Neuroendocrine regulation of reproduction by GnRH neurons: multidisciplinary studies using a small fish brain model.

After the discovery of GnRH, GnRH neurons have been considered to represent the final common pathway for the neural control of reproduction. There is now compelling data in mammals that two populations of kisspeptin neurons constitute two different systems to control the episodic and surge release of GnRH/LH for the control of different aspects of reproduction, follicular development and ovulation. However, accumulating evidence indicates that kisspeptin neurons in non-mammalian species do not serve as a regulator of reproduction, and the non-mammalian species are believed to show only surge release of GnRH to trigger ovulation. Therefore, the GnRH neurons in non-mammalian species may offer simpler models for the study of their functions in neuroendocrine regulation of reproduction, especially ovulation. Our research group has taken advantage of many unique technical advantages of small fish brain for the study of anatomy and physiology of GnRH neurons, which underlie regular ovulatory cycles during the breeding season. Here, recent advances in multidisciplinary study of GnRH neurons are reviewed, with a focus on studies using small teleost fish models.

Emerging roles for kisspeptin in metabolism.

Kisspeptin, a neuropeptide hormone, has been firmly established as a key regulator of the hypothalamic-pituitary-gonadal axis and mammalian reproductive behaviour. In recent years, a growing body of evidence has emerged suggesting a role for kisspeptin in regulating metabolic processes. This data suggest that kisspeptin exerts its metabolic effects indirectly via gonadal hormones and/or directly via the kisspeptin receptor in the brain, pancreas and brown adipose tissue. Kisspeptin receptor knockout studies indicate that kisspeptin may play sexually dimorphic roles in the physiological regulation of energy expenditure, food intake and body weight. Some, but not all, in vitro work demonstrates positive effects on glucose-stimulated insulin secretion, which is more marked at higher kisspeptin concentrations. Acute and chronic in vivo rodent, non-human primate and human studies reveal enhancement of glucose-stimulated insulin secretion in response to pharmacological doses of kisspeptin. Although significant progress has been made in elucidating the metabolic effects of kisspeptin, further mechanistic work and translational studies are required to address unanswered questions and establish the metabolic effects of kisspeptin in diverse human populations (including women, people with obesity and people with diabetes).

Effects of undernutrition and low energy availability on reproductive functions and their underlying neuroendocrine mechanisms.

It has been well established that undernutrition and low energy availability disturb female reproductive functions in humans and many animal species. These reproductive dysfunctions are mainly caused by alterations of some hypothalamic factors, and consequent reduction of gonadotrophin-releasing hormone (GnRH) secretion. Evidence from literature suggests that increased activity of orexigenic factors and decreased activity of anorexigenic/satiety-related factors in undernourished conditions attenuate GnRH secretion in an integrated manner. Likewise, the activity of kisspeptin neurons, which is a potent stimulator of GnRH, is also reduced in undernourished conditions. In addition, it has been suggested that gonadotrophin-inhibitory hormone, which has anti-GnRH and gonadotrophic effects, may be involved in reproductive dysfunctions under several kinds of stress conditions. It should be remembered that these alterations, i.e., promotion of feeding behavior and temporary suppression of reproductive functions, are induced to prioritize the survival of individual over that of species, and that improvements in metabolic and nutritional conditions should be considered with the highest priority.

Neuroendocrine mechanisms of reproductive dysfunctions in undernourished condition.

It is well known that undernourished conditions disturb female reproductive functions in many species, including humans. These alterations are mainly caused by a reduction in gonadotrophin-releasing hormone (GnRH) secretion from the hypothalamus. Evidence from the literature suggests that some hypothalamic factors play pivotal roles in the coordination of reproductive functions and energy homeostasis in response to environmental cues and internal nutritional status. Generally, anorexigenic/satiety-related factors, such as leptin, alpha-melanocyte-stimulating hormone, and proopiomelanocortin, promote GnRH secretion, whereas orexigenic factors, such as neuropeptide Y, agouti-related protein, orexin, and ghrelin, attenuate GnRH secretion. Conversely, gonadotrophin-inhibitory hormone, which exerts anti-GnRH and gonadotrophic effects, promotes feeding behavior in many species. In addition, the activity of kisspeptin, which is a potent stimulator of GnRH, is reduced by undernourished conditions. Under normal nutritional conditions, these factors are coordinated to maintain both feeding behavior and reproductive functions. However, in undernourished conditions their activity levels are markedly altered to promote feeding behavior and temporarily suppress reproductive functions, in order to prioritize the survival of the individual over that of the species.

