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Although there is a great demand for increased coronavirus disease 2019vaccination worldwide, rare side effects of the vaccines in susceptible individuals are attracting attention. We recently treated two patients who developed systemic lupus erythematosus after administration of a severe acute respiratory syndrome coronavirus 2 vaccine from Pfizer-BioNTech or Moderna. While causal relationships between vaccination and adverse events are difficult to discern due to both confounding and masking factors, our findings suggest that attention to possible adjuvant-related autoimmune diseases in certain individuals receiving severe acute respiratory syndrome coronavirus 2 vaccines is appropriate.

Many organisms living along the coastlines synchronize their reproduction with the lunar cycle. At the time of spring tide, thousands of grass puffers (Takifugu alboplumbeus) aggregate and vigorously tremble their bodies at the water's edge to spawn. To understand the mechanisms underlying this spectacular semilunar beach spawning, we collected the hypothalamus and pituitary from male grass puffers every week for 2 months. RNA sequencing (RNA-seq) analysis identified 125 semilunar genes, including genes crucial for reproduction (e.g., gonadotropin-releasing hormone 1 [gnrh1], luteinizing hormone β subunit [lhb]) and receptors for pheromone prostaglandin E (PGE). PGE

In multi-tree-structured Peer-to-Peer (P2P) video streaming, video streams are delivered from the source peer to subscribing peers in such a way that each stream is divided into several sub-streams and those sub-streams are delivered through different spanning trees. In this paper, we focus on the delivery of all sub-streams to [Formula: see text] subscribers through spanning trees of depth two each, which will be referred to as the 2-hop delivery of sub-streams. The main contribution of the current paper is the derivation of a tight lower bound on the upload capacity of homogeneous peers so that 2-hop delivery is possible if and only if the upload capacity of each peer is greater than or equal to the derived bound.

Many organisms living along the coastlines synchronize their reproduction with the lunar cycle. At the time of spring tide, thousands of grass puffers (Takifugu alboplumbeus) aggregate and vigorously tremble their bodies at the water’s edge to spawn. To understand the mechanisms underlying this spectacular semilunar beach spawning, we collected the hypothalamus and pituitary from male grass puffers every week for 2 months. RNA sequencing (RNA-seq) analysis identified 125 semilunar genes, including genes crucial for reproduction (e.g., gonadotropin-releasing hormone 1 [gnrh1], luteinizing hormone β subunit [lhb]) and receptors for pheromone prostaglandin E (PGE). PGE2 is secreted into the seawater during the spawning, and its administration activates olfactory sensory neurons and triggers trembling behavior of surrounding individuals. These results suggest that PGE2 synchronizes lunar-regulated beach-spawning behavior in grass puffers. To further explore the mechanism that regulates the lunar-synchronized transcription of semilunar genes, we searched for semilunar transcription factors. Spatial transcriptomics and multiplex fluorescent in situ hybridization showed co-localization of the semilunar transcription factor CCAAT/enhancer-binding protein δ (cebpd) and gnrh1, and cebpd induced the promoter activity of gnrh1. Taken together, our study demonstrates semilunar genes that mediate lunar-synchronized beach-spawning behavior.

FSH is a member of the pituitary glycoprotein hormone (GPH) family and consists of a non-covalently linked GPH α-subunit and a hormone-specific β-subunit. The GPH α- and FSH β-subunits contain a cysteine-knot motif and several N-linked glycosylation sites, both of which are important for biological activity. FSH as well as LH and CG have gonadotropic function and are thus grouped as gonadotropins. FSH binds to FSHR located on the plasma membrane of granulosa cells in the ovary and stimulates follicular growth. In the testis, FSH binds to FSHR in Sertoli cells and stimulates spermatogenesis. The synthesis and release of FSH are regulated by GnRH; gonadal proteins such as activin, inihibin, and follistatin; and gonadal steroids. FSH is implicated in hypergonadotropic ovarian dysgenesis, and its plasma levels are useful in the investigation of menstrual irregularities. Recombinant human FSH is used for treatment of infertility to stimulate follicular development.

