The dendron and episodic neuropeptide release

Abstract: The unexpected observation that the long processes of gonadotrophin‐releasing hormone (GnRH) neurons not only conducted action potentials, but also operated to integrate afferent information at their distal‐most extent gave rise to the concept of a blended dendritic‐axonal process termed the “dendron”. The proximal dendrites of the GnRH neuron function in a conventional manner, receiving synaptic inputs and initiating action potentials that are critical for the surge mode of GnRH secretion. The distal dendrons are regulated by both classical synapses and volume transmission and likely operate using subthreshold electrotonic propagation into the nearby axon terminals in the median eminence. Evidence indicates that neural processing at the distal dendron is responsible for the pulsatile patterning of GnRH secretion. Although the dendron remains unique to the GnRH neuron, data show that it exists in both mice and rats and may be a common feature of mammalian species in which GnRH neuron cell bodies do not migrate into the basal hypothalamus. This review outlines the discovery and function of the dendron as a unique neuronal structure optimised to generate episodic neuronal output

Morphological assessment of GABA and glutamate inputs to GnRH neurons in intact female mice using expansion microscopy

Funder: Wellcome TrustFunder: Health Research Council of New Zealand; Id: http://dx.doi.org/10.13039/501100001505Abstract: The roles GABAergic and glutamatergic inputs in regulating the activity of the gonadotrophin‐releasing hormone (GnRH) neurons at the time of the preovulatory surge remain unclear. We used expansion microscopy to compare the density of GABAergic and glutamatergic synapses on the GnRH neuron cell body and proximal dendrite in dioestrous and pro‐oestrous female mice. An evaluation of all synapses immunoreactive for synaptophysin revealed that the highest density of inputs to rostral preoptic area GnRH neurons occurred within the first 45 µm of the primary dendrite (approximately 0.19 synapses µm‐1) with relatively few synapses on the GnRH neuron soma or beyond 45 µm of the dendrite (0.05‐0.08 synapses µm‐1). Triple immunofluorescence labelling demonstrated a predominance of glutamatergic signalling with twice as many vesicular glutamate transporter 2 synapses detected compared to vesicular GABA transporter. Co‐labelling with the GABAA receptor scaffold protein gephyrin and the glutamate receptor postsynaptic density marker Homer1 confirmed these observations, as well as the different spatial distribution of GABA and glutamate inputs along the dendrite. Quantitative assessments revealed no differences in synaptophysin, GABA or glutamate synapses at the proximal dendrite and soma of GnRH neurons between dioestrous and pro‐oestrous mice. Taken together, these studies demonstrate that the GnRH neuron receives twice as many glutamatergic synapses compared to GABAergic synapses and that these inputs preferentially target the first 45 µm of the GnRH neuron proximal dendrite. These inputs appear to be structurally stable before the onset of pro‐oestrous GnRH surge

Highly redundant neuropeptide volume co-transmission underlying episodic activation of the GnRH neuron dendron.

The necessity and functional significance of neurotransmitter co-transmission remains unclear. The glutamatergic 'KNDy' neurons co-express kisspeptin, neurokinin B (NKB), and dynorphin and exhibit a highly stereotyped synchronized behavior that reads out to the gonadotropin-releasing hormone (GnRH) neuron dendrons to drive episodic hormone secretion. Using expansion microscopy, we show that KNDy neurons make abundant close, non-synaptic appositions with the GnRH neuron dendron. Electrophysiology and confocal GCaMP6 imaging demonstrated that, despite all three neuropeptides being released from KNDy terminals, only kisspeptin was able to activate the GnRH neuron dendron. Mice with a selective deletion of kisspeptin from KNDy neurons failed to exhibit pulsatile hormone secretion but maintained synchronized episodic KNDy neuron behavior that is thought to depend on recurrent NKB and dynorphin transmission. This indicates that KNDy neurons drive episodic hormone secretion through highly redundant neuropeptide co-transmission orchestrated by differential post-synaptic neuropeptide receptor expression at the GnRH neuron dendron and KNDy neuron

Highly redundant neuropeptide volume co-transmission underlying episodic activation of the GnRH neuron dendron.

