Exposure to Cadmium Alters the Population of Glial Cell Types and Disrupts the Regulatory Mechanisms of the HPG Axis in Prepubertal Female Rats.

Despite the fact that the brain is susceptible to neurotoxicity induced by cadmium (Cd), the effects of Cd on the neuroanatomical development in the hypothalamus and regulatory mechanisms of the hypothalamic-pituitary-gonadal (HPG) axis are not fully understood. To clarify this issue, we investigated the effects of 25 mg/kg BW/day cadmium chloride (CdCl2) on neuroanatomical alterations in the hypothalamus of prepubertal female rats. Twenty-four Sprague-Dawley rats were randomly assigned to two groups (n = 12), and CdCl2 was administered via gavage from postnatal days (PND) 21 to PND35. The results of the stereological analysis demonstrated that prepubertal exposure to Cd reduced the number of neurons and oligodendrocytes in the arcuate (ARC) and dorsomedial hypothalamus nucleus (DMH) nuclei. In contrast, Cd exposure increased the number of microglial cells in the ARC and DMH nuclei. Cd exposure decreased the mRNA levels of gonadotropin-releasing hormone (GnRH) and increased the mRNA levels of RFamide-related peptide (RFRP-3), but not kisspeptin (Kiss1) in the hypothalamus. Moreover, hormonal assay showed that Cd exposure caused a reduction in the concentration of gonadotropins: luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in serum. Immunohistochemical expression of RFRP-3 in neuronal cell bodies demonstrated that the mean number of RFRP-3 expressing neurons in the DMH nucleus of cadmium-treated rats was dramatically higher than the vehicle group. Overall, exposure to Cd during the prepubertal period alters the population of neurons and glial cell types in the hypothalamus. Additionally, Cd exposure disrupts the regulatory mechanisms of the HPG axis.

Advancing reproductive neuroendocrinology through research on the regulation of GnIH and on its diverse actions on reproductive physiology and behavior.

The discovery of gonadotropin-inhibitory hormone (GnIH) in 2000 has led to a new research era of reproductive neuroendocrinology because, for a long time, researchers believed that only gonadotropin-releasing hormone (GnRH) regulated reproduction as a neurohormone. Later studies on GnIH demonstrated that it acts as a new key neurohormone inhibiting reproduction in vertebrates. GnIH reduces gonadotropin release andsynthesis via the GnIH receptor GPR147 on gonadotropes and GnRH neurons. Furthermore, GnIH inhibits reproductive behavior, in addition to reproductive neuroendocrine function. The modification of the synthesis of GnIH and its release by the neuroendocrine integration of environmental and internal factors has also been demonstrated. Thus, the discovery of GnIH has facilitated advances in reproductive neuroendocrinology. Here, we describe the advances in reproductive neuroendocrinology driven by the discovery of GnIH, research on the effects of GnIH on reproductive physiology and behavior, and the regulatory mechanisms underlying GnIH synthesis and release.

Gonadotropin-inhibitory hormone (GnIH): A new key neurohormone controlling reproductive physiology and behavior.

The discovery of novel neurohormones is important for the advancement of neuroendocrinology. In early 1970s, gonadotropin-releasing hormone (GnRH), a hypothalamic neuropeptide that promotes gonadotropin release, was identified to be an endogenous neurohormone in mammals. In 2000, thirty years later, another hypothalamic neuropeptide, gonadotropin-inhibitory hormone (GnIH), that inhibits gonadotropin release, was found in quail. GnIH acts via GPR147 and inhibits gonadotropin release and synthesis and reproductive function in birds through actions on GnRH neurons in the hypothalamus and pituitary gonadotrophs. Later, GnIH was found in other vertebrates including humans. GnIH studies have advanced the progress of reproductive neuroendocrinology. Furthermore, recent GnIH studies have indicated that abnormal changes in GnIH expression may cause pubertal disorder and reproductive dysfunction. Here, we describe GnIH discovery and its impact on the progress of reproductive neuroendocrinology. This review also highlights advancement and perspective of GnIH studies on drug development for pubertal disorder and reproductive dysfunction. (149/150).

