Inhibition of metastin (kisspeptin-54)-GPR54 signaling in the arcuate nucleus-median eminence region during lactation in rats

Follicular development and ovulation are suppressed during lactation in various mammalian species, mainly due to the suppression of pulsatile GnRH/LH secretion. Metastin (kisspeptin-54), a KiSS-1 gene product, is an endogenous ligand for GPR54, a G-protein-coupled receptor, and suggested to play a critical role in regulating the gonadal axis. The present study therefore aims to determine whether metastin (kisspeptin-54)-GPR54 signaling in discrete brain areas is inhibited by the suckling stimulus that causes suppression of LH secretion in lactating rats. Quantitative RT-PCR revealed that the KiSS-1 mRNA level was significantly lower in the arcuate nucleus (ARC)-median eminence region in lactating ovariectomized (OVX) and estrogen-treated OVX rats than in nonlactating controls. KiSS-1 mRNA in the anteroventral periventricular nucleus was kept at a low level in both lactating and nonlactating rats despite estrogen treatment. GPR54 mRNA levels were significantly lower in lactating than nonlactating rats in the anteroventral periventricular nucleus, but the levels in lactating mothers of the preoptic area and ARC-median eminence were comparable with nonlactating controls. Although KiSS-1 mRNA-expressing cells or metastin (kisspeptin-54) immunoreactivities were densely located in the ARC of nonlactating controls, few were found in the ARC of lactating OVX animals. Various doses of metastin (kisspeptin-54) (0.02, 0.2, and 2 nmol) injected into the third ventricle caused a significant increase in LH secretion in both lactating and nonlactating OVX rats, suggesting that lactating rats are responsive to metastin (kisspeptin-54) stimulus. Thus, the present study demonstrated that KiSS-1 mRNA/metastin (kisspeptin-54) expression is inhibited in the ARC by the suckling stimulus, suggesting that the inhibition is most probably involved in suppressing LH secretion in lactating rats.     

KiSS-1 and GPR54 as new players in gonadotropin regulation and puberty

The recent identification of loss-of-function mutations in the gene encoding GPR54, the receptor for the KiSS-1-derived peptides, kisspeptins, has highlighted a previously unrecognized pathway in the physiologic regulation of puberty and reproduction. Patients with loss-of-function mutations in GPR54 have idiopathic hypogonadotropic hypogonadism, and mice lacking GPR54 similarly fail to undergo puberty and have immature reproductive organs and low levels of sex steroids and gonadotropins. These observations have led to the hypothesis that kisspeptins activate hypothalamic GnRH release, thereby serving as a pivotal factor in the pubertal activation of the reproductive cascade. This hypothesis is supported by subsequent studies in rodent and primate models that have demonstrated localization of KiSS-1 mRNA in the hypothalamus, colocalization of GPR54 in GnRH neurons, GnRH-dependent activation of LH and FSH release by intracerebroventricular or peripheral administration of kisspeptin, and increased hypothalamic KiSS-1 and GPR54 mRNA levels at the onset of puberty. Taken together, these findings weave a compelling case for a role of the kisspeptin-GPR54 system in the activation of GnRH neurons at the time of pubertal awakening of the reproductive axis.     

Changes in hypothalamic KiSS-1 system and restoration of pubertal activation of the reproductive axis by kisspeptin in undernutrition

Activation of the gonadotropic axis critically depends on sufficient body energy stores, and conditions of negative energy balance result in lack of puberty onset and reproductive failure. Recently, KiSS-1 gene-derived kisspeptin, signaling through the G protein-coupled receptor 54 (GPR54), has been proven as a pivotal regulator in the control of gonadotropin secretion and puberty. However, the impact of body energy status upon hypothalamic expression and function of this system remains unexplored. In this work, we evaluated the expression of KiSS-1 and GPR54 genes at the hypothalamus as well as the ability of kisspeptin-10 to elicit GnRH and LH secretion in prepubertal rats under short-term fasting. In addition, we monitored the actions of kisspeptin on food intake and the effects of its chronic administration upon puberty onset in undernutrition. Food deprivation induced a concomitant decrease in hypothalamic KiSS-1 and increase in GPR54 mRNA levels in prepubertal rats. In addition, LH responses to kisspeptin in vivo were enhanced, and its GnRH secretagogue action in vitro was sensitized, under fasting conditions. Central kisspeptin administration failed to change food intake patterns in animals fed ad libitum or after a 12-h fast. However, chronic treatment with kisspeptin was able to restore vaginal opening (in approximately 60%) and to elicit gonadotropin and estrogen responses in a model of undernutrition. In summary, our data are the first to show an interaction between energy status and the hypothalamic KiSS-1 system, which may constitute a target for disruption (and eventual therapeutic intervention) of pubertal development in conditions of negative energy balance.     

