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Beta-nerve growth factor stimulates spontaneous electrical activity of in vitro embryonic mouse GnRH neurons through a P75 mediated-mechanism.

ABSTRACT: The control of ovulation helps guarantee the success of reproduction and as such, contributes to the fitness of a species. In mammals, two types of ovulation are observed: induced and spontaneous ovulation. Recent work on camelids, that are induced ovulators, highlighted the role of a factor present in seminal plasma, beta Nerve Growth Factor (?-NGF), as the factor that triggers ovulation in a GnRH dependent manner. In the present work, we characterized alpaca ?-NGF (a?-NGF) and its 3D structure and compared it with human recombinant ?-NGF (h?-NGF). We showed that the ?-NGF enriched fraction of alpaca semen and the human recombinant protein, both stimulated spontaneous electrical activity of primary GnRH neurons derived from mouse embryonic olfactory placodes. This effect was dose-dependent and mediated by p75 receptor signaling. P75 receptors were found expressed in vitro by olfactory ensheathing cells (OEC) in close association with GnRH neurons and in vivo by tanycytes in close vicinity to GnRH fibers in adult mouse. Altogether, these results suggested that ?-NGF induced ovulation through an increase in GnRH secretion provoked by a glial dependent P75 mediated mechanism.

SUBMITTER: Pinet-Charvet C

PROVIDER: S-EPMC7326925 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

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Beta-nerve growth factor stimulates spontaneous electrical activity of in vitro embryonic mouse GnRH neurons through a P75 mediated-mechanism.

Pinet-Charvet Caroline C Fleurot Renaud R Derouin-Tochon Flavie F de Graaf Simon S Druart Xavier X Tsikis Guillaume G Taragnat Catherine C Teixeira-Gomes Ana-Paula AP Labas Valérie V Moreau Thierry T Cayla Xavier X Duittoz Anne H AH

Scientific reports 20200630 1

The control of ovulation helps guarantee the success of reproduction and as such, contributes to the fitness of a species. In mammals, two types of ovulation are observed: induced and spontaneous ovulation. Recent work on camelids, that are induced ovulators, highlighted the role of a factor present in seminal plasma, beta Nerve Growth Factor (β-NGF), as the factor that triggers ovulation in a GnRH dependent manner. In the present work, we characterized alpaca β-NGF (aβ-NGF) and its 3D structure ...[more]

PMID: 32606357

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Project description:Hypothalamic gonadotropin-releasing hormone (GnRH) neurons are required for fertility in all mammalian species studied to date. In rodents, GnRH neuron cell bodies reside in the rostral hypothalamus, and most extend a single long neuronal process in the caudal direction to terminate at the median eminence (ME), the site of hormone secretion. The molecular cues that GnRH neurites use to grow and navigate to the ME during development, however, remain poorly described. Reverse transcription-PCR (RT-PCR) identified mRNAs encoding Netrin-1, and its receptor, DCC, in the fetal preoptic area (POA) and mediobasal hypothalamus (MBH), respectively, from gestational day 12.5 (GD12.5), a time when the first GnRH neurites extend toward the MBH. Moreover, a subpopulation of GnRH neurons from GD14.5 through GD18.5 express the Netrin-1 receptor, DCC, suggesting a role for Netrin-1/DCC signaling in GnRH neurite growth and/or guidance. In support of this notion, when GD15.5 POA explants, containing GnRH neurons actively extending neurites, were grown in three-dimensional collagen gels and challenged with exogenous Netrin-1 (100 ng/ml or 400 ng/ml) GnRH neurite growth was stimulated. In addition, Netrin-1 provided from a fixed source was able to stimulate outgrowth, although it did not appear to chemoattract GnRH neurites. Finally, the effects of Netrin-1 on the outgrowth of GnRH neurites could be inhibited by blocking either L-type voltage-gated calcium channels (VGCCs) with nifedipine (10 µM), or ryanodine receptors with ryanodine (10 µM). This is consistent with the role of Ca2+ from extra- and intracellular sources in Netrin-1/DCC-dependent growth cone motility in other neurons. These results indicate that Netrin-1 directly stimulates the growth of a subpopulation of GnRH neurites that express DCC, provide further understanding of the mechanisms by which GnRH nerve terminals arrive at their site of hormone secretion, and identify an additional neuronal population whose neurites utilize Netrin-1/DCC signaling for their development.

Project description:RationaleAirway hyperresponsiveness (AHR) is classically found in asthma, and persistent AHR is associated with poor asthma control. Although airway smooth muscle (ASM) cells play a critical pathophysiologic role in AHR, the paracrine contributions of surrounding cells such as fibroblasts to the contractile phenotype of ASM cells have not been examined fully. This study addresses the hypothesis that nicotine promotes a contractile ASM cell phenotype by stimulating fibroblasts to increase nerve growth factor (NGF) secretion into the environment.MethodsPrimary lung fibroblasts isolated from wild type and α7 nicotinic acetylcholine receptor (α7 nAChR) deficient mice were treated with nicotine (50 µg/ml) in vitro for 72 hours. NGF levels were measured in culture media and in bronchoalveolar lavage (BAL) fluid from asthmatic, smoking and non-smoking subjects by ELISA. The role of the NFκB pathway in nicotine-induced NGF expression was investigated by measuring NFκB nuclear translocation, transcriptional activity, chromatin immunoprecipitation assays, and si-p65 NFκB knockdown. The ability of nicotine to stimulate a fibroblast-mediated, contractile ASM cell phenotype was confirmed by examining expression of contractile proteins in ASM cells cultured with fibroblast-conditioned media or BAL fluid.ResultsNGF levels were elevated in the bronchoalveolar lavage fluid of nicotine-exposed mice, current smokers, and asthmatic children. Nicotine increased NGF secretion in lung fibroblasts in vitro in a dose-dependent manner and stimulated NFκB nuclear translocation, p65 binding to the NGF promoter, and NFκB transcriptional activity. These responses were attenuated in α7 nAChR deficient fibroblasts and in wild type fibroblasts following NFκB inhibition. Nicotine-treated, fibroblast-conditioned media increased expression of contractile proteins in ASM cells.ConclusionNicotine stimulates NGF release by lung fibroblasts through α7 nAChR and NFκB dependent pathways. These novel findings suggest that the nicotine-α7 nAChR-NFκB- NGF axis may provide novel therapeutic targets to attenuate tobacco smoke-induced AHR.

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Beta-nerve growth factor stimulates spontaneous electrical activity of in vitro embryonic mouse GnRH neurons through a P75 mediated-mechanism.

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Beta-nerve growth factor stimulates spontaneous electrical activity of in vitro embryonic mouse GnRH neurons through a P75 mediated-mechanism.

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