ERα Signaling in GHRH/Kiss1 Dual-Phenotype Neurons Plays Sex-Specific Roles in Growth and Puberty.

Gonadal steroids modulate growth hormone (GH) secretion and the pubertal growth spurt via undefined central pathways. GH-releasing hormone (GHRH) neurons express estrogen receptor α (ERα) and androgen receptor (AR), suggesting changing levels of gonadal steroids during puberty directly modulate the somatotropic axis. We generated mice with deletion of ERα in GHRH cells (GHRHΔERα), which displayed reduced body length in both sexes. Timing of puberty onset was similar in both groups, but puberty completion was delayed in GHRHΔERα females. Lack of AR in GHRH cells (GHRHΔAR mice) induced no changes in body length, but puberty completion was also delayed in females. Using a mouse model with two reporter genes, we observed that, while GHRHtdTom neurons minimally colocalize with Kiss1hrGFP in prepubertal mice, ∼30% of GHRH neurons coexpressed both reporter genes in adult females, but not in males. Developmental analysis of Ghrh and Kiss1 expression suggested that a subpopulation of ERα neurons in the arcuate nucleus of female mice undergoes a shift in phenotype, from GHRH to Kiss1, during pubertal transition. Our findings demonstrate that direct actions of gonadal steroids in GHRH neurons modulate growth and puberty and indicate that GHRH/Kiss1 dual-phenotype neurons play a sex-specific role in the crosstalk between the somatotropic and gonadotropic axes during pubertal transition.SIGNIFICANCE STATEMENT Late maturing adolescents usually show delayed growth and bone age. At puberty, gonadal steroids have stimulatory effects on the activation of growth and reproductive axes, but the existence of gonadal steroid-sensitive neuronal crosstalk remains undefined. Moreover, the neural basis for the sex differences observed in the clinical arena is unknown. Lack of ERα in GHRH neurons disrupts growth in both sexes and causes pubertal delay in females. Deletion of androgen receptor in GHRH neurons only delayed female puberty. In adult females, not males, a subset of GHRH neurons shift phenotype to start producing Kiss1. Thus, direct estrogen action in GHRH/Kiss1 dual-phenotype neurons modulates growth and puberty and may orchestrate the sex differences in endocrine function observed during pubertal transition.

Leptin and inflammatory factors play a synergistic role in the regulation of reproduction in male mice through hypothalamic kisspeptin-mediated energy balance.

BACKGROUND: Energy balance is closely related to reproductive function, wherein hypothalamic kisspeptin mediates regulation of the energy balance. However, the central mechanism of kisspeptin in the regulation of male reproductive function under different energy balance states is unclear. Here, high-fat diet (HFD) and exercise were used to change the energy balance to explore the role of leptin and inflammation in the regulation of kisspeptin and the hypothalamic-pituitary-testis (HPT) axis. METHODS: Four-week-old male C57BL/6 J mice were randomly assigned to a normal control group (n = 16) or an HFD (n = 49) group. After 10 weeks of HFD feeding, obese mice were randomly divided into obesity control (n = 16), obesity moderate-load exercise (n = 16), or obesity high-load exercise (n = 17) groups. The obesity moderate-load exercise and obesity high-load exercise groups performed exercise (swimming) for 120 min/day and 120 min × 2 times/day (6 h interval), 5 days/week for 8 weeks, respectively. RESULTS: Compared to the mice in the normal group, in obese mice, the mRNA and protein expression of the leptin receptor, kiss, interleukin-10 (IL-10), and gonadotropin-releasing hormone (GnRH) decreased in the hypothalamus; serum luteinizing hormone (LH), follicle-stimulating hormone (FSH), and testosterone levels and sperm quality decreased; and serum leptin, estradiol, and tumor necrosis factor-α (TNF-α) levels and sperm apoptosis increased. Moderate- and high-load exercise effectively reduced body fat and serum leptin levels but had the opposite effects on the hypothalamus and serum IL-10 and TNF-α levels. Moderate-load exercise had anti-inflammatory effects accompanied by increased mRNA and protein expression of kiss and GnRH in the hypothalamus and increased serum FSH, LH, and testosterone levels and improved sperm quality. High-load exercise also promoted inflammation, with no significant effect on the mRNA and protein expression of kiss and GnRH in the hypothalamus, serum sex hormone level, or sperm quality. Moderate-load exercise improved leptin resistance and inflammation and reduced the inhibition of kisspeptin and the HPT axis in obese mice. The inflammatory response induced by high-load exercise may counteract the positive effect of improving leptin resistance on kisspeptin and HPT. CONCLUSION: During changes in energy balance, leptin and inflammation jointly regulate kisspeptin expression on the HPT axis.

