scholarly journals In vivo imaging of the GnRH pulse generator reveals a temporal order of neuronal activation and synchronization during each pulse

2021 ◽  
Author(s):  
Aleisha M Moore ◽  
Lique M Coolen ◽  
Michael N Lehman
Keyword(s):  
Temporal Order ◽  
Pulse Generator ◽  
Polycystic Ovary ◽  
Temporal Organization ◽  
Neuron Activity ◽  
Hypothalamic Amenorrhea ◽  
Freely Moving ◽  
Kndy Neurons ◽  
In Vivo Calcium Imaging

A hypothalamic pulse generator located in the arcuate nucleus controls episodic release of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) and is essential for reproduction. Recent evidence suggests this generator is comprised of arcuate 'KNDy' cells, the abbreviation based on co-expression of kisspeptin, neurokinin B, and dynorphin. However, direct visual evidence of KNDy neuron activity at a single-cell level during a pulse is lacking. Here, we use in vivo calcium imaging in freely moving female mice to show that individual KNDy neurons are synchronously activated in an episodic manner, and these synchronized episodes always precede LH pulses. Furthermore, synchronization among KNDy cells occurs in a temporal order, with some subsets of KNDy cells serving as 'leaders' and others as 'followers' during each synchronized episode. These results reveal an unsuspected temporal organization of activation and synchronization within the GnRH pulse generator, suggesting that different subsets of KNDy neurons are activated at pulse onset than afterward during maintenance and eventual termination of each pulse. Further studies to distinguish KNDy leader from follower cells is likely to have important clinical significance, since regulation of pulsatile GnRH secretion is essential for normal reproduction and disrupted in pathological conditions such as polycystic ovary syndrome and hypothalamic amenorrhea.

scholarly journals Suppression of the GnRH Pulse Generator by Neurokinin B Involves a κ-Opioid Receptor-Dependent Mechanism

Endocrinology ◽  
2012 ◽  
Vol 153 (10) ◽  
pp. 4894-4904 ◽  
Author(s):  
P. Grachev ◽  
X. F. Li ◽  
J. S. Kinsey-Jones ◽  
A. L. di Domenico ◽  
R. P. Millar ◽  
...  
Keyword(s):  
Opioid Receptor ◽  
Pulse Generator ◽  
Pulse Frequency ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Neurokinin B ◽  
Kndy Neurons ◽  
Lh Secretion ◽  
Dose Dependent

Abstract Neurokinin B (NKB) and its receptor (NK3R) are coexpressed with kisspeptin, Dynorphin A (Dyn), and their receptors [G-protein-coupled receptor-54 (GPR54)] and κ-opioid receptor (KOR), respectively] within kisspeptin/NKB/Dyn (KNDy) neurons in the hypothalamic arcuate nucleus (ARC), the proposed site of the GnRH pulse generator. Much previous research has employed intracerebroventricular (icv) administration of KNDy agonists and antagonists to address the functions of KNDy neurons. We performed a series of in vivo neuropharmacological experiments aiming to determine the role of NKB/NK3R signaling in modulating the GnRH pulse generator and elucidate the interaction between KNDy neuropeptide signaling systems, targeting our interventions to ARC KNDy neurons. First, we investigated the effect of intra-ARC administration of the selective NK3R agonist, senktide, on pulsatile LH secretion using a frequent automated serial sampling method to obtain blood samples from freely moving ovariectomized 17β-estradiol-replaced rats. Our results show that senktide suppresses LH pulses in a dose-dependent manner. Intra-ARC administration of U50488, a selective KOR agonist, also caused a dose-dependent, albeit more modest, decrease in LH pulse frequency. Thus we tested the hypothesis that Dyn/KOR signaling localized to the ARC mediates the senktide-induced suppression of the LH pulse by profiling pulsatile LH secretion in response to senktide in rats pretreated with nor-binaltorphimine, a selective KOR antagonist. We show that nor-binaltorphimine blocks the senktide-induced suppression of pulsatile LH secretion but does not affect LH pulse frequency per se. In order to address the effects of acute activation of ARC NK3R, we quantified (using quantitative RT-PCR) changes in mRNA levels of KNDy-associated genes in hypothalamic micropunches following intra-ARC administration of senktide. Senktide down-regulated expression of genes encoding GnRH and GPR54 (GNRH1 and Kiss1r, respectively), but did not affect the expression of Kiss1 (which encodes kisspeptin). We conclude that NKB suppresses the GnRH pulse generator in a KOR-dependent fashion and regulates gene expression in GnRH neurons.

