Gonadotropin-releasing Hormone Neurons Research Articles

Protein tyrosine phosphatase receptors N and N2 regulate gonadotropin-releasing hormone neuron function

Searchable abstracts of presentations at key conferences in endocrinology ISSN 1470-3947 (print) | ISSN 1479-6848 (online)

Review: Early and late determinants of puberty in ruminants and the role of nutrition.

This article reviews the scientific literature on puberty with a focus on ruminants and draws inference, where appropriate, from recent findings in transgenic mouse models and human pathology. Early genetic determinants of puberty have been discovered in humans suffering from hypogonadotropic hypogonadism or central precocious puberty. Transgenic mouse models selected on the basis of the causative defective genes helped in discovering the cellular and molecular mechanisms involved. Most of the genes found are involved in the development of neuroendocrine networks during embryo development and early postnatal life. Notwithstanding that the development of neuroendocrine networks takes place early in puberty, a delay or acceleration in the development of Gonadotropin Releasing Hormone (GnRH) neurons has an impact on puberty onset inducing a delay or an advance, respectively. Among the genes discovered in humans and laboratory models, only a few of them displayed polymorphisms associated with advanced sexual maturity, but also marbling, growth traits and callipygian conformation. This could be related to the fact that rather than puberty onset, most research monitored sexual maturity. Sexual maturity occurs after puberty onset and involves factors regulating the maturation of gonads and in the expression of sexual behaviour. The association with growth and metabolic traits is not surprising since nutrition is the major environmental factor that will act on late genetic determinants of puberty onset. However, a recent hypothesis emerged suggesting that it is the postnatal activation of the GnRH neuronal network that induces the acceleration of growth and weight gain. Hence, nutritional factors need the activation of GnRH neurons first before acting on late genetic determinants. Moreover, nutritional factors can also affect the epigenetic landscape of parental gamete's genome with the consequence of specific methylation of genes involved in GnRH neuron development in the embryo. Season is another important regulator of puberty onset in seasonal small ruminants and appears to involve the same mechanisms that are involved in seasonal transition in adults. The social environment is also an underestimated factor affecting puberty onset in domestic ruminants, most research studies focused on olfactory cues, but the genetic basis has not heretofore been adequately tackled by the scientific community. Additionally, there is some evidence to suggest transgenerational effects exist, in that nutritional and social cues to which parents were exposed, could affect the epigenetic landscape of parental gametes resulting in the epigenetic regulation of early genetic determinants of puberty onset in their offspring.

Review: Gestational and postnatal nutritional effects on the neuroendocrine control of puberty and subsequent reproductive performance in heifers.

Pubertal attainment is an intricate biological process that involves maturation of the reproductive neuroendocrine axis and increased pulsatile release of gonadotropin-releasing hormone (GnRH) and luteinizing hormone. Nutrition is a critical environmental factor controlling the timing of puberty attainment. Nutrient restriction during early postnatal development delays puberty, whereas increased feed intake and adiposity during this period hasten pubertal maturation by imprinting the hypothalamus. Moreover, the dam's nutrition during gestation can program the neuroendocrine system in the developing fetus and has the potential to advance or delay puberty in the offspring. Leptin, a hormone produced primarily by adipose cells, plays an important role in communicating energy status to the brain and regulating sexual maturation. Leptin's regulation of GnRH release is mediated by an upstream neuronal network since GnRH neurons do not contain the leptin receptor. Two groups of neurons located in the arcuate nucleus of the hypothalamus that express neuropeptide Y (NPY), an orexigenic peptide, and alpha melanocyte-stimulating hormone (αMSH), an anorexigenic peptide, are central elements of the neural circuitry that relay inhibitory (NPY) and excitatory (αMSH) inputs to GnRH neurons. Moreover, KNDy neurons, neurons in the arcuate nucleus that co-express kisspeptin, neurokinin B (NKB), and dynorphin, also play a role in the metabolic regulation of puberty. Our studies in beef heifers demonstrate that increased rates of BW gain during early postweaning (4-9 mo of age) result in reduced expression of NPY mRNA, increased expression of proopiomelanocortin and kisspeptin receptor mRNA, reduced NPY inhibitory inputs to GnRH neurons, and increased excitatory αMSH inputs to KNDy neurons. Finally, our most recent data demonstrate that nutrition of the cow during the last two trimesters of gestation can also induce transcriptional and structural changes in hypothalamic neurocircuitries in the heifer progeny that likely persist long-term after birth. Managerial approaches, such as supplementation of the dam during gestation (fetal programming), creep feeding, early weaning, and stair-step nutritional regimens have been developed to exploit brain plasticity and advance pubertal maturation in heifers.