Gonadotropin-inhibitory hormone (GnIH) distribution in the brain of the ancient fish Atractosteus tropicus (Holostei, Lepisosteiformes).

The Holostei group occupies a critical phylogenetic position as the sister group of the Teleostei. However, little is known about holostean pituitary anatomy or brain distribution of important reproductive neuropeptides, such as the gonadotropin-inhibitory hormone (GnIH). Thus, the present study set out to characterize the structure of the pituitary and to localize GnIH-immunoreactive cells in the brain of Atractosteus tropicus from the viewpoint of comparative neuroanatomy. Juveniles of both sexes were processed for general histology and immunohistochemistry. Based on the differences in cell organization, morphology, and staining properties, the neurohypophysis and three regions in the adenohypophysis were identified: the rostral and proximal pars distalis (PPD) and the pars intermedia. This last region was found to be innervated by the neurohypophysis. This organization, together with the presence of a saccus vasculosus, resembles the general teleost pituitary organization. A vast number of blood vessels were also recognized between the infundibulum floor of the hypothalamus and the PPD, evidencing the characteristic presence of a median eminence and a portal system. However, this well-developed pituitary portal system resembles that of tetrapods. As regards the immunohistochemical localization of GnIH, we found four GnIH-immunoreactive (GnIH-ir) populations in three hypothalamic nuclei (suprachiasmatic, retrotuberal, and tuberal nuclei) and one in the diencephalon (prethalamic nucleus), as well as a few scattered neurons throughout the olfactory bulbs, the telencephalon, and the intersection between them. GnIH-ir fibers showed a widespread distribution over almost all brain regions, suggesting that GnIH function is not restricted to reproduction only. In conclusion, the present study describes, for the first time, the pituitary of A. tropicus and the neuroanatomical localization of GnIH in a holostean fish that exhibits a similar distribution pattern to that of teleosts and other vertebrates, suggesting a high degree of phylogenetic conservation of this system.

Regulation of the hypothalamic GnRH-GnIH system by putrescine in adult female rats and GT1-7 neuronal cell line.

The gonadotropin-releasing hormone-gonadotropin inhibitor (GnRH-GnIH) system in the hypothalamus of mammals is the key factor that controls the entire reproductive system. The aim of this study was to immunolocalize GnIH (RFRP-3) in the hypothalamus during the estrous cycle and to study the effect of putrescine on the expression of GnRH-I and GnIH through both in vivo and in vitro (GT1-7 cells) approach and the circulatory levels of GnRH-I, GnIH, and gonadotropins were also investigated. The study also aims in analyzing all the immunofluorescence images by measuring the relative pixel count of an image. This study showed the effect of putrescine on the morphology of ovary, uterus, and the expression of the steroidogenic acute regulatory protein in the ovary. This study showed GnIH expression was intense during the diestrus and moderate during proestrus and estrus, whereas mild staining during the metestrus. The study further showed that putrescine supplementation to adult female rats increased both GnRH-I expression in the hypothalamus as well as the GnRH-I levels in circulation. The study, for the first time, also showed that putrescine supplementation decreased the expression and release of GnIH. These effects of upregulating GnRH-I expression and downregulating GnIH expression were confirmed by in vitro experiments using GT1-7 cells. Putrescine supplementation also increased the gonadotropin levels in the serum. To summarize, putrescine can regulate the hypothalamic-pituitary-gonadal axis by increasing the GnRH-I, luteinizing hormone, and follicle-stimulating hormone levels and suppressing GnIH levels. This is the first report showing the simultaneous effects of putrescine on the regulation of both GnRH-I and GnIH in the hypothalamus.

Light-at-night exposure affects brain development through pineal allopregnanolone-dependent mechanisms.