Expression of KiSS-1 in rat ovary: putative local regulator of ovulation?

Kisspeptins, the products of KiSS-1 gene, and their receptor, GPR54, have recently emerged as essential gatekeepers of reproduction, mainly through regulation of GnRH secretion at the hypothalamus. However, the profound hypogonadotropism linked to GPR54 inactivation is likely to mask additional functions of this system at other levels of the gonadal axis, in which expression of KiSS-1 and GPR54 has been preliminarily reported. We describe herein the expression of KiSS-1 gene and kisspeptin immunoreactivity (IR) in rat ovary and evaluate its developmental and hormonal regulation. KiSS-1 and GPR54 mRNAs were persistently detected in adult ovary along estrous cycle. Yet, contrary to GPR54, ovarian KiSS-1 levels fluctuated in a cyclic-dependent manner, with a robust increase in the afternoon of proestrus, i.e. preceding ovulation. In addition, kisspeptin-IR was observed in rat ovary, with strong signals in theca layers of growing follicles, corpora lutea, and interstitial gland, compartments in which modest GPR54-IR was also detected. Interestingly, the rise in ovarian KiSS-1 mRNA at proestrus was prevented by blockade of preovulatory gonadotropin surge and restored by replacement with human chorionic gonadotropin as superagonist of LH. In addition, immature ovaries showed low to negligible levels of KiSS-1 mRNA, which were significantly enhanced by gonadotropin priming. In summary, we present novel evidence for the developmental and hormonally regulated expression of the KiSS-1 gene, and the presence of kisspeptin-IR, in rat ovary. The ability of the LH surge to timely induce ovarian expression of KiSS-1 at the preovulatory period strongly suggests a previously unsuspected role of locally produced kisspeptin in the control of ovulation.     

Increased hypothalamic GPR54 signaling: a potential mechanism for initiation of puberty in primates

To further study the role of GPR54 signaling in the onset of primate puberty, we used the monkey to examine the ability of kisspeptin-10 to elicit the release of gonadotropin-releasing hormone (GnRH) precociously, and we describe the expression of GPR54 and KiSS-1 in the hypothalamus during the peripubertal period. Agonadal juvenile male monkeys were implanted with a lateral cerebroventricular cannula and a jugular vein catheter. The responsiveness of the juvenile pituitary to endogenous GnRH release was heightened with a chronic pulsatile i.v. infusion of synthetic GnRH before kisspeptin-10 (112-121) injection. Intracerebroventricular (30 microg or 100 microg) or i.v. (100 microg) bolus injections of kisspeptin-10 elicited a robust GnRH discharge, as reflected by luteinizing hormone secretion, which was abolished by pretreatment with a GnRH-receptor antagonist. RNA was isolated from the hypothalamus of agonadal males before (juvenile) and after (pubertal) the pubertal resurgence of pulsatile GnRH release and from juvenile, early pubertal, and midpubertal ovary-intact females. KiSS-1 mRNA levels detected by real-time PCR increased with puberty in both male and female monkeys. In intact females, but not in agonadal males, GPR54 mRNA levels in the hypothalamus increased approximately 3-fold from the juvenile to midpubertal stage. Hybridization histochemistry indicated robust KiSS-1 and GPR54 mRNA expression in the region of the arcuate nucleus. These findings are consistent with the hypothesis that GPR54 signaling by its cognate ligand in the primate hypothalamus may be activated at the end of the juvenile phase of development and may contribute to the pubertal resurgence of pulsatile GnRH release, the central drive for puberty.     