Diversity within olfactory sensory derivatives revealed by the contribution of Dbx1 lineages.

In vertebrates, the embryonic olfactory epithelium contains progenitors that will give rise to distinct classes of neurons, including olfactory sensory neurons (OSNs; involved in odor detection), vomeronasal sensory neurons (VSNs; responsible for pheromone sensing), and gonadotropin-releasing hormone (GnRH) neurons that control the hypothalamic-pituitary-gonadal axis. Currently, these three neuronal lineages are usually believed to emerge from uniform pools of progenitors. Here, we found that the homeodomain transcription factor Dbx1 is expressed by neurogenic progenitors in the developing and adult mouse olfactory epithelium. We demonstrate that Dbx1 itself is dispensable for neuronal fate specification and global organization of the olfactory sensory system. Using lineage tracing, we characterize the contribution of Dbx1 lineages to OSN, VSN, and GnRH neuron populations and reveal an unexpected degree of diversity. Furthermore, we demonstrate that Dbx1-expressing progenitors remain neurogenic in the absence of the proneural gene Ascl1. Our work therefore points to the existence of distinct neurogenic programs in Dbx1-derived and other olfactory lineages.

The initiation and maintenance of gonadotropin-releasing hormone neuron identity in congenital hypogonadotropic hypogonadism.

Neurons that secrete gonadotropin-releasing hormone (GnRH) drive vertebrate reproduction. Genetic lesions that disrupt these neurons in humans lead to congenital hypogonadotropic hypogonadism (CHH) and reproductive failure. Studies on CHH have largely focused on the disruption of prenatal GnRH neuronal migration and postnatal GnRH secretory activity. However, recent evidence suggests a need to also focus on how GnRH neurons initiate and maintain their identity during prenatal and postnatal periods. This review will provide a brief overview of what is known about these processes and several gaps in our knowledge, with an emphasis on how disruption of GnRH neuronal identity can lead to CHH phenotypes.

Short-Term Exposure to Benzo(a)Pyrene Causes Disruption of GnRH Network in Zebrafish Embryos.

Benzo(a)pyrene (BaP), a polycyclic aromatic hydrocarbon, is considered a common endocrine disrupting chemical (EDC) with mutagenic and carcinogenic effects. In this work, we evaluated the effects of BaP on the hypothalamo-pituitary-gonadal axis (HPG) of zebrafish embryos. The embryos were treated with 5 and 50 nM BaP from 2.5 to 72 hours post-fertilization (hpf) and obtained data were compared with those from controls. We followed the entire development of gonadotropin releasing hormone (GnRH3) neurons that start to proliferate from the olfactory region at 36 hpf, migrate at 48 hpf and then reach the pre-optic area and the hypothalamus at 72 hpf. Interestingly, we observed a compromised neuronal architecture of the GnRH3 network after the administration of 5 and 50 nM BaP. Given the toxicity of this compound, we evaluated the expression of genes involved in antioxidant activity, oxidative DNA damage and apoptosis and we found an upregulation of these pathways. Consequently, we performed a TUNEL assay and we confirmed an increment of cell death in brain of embryos treated with BaP. In conclusion our data reveal that short-term exposure of zebrafish embryos to BaP affects GnRH3 development likely through a neurotoxic mechanism.

Acetylcholine Inhibits Spontaneous Firing Activity of Terminal Nerve GnRH Neurons in Medaka.

