scholarly journals An inhibitory circuit from brainstem to GnRH neurons in male mice: a new role for the RFRP receptor

Endocrinology ◽  
2021 ◽  
Author(s):  
Stephanie Constantin ◽  
Katherine Pizano ◽  
Kaya Matson ◽  
Yufei Shan ◽  
Daniel Reynolds ◽  
...  
Keyword(s):  
Neuropeptide Y ◽  
Fluorescent Protein ◽  
Nucleus Tractus Solitarius ◽  
Brain Slices ◽  
Adult Brain ◽  
Energy Deficit ◽  
Gonadotropin Secretion ◽  
Gnrh Neurons ◽  
Gonadotropin Inhibitory Hormone ◽  
Green Fluorescent

Abstract RFamide-related peptides (RFRPs, mammalian orthologs of gonadotropin-inhibitory hormone) convey circadian, seasonal and social cues to the reproductive system. They regulate gonadotropin secretion by modulating gonadotropin-releasing hormone (GnRH) neurons via the RFRP receptor. Mice lacking this receptor are fertile but exhibit abnormal gonadotropin responses during metabolic challenges such as acute fasting, when the normal drop in gonadotropin levels is delayed. Although it is known that these food intake signals to the reproductive circuit originate in the nucleus tractus solitarius (NTS) in the brainstem, the phenotype of the neurons conveying the signal remains unknown. Given that neuropeptide FF (NPFF), another RFamide peptide, resides in the NTS and can bind to the RFRP receptor, we hypothesized that NPFF may regulate GnRH neurons. To address this question, we used a combination of techniques: cell-attached electrophysiology on GnRH-driven green fluorescent protein-tagged neurons in acute brain slices; calcium imaging on cultured GnRH neurons; and immunostaining on adult brain tissue. We found 1) NPFF inhibits GnRH neuron excitability via the RFRP receptor and its canonical signaling pathway (Gi/o protein and G protein-coupled inwardly-rectifying potassium channels), 2) NPFF-like fibers in the vicinity of GnRH neurons coexpress neuropeptide Y, 3) the majority of NPFF-like cell bodies in the NTS also coexpress neuropeptide Y, and 4) acute fasting increased NPFF-like immunoreactivity in the NTS. Together these data indicate that NPFF neurons within the NTS inhibit GnRH neurons, and thus reproduction, during fasting but prior to the energy deficit.

scholarly journals RFamide-Related Peptide-3, a Mammalian Gonadotropin-Inhibitory Hormone Ortholog, Regulates Gonadotropin-Releasing Hormone Neuron Firing in the Mouse

Endocrinology ◽  
2009 ◽  
Vol 150 (6) ◽  
pp. 2799-2804 ◽  
Author(s):  
Eric Ducret ◽  
Greg M. Anderson ◽  
Allan E. Herbison
Keyword(s):  
Firing Rate ◽  
Fluorescent Protein ◽  
Suppressive Effect ◽  
Small Population ◽  
Neuron Activity ◽  
Gonadotropin Secretion ◽  
Gnrh Neurons ◽  
Gonadotropin Inhibitory Hormone ◽  
Green Fluorescent ◽  
Prolactin Releasing Peptide

The recent discovery that an RFamide termed gonadotropin-inhibitory hormone is likely to be a hypophysiotrophic gonadotropin release-inhibiting hormone in birds has generated interest into the role of LPXRFamide neuropeptides in the control of gonadotropin secretion in mammals. Recent immunocytochemical studies in birds and mammals have suggested that neurons expressing the mammalian LPXRFamides, RFamide-related peptides (RFRPs) 1 and 3, may innervate and regulate GnRH neurons directly. We used cell-attached electrophysiology in adult male and female GnRH-green fluorescent protein-tagged neurons to examine whether RFRP-3 modulated the electrical excitability of GnRH neurons. RFRP-3 was found to exhibit rapid and repeatable inhibitory effects on the firing rate of 41% of GnRH neurons. A small population of GnRH neurons (12%) increased their firing rate in response to RFRP-3, and the remainder was unaffected. No difference was detected in the RFRP-3 responses of GnRH neurons from male, diestrous, or proestrus female mice. The suppressive effect of RFRP-3 was maintained when amino acid transmission was blocked, suggesting a possible direct effect of RFRP-3 upon GnRH neurons. To evaluate the effects of other RFamide neuropeptides on GnRH neurons, we tested the actions of prolactin-releasing peptide-20 and -31. Neither compounds altered the firing rate of GnRH neurons. These studies demonstrate that RFRP-3 has a likely direct suppressive action on the excitability of GnRH neurons, indicating a role for RFRPs in the regulation of gonadotropin secretion in mammals through modulation of GnRH neuron activity.