Effect of anti-Müllerian hormone on the regulation of pituitary gonadotropin subunit expression: roles of kisspeptin and its receptors in gonadotroph LßT2 cells.

Anti-Müllerian hormone (AMH) is primarily produced by ovarian granulosa cells and contributes to follicle development. AMH is also produced in other tissues, including the brain and pituitary; however, its roles in these tissues are not well understood. In this study, we examined the effect of AMH on pituitary gonadotrophs. We detected AMH and AMH receptor type 2 expression in LßT2 cells. In these cells, the expression of FSHß- but not α- and LHß-subunits increased significantly as the concentration of AMH increased. LßT2 cells expressed Kiss-1 and Kiss-1R. AMH stimulation resulted in decreases in both Kiss-1 and Kiss-1R. The siRNA-mediated knockdown of Kiss-1 in LßT2 cells did not alter the basal expression levels of α-, LHß-, and FSHß-subunits. In LßT2 cells overexpressing Kiss-1R, exogenous kisspeptin stimulation significantly increased the expression of all three gonadotropin subunits. However, kisspeptin-induced increases in these subunits were almost completely eliminated in the presence of AMH. In contrast, GnRH-induced increases in the three gonadotropin subunits were not modulated by AMH. Our observations suggested that AMH acts on pituitary gonadotrophs and induces FSHß-subunit expression with concomitant decreases in Kiss-1 and Kiss-1R gene expression. Kisspeptin, but not GnRH-induced gonadotropin subunit expression, was inhibited by AMH, suggesting that it functions in association with the kisspeptin/Kiss-1R system in gonadotrophs.

Short-term changes and correlations of plasma spexin, kisspeptin, and galanin levels after laparoscopic sleeve gastrectomy.

PURPOSE: To determine the circulating levels of spexin, kisspeptin, galanin, and the correlations between these peptides after laparoscopic sleeve gastrectomy (LSG). METHODS: The plasma levels of the spexin, kisspeptin, and galanin and metabolic parameters (body mass index, weight loss, % excess weight loss, body fat, fasting glucose, HbA1C, and cholesterol levels) were measured (baseline, 1 month, and 3 months) and correlated in thirty adult individuals with obesity (22 female and 8 male) after LSG. RESULTS: The body mass index (BMI), body fat, fasting glucose, total and low-density lipoprotein cholesterol decreased, while high-density lipoprotein cholesterol and % EWL (excess weight loss) increased at 3 months after surgery. The plasma spexin levels increased at 3 months, kisspeptin levels increased at 1 month and stabilized afterward, and galanin levels decreased at 3 months after LSG. Significant correlations were found between metabolic parameters with spexin, kisspeptin, and galanin. In addition, spexin and kisspeptin were negatively correlated with galanin, while spexin was positively correlated with kisspeptin. CONCLUSIONS: The biochemical data reveal evidence that LSG causes an increase in the levels of spexin, and kisspeptin and a decrease in galanin levels. Our findings, therefore, suggest a possible interaction between these novel peptides, which have potential roles in obesity and glucose metabolism.