The molecular mechanisms by which environmental light conditions affect cerebellar development are incompletely understood. We showed that circadian disruption by light-at-night induced Purkinje cell death through pineal allopregnanolone (ALLO) activity during early life in chicks. Light-at-night caused the loss of diurnal variation of pineal ALLO synthesis during early life and led to cerebellar Purkinje cell death, which was suppressed by a daily injection of ALLO. The loss of diurnal variation of pineal ALLO synthesis induced not only reduction in pituitary adenylate cyclase-activating polypeptide (PACAP), a neuroprotective hormone, but also transcriptional repression of the cerebellar Adcyap1 gene that produces PACAP, with subsequent Purkinje cell death. Taken together, pineal ALLO mediated the effect of light on early cerebellar development in chicks.

Molecular Mechanisms of Gonadotropin-Inhibitory Hormone (GnIH) Actions in Target Cells and Regulation of GnIH Expression.

Since gonadotropin-inhibitory hormone (GnIH) was discovered in 2000 as the first hypothalamic neuropeptide that actively inhibits gonadotropin release, researches conducted for the last 18 years have demonstrated that GnIH acts as a pronounced negative regulator of reproduction. Inhibitory effect of GnIH on reproduction is mainly accomplished at hypothalamic-pituitary levels; gonadotropin-releasing hormone (GnRH) neurons and gonadotropes are major targets of GnIH action based on the morphological interaction with GnIH neuronal fibers and the distribution of GnIH receptor. Here, we review molecular studies mainly focusing on the signal transduction pathway of GnIH in target cells, GnRH neurons, and gonadotropes. The use of well-defined cellular model systems allows the mechanistic study of signaling pathway occurring in target cells by demonstrating the direct cause-and-effect relationship. The insights gained through studying molecular mechanism of GnIH action contribute to deeper understanding of the mechanism of how GnIH communicates with other neuronal signaling systems to control our reproductive function. Reproductive axis closely interacts with other endocrine systems, thus GnIH expression levels would be changed by adrenal and thyroid status. We also briefly review molecular studies investigating the regulatory mechanisms of GnIH expression to understand the role of GnIH as a mediator between adrenal, thyroid and gonadal axes.

Brain mapping of the gonadotropin-inhibitory hormone-related peptide 2 with a novel antibody suggests a connection with emotional reactivity in the Japanese quail (Coturnix japonica, Temminck & Schlegel, 1849).

Gonadotropin-inhibitory hormone (GnIH) is a neuropeptide first discovered in the quail brain that is involved in the control of reproductive physiology and behaviors, and stress response. GnIH gene encodes a second peptide, GnIH-related peptide-2 (RP2), the distribution and function of which remain unknown. We therefore studied GnIH-RP2 distribution by immunohistochemistry using a novel antibody capable of discriminating between GnIH and GnIH-RP2. The overall distribution of GnIH-RP2 is similar to that of GnIH. The vast majority of labeled neurons is located in the paraventricular nucleus (PVN) of the hypothalamus. Labeling of fibers is conspicuous in the diencephalon, but present also in the mesencephalon and telencephalon. Several regions involved in the control of reproduction and stress response (the PVN, septum, bed nucleus of the stria terminalis and nucleus commissura pallii) showed a dense network of immunolabeled fibers. To investigate the potential function of GnIH-RP2 we compared its expression in two quail lines genetically selected for divergence in their emotional reactivity. A quantitative analysis in the above-mentioned brain regions showed that the density of fibers was similar in the two lines. However, the number of GnIH-RP2 labeled neurons was higher in the median portion of the PVN in birds with higher emotional reactivity. These results point to a possible involvement of GnRH-RP2 in modulating stress response and/or emotional reactivity.

Morphological relationship between GnIH and GnRH neurons in the brain of the neotropical cichlid fish Cichlasoma dimerus.