The necessity and functional significance of neurotransmitter co-transmission remains unclear. The glutamatergic 'KNDy' neurons co-express kisspeptin, neurokinin B (NKB), and dynorphin and exhibit a highly stereotyped synchronized behavior that reads out to the gonadotropin-releasing hormone (GnRH) neuron dendrons to drive episodic hormone secretion. Using expansion microscopy, we show that KNDy neurons make abundant close, non-synaptic appositions with the GnRH neuron dendron. Electrophysiology and confocal GCaMP6 imaging demonstrated that, despite all three neuropeptides being released from KNDy terminals, only kisspeptin was able to activate the GnRH neuron dendron. Mice with a selective deletion of kisspeptin from KNDy neurons failed to exhibit pulsatile hormone secretion but maintained synchronized episodic KNDy neuron behavior that is thought to depend on recurrent NKB and dynorphin transmission. This indicates that KNDy neurons drive episodic hormone secretion through highly redundant neuropeptide co-transmission orchestrated by differential post-synaptic neuropeptide receptor expression at the GnRH neuron dendron and KNDy neuron

Female sexual behavior in mice is controlled by kisspeptin neurons.

Sexual behavior is essential for the survival of many species. In female rodents, mate preference and copulatory behavior depend on pheromones and are synchronized with ovulation to ensure reproductive success. The neural circuits driving this orchestration in the brain have, however, remained elusive. Here, we demonstrate that neurons controlling ovulation in the mammalian brain are at the core of a branching neural circuit governing both mate preference and copulatory behavior. We show that male odors detected in the vomeronasal organ activate kisspeptin neurons in female mice. Classical kisspeptin/Kiss1R signaling subsequently triggers olfactory-driven mate preference. In contrast, copulatory behavior is elicited by kisspeptin neurons in a parallel circuit independent of Kiss1R involving nitric oxide signaling. Consistent with this, we find that kisspeptin neurons impinge onto nitric oxide-synthesizing neurons in the ventromedial hypothalamus. Our data establish kisspeptin neurons as a central regulatory hub orchestrating sexual behavior in the female mouse brain

Definition of Estrogen Receptor Pathway Critical for Estrogen Positive Feedback to Gonadotropin-Releasing Hormone Neurons and Fertility

SummaryThe mechanisms through which estrogen regulates gonadotropin-releasing hormone (GnRH) neurons to control mammalian ovulation are unknown. We found that estrogen positive feedback to generate the preovulatory gonadotropin surge was normal in estrogen receptor β knockout (ERβ) mutant mice, but absent in ERα mutant mice. An ERα-selective compound was sufficient to generate positive feedback in wild-type mice. As GnRH neurons do not express ERα, estrogen positive feedback upon GnRH neurons must be indirect in nature. To establish the cell type responsible, we generated a neuron-specific ERα mutant mouse line. These mice failed to exhibit estrogen positive feedback, demonstrating that neurons expressing ERα are critical. We then used a GnRH neuron-specific Pseudorabies virus (PRV) tracing approach to show that the ERα-expressing neurons innervating GnRH neurons are located within rostral periventricular regions of the hypothalamus. These studies demonstrate that ovulation is driven by estrogen actions upon ERα-expressing neuronal afferents to GnRH neurons

Defining a novel leptin–melanocortin–kisspeptin pathway involved in the metabolic control of puberty