scholarly journals Suppression of Basal Spontaneous Gonadotropin-Releasing Hormone Neuronal Activity during Lactation: Role of Inhibitory Effects of Neuropeptide Y

Endocrinology ◽  
2008 ◽  
Vol 150 (1) ◽  
pp. 333-340 ◽  
Author(s):  
Jing Xu ◽  
Melissa A. Kirigiti ◽  
Michael A. Cowley ◽  
Kevin L. Grove ◽  
M. Susan Smith
Keyword(s):  
Neuropeptide Y ◽  
Neuronal Activity ◽  
Fluorescent Protein ◽  
Resting Membrane Potential ◽  
Inhibitory Effects ◽  
Gnrh Neurons ◽  
Cell Current ◽  
Green Fluorescent ◽  
Hypothalamic Slices

Increased neuropeptide Y (NPY) activity drives the chronic hyperphagia of lactation and may contribute to the suppression of GnRH activity. The majority of GnRH neurons are contacted by NPY fibers, and GnRH cells express NPY Y5 receptor (Y5R). Therefore, NPY provides a neurocircuitry for information about food intake/energy balance to be directly transmitted to GnRH neurons. To investigate the effects of lactation on GnRH neuronal activity, hypothalamic slices were prepared from green fluorescent protein-GnRH transgenic rats. Extracellular loose-patch recordings determined basal GnRH neuronal activity from slices of ovariectomized control and lactating rats. Compared with controls, hypothalamic slices from lactating rats had double the number of quiescent GnRH neurons (14.51 ± 2.86 vs. 7.04 ± 2.84%) and significantly lower firing rates of active GnRH neurons (0.25 ± 0.02 vs. 0.37 ± 0.03 Hz). To study the NPY-postsynaptic Y5R system, whole-cell current-clamp recordings were performed in hypothalamic slices from control rats to examine NPY/Y5R antagonist effects on GnRH neuronal resting membrane potential. Under tetrodotoxin treatment, NPY hyperpolarized GnRH neurons from −56.7 ± 1.94 to −62.1 ± 1.83 mV; NPY’s effects were blocked by Y5R antagonist. To determine whether increased endogenous NPY tone contributes to GnRH neuronal suppression during lactation, hypothalamic slices were treated with Y5R antagonist. A significantly greater percentage of GnRH cells were activated in slices from lactating rats (52%) compared with controls (28%). These results suggest that: 1) basal GnRH neuronal activity is suppressed during lactation; 2) NPY can hyperpolarize GnRH neurons via postsynaptic Y5R; and 3) increased inhibitory NPY tone during lactation is a component of the mechanisms responsible for suppression of GnRH neuronal activity. Neuropeptide Y (NPY) directly hyperpolarizes GnRH neurons via postsynaptic NPY Y5 receptor. Increased inhibitory NPY tone during lactation is an important component of the suppression of GnRH neuronal activity.

scholarly journals Dose-Dependent Switch in Response of Gonadotropin-Releasing Hormone (GnRH) Neurons to GnRH Mediated through the Type I GnRH Receptor