Effects of KiSS-1 peptide, the natural ligand of GPR54, on follicle-stimulating hormone secretion in the rat

KiSS-1 was originally identified as a metastasis suppressor gene encoding an array of structurally related peptides, namely kisspeptins, which acting through the G protein-coupled receptor GPR54 are able to inhibit tumor progression. Unexpectedly, a reproductive facet of this newly discovered system has recently arisen, and characterization of the role of the KiSS-1/GPR54 system in the neuroendocrine control of gonadotropin secretion has been initiated. However, such studies have been so far mostly restricted to LH, and very little is known about the actual contribution of this system in the regulation of FSH release. To address this issue, the effects of KiSS-1 peptide on FSH secretion were monitored in vivo and in vitro under different experimental conditions. Intracerebroventricular administration of KiSS-1 peptide significantly stimulated FSH secretion in prepubertal and adult rats. Yet, dose-response analyses in vivo demonstrated an ED(50) value for the FSH-releasing effects of KiSS-1 of 400 pmol, i.e. approximately 100-fold higher than that of LH. In addition, systemic (ip and iv) injection of KiSS-1 significantly stimulated FSH secretion in vivo. However, KiSS-1 failed to elicit basal FSH release directly at the pituitary level, although it moderately enhanced GnRH-stimulated FSH secretion in vitro. Finally, mechanistic studies revealed that the ability of KiSS-1 to elicit FSH secretion was abolished by the blockade of endogenous GnRH actions, but it was persistently observed in different models of leptin insufficiency and after blockade of endogenous excitatory amino acid and nitric oxide pathways, i.e. relevant signals in the neuroendocrine control of gonadotropin secretion. In summary, our results extend previous recent observations on the role of KiSS-1 in the control of LH secretion and provide solid evidence for a stimulatory effect of KiSS-1 on FSH release, acting at central level. Overall, it is proposed that the KiSS-1/GPR54 system is a novel, pivotal downstream element in the neuroendocrine network governing gonadotropin secretion.     

GPR54 and KiSS-1: Role in the regulation of puberty and reproduction

The finding of inactivating mutations in GPR54 in IHH patients and the lack of reproductive maturation of the GPR54 null mouse have uncovered a previously unrecognized role for GPR54 and KiSS-1 in the physiologic regulation of puberty and reproduction. This newly identified function for GPR54 and its cognate ligand, kisspeptin, has led to additional studies that have localized GPR54 and KiSS-1 mRNA in the hypothalamus, colocalized GPR54 in GnRH neurons, demonstrated GnRH-dependent activation of LH and FSH release by kisspeptin, and shown increased hypothalamic KiSS-1 and GPR54 mRNA levels at the time of puberty. Taken together, these findings establish the role of the kisspeptin-GPR54 system in the stimulation of GnRH neurons during puberty. The mechanisms by which kisspeptin activates GnRH release, as well as the trigger for this pathway at the onset of puberty, are yet to be elucidated. In the future, modulators of GPR54 activity, including kisspeptin, may prove valuable in clinical applications in the fields of both cancer therapy and reproductive medicine.     

Minireview: kisspeptin neurons as central processors in the regulation of gonadotropin-releasing hormone secretion

The Kiss1 gene encodes a family of peptides called kisspeptins, which bind to the G protein-coupled receptor GPR54. Kisspeptin(s) and its receptor are expressed in the forebrain, and the discovery that mice and humans lacking a functional GPR54 fail to undergo puberty and exhibit hypogonadotropic hypogonadism implies that kisspeptin signaling plays an essential role in reproduction. Studies in several mammalian species have shown that kisspeptins stimulate the secretion of gonadotropins from the pituitary by stimulating the release of GnRH from the forebrain after the activation of GPR54, which is expressed by GnRH neurons. Kisspeptin is expressed abundantly in the arcuate nucleus (Arc) and the anteroventral periventricular nucleus (AVPV) of the forebrain. Both estradiol and testosterone regulate the expression of the Kiss1 gene in the Arc and AVPV; however, the response of the Kiss1 gene to these steroids is exactly opposite between these two nuclei. Estradiol and testosterone down-regulate Kiss1 mRNA in the Arc and up-regulate its expression in the AVPV. Thus, kisspeptin neurons in the Arc may participate in the negative feedback regulation of gonadotropin secretion, whereas kisspeptin neurons in the AVPV may contribute to generating the preovulatory gonadotropin surge in the female. Hypothalamic levels of Kiss1 and GPR54 mRNA increase dramatically at puberty, suggesting that kisspeptin signaling could mediate the neuroendocrine events that trigger the onset of puberty. Together, these observations demonstrate that kisspeptin-GPR54 signaling in the brain serves as an important conduit for controlling GnRH secretion in the developing and adult animal.     