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

2021 ◽  
Author(s):  
Charlotte Vanacker ◽  
R. Anthony DeFazio ◽  
Charlene M. Sykes ◽  
Suzanne M. Moenter
Keyword(s):  
Arcuate Nucleus ◽  
Designer Drugs ◽  
Neuron Activity ◽  
Gnrh Neurons ◽  
Hypothalamic Arcuate Nucleus ◽  
Lh Release ◽  
Main Regulator ◽  
Kndy Neurons

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

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.

scholarly journals Modulation of pulsatile GnRH dynamics across the ovarian cycle via changes in the network excitability and basal activity of the arcuate kisspeptin network

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Margaritis Voliotis ◽  
Xiao Feng Li ◽  
Ross Alexander De Burgh ◽  
Geffen Lass ◽  
Deyana Ivanova ◽  
...  
Keyword(s):  
Pulse Generator ◽  
Reproductive Function ◽  
Pulse Generation ◽  
Ovarian Cycle ◽  
Gonadal Steroids ◽  
Pulsatile Gnrh ◽  
Gnrh Release ◽  
Kndy Neurons ◽  
Stimulation Of

Pulsatile 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. To better understand these mechanisms we start by perturbing the KNDy system at different stages of the estrous cycle using optogenetics. We find that optogenetic stimulation of KNDy neurons stimulates pulsatile GnRH/LH secretion in estrous mice but inhibits it in diestrous mice. These in-vivo results in combination with mathematical modelling suggest that the transition between estrus and diestrus is underpinned by well-orchestrated changes in neuropeptide signalling and in the excitability of the KNDy population controlled via glutamate signalling. Guided by model predictions, we show that blocking glutamate signalling 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 pulse generator dynamics. Finally, our results could inform useful repurposing of drugs targeting the glutamate system in reproductive therapy.

scholarly journals In Vivo Multi-Day Calcium Imaging of CA1 Hippocampus in Freely Moving Rats Reveals a High Preponderance of Place Cells and No Detectable Photobleaching

2021 ◽  
Author(s):  
Hannah S Wirtshafter ◽  
John F Disterhoft
Keyword(s):  
Calcium Imaging ◽  
Place Cells ◽  
Freely Moving Rats ◽  
Calcium Indicators ◽  
Large Populations ◽  
Freely Moving ◽  
Cell Changes ◽  
Long Timescales ◽  
In Vivo Calcium Imaging

Calcium imaging using GCaMP calcium indicators and miniature microscopes has been used to image cellular populations during long timescales and in different task phases, as well as to determine neuronal circuit topology and organization. Because the hippocampus (HPC) is essential for many tasks of memory, spatial navigation, and learning, calcium imaging of large populations of HPC neurons can provide new insight on cell changes and organization over time during these tasks. To our knowledge, all reported HPC in vivo calcium imaging experiments have been done in mouse. However, rats have many behavioral and physiological experimental advantages over mice, and, due to their larger size, rats are able to support larger implants, thereby enabling the recording of a greater number of cells. In this paper, we present the first in vivo calcium imaging from CA1 hippocampus in freely moving rats. Using GCaMP7c and the UCLA Miniscope, we demonstrate that hundreds of cells (mean 240+-90 cells per session, maximum 428 cells) can reliably be visualized and held across weeks, and that calcium events in these cells are correlated with periods of movement. We additionally show proof of method by showing that an extremely high percent of place cells (82.3%+-8.1%, far surpassing the percent seen during mouse calcium imaging) can be recorded on a navigational task, and that these place cells enable accurately decoding of animal position. Finally, we show that calcium imaging is rats is not prone to photobleaching during hour-long recordings and that cells can be reliably recorded for an hour or more per session. A detailed protocol for this technique, including notes on the numerous parameter changes needed to use Ca2+ in rats, is included in the Materials and Methods section, and implications of these advancements are discussed.