The classic glycoprotein hormone (GPH) family consists of four members: FSH, LH, and TSH secreted from the pituitary, and CG secreted from the placenta. They are heterodimers consisting of a common α-subunit and a hormone-specific β-subunit. The α- and β-subunits contain a cysteine-knot motif and several N-linked glycosylation sites, which are important for biological activity of the hormone. FSH, LH, and CG have gonadotropic activity, and TSH has thyrotropic activity. Another heterodimeric GPH, thyrostimulin, which consists of glycoprotein α-subunit 2 (GPA2) and glycoprotein β-subunit 5 (GPB5), has been identified in both vertebrates and invertebrates. Thyrostimulin has thyrotropic activity in human. The synthesis and release of FSH and LH are regulated by hypothalamic neurohormones such as GnRH and GnIH, and the feedback effects of gonadal steroids. The synthesis and release of TSH are regulated by TRH and the feedback effects of thyroid hormones.

LH is a member of the pituitary glycoprotein hormone (GPH) family and consists of a non-covalently linked GPH α-subunit and hormone-specific β-subunit. The GPH α- and LH β-subunits contain a cysteine-knot motif and several N-linked glycosylation sites, both of which are important for biological activity. LH as well as FSH and CG has gonadotropic function and thus the grouping as gonadotropins. LH binds to LHR located on the plasma membrane of granulosa cells in the ovary and stimulates estrogen production, ovulation, and the development of the corpus luteum. In the testis, LH binds to LHR in Leydig cells and stimulates angrogen production and spermatogenesis. The synthesis and release of LH are regulated by GnRH and gonadal steroids. LH is implicated in hypogonadal dysgenesis and its urinary levels are useful in the prediction of ovulation. Recombinant human LH is used for treatment of infertility to stimulate follicular maturation.

TSH is a member of the pituitary glycoprotein hormone (GPH) family and consists of a non-covalently linked GPH α-subunit and hormone-specific β-subunit. The GPH α- and TSH β-subunits contain a cysteine-knot motif and several N -linked glycosylation sites, both of which are important for biological activity. TSH binds to the TSHR located on the membrane of thyroid follicular cells and stimulates thyroid growth and function, and the synthesis and release of thyroid hormones (T 3 and T 4 ). The synthesis and release of TSH are stimulated by TRH, while thyroid hormones inhibit the synthesis and release of TSH through binding to thyroid hormone receptors. TSH levels are measured as part of a thyroid function test in patients suspected of hyperthyroidism and hypothyroidism. Graves’ disease is an autoimmune thyroid disorder in which the body produces antibodies to TSHR that mimic the effects of TSH.

Animals regulate a variety of aspects of physiology according to environmental light conditions via nonvisual opsins such as melanopsin. In order to study photic regulation of fish physiology, expression changes of the genes for melanopsin (opn4xa and opn4xb) and effects of light on them were examined in juvenile grass puffer Takifugu alboplumbeus using quantitative real-time PCR. In the brain of juvenile fish, no significant diurnal nor circadian changes were observed in opn4x mRNA levels. On the other hand, in the eyes, the mRNA level of opn4xa showed a significant diurnal rhythm with a peak at Zeitgeber time (ZT) 4, while no apparent circadian changes were observed. The mRNA level of opn4xb in the eyes showed a diurnal change similar to that of opn4xa, while it showed a significant circadian change. Furthermore, continuous exposure to light during a subjective night significantly increased the mRNA levels of opn4xa in the eyes at ZT24, suggesting that light induces gene expression of opn4xa in the eyes and that the induction occurs only during the night-day transition period. These results suggest that Opn4xa and Opn4xb play differential roles in the eyes of juvenile grass puffer to mediate the physiological effects of environmental light information.

Tiger puffer, Takifugu rubripes, a commercially important long-distance migratory fish, return to specific spawning grounds for reproduction. To clarify reproductive neuroendocrine system of the tiger puffer, the changes in the expression levels of the genes encoding three gonadotropin-releasing hormones (GnRHs), gonadotropin-inhibitory hormone (GnIH), GnIH receptor (GnIH-R), kisspeptin and kisspeptin receptor in the brain and gonadotropin (GTH) subunits, growth hormone (GH) and prolactin (PRL) in the pituitary were examined in the tiger puffer captured in the wild at different reproductive stages, namely immature and mature fish of both sexes, and post-ovulatory females that were obtained by hormonal treatment. The amounts of three gnrh mRNAs, gnih, gnih-r, fshb and lhb were substantially increased in the mature fish compared to the immature fish, especially in the females, and these augmented expressions were drastically decreased in the post-ovulatory females. gh expression showed a slight increase in the mature males. In contrast, kiss2, kiss2r and prl did not show significant changes in the males but significantly decreased in the post-ovulatory females. The present results demonstrate the expression dynamics of the hypothalamo-pituitary-gonadal axis genes associated with the reproductive conditions and the possible involvement of the GnRH/GnIH/GTH system in the regulation of the sexual maturation and spawning in the wild tiger puffer.