The Origin of GnRH Pulse Generation: An Integrative Mathematical-Experimental Approach.

Fertility critically depends on the gonadotropin-releasing hormone (GnRH) pulse generator, a neural construct comprised of hypothalamic neurons coexpressing kisspeptin, neurokoinin-B and dynorphin. Here, using mathematical modeling and in vivo optogenetics we reveal for the first time how this neural construct initiates and sustains the appropriate ultradian frequency essential for reproduction. Prompted by mathematical modeling, we show experimentally using female estrous mice that robust pulsatile release of luteinizing hormone, a proxy for GnRH, emerges abruptly as we increase the basal activity of the neuronal network using continuous low-frequency optogenetic stimulation. Further increase in basal activity markedly increases pulse frequency and eventually leads to pulse termination. Additional model predictions that pulsatile dynamics emerge from nonlinear positive and negative feedback interactions mediated through neurokinin-B and dynorphin signaling respectively are confirmed neuropharmacologically. Our results shed light on the long-elusive GnRH pulse generator offering new horizons for reproductive health and wellbeing.SIGNIFICANCE STATEMENT The gonadotropin-releasing hormone (GnRH) pulse generator controls the pulsatile secretion of the gonadotropic hormones LH and FSH and is critical for fertility. The hypothalamic arcuate kisspeptin neurons are thought to represent the GnRH pulse generator, since their oscillatory activity is coincident with LH pulses in the blood; a proxy for GnRH pulses. However, the mechanisms underlying GnRH pulse generation remain elusive. We developed a mathematical model of the kisspeptin neuronal network and confirmed its predictions experimentally, showing how LH secretion is frequency-modulated as we increase the basal activity of the arcuate kisspeptin neurons in vivo using continuous optogenetic stimulation. Our model provides a quantitative framework for understanding the reproductive neuroendocrine system and opens new horizons for fertility regulation.

Kisspeptin induces LH release and ovulation in an induced ovulator†.

Kisspeptin has been implicated in the ovulatory process of several species of spontaneous ovulators but in only one induced ovulator. In contrast, NGF in semen is the principal trigger of ovulation in other species of induced ovulators-camelids. We tested the hypotheses that kisspeptin induces luteinizing hormone (LH) secretion in llamas through a hypothalamic mechanism, and kisspeptin neurons are the target of NGF in its ovulation-inducing pathway. In Experiment 1, llamas were given either NGF, kisspeptin, or saline intravenously, and LH secretion and ovulation were compared among groups. All llamas treated with NGF (5/5) or kisspeptin (5/5) had an elevation of LH blood concentrations after treatment and ovulated, whereas none of the saline group did (0/5). In Experiment 2, llamas were either pretreated with a gonadotropin-releasing hormone (GnRH) receptor antagonist or saline and treated 2 h later with kisspeptin. Llamas pretreated with saline had elevated plasma LH concentrations and ovulated (6/6) whereas llamas pretreated with cetrorelix did not (0/6). In Experiment 3, we evaluated the hypothalamic kisspeptin-GnRH neuronal network by immunohistochemistry. Kisspeptin neurons were detected in the arcuate nucleus, the preoptic area, and the anterior hypothalamus, establishing synaptic contacts with GnRH neurons. We found no colocalization between kisspeptin and NGF receptors by double immunofluorescence. Functional and morphological findings support the concept that kisspeptin is a mediator of the LH secretory pathway in llamas; however, the role of kisspeptins in the NGF ovulation-inducing pathway in camelids remains unclear since NGF receptors were not detected in kisspeptin neurons in the hypothalamus.

Synthesis of Kisspeptin-Mimicking Fragments and Investigation of their Skin Anti-Aging Effects.