scholarly journals Aversive emotion rapidly activates orexin neurons and increases heart rate in freely moving mice

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Akira Yamashita ◽  
Shunpei Moriya ◽  
Ryusei Nishi ◽  
Jun Kaminosono ◽  
Akihiro Yamanaka ◽  
...  
Keyword(s):  
Heart Rate ◽  
Neuronal Activity ◽  
Defense Response ◽  
Aversive Stimulus ◽  
Critical Role ◽  
Neuron Activity ◽  
Dual Channel ◽  
Arterial Blood ◽  
Freely Moving

AbstractThe perifornical area of the hypothalamus has been known as the center for the defense response, or fight-or-flight response, which is characterized by a concomitant rise in arterial blood pressure, heart rate, and respiratory frequency. It is well established that orexin neurons, which are located in this region, play a critical role in this response. In this study, we further examined this role by recording orexin neuronal activity and heart rate in freely moving mice using an original dual-channel fiber photometry system in vivo. Analysis of orexin neuron activity in relation to autonomic responses to aversive stimuli revealed a rapid increase in neuronal activity just prior to changes in heart rate. In addition, we examined whether orexin neurons would be activated by a conditioned neutral sound that was previously associated with aversive stimulus. We show that the memory of the aversive stimulus activated orexin neurons and increased heart rate. Our data suggest that orexin neurons are a key component linking aversive emotions to autonomic defense response. Our data also suggest that targeting orexin neurons may enable treatment of psychiatric disorders associated with chronic stress and traumatic memories.

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

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Charlotte Vanacker ◽  
Richard Anthony Defazio ◽  
Charlene M Sykes ◽  
Suzanne M Moenter
Keyword(s):  
Arcuate Nucleus ◽  
Designer Drugs ◽  
Neuron Activity ◽  
Gnrh Neurons ◽  
Hypothalamic Arcuate Nucleus ◽  
Lh Release ◽  
Main Regulator ◽  
Kndy Neurons

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

scholarly journals Novel nose poke-based temporal discrimination tasks with concurrent in vivo calcium imaging in freely moving mice

2019 ◽  
Vol 12 (1) ◽  
Author(s):  
William D. Marks ◽  
Hisayuki Osanai ◽  
Jun Yamamoto ◽  
Sachie K. Ogawa ◽  
Takashi Kitamura
Keyword(s):  
Calcium Imaging ◽  
Discrimination Task ◽  
Animal Movement ◽  
Temporal Discrimination ◽  
Cell Activity ◽  
Memory Formation ◽  
Nose Poke ◽  
Freely Moving ◽  
In Vivo Calcium Imaging

Abstract The hippocampus has been known to process temporal information as part of memory formation. While time cells have been observed in the hippocampus and medial entorhinal cortex, a number of the behavioral tasks used present potential confounds that may cause some contamination of time cell observations due to animal movement. Here, we report the development of a novel nose poke-based temporal discrimination task designed to be used with in vivo calcium imaging for the analysis of hippocampal time cells in freely moving mice. First, we developed a ten second held nose poke paradigm for use in mice to deliver a purer time metric for the analysis of time cell activity in hippocampus CA1. Second, we developed a temporal discrimination task that involves the association of held nose poke durations of differing lengths with differential spatial cues presented in arms on a linear I-maze. Four of five mice achieved successful temporal discrimination within three weeks. Calcium imaging has been successfully performed in each of these tasks, with time cell activity being detected in the 10s nose poke task, and calcium waves being observed in discrete components of the temporal discrimination task. The newly developed behavior tasks in mice serve as novel tools to accelerate the study of time cell activity and examine the integration of time and space in episodic memory formation.