Endocrinology ◽  
2004 ◽  
Vol 145 (2) ◽  
pp. 728-735 ◽  
Author(s):  
Chun Xu ◽  
Xu-Zhi Xu ◽  
Craig S. Nunemaker ◽  
Suzanne M. Moenter
Keyword(s):  
Firing Rate ◽  
Fluorescent Protein ◽  
Brain Slices ◽  
Dose Dependence ◽  
Gnrh Receptor ◽  
Neuron Activity ◽  
Type I ◽  
Pulsatile Release ◽  
Gnrh Neurons ◽  
Green Fluorescent

Abstract Pulsatile release of GnRH provides central control of reproduction. GnRH neuron activity is likely synchronized to produce hormone pulses, but the mechanisms are largely unknown. One candidate for communication among these neurons is GnRH itself. Cultured embryonic and immortalized GnRH neurons express GnRH receptor type I (GnRHR-1), but expression has not been shown in adult GnRH neurons. Using mice that express green fluorescent protein (GFP) in GnRH neurons, we tested whether adult GnRH neurons express GnRHR-1. GFP-positive (n = 42) and -negative neurons (n = 22) were harvested from brain slices, and single-cell RT-PCR was performed with cell contents. Fifty-two percent of the GnRH neurons tested expressed GnRHR-1, but only 9% of non-GnRH hypothalamic neurons expressed GnRHR-1; no false harvest controls (n = 13) were positive. GnRHR-1 expression within GnRH neurons suggested a physiological ultrashort loop feedback role for GnRH. Thus, we examined the effect of GnRH on the firing rate of GnRH neurons. Low-dose GnRH (20 nm) significantly decreased firing rate in 12 of 22 neurons (by 42 ± 4%, P < 0.05), whereas higher doses increased firing rate (200 nm, five of 10 neurons, 72 ± 26%; 2000 nm, nine of 13 neurons, 53 ± 8%). Interestingly, the fraction of GnRH neurons responding was similar to the fraction in which GnRHR-1 was detected. Together, these data demonstrate that a subpopulation of GnRH neurons express GnRHR-1 and respond to GnRH with altered firing. The dose dependence suggests that this autocrine control of GnRH neurons may be not only a mechanism for generating and modulating pulsatile release, but it may also be involved in the switch between pulse and surge modes of release.

scholarly journals Neurosteroids Alter γ-Aminobutyric Acid Postsynaptic Currents in Gonadotropin-Releasing Hormone Neurons: A Possible Mechanism for Direct Steroidal Control

Endocrinology ◽  
2003 ◽  
Vol 144 (10) ◽  
pp. 4366-4375 ◽  
Author(s):  
Shannon D. Sullivan ◽  
Suzanne M. Moenter
Keyword(s):  
Gabaa Receptor ◽  
Decay Time ◽  
Chloride Channels ◽  
Fluorescent Protein ◽  
Brain Slices ◽  
Cell Voltage ◽  
Dehydroepiandrosterone Sulfate ◽  
Postsynaptic Currents ◽  
Gnrh Neurons ◽  
Green Fluorescent

Pulsatile GnRH release is required for fertility and is regulated by steroid feedback. Whether or not steroids or their metabolites act directly on GnRH neurons is not well established. In some neurons, steroid metabolites known as neurosteroids modulate the function of the GABAA receptor. Specifically, the progesterone derivative allopregnanolone is an allosteric agonist at this receptor, whereas the androgen dehydroepiandrosterone sulfate (DHEAS) is an allosteric antagonist. We hypothesized these metabolites act similarly on GnRH neurons to modify the response to GABA. Whole-cell voltage-clamp recordings of GABAergic miniature postsynaptic currents (mPSCs) were made from green fluorescent protein-identified GnRH neurons in brain slices from diestrous mice. Glutamatergic currents were blocked with antagonists and action potentials blocked with tetrodotoxin, minimizing presynaptic effects of treatments. Allopregnanolone (5 μm) increased mPSC rate of rise, amplitude and decay time by 15.9 ± 6.1%, 16.5 ± 6.3%, and 58.3 ± 18.6%, respectively (n = 7 cells). DHEAS (5 μm) reduced mPSC rate of rise (32.1 ± 5.7%) and amplitude (27.6 ± 4.3%) but did not alter decay time (n = 8). Effects of both neurosteroids were dose dependent between 0.1 and 10 μm. In addition to independent actions, DHEAS also reversed effects of allopregnanolone on rate of rise and amplitude so that these parameters were returned to pretreatment baseline values (n = 6). These data indicate allopregnanolone increases and DHEAS decreases responsiveness of GnRH neurons to activation of GABAA receptors by differentially modulating current flow through GABAA receptor chloride channels. This provides one mechanism for direct steroid feedback to GnRH neurons.

