Development of Gonadotropin-Releasing Hormone-Secreting Neurons from Human Pluripotent Stem Cells.
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ABSTRACT:
SUBMITTER: Lund C
PROVIDER: S-EPMC4982984 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Human pluripotent stem cells (hPSCs) provide an unlimited cell source for regenerative medicine. Hormone-producing cells are particularly suitable for cell therapy, and hypopituitarism, a defect in pituitary gland function, represents a promising therapeutic target. Previous studies have derived pituitary lineages from mouse and human ESCs using 3D organoid cultures that mimic the complex events underlying pituitary gland development in vivo. Instead of relying on unknown cellular signals, we present a simple and efficient strategy to derive human pituitary lineages from hPSCs using monolayer culture conditions suitable for cell manufacturing. We demonstrate that purified placode cells can be directed into pituitary fates using defined signals. hPSC-derived pituitary cells show basal and stimulus-induced hormone release in vitro and engraftment and hormone release in vivo after transplantation into a murine model of hypopituitarism. This work lays the foundation for future cell therapy applications in patients with hypopituitarism.
| S-EPMC4912387 | biostudies-literature
Project description:Hypophysiotropic projections of gonadotropin-releasing hormone (GnRH)-synthesizing neurons form the final common output way of the hypothalamus in the neuroendocrine control of reproduction. Several peptidergic neuronal systems of the medial hypothalamus innervate human GnRH cells and mediate crucially important hormonal and metabolic signals to the reproductive axis, whereas much less is known about the contribution of the lateral hypothalamic area to the afferent control of human GnRH neurons. Orexin (ORX)- and melanin-concentrating hormone (MCH)-synthesizing neurons of this region have been implicated in diverse behavioral and autonomic processes, including sleep and wakefulness, feeding and other functions. In the present immunohistochemical study, we addressed the anatomical connectivity of these neurons to human GnRH cells in post-mortem hypothalamic samples obtained from autopsies. We found that 38.9 ± 10.3% and 17.7 ± 3.3% of GnRH-immunoreactive (IR) perikarya in the infundibular nucleus of human male subjects received ORX-IR and MCH-IR contacts, respectively. On average, each 1 mm segment of GnRH dendrites received 7.3 ± 1.1 ORX-IR and 3.7 ± 0.5 MCH-IR axo-dendritic appositions. Overall, the axo-dendritic contacts dominated over the axo-somatic contacts and represented 80.5 ± 6.4% of ORX-IR and 76.7 ± 4.6% of MCH-IR inputs to GnRH cells. Based on functional evidence from studies of laboratory animals, the direct axo-somatic and axo-dendritic input from ORX and MCH neurons to the human GnRH neuronal system may convey critical metabolic and other homeostatic signals to the reproducive axis. In this study, we also report the generation and characterization of new antibodies for immunohistochemical detection of GnRH neurons in histological sections.
| S-EPMC4559643 | biostudies-literature
Project description:Most central neurons in the mammalian brain possess an appendage called a primary cilium that projects from the soma into the extracellular space. The importance of these organelles is highlighted by the fact that primary cilia dysfunction is associated with numerous neuropathologies, including hyperphagia-induced obesity, hypogonadism, and learning and memory deficits. Neuronal cilia are enriched for signaling molecules, including certain G protein-coupled receptors (GPCRs), suggesting that neuronal cilia sense and respond to neuromodulators in the extracellular space. However, the impact of cilia on signaling to central neurons has never been demonstrated. Here, we show that the kisspeptin receptor (Kiss1r), a GPCR that is activated by kisspeptin to regulate the onset of puberty and adult reproductive function, is enriched in cilia projecting from mouse gonadotropin-releasing hormone (GnRH) neurons. Interestingly, GnRH neurons in adult animals are multiciliated and the percentage of GnRH neurons possessing multiple Kiss1r-positive cilia increases during postnatal development in a progression that correlates with sexual maturation. Remarkably, disruption of cilia selectively on GnRH neurons leads to a significant reduction in kisspeptin-mediated GnRH neuronal activity. To our knowledge, this result is the first demonstration of cilia disruption affecting central neuronal activity and highlights the importance of cilia for proper GPCR signaling.
| S-EPMC4104922 | biostudies-literature
Project description:Given the critical central role of gonadotropin-releasing hormone (GnRH) neurons in fertility, it is not surprising that the GnRH neural network is implicated in the pathology of polycystic ovary syndrome (PCOS), the most common cause of anovulatory infertility. Although many symptoms of PCOS relate most proximately to ovarian dysfunction, the central reproductive neuroendocrine system ultimately drives ovarian function through its regulation of anterior pituitary gonadotropin release. The typical cyclical changes in frequency of GnRH release are often absent in women with PCOS, resulting in a persistent high-frequency drive promoting gonadotropin changes (i.e., relatively high luteinizing hormone and relatively low follicle-stimulating hormone concentrations) that contribute to ovarian hyperandrogenemia and ovulatory dysfunction. However, the specific mechanisms underpinning GnRH neuron dysfunction in PCOS remain unclear. Here, we summarize several preclinical and clinical studies that explore the causes of aberrant GnRH secretion in PCOS and the role of disordered GnRH secretion in PCOS pathophysiology.
