RT-qPCR analysis.It was also shown that exogenous obestatin changes the selected gene expression of GnRH pulse generator, decreases the secretory activity of GnRH neurons, resulting from the inhibition of GnRH release from median eminence terminal nerves, and also decreases the GnRH receptor gene expression in pituitary. On the basis of the obtained results it can be concluded that obestatin may be involved in the modulation of reproduction processes in animals at the level of the central nervous system. However, the mechanism of its action requires further research, especially identifying the obestatin receptor itself.

scholarly journals Morphological Characterization of the Action Potential Initiation Segment in GnRH Neuron Dendrites and Axons of Male Mice

Endocrinology ◽  
2015 ◽  
Vol 156 (11) ◽  
pp. 4174-4186 ◽  
Author(s):  
Michel K. Herde ◽  
Allan E. Herbison
Keyword(s):  
Action Potential ◽  
Cell Body ◽  
Brain Slices ◽  
Gnrh Neuron ◽  
Male Mice ◽  
Single Action ◽  
Ankyrin G ◽  
Gnrh Neurons ◽  
Action Potential Initiation ◽  
Potential Initiation

GnRH neurons are the final output neurons of the hypothalamic network controlling fertility in mammals. In the present study, we used ankyrin G immunohistochemistry and neurobiotin filling of live GnRH neurons in brain slices from GnRH-green fluorescent protein transgenic male mice to examine in detail the location of action potential initiation in GnRH neurons with somata residing at different locations in the basal forebrain. We found that the vast majority of GnRH neurons are bipolar in morphology, elaborating a thick (primary) and thinner (secondary) dendrite from opposite poles of the soma. In addition, an axon-like process arising predominantly from a proximal dendrite was observed in a subpopulation of GnRH neurons. Ankyrin G immunohistochemistry revealed the presence of a single action potential initiation zone ∼27 μm in length primarily in the secondary dendrite of GnRH neurons and located 30 to 140 μm distant from the cell soma, depending on the type of process and location of the cell body. In addition to dendrites, the GnRH neurons with cell bodies located close to hypothalamic circumventricular organs often elaborated ankyrin G–positive axon-like structures. Almost all GnRH neurons (>90%) had their action potential initiation site in a process that initially, or ultimately after a hairpin loop, was coursing in the direction of the median eminence. These studies indicate that action potentials are initiated in different dendritic and axonal compartments of the GnRH neuron in a manner that is dependent partly on the neuroanatomical location of the cell body.

scholarly journals Indirect assessment of pulsatile gonadotropin-releasing hormone release in agonadal prepubertal rhesus monkeys (Macaca mulatta)

1999 ◽  
Vol 160 (1) ◽  
pp. 35-41 ◽  
Author(s):  
KJ Suter ◽  
CR Pohl ◽  
TM Plant
Keyword(s):  
Pulse Generator ◽  
Intermittent Infusion ◽  
Open Loop ◽  
Remote Access ◽  
Striking Difference ◽  
Major Purpose ◽  
Venous Circulation ◽  
Pulsatile Gnrh ◽  
Gnrh Release ◽  
Lh Secretion

The major purpose of this study was to characterize the open-loop frequency of pulsatile GnRH release in the female rhesus monkey at an age (15-20 months) when the prepubertal restraint on the hypothalamic-pituitary axis is maximally imposed. Additionally, evidence for pulsatile GnRH release in agonadal males of comparable age was also sought. Episodic LH secretion from the pituitary was used as an indirect index of GnRH discharges. In order to maximize the sensitivity of this in situ bioassay, the responsiveness of the pituitary gonadotrophs was usually first heightened by an i.v. intermittent infusion of the synthetic peptide. Monkeys (five females, three males) were castrated between 9 and 14 months of age, implanted with indwelling venous catheters, fitted with nylon jackets and housed in specialized cages that permitted remote access to the venous circulation with minimal restraint and without interruption of the light-darkness cycle. In females, LH secretion was generally assessed at 20-day intervals during alternate nighttime (1900-0200 h) and daytime (0700-1400 h) windows. In males, LH was assessed less frequently and only at night. The mean frequency of pulsatile LH release in agonadal prepubertal females was 4 pulses/7 h during the night and 2 pulses/7 h during the day. These findings indicate that, prior to puberty in the female monkey, the GnRH pulse generator operates at a relatively slow frequency and is subjected to diurnal modulation. In males, evidence for robust pulsatile GnRH release was not observed. The striking difference in activity of the GnRH pulse generator in agonadal prepubertal male and female monkeys reinforces the view that the ontogeny of the hypothalamic drive to the pituitary-gonadal axis in higher primates, including man, is sexually differentiated.

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

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.

Sign in / Sign up

Export Citation Format

Share Document