scholarly journals Origin of Neuropeptide Y-Containing Afferents to Gonadotropin-Releasing Hormone Neurons in Male Mice

Endocrinology ◽  
2003 ◽  
Vol 144 (11) ◽  
pp. 4967-4974 ◽  
Author(s):  
Gergely F. Turi ◽  
Zsolt Liposits ◽  
Suzanne M. Moenter ◽  
Csaba Fekete ◽  
Erik Hrabovszky
Keyword(s):  
Green Fluorescent Protein ◽  
Neuropeptide Y ◽  
Enhanced Green Fluorescent Protein ◽  
Fluorescent Protein ◽  
Monosodium Glutamate ◽  
Male Mice ◽  
Combined Application ◽  
Gnrh Neurons ◽  
Arcuate Nuclei ◽  
Green Fluorescent

Abstract The origin of neuropeptide Y (NPY) afferents to GnRH neurons was investigated in male mice. Neonatal lesioning of the hypothalamic arcuate nuclei (ARC) with monosodium glutamate markedly reduced the number of NPY fibers in the preoptic area as well as the frequency of their contacts with perikarya and proximal dendrites of GnRH neurons. Dual-label immunofluorescence studies to determine the precise contribution of the ARC to the innervation of GnRH neurons by NPY axons were carried out on transgenic mice in which enhanced green fluorescent protein was expressed under the control of the GnRH promoter (GnRH-enhanced green fluorescent protein mice). The combined application of red Cy3 and blue AMCA fluorochromogenes established that 49.1 ± 7.3% of NPY axons apposed to green GnRH neurons also contained agouti-related protein (AGRP), a selective marker for NPY axons arising from the ARC. Immunoelectronmicroscopic analysis detected symmetric synapses between AGRP fibers and GnRH-immunoreactive perikarya. Additional triple-fluorescence experiments revealed the presence of dopamine-β-hydroxylase immunoreactivity within 25.4 ± 3.3% of NPY afferents to GnRH neurons. This enzyme marker enabled the selective labeling of NPY pathways ascending from noradrenergic/adrenergic cell populations of the brain stem, thus defining a second important source for NPY-containing fibers regulating GnRH cells. The absence of both topographic markers (AGRP and dopamine-β-hydroxylase) within 26% of NPY contacts suggests that additional sources of NPY fibers to GnRH neurons exist. Future studies will address distinct functions of the two identified NPY systems in the afferent neuronal regulation of the GnRH system.

scholarly journals Norepinephrine Suppresses Gonadotropin-Releasing Hormone Neuron Excitability in the Adult Mouse

Endocrinology ◽  
2007 ◽  
Vol 149 (3) ◽  
pp. 1129-1135 ◽  
Author(s):  
Seong-Kyu Han ◽  
Allan E. Herbison
Keyword(s):  
Adrenergic Receptors ◽  
Adult Mouse ◽  
Fluorescent Protein ◽  
Receptor Activation ◽  
Electrical Excitability ◽  
Gonadotropin Secretion ◽  
Membrane Hyperpolarization ◽  
Gnrh Neurons ◽  
Green Fluorescent ◽  
Β Adrenergic Receptors