| S-EPMC9232905 | biostudies-literature
Project description:Gonadotropin-releasing hormone (GnRH) is a hypothalamic decapeptide essential for fertility in vertebrates. Human male patients lacking GnRH and treated with hormone therapy can remain fertile after cessation of treatment suggesting that new GnRH neurons can be generated during adult life. We used zebrafish to investigate the neurogenic potential of the adult hypothalamus. Previously we have characterized the development of GnRH cells in the zebrafish linking genetic pathways to the differentiation of neuromodulatory and endocrine GnRH cells in specific regions of the brain. Here, we developed a new method to obtain neural progenitors from the adult hypothalamus in vitro. Using this system, we show that neurospheres derived from the adult hypothalamus can be maintained in culture and subsequently differentiate glia and neurons. Importantly, the adult derived progenitors differentiate into neurons containing GnRH and the number of cells is increased through exposure to either testosterone or GnRH, hormones used in therapeutic treatment in humans. Finally, we show in vivo that a neurogenic niche in the hypothalamus contains GnRH positive neurons. Thus, we demonstrated for the first time that neurospheres can be derived from the hypothalamus of the adult zebrafish and that these neural progenitors are capable of producing GnRH containing neurons.
| S-EPMC4582115 | biostudies-literature
Project description:Acquisition of a mature pattern of gonadotropin-releasing hormone (GnRH) secretion from the CNS is a hallmark of the pubertal process. Little is known about GnRH release during sexual maturation, but it is assumed to be minimal before later stages of puberty. We studied spontaneous GnRH secretion in brain slices from male mice during perinatal and postnatal development using fast-scan cyclic voltammetry (FSCV) to detect directly the oxidation of secreted GnRH. There was good correspondence between the frequency of GnRH release detected by FSCV in the median eminence of slices from adults with previous reports of in vivo luteinizing hormone (LH) pulse frequency. The frequency of GnRH release in the late embryonic stage was surprisingly high, reaching a maximum in newborns and remaining elevated in 1-week-old animals despite low LH levels. Early high-frequency GnRH release was similar in wild-type and kisspeptin knock-out mice indicating that this release is independent of kisspeptin-mediated excitation. In vivo treatment with testosterone or in vitro treatment with gonadotropin-inhibitory hormone (GnIH) reduced GnRH release frequency in slices from 1-week-old mice. RF9, a putative GnIH antagonist, restored GnRH release in slices from testosterone-treated mice, suggesting that testosterone inhibition may be GnIH-dependent. At 2-3 weeks, GnRH release is suppressed before attaining adult patterns. Reduction in early life spontaneous GnRH release frequency coincides with the onset of the ability of exogenous GnRH to induce pituitary LH secretion. These findings suggest that lack of pituitary secretory response, not lack of GnRH release, initially blocks downstream activation of the reproductive system.
| S-EPMC4220034 | biostudies-literature
Project description:Fibroblast growth factor 8 (FGF8) is a potent morphogen that regulates the ontogenesis of gonadotropin-releasing hormone (GnRH) neurons, which control the hypothalamus-pituitary-gonadal (HPG) axis, and therefore reproductive success. Indeed, FGF8 and FGFR1 deficiency severely compromises vertebrate reproduction in mice and humans and is associated with Kallmann Syndrome (KS), a congenital disease characterized by hypogonadotropic hypogonadism associated with anosmia. Our laboratory demonstrated that FGF8 signaling through FGFR1, both of which are KS-related genes, is necessary for proper GnRH neuron development in mice and humans. Here, we investigated the possibility that non-genetic factors, such as the epigenome, may contribute to KS onset. For this purpose, we developed an embryonic explant model, utilizing the mouse olfactory placode (OP), the birthplace of GnRH neurons. We show that TET1, which converts 5-methylcytosine residues (5mC) to 5-hydroxymethylated cytosines (5hmC), controls transcription of Fgf8 during GnRH neuron ontogenesis. Through MeDIP and ChIP RT-qPCR we found that TET1 bound to specific CpG islands on the Fgf8 promoter. We found that the temporal expression of Fgf8 correlates with not only TET1 binding, but also with 5hmC enrichment. siRNA knockdown of Tet1 reduced Fgf8 and Fgfr1 mRNA expression. During this time period, Fgf8 also switched histone status, most likely via recruitment of EZH2, a major component of the polycomb repressor complex-2 (PRC2) at E13.5. Together, these studies underscore the significance of epigenetics and chromatin modifications to temporally regulated genes involved in KS.