Insulin-like growth factor (IGF)-I is a growth promoting hormone that exerts its actions through endocrine, paracrine and autocrine modes. Local IGF-I is essential for normal growth, whereas circulating IGF-I plays a crucial role in regulating the production and secretion of growth hormone (GH) by the pituitary gland. These actions of IGF-I are modulated by six insulin-like growth factor binding proteins (IGFBPs). In teleosts, two subtypes of each IGFBP are present due to an extra round of whole-genome duplication. IGFBP-1 is generally inhibitory to IGF-I action under catabolic conditions such as fasting and stress. In salmon, IGFBP-1a and -1b are two of three major circulating IGFBPs and assumed to affect growth through modulating IGF-I action. However, exact functions of salmon IGFBP-1 subtypes on growth regulation are not known due to the lack of purified or recombinant protein. We expressed recombinant salmon (rs) IGFBP-1a and -1b with a fusion protein (thioredoxin, Trx) and a His-tag using the pET-32a(+) vector expression system in Escherichia coli. Trx.His.rsIGFBP-1s were isolated by Ni-affinity chromatography, enzymatically cleaved by enterokinase to remove the fusion partners and further purified by reversed-phase HPLC. We next examined effects of rsIGFBP-1a and -1b in combination with human IGF-I on GH release from cultured masu salmon (Oncorhynchus masou) pituitary cells. Unexpectedly, IGF-I increased GH release and an addition of rsIGFBP-1a, but not rsIGFBP-1b, restored GH levels. The results suggest that IGFBP-1a can inhibit IGF-I action on the pituitary in masu salmon. Availability of recombinant salmon IGFBP-1s should facilitate further functional analyses and assay development.

Water temperature alone can affect the growth, metabolic rates and physiological responses of aquatic organisms. Our earlier study reported that higher temperature affects cellular and hemato-biochemical responses in rohu, Labeo rohita. In this backdrop, the present study assessed the effect of higher acclimation temperature on the regulatory mechanisms of growth and stress responses of juvenile L. rohita acclimatized in three temperature conditions (30 °C, 33 °C, and 36 °C) for a period of 30 days. The relative expression of genes for growth hormone (GH), insulin-like growth factors (IGF-1 and IGF-2) and heat shock proteins (hsp70 and hsp90) were measured by real-time quantitative PCR. The results revealed that the highest acclimation temperature (36 °C) significantly decreased the weight gain (WG) and specific growth rate (SGR), and increased the feed conversion ratio (FCR) compared to 30 °C (control), while increased WG, SGR and lowered FCR were observed in fish reared at the intermediate temperature (33 °C) compared to 30 °C. Similarly, the GH gene expression in the pituitary was significantly decreased and increased at 36 °C and 33 °C, respectively as compared to 30 °C. A significantly lower expression of IGF-1 and IGF-2, and higher expression of hsp70 and hsp90 were observed in the liver of fish at 36 °C. The results of the present study indicate that although slightly elevated temperature promotes the growth of juvenile L. rohita, the higher acclimation temperature may induce stress response and impair growth performance by suppressing GH/IGF system.

Gonadotropin-inhibitory hormone (GnIH) is a multifunctional hypophysiotropic neurohormone and has a stimulatory role in the control of reproduction in the grass puffer. To clarify the neuroendocrine mechanisms underlying the effect of changes in water temperature on reproduction in fish, we previously revealed that, in parallel to gonadal regression, both low and high temperature significantly decreased the expressions of the genes encoding kisspeptin (kiss2), kisspeptin receptor (kiss2r), gonadotropin-releasing hormone 1 (gnrh1) in the brain and gonadotropin (GTH) subunits (fshb and lhb) in the pituitary of sexually mature male grass puffer. In this study, we examined the changes in expression of gnih and GnIH receptor gene (gnihr) in the brain and pituitary along with the genes for growth hormone (gh) and prolactin (prl) in the pituitary of male grass puffer exposed to low temperature (14 °C), normal temperature (21 °C, as initial control) and high temperature (28 °C) conditions for 7 days. The levels of gnih and gnihr mRNAs were significantly decreased in both low and high temperature conditions compared to normal temperature in the brain and pituitary. Similarly, the gh mRNA levels were significantly decreased in both low and high temperature conditions. The prl mRNAs showed no significant changes at high temperature, whereas drastically decreased at low temperature possibly by dysfunctional cold stress. Taken together, the present results suggest that, in addition to the inhibitory effect of temperature changes on the Kiss2/GnRH1/GTH system, the suppression of GnIH/GH system may also be involved in the termination of reproduction by high temperature at the end of breeding season.