Repetitive activation of hypothalamic G protein-coupled receptor 54 with intravenous pulses of kisspeptin in the juvenile monkey (Macaca mulatta) elicits a sustained train of gonadotropin-releasing hormone discharges

The purpose of the present study was to further examine the hypothesis that activation of G protein-coupled receptor 54 (GPR54) signaling at the end of the juvenile phase of primate development is responsible for initiation of gonadarche and the onset of puberty. Accordingly, we determined whether repetitive iv administration of the GPR54 receptor agonist kisspeptin-10 (2 microg as a brief 1-min infusion once every hour for 48 h) to the juvenile male rhesus monkey would prematurely elicit sustained, pulsatile release of hypothalamic GnRH, the neuroendocrine trigger for gonadarche. GnRH release was monitored indirectly by measuring LH secretion from the in situ pituitary, the GnRH responsiveness of which had been heightened before the experiment with an intermittent iv infusion of synthetic GnRH. Agonadal animals (n = 4) were employed to eliminate any confounding and secondary effects of changing feedback signals from the testis. The first brief infusion of kisspeptin-10 evoked an LH discharge that mimicked those produced by GnRH priming, and this was followed by a train of similar LH discharges in response to hourly activation of GPR54 by repetitive kisspeptin-10 administration. Concomitant treatment with a GnRH receptor antagonist, acyline, abolished kisspeptin-10-induced LH release. Repetitive kisspeptin-10 administration also provided a GnRH-dependent signal to FSH secretion. These findings are consistent with the notion that, in primates, the transition from the juvenile (attenuated GnRH release) to pubertal (robust GnRH release) state is controlled by activation of GPR54 resulting from increased expression of hypothalamic KiSS-1 and release of kisspeptin in this region of the brain.     

Characterization of the potent luteinizing hormone-releasing activity of KiSS-1 peptide, the natural ligand of GPR54

Loss-of-function mutations of the gene encoding GPR54, the putative receptor for the KiSS-1-derived peptide metastin, have been recently associated with hypogonadotropic hypogonadism, in both rodents and humans. Yet the actual role of the KiSS-1/GPR54 system in the neuroendocrine control of gonadotropin secretion remains largely unexplored. To initiate such analysis, the effects of KiSS-1 peptide on LH secretion were monitored using in vivo and in vitro settings under different experimental conditions. Central intracerebroventricular administration of KiSS-1 peptide potently elicited LH secretion in vivo over a range of doses from 10 pmol to 1 nmol. The effect of centrally injected KiSS-1 appeared to be mediated via the hypothalamic LHRH. However, no effect of central administration of KiSS-1 was detected on relative LHRH mRNA levels. Likewise, systemic (i.p. and i.v.) injection of KiSS-1 markedly stimulated LH secretion. This effect was similar in terms of maximum response to that of central administration of KiSS-1 and might be partially attributed to its ability to stimulate LH secretion directly at the pituitary. Finally, the LH-releasing activity of KiSS-1 was persistently observed after blockade of endogenous excitatory amino acid and nitric oxide pathways, i.e. relevant neurotransmitters in the neuroendocrine control of LH secretion. In summary, our results provide solid evidence for a potent stimulatory effect of KiSS-1 on LH release, acting at central levels (likely the hypothalamus) and eventually at the pituitary, and further document a novel role of the KiSS-1/GPR54 system as a relevant downstream element in the neuroendocrine network governing LH secretion.     