In recent years, a number of active materials have been developed to provide anti-aging benefits for skin and, among them, peptides have been considered the most promising candidate due to their remarkable and long-lasting anti-wrinkle activity. Recent studies have begun to elucidate the relationship between the secretion of emotion-related hormones and skin aging. Kisspeptin, a neuropeptide encoded by the KISS1 gene, has gained attention in reproductive endocrinology since it stimulates the reproductive axis in the hypothalamus; however, the effects of Kisspeptin on skin have not been studied yet. In this study, we synthesized Kisspeptin-10 and Kisspeptin-E, which are biologically active fragments, to mimic the action of Kisspeptin. Next, we demonstrated the anti-aging effects of the Kisspeptin-mimicking fragments using UV-induced skin aging models, such as UV-induced human dermal fibroblasts (Hs68) and human skin explants. Kisspeptin-E suppressed UV-induced 11 beta-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) stimulation leading to a regulation of skin aging related genes, including type I procollagen, matrix metalloproteinases-1 (MMP-1), interleukin-6 (IL-6), and IL-8, and rescued the skin integrity. Taken together, these results suggest that Kisspeptin-E could be useful to improve UV-induced skin aging by modulating expression of stress related genes, such as 11ß-HSD1.

Expression of the kisspeptin/gonadotropin-releasing hormone (GnRH) system in the brain of female Chinese sucker (Myxocyprinus asiaticus) at the onset of puberty.

The kisspeptin-kisspeptin receptor (kissr)-gonadotropin-releasing hormone (GnRH) system plays a key role in regulating the onset of puberty in mammals. However, the role of this system in fish is still unclear. We examined the relative gene expression patterns for kiss1, kiss2, kissr2, sGnRH, and pjGnRH in all parts of the brains of Chinese sucker (Myxocyprinus asiaticus) females at the prepubertal and pubertal stages by using real-time PCR. We also analyzed the expression of kiss1 and GnRH1 via immunofluorescence. Two variants of kisspeptin; a variant of kissr (kissr2); and two variants of GnRH, pjGnRH (GnRH1), and sGnRH (GnRH3), were expressed in all parts of the brain. The mRNA expression of kiss1 was higher in the telencephalon, mesencephalon, and diencephalon at the pubertal stage than at the prepubertal stage, and the expression of kiss2 was higher in only the telencephalon. The expression of kissr2 was higher in all parts of the brain, except the medulla, at the pubertal stage than at the prepubertal stage. pjGnRH was highly expressed in all parts of the brain at the pubertal stage, whereas sGnRH expression showed no distinct changes, except in the epencephalon. Strong kiss1 and weak GnRH-1 immunoreactivity was observed in the pineal gland, lateral tuberal nucleus (NLT), and ventral part of the NLT in the diencephalon of the Chinese sucker females at the pubertal stage. Our results suggest that the kiss1-kissr2-pjGnRH system was expressed highly at the onset of pubertal female Chinese sucker.

[Research progress of kisspeptin in female reproductive endocrine and assisted reproductive techniques].

In recent years, it has been found that kisspeptin plays some key roles in the physiological processes of the brain, such as gender differentiation, positive and negative feedback of sex hormones, onset of puberty, and transduction of energy signals in the body, which suggests that kisspeptin may be a key molecule for the maturation and regulation of female reproductive function. In addition to the systemic roles of the kisspeptin, its local roles in reproductive organs are constantly being discovered. With the discovery that kisspeptin is involved in the pathological process of reproductive endocrine diseases such as isolated hypogonadotropic hypogonadism (IHH), polycystic ovary syndrome (PCOS), premature ovarian failure (POF) and pathological hyperprolactinemia, exogenous application of kisspeptin to solve reproductive problems has become a new hot topic. The review focuses on the research progress of kisspeptin in the female reproductive system, especially on its application in assisted reproduction.

Gonadotropin-Releasing Hormone (GnRH) Neuron Excitability Is Regulated by Estradiol Feedback and Kisspeptin.