Objective Puberty is a key developmental phenomenon highly sensitive to metabolic modulation. Worrying trends of changes in the timing of puberty have been reported in humans. These might be linked to the escalating prevalence of childhood obesity and could have deleterious impacts on later (cardio-metabolic) health, but their underlying mechanisms remain unsolved. The neuropeptide α-MSH, made by POMC neurons, plays a key role in energy homeostasis by mediating the actions of leptin and likely participates in the control of reproduction. However, its role in the metabolic regulation of puberty and interplay with kisspeptin, an essential puberty-regulating neuropeptide encoded by Kiss1, remain largely unknown. We aim here to unveil the potential contribution of central α-MSH signaling in the metabolic control of puberty by addressing its role in mediating the pubertal effects of leptin and its potential interaction with kisspeptin. Methods Using wild type and genetically modified rodent models, we implemented pharmacological studies, expression analyses, electrophysiological recordings, and virogenetic approaches involving DREADD technology to selectively inhibit Kiss1 neurons, in order to interrogate the physiological role of a putative leptin→α-MSH→kisspeptin pathway in the metabolic control of puberty. Results Stimulation of central α-MSH signaling robustly activated the reproductive axis in pubertal rats, whereas chronic inhibition of melanocortin receptors MC3/4R, delayed puberty, and prevented the permissive effect of leptin on puberty onset. Central blockade of MC3/4R or genetic elimination of kisspeptin receptors from POMC neurons did not affect kisspeptin effects. Conversely, congenital ablation of kisspeptin receptors or inducible, DREADD-mediated inhibition of arcuate nucleus (ARC) Kiss1 neurons resulted in markedly attenuated gonadotropic responses to MC3/4R activation. Furthermore, close appositions were observed between POMC fibers and ARC Kiss1 neurons while blockade of α-MSH signaling suppressed Kiss1 expression in the ARC of pubertal rats. Conclusions Our physiological, virogenetic, and functional genomic studies document a novel α-MSH→kisspeptin→GnRH neuronal signaling pathway involved in transmitting the permissive effects of leptin on pubertal maturation, which is relevant for the metabolic (and, eventually, pharmacological) regulation of puberty onsetThis work was supported by grants BFU2011-025021 & BFU2014-57581-P (Ministerio de Economía y Competitividad, Spain; co-funded with EU funds from FEDER Program); project PIE-00005 (Flexi-Met, Instituto de Salud Carlos III, Ministerio de Sanidad, Spain); Projects P08-CVI-03788 and P12-FQM-01943 (Junta de Andalucía, Spain); EU research contract DEER FP7-ENV-2007-1 and the New Zealand Health Research Council. CIBER Fisiopatología de la Obesidad y Nutrición is an initiative of Instituto de Salud Carlos III. Senior authors are indebted with Dr. R.A. Steiner (University of Washington, Seattle, USA) and Dr. U. Boehm (University of Saarland School of Medicine, Homburg, Germany) for provision of relevant mouse lines, essential for conduction of some of the experiments included in this studyS

Mechanism of arcuate kisspeptin neuron synchronization in acute brain slices from female mice

The mechanism by which arcuate kisspeptin (ARNKISS) neurons co-expressing glutamate, neurokinin B, and dynorphin intermittently synchronize their activity to drive pulsatile hormone secretion remains unclear in females. In order to study spontaneous synchronization within the ARNKISS neuron network, acute brain slices were prepared from adult female Kiss1-GCaMP6 mice. Analysis of both spontaneous synchronizations and those driven by high frequency stimulation of individual ARNKISS neurons revealed that the network exhibits semi-random emergent excitation dependent upon glutamate signaling through AMPA receptors. No role for NMDA receptors was identified. In contrast to male mice, ongoing tachykinin receptor tone within the slice operated to promote spontaneous synchronizations in females. As previously observed in males, we found that ongoing dynorphin transmission in the slice did not contribute to synchronization events. These observations indicate that a very similar AMPA receptor-dependent mechanism underlies ARNKISS neuron synchronizations in the female mouse supporting the “glutamate two-transition” model for kisspeptin neuron synchronization. However, a potentially important sex difference appears to exist with a more prominent facilitatory role for tachykinin transmission in the female