Vertebrates generally possess hypophysiotropic and non-hypophysiotropic gonadotropin releasing hormone (GnRH) neurons. The terminal nerve (TN) GnRH neurons are known to belong to the non-hypophysiotropic neurons and have been suggested to modulate sexual behaviors. These neurons show spontaneous pacemaker firing activity and release neuropeptides GnRH and neuropeptide FF. Since the spontaneous firing activities of peptidergic neurons, including GnRH neurons, are believed to play important roles in the release of neuropeptides, understanding the regulatory mechanisms of these spontaneous firing activities is important. Here, we analyzed firing activities of the TN-GnRH neurons in medaka during application of acetylcholine (ACh), which is one of the essential neuromodulators in the brain. Whole cell patch clamp recording of TN-GnRH neurons demonstrated that ACh induces hyperpolarization and inhibits their pacemaker firing. Electrophysiological analysis using an antagonist for acetylcholine receptors and in situ hybridization analysis showed that firing of TN-GnRH neurons is inhibited via M2-type muscarinic acetylcholine receptor. These findings, taken together with literature from several other fish species (including teleosts and elasmobranchs), indicate that ACh may generally play an inhibitory role in modulating spontaneous activities of TN-GnRH neurons and thereby sexual behaviors in fish.

Neural Control of Sexual Behavior in Fish.

Many vertebrate species show breeding periods and exhibit series of characteristic species-specific sexual behaviors only during the breeding period. Here, secretion of gonadal sex hormones from the mature gonads has been considered to facilitate sexual behaviors. Thus, the sexual behavior has long been considered to be regulated by neural and hormonal mechanisms. In this review, we discuss recent progress in the study of neural control mechanisms of sexual behavior with a focus on studies using fish, which have often been the favorite animals used by many researchers who study instinctive animal behaviors. We first discuss control mechanisms of sexual behaviors by sex steroids in relation to the anatomical studies of sex steroid-concentrating neurons in various vertebrate brains, which are abundantly distributed in evolutionarily conserved areas such as preoptic area (POA) and anterior hypothalamus. We then focus on another brain area called the ventral telencephalic area, which has also been suggested to contain sex steroid-concentrating neurons and has been implicated in the control of sexual behaviors, especially in teleosts. We also discuss control of sex-specific behaviors and sexual preference influenced by estrogenic signals or by olfactory/pheromonal signals. Finally, we briefly summarize research on the modulatory control of motivation for sexual behaviors by a group of peptidergic neurons called terminal nerve gonadotropin-releasing hormone (TN-GnRH) neurons, which are known to be especially developed in fishes among various vertebrate species.

Hypothalamic Kisspeptin Neurons as a Target for Whole-Cell Patch-Clamp Recordings.

Kisspeptins are essential for the maturation of the hypothalamic-pituitary-gonadal (HPG) axis and fertility. Hypothalamic kisspeptin neurons located in the anteroventral periventricular nucleus and rostral periventricular nucleus, as well as the arcuate nucleus of the hypothalamus, project to gonadotrophin-releasing hormone (GnRH) neurons, among other cells. Previous studies have demonstrated that kisspeptin signaling occurs through the Kiss1 receptor (Kiss1r), ultimately exciting GnRH neuron activity. In humans and experimental animal models, kisspeptins are sufficient for inducing GnRH secretion and, consequently, luteinizing hormone (LH) and follicle stimulant hormone (FSH) release. Since kisspeptins play an essential role in reproductive functions, researchers are working to assess how the intrinsic activity of hypothalamic kisspeptin neurons contributes to reproduction-related actions and identify the primary neurotransmitters/neuromodulators capable of changing these properties. The whole-cell patch-clamp technique has become a valuable tool for investigating kisspeptin neuron activity in rodent cells. This experimental technique allows researchers to record and measure spontaneous excitatory and inhibitory ionic currents, resting membrane potential, action potential firing, and other electrophysiological properties of cell membranes. In the present study, crucial aspects of the whole-cell patch-clamp technique, known as electrophysiological measurements that define hypothalamic kisspeptin neurons, and a discussion of relevant issues about the technique, are reviewed.

Defective jagged-1 signaling affects GnRH development and contributes to congenital hypogonadotropic hypogonadism.