scholarly journals Prenatal androgen treatment does not alter the firing activity of hypothalamic arcuate kisspeptin neurons in female mice

2021 ◽  
Author(s):  
Amanda G Gibson ◽  
Jennifer Jaime ◽  
Laura L Burger ◽  
Suzanne M Moenter
Keyword(s):  
Firing Rate ◽  
Polycystic Ovary ◽  
Developmental Trajectory ◽  
Dependent Manner ◽  
Gnrh Neuron ◽  
Neuron Firing ◽  
Firing Rates ◽  
Gnrh Neurons ◽  
Kndy Neurons

Neuroendocrine control of reproduction is disrupted in many individuals with polycystic ovary syndrome, who present with increased luteinizing hormone (LH), and presumably gonadotropin-releasing hormone (GnRH), release frequency, and high androgen levels. Prenatal androgenization (PNA) recapitulates these phenotypes in primates and rodents. Female offspring of mice injected with dihydrotestosterone (DHT) on gestational D16-18 exhibit disrupted estrous cyclicity, increased LH and testosterone, and increased GnRH neuron firing rate as adults. PNA also alters the developmental trajectory of GnRH neuron firing rates, markedly blunting the prepubertal peak in firing that occurs in 3wk-old controls. GnRH neurons do not express detectable androgen receptors and are thus probably not the direct target of DHT. Rather, PNA likely alters GnRH neuronal activity by modulating upstream neurons, such as hypothalamic arcuate neurons co-expressing kisspeptin, neurokinin B (gene Tac2), and dynorphin, aka KNDy neurons. We hypothesized PNA treatment changes firing rates of KNDy neurons in a similar age-dependent manner as GnRH neurons. We conducted targeted extracellular recordings (0.5-2h) of Tac2-identified KNDy neurons from control and PNA mice at 3wks of age and in adulthood. About half of neurons were quiescent (<0.005Hz). Long-term firing rates of active cells varied, suggestive of episodic activity, but were not different among groups. Short-term burst firing was also similar. We thus reject the hypothesis that PNA alters the firing rate of KNDy neurons. This does not preclude altered neurosecretory output of KNDy neurons, involvement of other neuronal populations, or in-vivo networks as critical drivers of altered GnRH firing rates in PNA mice.

scholarly journals Hippocampal Interneurons are Required for Trace Eyeblink Conditioning in Mice

2021 ◽  
Author(s):  
Wei-Wei Zhang ◽  
Rong-Rong Li ◽  
Jie Zhang ◽  
Jie Yan ◽  
Qian-Hui Zhang ◽  
...  
Keyword(s):  
Eyeblink Conditioning ◽  
Pyramidal Cells ◽  
Conditioned Eyeblink ◽  
Cellular Mechanisms ◽  
Sustained Activity ◽  
Early Acquisition ◽  
Freely Moving ◽  
Trace Eyeblink Conditioning

AbstractWhile the hippocampus has been implicated in supporting the association among time-separated events, the underlying cellular mechanisms have not been fully clarified. Here, we combined in vivo multi-channel recording and optogenetics to investigate the activity of hippocampal interneurons in freely-moving mice performing a trace eyeblink conditioning (tEBC) task. We found that the hippocampal interneurons exhibited conditioned stimulus (CS)-evoked sustained activity, which predicted the performance of conditioned eyeblink responses (CRs) in the early acquisition of the tEBC. Consistent with this, greater proportions of hippocampal pyramidal cells showed CS-evoked decreased activity in the early acquisition of the tEBC. Moreover, optogenetic suppression of the sustained activity in hippocampal interneurons severely impaired acquisition of the tEBC. In contrast, suppression of the sustained activity of hippocampal interneurons had no effect on the performance of well-learned CRs. Our findings highlight the role of hippocampal interneurons in the tEBC, and point to a potential cellular mechanism subserving associative learning.

Sign in / Sign up

Export Citation Format

Share Document