Kisspeptin expression in the brain: Catalyst for the initiation of puberty

In 2003, two independent groups of researchers discovered almost simultaneously that inactivating mutations of the G protein coupled receptor, GPR54, cause hypogonadotropic hypogonadism in mice and men. Since this discovery, kisspeptins, the natural ligands for GPR54, have been thrust into the reproductive neuroendocrine spotlight, as major regulators of GnRH function. Kisspeptins are the peptide products of the KiSS-1 gene, and potently stimulate gonadotrophin secretion when administered either centrally or peripherally. Expression of KiSS-1 has been localised to specific regions of the hypothalamus in many species and is regulated by gonadal steroids and across the estrous cycle. It appears that kisspeptin transmits steroid feedback signals to GnRH cells, especially the positive feedback effect of estrogen that causes the preovulatory GnRH/LH surge. Importantly, kisspeptin function appears to be fundamental to the initiation of puberty.     

The role of KiSS-1 in the regulation of puberty in higher primates

Puberty in higher primates is triggered by resurgence in the pulsatile secretion of hypothalamic GnRH after a hiatus in the robust release of this hypophysiotropic signal during childhood and juvenile development. Interestingly, the prepubertal decline in GnRH release is not associated with a marked reduction in the expression of either the gene that codes for GnRH (GnRH-1) or the decapeptide itself, and the network of GnRH neurons in the hypothalamus of the juvenile may by activated prematurely and with surprising ease by intermittent neurochemical stimulation with N-methyl-d-aspartate (NMDA), a glutamate receptor agonist. KiSS-1, a gene that encodes for kisspeptin-121, which is proteolytically cleaved to a 54 amino acid peptide, metastin, was initially studied in the context of tumor suppression. In 2003, however, inactivating mutations in the metastin receptor, GPR54, were reported to be associated with hypogonadotropic hypogonadism and absent puberty in man. Subsequent studies in the rhesus monkey have shown that GPR54 and KiSS-1 are expressed in the mediobasal hypothalamus (MBH), KiSS-1 expression in the MBH increases at the time of the pubertal resurgence in GnRH release and pulsatile, but not continuous, i.v. administration of metastin 45-54 in the juvenile male monkey elicits sustained GnRH release precociously. The significance of these findings in the context of the initiation of the onset of puberty is discussed.     

KiSS-1 and reproduction: focus on its role in the metabolic regulation of fertility

Unraveling of the master role of kisspeptins, the products of the KiSS-1 gene, and their receptor, GPR54, in the control of reproduction has been a major breakthrough in contemporary neuroendocrinology. Indeed, since the disclosure of their reproductive dimension in late 2003, an ever-growing number of genetic, molecular, physiologic and pharmacological studies have defined the crucial role of KiSS-1 neurons as central processors for the dynamic regulation of the gonadotropic axis and its full activation at puberty. Yet, the potential role of the hypothalamic KiSS-1 system as an intermediary factor for the well-known interplay between energy status and reproduction initially received little attention. Recent data, however, strongly suggest a prominent role of KiSS-1 in the metabolic control of fertility, as expression of KiSS-1 gene at the hypothalamus is down-regulated in conditions of negative energy balance and kisspeptin administration is capable of overcoming the hypogonadotropic state observed in undernutrition and disturbed metabolic conditions. Leptin, the adipocyte hormone signaling the size of body energy stores, is likely to play a pivotal role in the metabolic control of the KiSS-1 system, since kisspeptin neurons express leptin receptors and leptin is able to normalize defective KiSS-1 gene expression in models of impaired gonadotropin secretion linked to hypoleptinemia, such as the ob/ob mouse and streptozotocin-induced diabetic rat. In sum, these data provide strong evidence for a central role of kisspeptins and GPR54 as molecular conduits for the metabolic regulation of reproductive function - a phenomenon with potential physiopathologic and therapeutic implications.     

Hypothalamic expression of KiSS-1 system and gonadotropin-releasing effects of kisspeptin in different reproductive states of the female Rat