In vertebrate species, fertility is controlled by gonadotropin-releasing hormone (GnRH) neurons. GnRH cells arise outside the central nervous system, in the developing olfactory pit, and migrate along olfactory/vomeronasal/terminal nerve axons into the forebrain during embryonic development. Congenital hypogonadotropic hypogonadism (CHH) and Kallmann syndrome are rare genetic disorders characterized by infertility, and they are associated with defects in GnRH neuron migration and/or altered GnRH secretion and signaling. Here, we documented the expression of the jagged-1/Notch signaling pathway in GnRH neurons and along the GnRH neuron migratory route both in zebrafish embryos and in human fetuses. Genetic knockdown of the zebrafish ortholog of JAG1 (jag1b) resulted in altered GnRH migration and olfactory axonal projections to the olfactory bulbs. Next-generation sequencing was performed in 467 CHH unrelated probands, leading to the identification of heterozygous rare variants in JAG1. Functional in vitro validation of JAG1 mutants revealed that 7 out of the 9 studied variants exhibited reduced protein levels and altered subcellular localization. Together our data provide compelling evidence that Jag1/Notch signaling plays a prominent role in the development of GnRH neurons, and we propose that JAG1 insufficiency may contribute to the pathogenesis of CHH in humans.

Acetylcholine regulation of GnRH neuronal activity: A circuit in the medial septum.

In vertebrates, gonadotropin-releasing hormone (GnRH)-secreting neurons control fertility by regulating gonadotrophs in the anterior pituitary. While it is known that acetylcholine (ACh) influences GnRH secretion, whether the effect is direct or indirect, and the specific ACh receptor (AChR) subtype(s) involved remain unclear. Here, we determined 1) whether ACh can modulate GnRH cellular activity and 2) a source of ACh afferents contacting GnRH neurons. Calcium imaging was used to assay GnRH neuronal activity. With GABAergic and glutamatergic transmission blocked, subtype-specific AChR agonists and antagonists were applied to identify direct regulation of GnRH neurons. ACh and nicotine caused a rise in calcium that declined gradually back to baseline after 5-6min. This response was mimicked by an alpha3-specific agonist. In contrast, muscarine inhibited GnRH calcium oscillations, and blocking M2 and M4 together prevented this inhibition. Labeling for choline acetyltransferase (ChAT) and GnRH revealed ChAT fibers contacting GnRH neurons, primarily in the medial septum (MS), and in greater number in females than males. ChAT positive cells in the MS are known to express p75NGFRs. Labeling for p75NGFR, ChAT and GnRH indicated that ChAT fibers contacting GnRH cells originate from cholinergic cells within these same rostral areas. Together, these results indicate that cholinergic cells in septal areas can directly regulate GnRH neurons.

Circadian and kisspeptin regulation of the preovulatory surge

Fertility in mammals is ultimately controlled by a small population of neurons – the gonadotropin-releasing hormone (GnRH) neurons – located in the ventral forebrain. GnRH neurons control gonadal function through the release of GnRH, which in turn stimulates the secretion of the anterior pituitary gonadotropins luteinizing hormone (LH) and follicle-stimulating hormone (FSH). In spontaneous ovulators, ovarian follicle maturation eventually stimulates, via sex steroid feedback, the mid-cycle surge in GnRH and LH secretion that causes ovulation. The GnRH/LH surge is initiated in many species just before the onset of activity through processes controlled by the central circadian clock, ensuring that the neuroendocrine control of ovulation and sex behavior are coordinated. This review aims to give an overview of anatomical and functional studies that collectively reveal some of the mechanisms through which the central circadian clock regulates GnRH neurons and their afferent circuits to drive the preovulatory surge.

Does kisspeptin exert a local modulatory effect on bovine ovarian steroidogenesis?