Reproduction is regulated by the hypothalamic-pituitary-gonadal axis. The first neuropeptide identified that regulates this function was the decapeptide gonadotropin-releasing hormone (GnRH). Nowadays, in gnatostomates, a number of GnRH variants have been identified and classified into three different types: GnRH1, GnRH2, and GnRH3. Almost 30 years later, a new peptide that inhibits gonadotropin synthesis and secretion was discovered and thus named as gonadotropin-inhibitory hormone (GnIH). In avians and mammals, the interaction and regulation between GnRH and GnIH neurons has been widely studied; however, in other vertebrate groups there is little information about the relationship between these neurons. In previous works, three GnRH variants and a GnIH propeptide were characterized in Cichlasoma dimerus, and it was demonstrated that GnIH inhibited gonadotropins release in this species. Because no innervation was detected at the pituitary level, we speculate that GnIH would inhibit gonadotropins via GnRH. Thus, the aim of the present study was to evaluate the anatomical relationship between neurons expressing GnIH and the three GnRH variants by double labelling confocal immunofluorescence in adults of C. dimerus. Our results showed no apparent contacts between GnIH and GnRH1, fiber to fiber interactions between GnIH and GnRH2, and co-localization of GnIH and GnRH3 variant in neurons of the nucleus olfacto-retinalis. In conclusion, whether GnIH regulates the expression or secretion of GnRH1 in this species, an indirect modulation seems more plausible. Moreover, the present results suggest an interaction between GnIH and GnRH2 systems. Finally, new clues were provided to investigate the role of nucleus olfacto-retinalis cells and putative GnIH and GnRH3 interactions in the modulation of the reproductive network in teleost fish.

Kobayashi award: Discovery of cerebellar and pineal neurosteroids and their biological actions on the growth and survival of Purkinje cells during development (review).

The brain has traditionally been considered to be a target site of peripheral steroid hormones. On the other hand, extensive studies over the past thirty years have demonstrated that the brain is a site of biosynthesis of several steroids. Such steroids synthesized de novo from cholesterol in the brain are called neurosteroids. To investigate the biosynthesis and biological actions of neurosteroids in the brain, data on the regio- and temporal-specific synthesis of neurosteroids are needed. In the mid 1990s, the Purkinje cell, an important cerebellar neuron, was discovered as a major cell producing neurosteroids in the brain of vertebrates. It was the first demonstration of de novo neuronal biosynthesis of neurosteroids in the brain. Subsequently, neuronal biosynthesis of neurosteroids and biological actions of neurosteroids have become clear by the follow-up studies using the Purkinje cell as an excellent cellular model. Progesterone and estradiol, which are known as sex steroid hormones, are actively synthesized de novo from cholesterol in the Purkinje cell during development, when cerebellar neuronal circuit formation occurs. Importantly, progesterone and estradiol synthesized in the Purkinje cell promote dendritic growth, spinogenesis and synaptogenesis via their cognate nuclear receptors in the Purkinje cell. Neurotrophic factors may mediate these neurosteroid actions. Futhermore, allopregnanolone (3α,5α-tetrahydroprogesterone), a progesterone metabolite, is also synthesized in the cerebellum and acts on the survival of Purkinje cells. On the other hand, at the beginning of 2010s, the pineal gland, an endocrine organ located close to the cerebellum, was discovered as an important site of the biosynthesis of neurosteroids. Allopregnanolone, a major pineal neurosteroid, acts on the Purkinje cell for the survival of Purkinje cells by suppressing the expression of caspase-3, a crucial mediator of apoptosis. I as a recipient of Kobayashi Award from the Japan Society for Comparative Endocrinology in 2016 summarize the discovery of cerebellar and pineal neurosteroids and their biological actions on the growth and survival of Purkinje cells during development.

Role of RFRP-3 in the Regulation of Kiss-1 Gene Expression in the AVPV Hypothalamic Cell Model mHypoA-50.