| S-EPMC6667164 | biostudies-literature
Project description:: Inflammation has a well-known suppressive effect on fertility. The function of gonadotropin-releasing hormone (GnRH) neurons, the central regulator of fertility is substantially altered during inflammation in females. In our review we discuss the latest results on how the function of GnRH neurons is modified by inflammation in females. We first address the various effects of inflammation on GnRH neurons and their functional consequences. Second, we survey the possible mechanisms underlying the inflammation-induced actions on GnRH neurons. The role of several factors will be discerned in transmitting inflammatory signals to the GnRH neurons: cytokines, kisspeptin, RFamide-related peptides, estradiol and the anti-inflammatory cholinergic pathway. Since aging and obesity are both characterized by reproductive decline our review also focuses on the mechanisms and pathophysiological consequences of the impact of inflammation on GnRH neurons in aging and obesity.
| S-EPMC7014424 | biostudies-literature
Project description:Kisspeptin and its cognate receptor, GPR54, are critical for reproductive development and for the regulation of gonadotropin-releasing hormone (GnRH) secretion. Although kisspeptin has been found to depolarize GnRH neurons, the underlying ionic mechanism has not been elucidated. Presently, we found that kisspeptin depolarized GnRH neurons in a concentration-dependent manner with a maximum depolarization of 22.6 +/- 0.6 mV and EC(50) of 2.8 +/- 0.2 nM. Under voltage-clamp conditions, kisspeptin induced an inward current of 18.2 +/- 1.6 pA (V(hold) = -60 mV) that reversed near -115 mV in GnRH neurons. The more negative reversal potential than E(K)(+) (-90 mV) was caused by the concurrent inhibition of barium-sensitive, inwardly rectifying (Kir) potassium channels and activation of sodium-dependent, nonselective cationic channels (NSCCs). Indeed, reducing extracellular Na(+) (to 5 mM) essentially eliminated the kisspeptin-induced inward current. The current-voltage relationships of the kisspeptin-activated NSCC currents exhibited double rectification with negative slope conductance below -40 mV in the majority of the cells. Pharmacological examination showed that the kisspeptin-induced inward currents were blocked by TRPC (canonical transient receptor potential) channel blockers 2-APB (2-aminoethyl diphenylborinate), flufenamic acid, SKF96365 (1-[beta-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenethyl]-1H-imidazole hydrochloride), and Cd(2+), but not by lanthanum (100 microM). Furthermore, single-cell reverse transcription-PCR analysis revealed that TRPC1, TRPC3, TRPC4, TRPC5, TRPC6, and TRPC7 subunits were expressed in GnRH neurons. Therefore, it appears that kisspeptin depolarizes GnRH neurons through activating TRPC-like channels and, to a lesser extent, inhibition of Kir channels. These actions of kisspeptin contribute to the pronounced excitation of GnRH neurons that is critical for mammalian reproduction.
| S-EPMC6670958 | biostudies-literature
Project description:T-type calcium channels are responsible for generating low-threshold spikes that facilitate burst firing and neurotransmitter release in neurons. Gonadotropin-releasing hormone (GnRH) neurons exhibit burst firing, but the underlying conductances are not known. Previously, we found that 17beta-estradiol (E2) increases T-type channel expression and excitability of hypothalamic arcuate nucleus neurons. Therefore, we used ovariectomized oil- or E2-treated EGFP (enhanced green fluorescent protein)-GnRH mice to explore the expression and E2 regulation of T-type channels in GnRH neurons. Based on single-cell reverse transcriptase-PCR and real-time PCR quantification of the T-type channel alpha(1) subunits, we found that all three subunits were expressed in GnRH neurons, with expression levels as follows: Cav3.3 > or = Cav3.2 > Cav3.1. The mRNA expression of the three subunits was increased with surge-inducing levels of E2 during the morning. During the afternoon, Cav3.3 mRNA expression remained elevated, whereas Cav3.1 and Cav3.2 were decreased. The membrane estrogen receptor agonist STX increased the expression of Cav3.3 but not Cav3.2 in GnRH neurons. Whole-cell patch recordings in GnRH neurons revealed that E2 treatment significantly augmented T-type current density at both time points and increased the rebound excitation during the afternoon. Although E2 regulated the mRNA expression of all three subunits in GnRH neurons, the increased expression combined with the slower inactivation kinetics of the T-type current indicates that Cav3.3 may be the most important for bursting activity associated with the GnRH/LH (luteinizing hormone) surge. The E2-induced increase in mRNA expression, which depends in part on membrane-initiated signaling, leads to increased channel function and neuronal excitability and could be a mechanism by which E2 facilitates burst firing and cyclic GnRH neurosecretion.
| S-EPMC2774467 | biostudies-literature