Kisspeptin, a hypothalamic neuropeptide encoded by the KISS1 gene, has a pivotal role in promoting GnRH secretion in mammals. Kisspeptin and its receptor (KISS1R) are also expressed in certain peripheral tissues including gonads, suggesting intra-gonadal roles. Such actions at the level of the bovine ovary have not been explored previously. The current aims were to determine whether KISS1 and its receptor (KISS1R) are expressed in the bovine ovary and whether kisspeptin or a kisspeptin antagonist can modulate ovarian steroid production by cultured ovarian cells. Granulosa (GC) and theca interna (TC) were collected from antral follicles (3-18 mm) categorized into five class sizes. Early, mid and regressing corpora lutea (CL) were also collected for RT-qPCR analysis of KISS1 and KISS1R expression. Bovine TC and GC cultured under both non-luteinizing (serum-free) and luteinizing (serum-supplemented) conditions were treated for 4 days with kisspeptin-10 (10-10-10-6M) or kisspeptin antagonist (p234; 10-10-10-6M), alone and in combination with either FSH (GC), LH (TC) or forskolin (luteinized GC/TC). Steroid secretion (GC: oestradiol, progesterone; TC: androstenedione, progesterone; luteinized GC/TC: progesterone) was measured by ELISA and viable cell number determined by neutral red uptake assay. KISS1 and KISS1R transcripts were detected in TC, GC and CL with significant differences between follicle categories and CL stages. However, neither kisspeptin-10 nor kisspeptin antagonist affected steroid secretion or viable cell number in any of the four ovarian cell culture models. As such, the hypothesis that kisspeptin has a direct intra-ovarian role to modulate follicular or luteal steroidogenesis, or cell proliferation/survival, is not supported.

Hypothalamic GnRH expression and pulsatility depends on a balance of prolactin receptors in the plains vizcacha, Lagostomus maximus.

In mammals, gestation is considered a physiological hyperprolactinemia status. Prolactin (PRL) is one of the modulators of gonadotropin-releasing hormone (GnRH) neurons function. The South American plains vizcacha (Lagostomus maximus) is a unique model to study the regulation of hypothalamic GnRH neurons by direct and indirect steroid-dependent pathways. The aim was to characterize the hypothalamic expression of endocrine markers in vizcacha during gestation as well as their response to experimental induced hyperprolactinemia. The possible involvement of PRL regulatory pathways on GnRH in the context of hypothalamic and pituitary reactivation in mid-gestating vizcachas was discussed. Using two in vivo approaches, we determined changes in the hypothalamic expression and distribution of prolactin receptor (PRLR), tyrosine hydroxylase (TH), and dopamine type 2 receptor. A significant increment in the number of tuberoinfundibular dopaminergic (TIDA) neurons was determined in the arcuate nucleus from early to term pregnancy. On the other hand, at preoptic area, the number of both TH+PRLR+ and GnRH+PRLR+ double-labeled neurons significantly decreased at mid-pregnancy probably allowing the recovery of GnRH expression indicating that both types of neurons may represent the key points of PRL indirect and direct pathways modulating GnRH. Moreover, in a model of induced hyperprolactinemic vizcachas, the inhibitory effect of PRL on GnRH at both expression and delivery levels were confirmed. These results suggest the concomitant participation of both PRL regulatory pathways on GnRH modulation and pinpoint the key role of PRL on GnRH expression enabling the recovery of the hypothalamic activity during the gestation in this species.

A functional spectrum of PROKR2 mutations identified in isolated hypogonadotropic hypogonadism.

Isolated hypogonadotropic hypogonadism (IHH) is a rare disease with hypogonadism and infertility caused by the defects in embryonic migration of hypothalamic gonadotropin-releasing hormone (GnRH) neurons, hypothalamic GnRH secretion or GnRH signal transduction. PROKR2 gene, encoding a G-protein coupled receptor PROKR2, is one of the most frequently mutated genes identified in IHH patients. However, the functional consequences of several PROKR2 mutants remain elusive. In this study, we systematically analyzed the Gαq, Gαs and ERK1/2 signaling of 23 IHH-associated PROKR2 mutations which are yet to be functionally characterized. We demonstrate that blockage of Gαq, instead of MAPK/ERK pathway, inhibited PROK2-induced migration of PROKR2-expressing cells, implying that PROKR2-related IHH results primarily due to Gαq signaling pathway disruption. Combined with previous reports, we categorized a total of 63 IHH-associated PROKR2 mutations into four distinct groups according Gαq pathway functionality: (i) neutral (N, >80% activity); (ii) low pathogenicity (L, 50-80% activity); (iii) medium pathogenicity (M, 20-50% activity) and (iv) high pathogenicity (H, <20% activity). We further compared the cell-based functional results with in silico mutational prediction programs. Our results indicated that while Sorting Intolerant from Tolerant predictions were accurate for transmembrane region mutations, mutations localized in the intracellular and extracellular domains were accurately predicted by the Combined Annotation Dependent Depletion prediction tool. Our results thus provide a functional database that can be used to guide diagnosis and appropriate genetic counseling in IHH patients with PROKR2 mutations.