Kisspeptins, products of the KiSS-1 gene with ability to bind G protein-coupled receptor 54 (GPR54), have been recently identified as major gatekeepers of reproductive function with ability to potently activate the GnRH/LH axis. Yet, despite the diversity of functional states of the female gonadotropic axis, pharmacological characterization of this effect has been mostly conducted in pubertal animals or adult male rodents, whereas similar studies have not been thoroughly conducted in the adult female. In this work, we evaluated maximal LH and FSH secretory responses to kisspeptin-10, as well as changes in sensitivity and hypothalamic expression of KiSS-1 and GPR54 genes, in different physiological and experimental models in the adult female rat. Kisspeptin-10 (1 nmol, intracerebroventricular) was able to elicit robust LH bursts at all phases of the estrous cycle, with maximal responses at estrus; yet, in diestrus LH, responses to kisspeptin were detected at doses as low as 0.1 pmol. In contrast, high doses of kisspeptin only stimulated FSH secretion at diestrus. Removal of ovarian sex steroids did not blunt the ability of kisspeptin to further elicit stimulated LH and FSH secretion, but restoration of maximal responses required replacement with estradiol and progesterone. Finally, despite suppressed basal levels, LH and FSH secretory responses to kisspeptin were preserved in pregnant and lactating females, although the magnitude of LH bursts and the sensitivity to kisspeptin were much higher in pregnant dams. Interestingly, hypothalamic KiSS-1 gene expression significantly increased during pregnancy, whereas GPR54 mRNA levels remained unaltered. In summary, our current data document for the first time the changes in hypothalamic expression of KiSS-1 system and the gonadotropic effects (maximal responses and sensitivity) of kisspeptin in different functional states of the female reproductive axis. The present data may pose interesting implications in light of the potential therapeutic use of kisspeptin analogs in the pharmacological manipulation of the gonadotropic axis in the female.     

Developmental and hormonally regulated messenger ribonucleic acid expression of KiSS-1 and its putative receptor, GPR54, in rat hypothalamus and potent luteinizing hormone-releasing activity of KiSS-1 peptide

The gonadotropic axis is centrally controlled by a complex regulatory network of excitatory and inhibitory signals that is activated at puberty. Recently, loss of function mutations of the gene encoding G protein-coupled receptor 54 (GPR54), the putative receptor for the KiSS-1-derived peptide metastin, have been associated with lack of puberty onset and hypogonadotropic hypogonadism. Yet the pattern of expression and functional role of the KiSS-1/GPR54 system in the rat hypothalamus remain unexplored to date. In the present work, expression analyses of KiSS-1 and GPR54 genes were conducted in different physiological and experimental settings, and the effects of central administration of KiSS-1 peptide on LH release were assessed in vivo. Persistent expression of KiSS-1 and GPR54 mRNAs was detected in rat hypothalamus throughout postnatal development, with maximum expression levels at puberty in both male and female rats. Hypothalamic expression of KiSS-1 and GPR54 genes changed throughout the estrous cycle and was significantly increased after gonadectomy, a rise that was prevented by sex steroid replacement both in males and females. Moreover, hypothalamic expression of the KiSS-1 gene was sensitive to neonatal imprinting by estrogen. From a functional standpoint, intracerebroventricular administration of KiSS-1 peptide induced a dramatic increase in serum LH levels in prepubertal male and female rats as well as in adult animals. In conclusion, we provide novel evidence of the developmental and hormonally regulated expression of KiSS-1 and GPR54 mRNAs in rat hypothalamus and the ability of KiSS-1 peptide to potently stimulate LH secretion in vivo. Our current data support the contention that the hypothalamic KiSS-1/GPR54 system is a pivotal factor in central regulation of the gonadotropic axis at puberty and in adulthood.     

Regulation of Kiss1 gene expression in the brain of the female mouse

The Kiss1 gene encodes a family of neuropeptides called kisspeptins, which activate the receptor G protein-coupled receptor-54 and play a role in the neuroendocrine regulation of GnRH secretion. We examined whether estradiol (E2) regulates KiSS-1 in the forebrain of the female mouse by comparing KiSS-1 mRNA expression among groups of ovary-intact (diestrus), ovariectomized (OVX), and OVX plus E2-treated mice. In the arcuate nucleus (Arc), KiSS-1 expression increased after ovariectomy and decreased with E2 treatment. Conversely, in the anteroventral periventricular nucleus (AVPV), KiSS-1 expression was reduced after ovariectomy and increased with E2 treatment. To determine whether the effects of E2 on KiSS-1 are mediated through estrogen receptor (ER)alpha or ERbeta, we evaluated the effects of E2 in OVX mice that lacked functional ERalpha or ERbeta. In OVX mice that lacked functional ERalpha, KiSS-1 mRNA did not respond to E2 in either the Arc or AVPV, suggesting that ERalpha is essential for mediating the inhibitory and stimulatory effects of E2. In contrast, KiSS-1 mRNA in OVX mice that lacked functional ERbeta responded to E2 exactly as wild-type animals. Double-label in situ hybridization revealed that virtually all KiSS-1-expressing neurons in the Arc and AVPV coexpress ERalpha, suggesting that the effects of E2 are mediated directly through KiSS-1 neurons. We conclude that KiSS-1 neurons in the Arc, which are inhibited by E2, may play a role in the negative feedback regulation of GnRH secretion, whereas KiSS-1 neurons in the AVPV, which are stimulated by E2, may participate in the positive feedback regulation of GnRH secretion.     