Radionuclide concentrations in wharf roaches inhabiting coastal areas of Honshu, Japan, were investigated in October 2011 and June 2012. Relative high concentrations of 110mAg (2.1–127 Bq kg-wet−1), 134Cs (2.6–61 Bq kg-wet−1), and 137Cs (3.5–92 Bq kg-wet−1) were detected in specimens from the eastern Honshu areas. Significantly lower 137Cs concentrations (0.7–1.6 Bq kg-wet−1) were detected in specimens from western and northern Honshu. The decay-corrected 137Cs concentration was significantly inversely correlated with the distance from the Fukushima Dai-ichi Nuclear Power Plant. Thus, wharf roach may serve as a good bioindicator for monitoring radioactive contamination of its habitats.

Gonadotropin-inhibitory hormone (GnIH) plays as a multifunctional neurohormone that controls reproduction in birds and mammals. LPXRFamide (LPXRFa) peptide, the fish ortholog of GnIH, has been shown to regulate the secretion of not only gonadotropin (GTH) but also growth hormone (GH) and prolactin (PRL), which are potentially important for gonadal function. To investigate the role of LPXRFa peptide on reproduction of the grass puffer, which spawns in semilunar cycles, we examined changes in the levels of gh and prl expression over the several months during the reproductive cycle, and the effects of goldfish LPXRFa peptide-1 (gfLPXRFa-1) on their expression were examined using primary pituitary cultures. The expression levels of both gh and prl showed significant changes during the reproductive cycle in both sexes with one peak in the spawning and pre-spawning periods for gh and prl, respectively. Particularly, gh showed substantial increase in expression in the spawning and post-spawning periods, indicative of its essentiality in the advanced stage of reproduction. gfLPXRFa-1 stimulated the expression of both gh and prl but there was a marked difference in response between them: gfLPXRFa-1 stimulated gh expression at a relatively low dose but little effect was observed on prl. Combined with the previous results of daily and circadian oscillations of lpxrfa expression, the present results suggest that LPXRFa peptide is important in the control of the cyclic reproduction by serving as a multifunctional hypophysiotropic factor that regulates the expression of gh and prl as well as GTH subunit genes.

In seasonally breeding animals, the circadian and photoperiodic regulation of neuroendocrine system is important for precisely-timed reproduction. Kisspeptin, encoded by the Kiss1 gene, acts as a principal positive regulator of the reproductive axis by stimulating gonadotrophin-releasing hormone (GnRH) neurone activity in vertebrates. However, the precise mechanisms underlying the cyclic regulation of the kisspeptin neuroendocrine system remain largely unknown. The grass puffer, Takifugu niphobles, exhibits a unique spawning rhythm: spawning occurs 1.5-2 h before high tide on the day of spring tide every 2 weeks, and the spawning rhythm is connected to circadian and lunar-/tide-related clock mechanisms. The grass puffer has only one kisspeptin gene (kiss2), which is expressed in a single neural population in the preoptic area (POA), and has one kisspeptin receptor gene (kiss2r), which is expressed in the POA and the nucleus dorsomedialis thalami. Both kiss2 and kiss2r show diurnal variations in expression levels, with a peak at Zeitgeber time (ZT) 6 (middle of day time) under the light/dark conditions. They also show circadian expression with a peak at circadian time 15 (beginning of subjective night-time) under constant darkness. The synchronous and diurnal oscillations of kiss2 and kiss2r expression suggest that the action of Kiss2 in the diencephalon is highly dependent on time. Moreover, midbrain GnRH2 gene (gnrh2) but not GnRH1 or GnRH3 genes show a unique semidiurnal oscillation with two peaks at ZT6 and ZT18 within a day. The cyclic expression of kiss2, kiss2r and gnrh2 may be important in the control of the precisely-timed diurnal and semilunar spawning rhythm of the grass puffer, possibly through the circadian clock and melatonin, which may transmit the photoperiodic information of daylight and moonlight to the reproductive neuroendocrine centre in the hypothalamus.