Kisspeptin, encoded by the Kiss-1 gene, plays a crucial role in reproductive function by governing the hypothalamic-pituitary-gonadal axis. The recently established Kiss-1-expressing cell model mHypoA-50 displays characteristics of neuronal cells of the anteroventral periventricular (AVPV) region of the mouse hypothalamus. Because Kiss-1 gene expression in these cells is upregulated by estradiol (E2), mHypoA-50 cells are regarded as a valuable model for the study of Kiss-1-expressing neurons in the AVPV region. These cells also express RFamide-related peptide-3 (RFRP-3), a mammalian homolog of gonadotropin inhibitory hormone. The RFRP-3 expression in mHypoA-50 cells was increased by melatonin stimulation. In addition, E2 stimulation increased RFRP-3 expression in these cells. Treatment of the mHypoA-50 cells with exogenous RFRP-3 resulted in the increase of Kiss-1 messenger RNA expression within the cells; however, RFRP-3 did not modify gonadotropin-releasing hormone or kisspeptin-induced Kiss-1 gene expression in these cells. In addition, we found that RFRP-3 stimulation increased the expression of corticotropin-releasing hormone, which may be involved in E2-induced positive feedback in mHypoA-50 cells. Our observations suggest that RFRP-3 might be involved in positive feedback regulation by directly or indirectly increasing Kiss-1 gene expression.

Action of neurotensin, corticotropin-releasing hormone, and RFamide-related peptide-3 in E2-induced negative feedback control: studies using a mouse arcuate nucleus hypothalamic cell model.

The recently established immortalized hypothalamic cell model mHypoA-55 possesses characteristics similar to those of Kiss-1 neurons in the arcuate nucleus (ARC) region of the hypothalamus. Here, we show that Kiss-1 gene expression in these cells was downregulated by 17ß-estradiol (E2) under certain conditions. Both neurotensin (NT) and corticotropin-releasing hormone (CRH) were expressed in these cells and upregulated by E2. Stimulation of mHypoA-55 cells with NT and CRH significantly decreased Kiss-1 mRNA expression. A mammalian gonadotropin-inhibitory hormone homolog, RFamide-related peptide-3 (RFRP-3), was also found to be expressed in mHypoA-55 cells, and RFRP-3 expression in these cells was increased by exogenous melatonin stimulation. E2 stimulation also upregulated RFRP-3 expression in these cells. Stimulation of mHypoA-55 cells with RFRP-3 significantly increased the expression of NT and CRH. Furthermore, melatonin stimulation resulted in the increase of both NT and CRH mRNA expression in mHypoA-55 cells. On the other hand, in experiments using mHypoA-50 cells, which were originally derived from hypothalamic neurons in the anteroventral periventricular nucleus, Kiss-1 gene expression was upregulated by both NT and CRH, although E2 increased both NT and CRH expression, similarly to the mHypoA-55 cells. Our observations using the hypothalamic ARC cell model mHypoA-55 suggest that NT and CRH have inhibitory effects on Kiss-1 gene expression under the influence of E2 in association with RFRP-3 expression. Thus, these neuropeptides might be involved in E2-induced negative feedback mechanisms.

Strain differences in intermale aggression and possible factors regulating increased aggression in Japanese quail.

The National Institute for Environmental Studies (NIES) of Japan established a strain of Japanese quail (Coturnix japonica) known as NIES-L by rotation breeding in a closed colony for over 35years; accordingly, the strain has highly inbred-like characteristics. Another strain called NIES-Brn has been maintained by randomized breeding in a closed colony to produce outbred-like characteristics. The current study aimed to characterize intermale aggressive behaviors in both strains and to identify possible factors regulating higher aggression in the hypothalamus, such as sex hormone and neuropeptide expression. Both strains displayed a common set of intermale aggressive behaviors that included pecking, grabbing, mounting, and cloacal contact behavior, although NIES-Brn quail showed significantly more grabbing, mounting, and cloacal contact behavior than did NIES-L quail. We examined sex hormone levels in the blood and diencephalon in both strains. Testosterone concentrations were significantly higher in the blood and diencephalon of NIES-Brn quail compared to NIES-L quail. We next examined gene expression in the hypothalamus of both strains using an Agilent gene expression microarray and real-time RT-PCR and found that gene expression of mesotocin (an oxytocin homologue) was significantly higher in the hypothalamus of NIES-Brn quail compared to NIES-L quail. Immunohistochemistry of the hypothalamus revealed that numbers of large cells (cell area>500µm2) expressing mesotocin were significantly higher in the NIES-Brn strain compared to the NIES-L strain. Taken together, our findings suggest that higher testosterone and mesotocin levels in the hypothalamus may be responsible for higher aggression in the NIES-Brn quail strain.