Suppression of neurotransmission on gonadotropin-releasing hormone neurons in letrozole-induced polycystic ovary syndrome: A mouse model.

Polycystic ovarian syndrome (PCOS) is a heterogeneous endocrine disorder in reproductive-age women, characterized by the accretion of small cystic follicles in the ovary associated with chronic anovulation and overproduction of androgens. Ovarian function in all mammals is controlled by gonadotropin-releasing hormone (GnRH) neurons, which are the central regulator of the hypothalamic-pituitary-gonadal (HPG) axis. However, the impact on the neurotransmitter system regulating GnRH neuronal function in the letrozole-induced PCOS mouse model remains unclear. In this study, we compared the response of various neurotransmitters and neurosteroids regulating GnRH neuronal activities between letrozole-induced PCOS and normal mice via electrophysiological techniques. Response to neurotransmitter systems like GABAergic, glutamatergic and kisspeptinergic were suppressed in letrozole-fed compared to normal mice. In addition, neurosteroids tetrahydrodeoxycorticosterone (THDOC) and 4,5,6,7-tetrahydroisoxazolo[5,4-c] pyridine-3-ol (THIP) mediated response on GnRH neurons were significantly smaller on letrozole-fed mice compared to normal mice. Furthermore, we also found that letrozole-fed mice showed irregularity in the estrous cycle, increased body weight, and anovulation in female mice. These findings suggest that PCOS is an endocrine disorder that may directly affect the neurotransmitter system regulating GnRH neuronal activity at the hypothalamic level and impact reproductive physiology.

Sex differences in the coexpression of prokineticin receptor 2 and gonadal steroids receptors in mice.

Loss-of-function mutations in prokineticin 2 (PROK2) and the cognate receptor prokineticin receptor 2 (PROKR2) genes have been implicated in reproductive deficits characteristic of Kallmann Syndrome (KS). Knock out of Prokr2 gene produces the KS-like phenotype in mice resulting in impaired migration of gonadotropin releasing hormone (GnRH) neurons, olfactory bulb dysgenesis, and infertility. Beyond a developmental role, pharmacological and genetic studies have implicated PROKR2 in the control of the estrous cycle in mice. However, PROKR2 is expressed in several reproductive control sites but the brain nuclei associated with reproductive control in adult mice have not been defined. We set out to determine if ProkR2 neurons in both male and female mouse brains directly sense changes in the gonadal steroids milieu. We focused on estrogen receptor α (ERα) and androgen receptor (AR) due to their well-described function in reproductive control via actions in the brain. We found that the ProkR2-Cre neurons in the posterior nucleus of the amygdala have the highest colocalization with ERα and AR in a sex-specific manner. Few colocalization was found in the lateral septum and in the bed nucleus of the stria terminalis, and virtually no colocalization was observed in the medial amygdala. Our findings indicate that the posterior nucleus of the amygdala is the main site where PROKR2 neurons may regulate aspects of the reproductive function and social behavior in adult mice.

A classification of genes involved in normal and delayed male puberty.

Puberty is a pivotal biological process that completes sexual maturation to achieve full reproductive capability. It is a major transformational period of life, whose timing is strongly affected by genetic makeup of the individual, along with various internal and external factors. Although the exact mechanism for initiation of the cascade of molecular events that culminate in puberty is not yet known, the process of pubertal onset involves interaction of numerous complex signaling pathways of hypothalamo-pituitary-testicular (HPT) axis. We developed a classification of the mechanisms involved in male puberty that allowed placing many genes into physiological context. These include (i) hypothalamic development during embryogenesis, (ii) synaptogenesis where gonadotropin releasing hormone (GnRH) neurons form neuronal connections with suprahypothalamic neurons, (iii) maintenance of neuron homeostasis, (iv) regulation of synthesis and secretion of GnRH, (v) appropriate receptors/proteins on neurons governing GnRH production and release, (vi) signaling molecules activated by the receptors, (vii) the synthesis and release of GnRH, (viii) the production and release of gonadotropins, (ix) testicular development, (x) synthesis and release of steroid hormones from testes, and (xi)the action of steroid hormones in downstream effector tissues. Defects in components of this system during embryonic development, childhood/adolescence, or adulthood may disrupt/nullify puberty, leading to long-term male infertility and/or hypogonadism. This review provides a list of 598 genes involved in the development of HPT axis and classified according to this schema. Furthermore, this review identifies a subset of 75 genes for which genetic mutations are reported to delay or disrupt male puberty.