Gonadotropin-releasing hormone (GnRH) neurons produce the central output controlling fertility and are regulated by steroid feedback. A switch from estradiol negative to positive feedback initiates the GnRH surge, ultimately triggering ovulation. This occurs on a daily basis in ovariectomized, estradiol-treated (OVX+E) mice; GnRH neurons are suppressed in the morning and activated in the afternoon. To test the hypotheses that estradiol and time of day signals alter GnRH neuron responsiveness to stimuli, GFP-identified GnRH neurons in brain slices from OVX+E or OVX female mice were recorded during the morning or afternoon. No differences were observed in baseline membrane potential. Current-clamp revealed GnRH neurons fired more action potentials in response to current injection during positive feedback relative to all other groups, which were not different from each other despite reports of differing ionic conductances. Kisspeptin increased GnRH neuron response in cells from OVX and OVX+E mice in the morning but not afternoon. Paradoxically, excitability in kisspeptin knock-out mice was similar to the maximum observed in control mice but was unchanged by time of day or estradiol. A mathematical model applying a Markov Chain Monte Carlo method to estimate probability distributions for estradiol- and time of day-dependent parameters was used to predict intrinsic properties underlying excitability changes. A single identifiable distribution of solutions accounted for similar GnRH neuron excitability in all groups other than positive feedback despite different underlying conductance properties; this was attributable to interdependence of voltage-gated potassium channel properties. In contrast, redundant solutions may explain positive feedback, perhaps indicative of the importance of this state for species survival.SIGNIFICANCE STATEMENT Infertility affects 15%-20% of couples; failure to ovulate is a common cause. Understanding how the brain controls ovulation is critical for new developments in both infertility treatment and contraception. Gonadotropin-releasing hormone (GnRH) neurons are the final common pathway for central neural control of ovulation. We studied how estradiol feedback regulates GnRH excitability, a key determinant of neural firing rate using laboratory and computational approaches. GnRH excitability is upregulated during positive feedback, perhaps driving increased neural firing rate at this time. Kisspeptin increased GnRH excitability and was essential for estradiol regulation of excitability. Modeling predicts that multiple combinations of changes to GnRH intrinsic conductances can produce the firing response in positive feedback, suggesting the brain has many ways to induce ovulation.

Glutamatergic Transmission to Hypothalamic Kisspeptin Neurons Is Differentially Regulated by Estradiol through Estrogen Receptor α in Adult Female Mice.

Estradiol feedback regulates gonadotropin-releasing hormone (GnRH) neurons and subsequent luteinizing hormone (LH) release. Estradiol acts via estrogen receptor α (ERα)-expressing afferents of GnRH neurons, including kisspeptin neurons in the anteroventral periventricular (AVPV) and arcuate nuclei, providing homeostatic feedback on episodic GnRH/LH release as well as positive feedback to control ovulation. Ionotropic glutamate receptors are important for estradiol feedback, but it is not known where they fit in the circuitry. Estradiol-negative feedback decreased glutamatergic transmission to AVPV and increased it to arcuate kisspeptin neurons; positive feedback had the opposite effect. Deletion of ERα in kisspeptin cells decreased glutamate transmission to AVPV neurons and markedly increased it to arcuate kisspeptin neurons, which also exhibited increased spontaneous firing rate. KERKO mice had increased LH pulse frequency, indicating loss of negative feedback. These observations indicate that ERα in kisspeptin cells is required for appropriate differential regulation of these neurons and neuroendocrine output by estradiol.SIGNIFICANCE STATEMENT The brain regulates fertility through gonadotropin-releasing hormone (GnRH) neurons. Ovarian estradiol regulates the pattern of GnRH (negative feedback) and initiates a surge of release that triggers ovulation (positive feedback). GnRH neurons do not express the estrogen receptor needed for feedback (estrogen receptor α [ERα]); kisspeptin neurons in the arcuate and anteroventral periventricular nuclei are postulated to mediate negative and positive feedback, respectively. Here we extend the network through which feedback is mediated by demonstrating that glutamatergic transmission to these kisspeptin populations is differentially regulated during the reproductive cycle and by estradiol. Electrophysiological and in vivo hormone profile experiments on kisspeptin-specific ERα knock-out mice demonstrate that ERα in kisspeptin cells is required for appropriate differential regulation of these neurons and for neuroendocrine output.