The Arg-Phe-amide peptide 26RFa/glutamine RF-amide peptide and its receptor: IUPHAR Review 24.

The RFamide neuropeptide 26RFa was first isolated from the brain of the European green frog on the basis of cross-reactivity with antibodies raised against bovine neuropeptide FF (NPFF). 26RFa and its N-terminally extended form glutamine RF-amide peptide (QRFP) have been identified as cognate ligands of the former orphan receptor GPR103, now renamed glutamine RF-amide peptide receptor (QRFP receptor). The 26RFa/QRFP precursor has been characterized in various mammalian and non-mammalian species. In the brain of mammals, including humans, 26RFa/QRFP mRNA is almost exclusively expressed in hypothalamic nuclei. The 26RFa/QRFP transcript is also present in various organs especially in endocrine glands. While humans express only one QRFP receptor, two isoforms are present in rodents. The QRFP receptor genes are widely expressed in the CNS and in peripheral tissues, notably in bone, heart, kidney, pancreas and testis. Structure-activity relationship studies have led to the identification of low MW peptidergic agonists and antagonists of QRFP receptor. Concurrently, several selective non-peptidic antagonists have been designed from high-throughput screening hit optimization. Consistent with the widespread distribution of QRFP receptor mRNA and 26RFa binding sites, 26RFa/QRFP exerts a large range of biological activities, notably in the control of energy homeostasis, bone formation and nociception that are mediated by QRFP receptor or NPFF2. The present report reviews the current knowledge concerning the 26RFa/QRFP-QRFP receptor system and discusses the potential use of selective QRFP receptor ligands for therapeutic applications.

Direct effects of RFRP-1, a mammalian GnIH ortholog, on ovarian activities of the cyclic mouse.

Arg(R)-Phe(F)-amide related peptide-1 (RFRP-1) and -3 (RFRP-3) are known as mammalian orthologs of gonadotropin-inhibitory hormone (GnIH). In mammals, these RFRPs are expressed not only in the hypothalamus and but also in gonads. Inhibitory roles of the hypothalamic and gonadal RFRP-3 in reproduction have been documented in mammals. However, functional roles of the hypothalamic and gonadal RFRP-1 in reproduction are still unclear in mammals. Therefore, in vitro studies were conducted to elucidate the direct effect of RFRP-1, a mammalian GnIH ortholog, on ovarian activities, such as steroidogenesis, apoptosis, cell proliferation and metabolism in the cyclic mouse. The ovaries collected from the proestrus mice were cultured in vitro with different doses (Control, 1ng/ml, 10ng/ml and 100ng/ml) of RFRP-1 for 24h at 37°C. A significant dose-dependent increase in estradiol release from the ovary was detected after the treatment of RFRP-1. Therefore, changes in the ovarian activities, such as steroidogenic markers (luteinizing hormone receptors; LH-R and 3ß-hydroxysteroid dehydrogenase; 3ß-HSD), apoptotic markers [Poly(ADP-ribose) polymerase-1; PARP-1 and cysteine-aspartic protease; caspase-3], a cell proliferation marker (proliferating cell nuclear antigen; PCNA) and metabolic markers (GLUT-4; glucose uptake) were assessed by the treatment of RFRP-1 in the proestrus ovary. The densitometry analysis showed the treatment of RFRP-1 significantly increased the expressions of LH-R and 3ß-HSD, steroidogenic markers. In contrast, the expressions of PCNA, a cell proliferation maker; PARP-1 and caspase-3, apoptotic markers were significantly decreased. Interestingly, RFRP-1 treatment further increases significantly glucose uptake and GLUT-4 receptor expression. These findings indicate that RFRP-1 possesses a stimulatory effect on ovarian steroidogenesis in the proestrus mouse. This is the first evidence showing the direct action of RFRP-1 on steroidogenesis in any vertebrate. In addition, RFRP-1 may also act directly on ovarian folliculogenesis as an inhibitory factor.