Water temperature is an environmental factor of primary importance that influences reproductive function in fish. To understand the molecular and physiological mechanisms underlying the regulation of reproduction by temperature, we examined changes in expression of genes encoding kisspeptin (kiss2), kisspeptin receptor (kiss2r) and three gonadotropin-releasing hormones (gnrh1, gnrh2 and gnrh3) in the brain and genes encoding gonadotropin (GTH) subunits (gpa, fshb and lhb) in the pituitary of grass puffer exposed to a low temperature (14°C), normal temperature (21°C) and high temperature (28°C) for 7days. In addition, the plasma levels of cortisol were examined after exposed to three temperature conditions. The gonadosomatic index was significantly decreased in both low and high temperature conditions. The levels of kiss2 and kiss2r mRNAs were significantly decreased at both low and high temperature conditions compared to normal temperature (control) condition. gnrh1 but not gnrh2 were significantly decreased in both temperature conditions, while gnrh3 showed a decreasing tendency in low temperature. Consequently, the levels of fshb and lhb mRNAs were significantly decreased in both low and high temperature conditions. Interestingly, the plasma levels of cortisol were significantly increased in low temperature but remain unchanged in high temperature, suggesting that the fish were under stress in the low temperature conditions but not in the high temperature conditions. Taken together, the present results indicate that anomalous temperature have an inhibitory effect on reproductive function through suppressing kiss2/kiss2r/gnrh1/fshb and lhb expression and these changes may occur in a normal physiological response as well as in a malfunctional stress response.

TRH was the first hypothalamic hypophysiotropic neurohormone to be identified. It consists of the tripeptide pGlu-His-Pro-NH2, which is fully conserved in the animal kingdom. TRH is synthesized from a precursor that contains multiple copies of the TRH progenitor sequence Gln-His-Pro-Gly. The hypophysiotropic TRH neurons are localized in the paraventricular nucleus (PVN) and send fibers to the external zone of the median eminence, where TRH is released into the hypophysial portal circulation. TRH stimulates the secretion of TSH, PRL, GH, and α-MSH from the pituitary differently depending on the species. TRH-expressing cells are widely distributed in the hypothalamus, thalamus, and lower brainstem regions, and also in peripheral tissues. TRH functions as a neurotransmitter and neuromodulator and is involved in various neurobehavioral functions, including sleep, anxiety, depression, learning, and memory. TRH action is mediated via a membrane receptor mainly coupled to Gq/11 protein.

Somatostatin (SS) was originally identified as a growth hormone (GH) inhibiting factor. Two forms of cyclic peptides, SS-14 and SS-28, are produced from a precursor encoded by the somatostatin 1 gene (SST). SS-14 is the predominant form in the brain, while SS-28 is mainly produced by intestinal enteroendocrine cells. Six SS genes have been identified in vertebrates, and two SS-related peptides, cortistatin (CST) and neuronostatin, are derived from SS family genes. SS exhibits a large variety of neuroendocrine, neuromodulatory, and gastrointestinal actions through binding to SS receptors that belong to the family of GPCRs. SS inhibits the release of GH and thyroid-stimulating hormone, and has various neuromodulatory roles in learning, cognitive functions, locomotor activity, anxiety, and depression. Moreover, SS exerts inhibitory effects on various gastrointestinal functions, including gastric acid secretion, gastric emptying, intestinal motility, and release of various gastrointestinal hormones.

The gonads are endocrine organs that secrete sex steroid hormones and protein hormones. The gonadal protein hormones include inhibin, activin, follistatin, and anti-Mullerian hormone (AMH). Inhibin, activin, and AMH are dimeric glycoproteins that belong to the TGF-β superfamily. Activin stimulates FSH secretion, while inhibin and follistatin have adverse effects in the pituitary. Likewise, activin is produced in many tissues and acts as an autocrine and/or paracrine growth factor during development and in reproductive physiology. Inhibin and follistatin inhibit activin actions through binding to activin receptors and activin, respectively. Activin signaling is mediated through type I and type II receptors. Activin binding to activin receptors leads to the activation of the Smad signaling pathway to regulate transcription of target genes. AMH causes regression of Mullerian ducts during male differentiation and, after birth, AMH is involved in the regulation of gonadal development.