Kisspeptin regulation of human decidual stromal cells motility via FAK-Src intracellular tyrosine kinases.

STUDY QUESTION: Can of Clinical Genetics, Maastricht University Medical Centre, Maastricht kisspeptin and its analogues regulate the motility of human decidual stromal cells and what intracellular signaling pathways are involved? SUMMARY ANSWER: Kisspeptin analogue-mediated cell motility in human decidual stromal cells via the focal adhesion kinase (FAK)-steroid receptor coactivator (Src) pathway suggesting that kisspeptin may modulate embryo implantation and decidual programming in human pregnancy. WHAT IS KNOWN ALREADY: The extravillous trophoblast invades the maternal decidua during embryo implantation and placentation. The motile behavior and invasive potential of decidual stromal cells regulate embryo implantation and programming of human pregnancy. STUDY DESIGN, SIZE, DURATION: Human decidual stromal cells were isolated from healthy women undergoing elective termination of a normal pregnancy at 6- to 12-week gestation, after informed consent. PARTICIPANTS/MATERIALS, SETTING, METHODS: Kisspeptin analogues were synthetic peptides. Cell motility was estimated by an invasion and migration assay. Immunoblot analysis was performed to investigate the expression of kisspeptin receptor and the effects of kisspeptin analogues on the phosphorylation of FAK and Src. Small interfering RNAs (siRNAs) were used to knock down the expression of kisspeptin receptor, FAK, Src, matrix metallo-proteinases (MMPs) 2 and 9, and extracellular signal-regulated protein kinase (ERK) 1/2. MAIN RESULTS AND THE ROLE OF CHANCE: The kisspeptin receptor was expressed in human decidual stromal cells. Kisspeptin agonist decreased, but antagonist increased, cell motility. Kisspeptin agonist decreased the phosphorylation of FAK and Src tyrosine kinases, whereas antagonist increased it. These effects on phosphorylation were abolished by kisspeptin receptor siRNA. The activation of cell motility by kisspeptin analogues was suppressed by siRNA knockdown of endogenous FAK (decreased 66%), Src (decreased 60%), kisspeptin receptor (decreased 26%), MMP-2 (decreased 36%), MMP-9 (decreased 23%), and ERK 1/2 inhibitor (decreased 27%). LIMITATIONS, REASONS FOR CAUTION: Human decidual stromal cells were obtained from women having terminations after 6-12 weeks of pregnancy and differences in timing could affect their properties. WIDER IMPLICATIONS OF THE FINDINGS: Kisspeptin acting within the endometrium has a potential modulatory role on embryo implantation and decidual programming of human pregnancy. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by grant NSC-104-2314-B-182A-146-MY2 (to H.-M.W.) from the Ministry of Science and Technology, Taiwan, and grants CMRPG3E0401 and CMRPG3E0402 (to H.-M.W.). This work was also supported by grants from the Canadian Institutes of Health Research to P.C.K.L. P.C.K.L. is the recipient of a Child & Family Research Institute Distinguished Investigator Award. The authors have no conflicts of interest to disclose. TRIAL REGISTRATION NUMBER: N/A.

Reproductive functions of Kisspeptin/KISS1R Systems in the Periphery.

Kisspeptin and its G protein-coupled receptor KISS1R play key roles in mammalian reproduction due to their involvement in the onset of puberty and control of the hypothalamic-pituitary-gonadal axis. However, recent studies have indicated a potential role of extra-hypothalamic kisspeptin in reproductive function. Here, we summarize recent advances in our understanding of the physiological significance of kisspeptin/KISS1R in the peripheral reproductive system (including the ovary, testis, uterus, and placenta) and the potential role of kisspeptin/KISS1R in reproductive diseases. A comprehensive understanding of the expression, function, and potential molecular mechanisms of kisspeptin/KISS1R in the peripheral reproductive system will contribute to the diagnosis, treatment and prevention of reproductive diseases.

Medial Amygdala Kiss1 Neurons Mediate Female Pheromone Stimulation of Luteinizing Hormone in Male Mice.