Thyroid disrupting pesticides impair the hypothalamic-pituitary-testicular axis of a wildlife bird, Amandava amandava.

The effect of two thyroid disrupting pesticides (TDPs) mancozeb (MCZ) and imidacloprid (IMI) on the hypothalamic-pituitary-gonadal/testicular (HPG) axis of a seasonally breeding bird, Amandava amandava has been evaluated. Male birds (n=8/group) were exposed to each of the pesticide (0.25% LD50 of respective pesticide) as well as to their two equimixture doses (0.25% of LD50 of each and 0.5% LD50 of each) through food for 30d during pre-breeding stage of the reproductive cycle. Reduction in weight, volume and other histopathological features revealed testicular regression. Suppression of gonadotropin releasing hormone, increased expression of gonadotropin inhibitory hormone in the hypothalamus of exposed groups as well as impairment of plasma levels of the reproduction related hormones indicated the disruption of the HPG axis. The pesticides interference of the thyroid function during the critical phase of reproductive development impaired the HPG axis; more significantly in co-exposed groups suggesting the cumulative toxicity.

Endocrine disrupting pesticides impair the neuroendocrine regulation of reproductive behaviors and secondary sexual characters of red munia (Amandava amandava).

The exposure effects of two endocrine disrupting pesticides (EDPs), mancozeb/MCZ and imidacloprid/IMI of the group dithiocarbamate and neonicotinoid respectively, on reproductive behaviors and secondary sexual characters have been studied in a seasonally breeding wildlife bird, red munia (Amandava amandava). Adult male birds were exposed to both the pesticides individually (0.25% LD50 of each) as well as co-exposed (MIX-I: 0.25% LD50 of each and MIX-II: 0.5% LD50 of each) through food for 30d in preparatory (July-August) and breeding (September-October) phase of reproductive cycle. Singing and pairing patterns started decreasing from 2nd week to complete disappearance during 4th week of pesticides exposures at both the phases of reproductive cycles. Similar trend was observed in the disappearance of spots on the plumage as well as color of both plumage and beak which turned black/gray from red. Pesticides caused impairment of the lactotropic as well as hypothalamic-pituitary-testicular (HPT) axes as there was increased plasma PRL and decreased LH, FSH and testosterone levels. Testicular expressions of GnRH and androgen receptor/AR were significantly decreased but that of GnIH significantly increased as compared to control. Significant differences among individually- and co-exposed groups were also present. Abnormalities in sexual behaviors and secondary sexual characteristics could be linked to inhibition of HPT axis and/or direct toxicity at the level of hypothalamus, pituitary and testis. In addition, pesticide-induced hyperprolactinemia as well as impaired thyroid hormones might have also affected maintenance of reproductive behaviors. On co-exposures, the more distinct impairments might be due to cumulative toxicity of pesticides.

Gonadotropin-inhibitory hormone (GnIH) in the amphibian brain and its relationship with the gonadotropin releasing hormone (GnRH) system: An overview.