GnRH is a decapeptide neurohormone that is produced in neurosecretory cells within the hypothalamus. Multiple GnRH isoforms, which are classified into four groups, are present in vertebrate species. GnRH1 secreted into the pituitary portal circulation stimulates the synthesis and release of LH and FSH. The midbrain GnRH2 neurons and the terminal nerve GnRH3 neurons send fibers throughout the brain and regulate reproductive behavior. GnRH action is mediated via a membrane receptor (GnRH-R) coupled to Gq/11 protein. The regulation of GnRH secretion is critical for reproductive success and rhythmicity, and involves many stimulatory and inhibitory factors, including gonadal steroids, neuropeptides, and neurotransmitters. A number of GnRH analogs are used not only as a fertility drug but also as an antifertility drug, and also to treat sex steroid hormone-responsive conditions such as prostate and breast cancers and endometriosis.

Full length cDNA and gene encoding ghrelin precursor and mature ghrelin peptide were identified from the stomach of Pacific bluefin tuna, Thunnus orientalis , which has unique metabolic physiology and high commercial value at fishery markets. Quantitative expression analysis was conducted for the gastric ghrelin and pepsinogen 2 genes during the early stage of somatic growth from the underyearling to yearling fish. The full length cDNA of bluefin tuna ghrelin precursor has a length of 470 bp and the deduced precursor is composed of 107 amino acids. The ghrelin gene is 1.9 kbp in length and has a 4 exon-3 intron structure. The major form of mature ghrelin in the stomach was an octanoylated 20-amino acid peptide with C-terminal amidation, while overall 12 different forms of ghrelin peptides, including short form of 18-amino acid peptide and seven kinds of acyl modifications were identified. The expression profiles of the gastric ghrelin and pepsinogen 2 genes showed no significant changes related to the early growth stages. The present results suggest that digestive physiology has already been functional in this growth stage of the juvenile bluefin tuna and ghrelin may have a role in the sustained digestive and metabolic activities.

The decapeptide gonadotropin-releasing hormone (GnRH) is the primary factor responsible for the hypothalamic control of gonadotropin (GTH) secretion. This review focuses on a family of neuropeptides, LPXRFamide (LPXRFa) peptides, which have been implicated in the regulation of GTH secretion. LPXRFa acts on the pituitary via a G protein-coupled receptor, LPXRFa-R, to enhance gonadal development and maintenance by increasing gonadotropin release and synthesis. Because LPXRFa exists and functions in several fish species, LPXRFa is considered to be a key neurohormone in fish reproduction control. The precursors to LPXRFamide peptides encoded plural LPXRFamide peptides and were highly divergent in vertebrates, particularly in lower vertebrates. Tissue distribution analyses indicated that LPXRFamide peptides were highly concentrated in the hypothalamus and other brainstem regions. In view of the localization and expression of LPXRFamide peptides in the hypothalamo-hypophysial system, LPXRFamide peptide in fish increase GTH release in vitro and in vivo. This review summarizes the advances made in our understanding of the biosynthesis, mode of action and functional significance of LPXRFa, a newly discovered key neurohormone.

When and how newborn neurons are organized to form a functional network in the developing brain remains poorly understood. An attractive model is the gonadotropin-releasing hormone (GnRH) neuron system, master regulator of the reproductive axis. Here we show that blockage of IGF signaling, a central growth-promoting signaling pathway, by the induced expression of a dominant-negative form of IGF1 receptor (IGF1R) or specific IGF1R inhibitors delayed the emergence of GnRH2 neurons in the midbrain and GnRH3 neurons in the olfactory bulb region. Blockage of IGF signaling also resulted in an abnormal appearance of GnRH3 neurons outside of the olfactory bulb region, although it did not change the locations of other olfactory neurons, GnRH2 neurons, or brain patterning. This IGF action is developmental stage-dependent because the blockade of IGF signaling in advanced embryos had no such effect. An application of phosphatidylinositol 3-kinase (PI3K) inhibitors phenocopied the IGF signaling deficient embryos, whereas the MAPK inhibitors had no effect, suggesting that this IGF action is mediated through the PI3K pathway. Real-timein vivoimaging studies revealed that the ectopic GnRH3 neurons emerged at the same time as the normal GnRH3 neurons in IGF-deficient embryos. Further experiments suggest that IGF signaling affects the spatial distribution of newborn GnRH3 neurons by influencing neural crest cell migration and/or differentiation. These results suggest that the IGF-IGF1R-PI3K pathway regulates the precise temporal and spatial organization of GnRH neurons in zebrafish and provides new insights into the regulation of GnRH neuron development.