Norepinephrine (NE) is considered to exert an important modulatory influence upon the activity of GnRH neurons. In the present study, we used a transgenic GnRH-green fluorescent protein mouse model to examine the effects of NE on the electrical excitability of GnRH neurons in male and female mice. Gramicidin-perforated patch recordings demonstrated that NE (10–100 μm) exerted a robust membrane hyperpolarization, with associated suppression of firing, in more than 85% of male prepubertal and adult GnRH neurons (n = 25). The same hyperpolarizing action was observed in female GnRH neurons from diestrous (91%, n = 11), proestrous (50%, n = 14), estrous (77%, n = 13), and ovariectomized (82%, n = 11) mice. A subpopulation (<10%) of silent GnRH neurons in all groups responded to NE with hyperpolarization followed by the initiation of firing upon NE washout. The hyperpolarizing actions of NE were mimicked by α1-adrenergic (phenylephrine) and β-adrenergic (isoproterenol) receptor agonists, but α2 receptor activation (guanabenz) had no effect. Approximately 75% of the NE-evoked hyperpolarization was blocked by the α1 receptor antagonist prazosin, and 75% of GnRH neurons responded to both phenylephrine and isoproterenol. These findings indicate that NE acts through both α1- and β-adrenergic receptors located on the soma/dendrites of GnRH neurons to directly suppress their excitability throughout the estrous cycle and after ovariectomy. These data force a reanalysis of existing models explaining the effects of NE on gonadotropin secretion.

scholarly journals Adiponectin receptors are expressed in hypothalamus and colocalized with proopiomelanocortin and neuropeptide Y in rodent arcuate neurons

2008 ◽  
Vol 200 (1) ◽  
pp. 93-105 ◽  
Author(s):  
E Guillod-Maximin ◽  
A F Roy ◽  
C M Vacher ◽  
A Aubourg ◽  
V Bailleux ◽  
...  
Keyword(s):  
Neuropeptide Y ◽  
Energy Homeostasis ◽  
Fluorescent Protein ◽  
Peripheral Tissues ◽  
Rat Hypothalamus ◽  
Ampk Phosphorylation ◽  
Hypothalamic Nuclei ◽  
Green Fluorescent ◽  
Amp Activated Protein Kinase

Adiponectin is involved in the control of energy homeostasis in peripheral tissues through Adipor1 and Adipor2 receptors. An increasing amount of evidence suggests that this adipocyte-secreted hormone may also act at the hypothalamic level to control energy homeostasis. In the present study, we observed the gene and protein expressions of Adipor1 and Adipor2 in rat hypothalamus using different approaches. By immunohistochemistry, Adipor1 expression was ubiquitous in the rat brain. By contrast, Adipor2 expression was more limited to specific brain areas such as hypothalamus, cortex, and hippocampus. In arcuate and paraventricular hypothalamic nuclei, Adipor1, and Adipor2 were expressed by neurons and astrocytes. Furthermore, using transgenic green fluorescent protein mice, we showed that Adipor1 and Adipor2 were present in pro–opiomelanocortin (POMC) and neuropeptide Y (NPY) neurons in the arcuate nucleus. Finally, adiponectin treatment by intracerebroventricular injection induced AMP-activated protein kinase (AMPK) phosphorylation in the rat hypothalamus. This was confirmed byin vitrostudies using hypothalamic membrane fractions. In conclusion, Adipor1 and Adipor2 are both expressed by neurons (including POMC and NPY neurons) and astrocytes in the rat hypothalamic nuclei. Adiponectin is able to increase AMPK phosphorylation in the rat hypothalamus. These data reinforced a potential role of adiponectin and its hypothalamic receptors in the control of energy homeostasis.

scholarly journals Effects of acute and chronic nicotine on catecholamine neurons of the nucleus of the solitary tract

2019 ◽  
Vol 316 (1) ◽  
pp. R38-R49
Author(s):  
Stephen J. Page ◽  
Mingyan Zhu ◽  
Suzanne M. Appleyard
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Fluorescent Protein ◽  
Brain Slices ◽  
Nicotine Withdrawal ◽  
Cardiovascular Reflexes ◽  
Solitary Tract ◽  
Catecholamine Neurons ◽  
Green Fluorescent ◽  
Tyrosine Hydroxylase Promoter