BACKGROUND/AIMS: The medial amygdala (MeA) responds to olfactory stimuli and alters reproductive physiology. However, the neuronal circuit that relays signals from the MeA to the reproductive axis remains poorly defined. This study aimed to test whether MeA kisspeptin (MeAKiss) neurons in male mice are sensitive to sexually relevant olfactory stimuli and transmit signals to alter reproductive physiology. We also investigated whether MeAKiss neurons have the capacity to elaborate glutamate and GABA neurotransmitters and potentially contribute to reproductive axis regulation. METHODS: Using female urine as a pheromone stimulus, MeAKiss neuronal activity was analysed and serum luteinizing hormone (LH) was measured in male mice. Next, using a chemogenetic approach, MeAKiss neurons were bi-directionally modulated to measure the effect on serum LH and evaluate the activation of the preoptic area. Lastly, using in situ hybridization, we identified the proportion of MeAKiss neurons that express markers for GABAergic (Vgat) and glutamatergic (Vglut2) neurotransmission. RESULTS: Male mice exposed to female urine showed a two-fold increase in the number of c-Fos-positive MeAKiss neurons concomitant with raised LH. Chemogenetic activation of MeAKiss neurons significantly increased LH in the absence of urine exposure, whereas inhibition of MeAKiss neurons did not alter LH. In situ hybridization revealed that MeAKiss neurons are a mixed neuronal population in which 71% express Vgat mRNA, 29% express Vglut2 mRNA, and 6% express both. CONCLUSIONS: Our results uncover, for the first time, that MeAKiss neurons process sexually relevant olfactory signals to influence reproductive hormone levels in male mice, likely through a complex interplay of neuropeptide and neurotransmitter signalling.

Effects of kisspeptin on pathogenesis and energy metabolism in polycystic ovarian syndrome (PCOS).

Kisspeptin has been shown to participate in the regulation of pituitary hormone secretion and energy metabolism. In PCOS patients, there are disorders in pituitary hormone secretion and energy metabolism. The aim of this study was to investigate the serum kisspeptin and its relationship with abnormal metabolism in PCOS. This restrospective case-control study included 73 cases with PCOS and 63 control cases. All subjects were divided into obese and nonobese groups based on BMI. The serum kisspeptin levels, Cor, DHEA-S, plasma concentrations of glucose were tested. We found that the level of kisspeptin in PCOS group was higher than it in control group. The kisspeptin levels in nonobese PCOS group increased most obviously over than the other groups. The kisspeptin levels of all the subjects were positively correlated with LH levels, negatively correlated with the glucose-AUC, the insulin-AUC, and triglyceride levels. The findings of this study suggest that kisspeptin may play an important role in ovulation disorders in PCOS patients through regulating the level of LH and it could regulate the body's energy metabolism by regulating glucose and lipid metabolism.

Role of kisspeptin neurons as a GnRH surge generator: Comparative aspects in rodents and non-rodent mammals.

Ovulation is an essential phenomenon for reproduction in mammalian females along with follicular growth. It is well established that gonadal function is controlled by the neuroendocrine system called the hypothalamus-pituitary-gonadal (HPG) axis. Gonadotropin-releasing hormone (GnRH) neurons, localized in the hypothalamus, had been considered to be the head in governing the HPG axis for a long time until the discovery of kisspeptin. In females, induction of ovulation and folliculogenesis has been linked to a surge mode and pulse mode of GnRH releases, respectively. The mechanisms of how the two modes of GnRH are differently regulated had long remained elusive. The discovery of kisspeptin neurons, distributed in two hypothalamic nuclei, such as the arcuate nucleus in the caudal hypothalamus and preoptic area or the anteroventral periventricular nucleus in the rostral hypothalamic regions, and analyses of the detailed functions of kisspeptin neurons have led marked progress on the understanding of different mechanisms regulating GnRH surges (ovulation) and GnRH pulses (folliculogenesis). The present review will focus on the role of kisspeptin neurons as the GnRH surge generator, including the sexual differentiation of the surge generation system and factors that regulate the surge generator. Comparative aspects between mammalian species are especially focused on.