Adolescence is a sensitive period for acrylamide-induced sex hormone disruption: Evidence from NHANES populations and experimental mice

Acrylamide (AA) is widely contaminated in environment and diet. However, the association of AA and sex hormones has rarely been investigated, especially in adolescents, a period of particular susceptibility to sex hormone disruption. In this study, survey-weighted multivariate linear regression models were conducted to determine the association between AA Hb biomarkers [HbAA and glycidamide (HbGA)] and sex hormones [total testosterone (TT) and estradiol (E2)] in a total of 3268 subjects from National Health and Nutrition Examination Survey (NHANES) 2013–2016 waves. Additionally, adult and pubertal mice were treated with AA to assess the effect of AA on sex hormones and to explore the potential mechanisms. Among all the subjects, significant negative patterns for HbGA and sex hormones were identified only in youths (6–19 years old), with the lowest β being − 0.53 (95% CI: −0.80 to −0.26) for TT in males and − 0.58 (95% CI: −0.93 to −0.23) for E2 in females. Stratified analysis further revealed significant negative associations between HbGA and sex hormones in adolescents, with the lowest β being − 0.58 (95% CI: −1.02 to −0.14) for TT in males and − 0.54 (95% CI: −1.03 to −0.04) for E2 in females, while there were no significant differences between children or late adolescents. In mice, the levels of TT and E2 were dramatically reduced in AA-treated pubertal mice but not in adult mice. AA disturbed the expression of genes in the hypothalamic–pituitary–gonadal (HPG) axis, induced apoptosis of hypothalamus-produced gonadotropin-releasing hormone (GnRH) neurons in the hypothalamus and reduced serum and hypothalamic GnRH levels in pubertal mice. Our study indicates AA could reduce TT and E2 levels by injuring GnRH neurons and disrupting the HPG axis in puberty, which manifested as severe endocrine disruption on adolescents. Our findings reinforce the idea that adolescence is a vulnerable stage in AA-induced sex hormone disruption.

Effects of benzo[a]pyrene on the reproductive axis: Impairment of kisspeptin signaling in human gonadotropin-releasing hormone primary neurons

The neuroendocrine control of reproduction is strictly coordinated at the central level by the pulsatile release of gonadotropin-releasing hormone (GnRH) by the hypothalamic GnRH neurons. Alterations of the GnRH-network, especially during development, lead to long-term reproductive and systemic consequences, also causing infertility. Recent evidence shows that benzo[a]pyrene (BaP), a diffuse pollutant that can play a role as an endocrine disruptor, affects gonadal function and gamete maturation, whereas data demonstrating its impact at hypothalamic level are very scarce. This study investigated the effects of BaP (10 μM) in a primary cell culture isolated from the human fetal hypothalamus (hfHypo) and exhibiting a clear GnRH neuron phenotype. BaP significantly decreased gene and protein expression of both GnRH and kisspeptin receptor (KISS1R), the master regulator of GnRH neuron function. Moreover, BaP exposure increased phospho-ERK1/2 signaling, a well-known mechanism associated with KISS1R activation. Interestingly, BaP altered the electrophysiological membrane properties leading to a significant depolarizing effect and it also significantly increased GnRH release, with both effects being not affected by kisspeptin addition. In conclusion, our findings demonstrate that BaP may alter GnRH neuron phenotype and function, mainly interfering with KISS1R signaling and GnRH secretion and therefore with crucial mechanisms implicated in the central neuroendocrine control of reproduction.