Nicotine is an addictive drug that has broad effects throughout the brain. One site of action is the nucleus of the solitary tract (NTS), where nicotine initiates a stress response and modulates cardiovascular and gastric function through nicotinic acetylcholine receptors (nAChRs). Catecholamine (CA) neurons in the NTS influence stress and gastric and cardiovascular reflexes, making them potential mediators of nicotine’s effects; however nicotine’s effect on these neurons is unknown. Here, we determined nicotine’s actions on NTS-CA neurons by use of patch-clamp techniques in brain slices from transgenic mice expressing enhanced green fluorescent protein driven by the tyrosine hydroxylase promoter (TH-EGFP). Picospritzing nicotine both induced a direct inward current and increased the frequency of spontaneous excitatory postsynaptic currents (sEPSCs) in NTS-CA neurons, effects blocked by nonselective nAChR antagonists TMPH and MLA. The increase in sEPSC frequency was mimicked by nAChRα7 agonist AR-R17779 and blocked by nAChRα7 antagonist MG624. AR-R17779 also increased the firing of TH-EGFP neurons, an effect dependent on glutamate inputs, as it was blocked by the glutamate antagonist NBQX. In contrast, the nicotine-induced current was mimicked by nAChRα4β2 agonist RJR2403 and blocked by nAChRα4β2 antagonist DHβE. RJR2403 also increased the firing rate of TH-EGFP neurons independently of glutamate. Finally, both somatodendritic and sEPSC nicotine responses from NTS-CA neurons were larger in nicotine-dependent mice that had under gone spontaneous nicotine withdrawal. These results demonstrate that 1) nicotine activates NTS-CA neurons both directly, by inducing a direct current, and indirectly, by increasing glutamate inputs, and 2) NTS-CA nicotine responsiveness is altered during nicotine withdrawal.

scholarly journals Activation of Neurokinin 3 Receptors Stimulates GnRH Release in a Location-Dependent but Kisspeptin-Independent Manner in Adult Mice

Endocrinology ◽  
2013 ◽  
Vol 154 (11) ◽  
pp. 3984-3989 ◽  
Author(s):  
Garrett T. Gaskins ◽  
Katarzyna M. Glanowska ◽  
Suzanne M. Moenter
Keyword(s):  
Green Fluorescent Protein ◽  
Carbon Fiber ◽  
Fluorescent Protein ◽  
Hypogonadotropic Hypogonadism ◽  
Brain Slices ◽  
Dependent Manner ◽  
Gnrh Secretion ◽  
Gnrh Release ◽  
Carbon Fiber Microelectrodes ◽  
Green Fluorescent

GnRH neurons form the final common pathway for the central control of reproduction. GnRH release occurs from terminals in the external layer of the median eminence (ME) for neuroendocrine control of the pituitary, and near GnRH-GnRH fiber appositions within the preoptic area (POA). Whether or not control of GnRH secretion by neuromodulators is different in these 2 areas is unknown. Mutations in neurokinin B (NKB) or the neurokinin-3 receptor (NK3R) are linked to hypogonadotropic hypogonadism in humans, suggesting that NKB may regulate GnRH secretion. Using fast scan cyclic voltammetry through carbon-fiber microelectrodes, we examined real-time GnRH release in response to the NK3R agonist senktide in the ME and POA. Coronal brain slices were acutely prepared from adult gonad-intact GnRH-green fluorescent protein male mice, and carbon-fiber microelectrodes were placed either within green fluorescent protein-positive terminal fields of the ME or near GnRH-GnRH fiber appositions in the POA. Senktide induced GnRH release consistently in the ME but not the POA, indicating that GnRH release is differentially regulated by NKB in a location-dependent manner. Senktide also induced GnRH secretion in the ME of kisspeptin-knockout (Kiss1 knockout) mice. Interestingly, release amplitude was lower compared with wild-type mice. These data indicate regulation of GnRH release by NK3R agonists is site specific and suggest that kisspeptin is not a required mediator between NK3R activation and GnRH secretion in the ME. This information will be useful for informing future models of afferent regulation of GnRH release.

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