It is well known that the hypothalamic neuropeptide gonadotropin-releasing hormone (GnRH) plays an important role as a primary factor regulating gonadotropin secretion in reproductive processes in vertebrates. The discovery of the presence of a gonadotropin-inhibitory hormone (GnIH) in the brains of birds has further contributed to our understanding of the reproduction control by the brain. GnIH plays a key role in inhibition of reproduction and acts on the pituitary gland and GnRH neurons via a novel G protein-coupled receptor (GPR147). GnIH decreases gonadotropin synthesis and release, thus inhibiting gonadal development and maintenance. The GnRH and GnIH neuronal peptidergic systems are well reported in mammals and birds, but limited information is available regarding their presence and localization in the brains of other vertebrate species, such as reptiles, amphibians and fishes. The aim of this review is to compile and update information on the localization of GnRH and GnIH neuronal systems, with a particular focus on amphibians, summarizing the neuroanatomical distribution of GnIH and GnRH and emphasizing the discovery of GnIH based on RFamide peptides and GnIH orthologous peptides found in other vertebrates and their functional significance.

Apoptosis-mediated testicular alteration in Japanese quail (Coturnix coturnix japonica) in response to temporal phase relation of serotonergic and dopaminergic oscillations.

Reproductive performance of many avian species, including Japanese quail, is reported to be modulated by specific temporal phase relation of serotonergic and dopaminergic oscillations. Accordingly, it has been shown that the serotonin precursor 5-HTP and the dopamine precursor l-DOPA given 8 h apart induce gonadal suppression and given 12 h apart lead to gonadal stimulation, while other temporal relationships were found to be ineffective. In the present study, we investigated the effects of 8- and 12-h phase relation of neural oscillations on testicular responses including expression of GnRH-I, GnIH, pro-apoptotic proteins (p53 and Bax), inactive and active executioner caspase-3, and the uncleaved DNA repair enzyme PARP-1. Testicular volume and mass decreased significantly in 8-h quail and increased in 12-h quail compared with controls. Expression of ir-GnIH, p53, Bax and active-caspase-3 increased and that of GnRH-I, pro-caspase-3 and uncleaved PARP-1 decreased in 8-h quail compared with controls. A TUNEL assay also confirmed testicular regression in these quail. Testes of 12-h quail exhibited significantly increased expression of GnRH-I, pro-caspase-3 and uncleaved PARP-1 compared with the control group. Our findings suggest that differential response of avian testes to 8- and 12-h phase relation of serotonergic and dopaminergic neural oscillations may be attributed to autocrine/paracrine action of GnIH expression, which is upregulated in regressed testes, leading to apoptotic changes, and downregulated in developed testes, causing apoptotic inhibition. It is concluded that specific phase relation of neural oscillations may modulate the local testicular GnRH-GnIH system and alter the apoptotic mechanism in quail testes. Moreover, these findings highlight the physiological effects of time-dependent drug delivery, including the specific time intervals between two drugs.

The ecological and physiological bases of variation in the phenology of gonad growth in an urban and desert songbird.

Birds often adjust to urban areas by advancing the timing (phenology) of vernal gonad growth. However, the ecological and physiological bases of this adjustment are unclear. We tested whether the habitat-related disparity in gonad growth phenology of male Abert's towhees, Melozone aberti, is due to greater food availability in urban areas of Phoenix, Arizona USA or, alternatively, a habitat-related difference in the phenology of key food types. To better understand the physiological mechanism underlying variation in gonad growth phenology, we compared the activity of the reproductive system at all levels of hypothalamo-pituitary-gonadal (HPG) axis. We found no habitat-associated difference in food availability (ground arthropod biomass), but, in contrast to the seasonal growth of leaves on desert trees, the leaf foliage of urban trees was already developed at the beginning of our study. Multiple estimates of energetic status did not significantly differ between the non-urban and urban towhees during three years that differed in the habitat-related disparity in gonad growth and winter precipitation levels. Thus, our results provide no support for the hypothesis that greater food abundance in urban areas of Phoenix drives the habitat-related disparity in gonad growth phenology in Abert's towhees. By contrast, they suggest that differences in the predictability and magnitude of change in food availability between urban and desert areas of Phoenix contribute to the observed habitat-related disparity in gonad growth. Endocrine responsiveness of the gonads may contribute to this phenomenon as desert - but not urban - towhees had a marked plasma testosterone response to GnRH challenge.