KiSS-1 system and reproduction: comparative aspects and roles in the control of female gonadotropic axis in mammals

In late 2003, inactivating mutations of the G protein-coupled receptor GPR54 were found in patients suffering hypogonadotropic hypogonadism. This observation led to the proposal that this receptor and its putative ligands (kisspeptins, encoded by the KiSS-1 gene) are essential in the control of reproduction; a contention that has been now substantiated by an ever growing number of experimental studies. However, most (if not all) of this work has been carried out in mammals (human, sheep and laboratory rodents). Moreover, characterization of gonadotropin responses to kisspeptin was conducted in males, whereas its actions on the female gonadotropic axis initially received much less attention. Notwithstanding, recent experimental data have unveiled very prominent roles of the KiSS-1 system in the control of key aspects of female reproduction, which include not only the timing puberty onset and its modulation by metabolic factors, but also the dynamic regulation of the gonadotropic axis in adulthood. On the latter, the KiSS-1 neuron has been proposed as key intermediary element for the negative and positive feedback effects of sex steroids on gonadotropin secretion. Moreover, expression of KiSS-1 (mRNA and peptide) and its receptor have been recently reported in the ovary, adding further complexity to the potential actions of this system in the female. In sum, compelling experimental evidence, obtained in mammals, has recently defined the pivotal role of the KiSS-1/GPR54 system in the control of essential aspects of female reproduction, from puberty to ovulation. While characterization of its role in non-mammalian species remains largely unexplored, the presence of GPR54 in GnRH neurons and the changes in its expression during pubertal development, reported recently in fish species, are suggestive of a conserved function of the KiSS-1/GPR54 system in the control of reproduction during evolution.     

Regulation of hypothalamic expression of KiSS-1 and GPR54 genes by metabolic factors: analyses using mouse models and a cell line

It is well established that reproductive function is metabolically gated. However, the mechanisms whereby energy stores and metabolic cues influence fertility are yet to be completely deciphered. Recently, the hypothalamic KiSS-1/GPR54 system has emerged as a fundamental regulator of the gonadotropic axis, which conveys the modulatory actions of sex steroids to GnRH neurons. Evidence is also mounting that KiSS-1 neurons may also represent the link between systemic metabolic signals and central control of reproduction. To further explore this possibility, we examined the impact of changes in energy status and key metabolic regulators on the hypothalamic expression of KiSS-1 and GPR54 genes, using different mouse models and the hypothalamic cell line N6. Time-course analysis of the effects of short-term fasting revealed a rapid (12- and 24-h) decline in KiSS-1 and GPR54 mRNA levels, which preceded that of GnRH (48 h). In contrast, diet-induced obesity or obesity associated with leptin deficiency (ob/ob vs. wild-type mice) failed to induce overt changes in hypothalamic expression of KiSS-1 and GPR54 genes. However, leptin infusion of ob/ob mice evoked a significant increase in KiSS-1 and GPR54 mRNA levels compared with pair-fed controls. Moreover, leptin, but not insulin or IGF-I, stimulated KiSS-1 mRNA expression in the mouse hypothalamic cell line N6. In addition, neuropeptide Y (NPY) null mice showed decreased KiSS-1 mRNA levels at the hypothalamus, whereas exposure to NPY increased expression of KiSS-1 in hypothalamic N6 cells. In sum, our present data further characterize the functional relevance and putative key mediators (such as leptin and NPY) of the metabolic regulation of the hypothalamic KiSS-1 system in the mouse.