Feeding partridges with organic or conventional grain triggers cascading effects in life-history traits

International audienceFarmland birds are declining across Europe and North America and the research of factors behind is the subject of extensive researches. Agricultural intensification is now recognized as a major factor governing the loss of biodiversity with strong evidence that pesticides induced direct bird mortality at a high dose. However, less attention has been given to the long-term effects of chronic exposure to low dose of pesticides. Here, we used an experimental procedure in which grey partridges were fed with untreated grains obtained from either organic (no pesticide) or conventional agriculture (with pesticide) for 26 weeks, thus strictly mimicking wild birds foraging on fields. We then examined a suite of life-history traits (ecophysiological and behavioural) that may ultimately, influence population dynamics. We show for the first time that ingesting low pesticide doses over a long period has long-term consequences on several major physiological pathways without inducing differential mortality. Compared to control partridges, birds exposed to chronic doses i) had less developed carotenoid-based ornaments due to lower concentrations of plasmatic carotenoids, ii) had higher activated immune system, iii) showed signs of physiological stress inducing a higher intestinal parasitic load, iv) had higher behavioural activity and body condition and v) showed lower breeding investment. Our results are consistent with a hormetic effect, in which exposure to a low dose of a chemical agent may induce a positive response, but our results also indicate that breeding adults may show impaired fitness traits bearing population consequences through reduced breeding investment or productivity. Given the current scale of use of pesticides in agrosystems, we suggest that such shifts in life-history traits may have a negative long-term impact on wild bird populations across agrosystems. We stress that long-term effects should no longer be ignored in pesticide risk assessment, where currently, only short-term effects are taken into account

The beta Nerve Growth Factor triggers ovulation in the mouse by acting upstream of GnRH neurons

International audienceThe timed induction of ovulation requires a cascade of events in which GnRH neurons play a key role. In mammals, ovulation can take place either spontaneously or be induced by mating. For induced ovulators, β-Nerve Growth Factor (βNGF) was described as an important stimulatory factor. We tested whether βNGF could induce ovulation in a spontaneous ovulator, the mouse, and investigated if GnRH is involved in its mechanism of action. Fifty mice (21 days) received 5IU of PMSG and, 48 hours later, one of the following treatments: 0.9% NaCl, 5IU of hCG, or βNGF (0.1µg, 1µg, or 10µg/mouse). All three βNGF doses induced ovulation (p <0.001) with an efficacy similar to that of hCG: ovulation rates were 80% for hCG or βNGF (0.1µg and 1µg), 100% for βNGF (10µg), and 10% for NaCl. To check if the effect of βNGF on ovulation required GnRH receptor activation, we tested responses in mice pre-treated with a GnRH receptor antagonist (Cetrorelix). Prepubertal mice (128) were allocated to five groups of treatment: βNGF (1 µg), Cetrorelix (50 ng) + βNGF (1 µg), GnRH (50 ng), Cetrorelix (50 ng) + GnRH (50 ng), or 0.9% NaCl. Both βNGF and GnRH triggered ovulations and this effect was blocked by Cetrorelix co-administration (ovulation rate: Cetrorelix + βNGF vs βNGF, p<0.05). These results suggest that βNGF requires the action of GnRH to trigger ovulation. To search for the central site of βNGF action we performed a series of double immunohistochemical labelling reactions in the hypothalamus of adult OVX+E2 female mice to localize the βNGF receptor p75 NTR. We found that p75 NTR is expressed in organum vasculosum of the lamina terminalis (OVLT), hypothalamic arcuate nucleus (ARC) and Median Eminence (ME ). In these regions, p75 NTR was found in neurons and tanycytes, but was not expressed by GnRH or Kisspeptin immunoreactive neurons. In conclusion, we have demonstrated that βNGF induces ovulation in the mouse and that its action requires the release of GnRH. By using Kiss and Gpr54 KO mice, we are currently investigating if the Kisspeptin signalling is also involved in mediating the effect of βNGF on ovulation

Beta-Nerve Growth Factor acts upstream of GnRH neurons to trigger ovulation in mice

International audienceTimed induction of ovulation requires a cascade of events in which GnRH and Kisspeptin (Kp) neurons play a key role. In mammals, ovulation is either spontaneous or induced by mating. In species with induced ovulations, β-Nerve Growth Factor (βNGF) is acknowledged as an important stimulatory factor whereas this role is attributed to Kp in species with spontaneous ovulations. However, recent studies suggested that the βNGF could also contribute in spontaneous ovulatory species. We tested whether βNGF induced ovulation in mice, a spontaneous ovulatory species, and investigated if GnRH and Kp systems are involved in its effect.Fifty mice (21 days) received 5IU of PMSG and, 48 hours later, one of the following treatments: 0.9% NaCl, 5 IU of hCG, or βNGF (0.1µg, 1 µg, or 10 µg/mouse). All three βNGF doses induced ovulation (p <0.001) with an efficacy similar to that of hCG. Ovulation rates were 80% for hCG or βNGF (0.1 µg and 1µg), 100% for βNGF (10 µg), and 10% for NaCl. To determine if βNGF action on ovulation required GnRH receptor activation, we tested responses in mice pre-treated with Cetrorelix, a GnRH receptor antagonist. Prepubertal mice (128) were allocated to five treatment groups: βNGF (1 µg), Cetrorelix (50 ng) + βNGF (1 µg), GnRH (50 ng), Cetrorelix (50 ng) + GnRH (50 ng), or 0.9% NaCl. Both βNGF and GnRH triggered ovulations and this effect was blocked by Cetrorelix (ovulation rate: Cetrorelix + βNGF vs βNGF, p<0.05). These results suggest that the GnRH is implicated in the βNGF-induced ovulation pass through.We then tested responses in Kiss1 and Kiss1r null mice at two ages: adult (KO Kiss1: 23; KO Kiss1r: 16) and prepubertal (KO Kiss:5 ; KO Kiss1r: 9) mice. Each type of mice was allocated into two groups of treatment: βNGF (0.2 µg) or hCG (5UI). In all groups treated with βNGF, the rate of ovulation and the number of corpora lutea per ovary was significantly? lower than in groups treated with hCG. These results suggest that βNGF requires the action of Kp to trigger ovulation.Using double immunohistochemical procedures on adult OVX+E2 female mice, we found both βNGF receptors, p75 NTR and TrkA, localized in the organum vasculosum of the lamina terminalis (OVLT). p75 NTR was also expressed in the arcuate nucleus (ARC) and Median Eminence (ME ). p75 NTR was found in neurons and tanycytes, but was not expressed by GnRH or Kp immunoreactive neurons suggesting the involvement of interneurons. In conclusion, βNGF induces ovulation in the mouse and this probably involves the release of GnRH and Kp. However, because none of these neurons expresses p75 NTR, the transduction of βNGF information is relayed either by interneurons or by glial cells. Among them, tanycytes appear as interesting candidates

Décryptage des possibles mécanismes centraux d’action du β Nerve Growth Factor (βNGF) dans l’induction de l’ovulation chez la souris

International audienceLe déclanchement de l’ovulation requiert une cascade complexe d’évènements où les neurones à GnRH jouent un rôle clé. Chez les mammifères, l’ovulation peut avoir lieu de façon spontanée ou être induite par un accouplement. Le β Nerve Growth Factor (βNGF) a été récemment décrit, comme étant le facteur déclenchant l’ovulation chez les espèces à ovulation induite. En utilisant le modèle murin, nous cherchons à vérifier si le βNGF peut déclencher l’ovulation chez les espèces à ovulation spontanée et à comprendre les mécanismes sous-jacents. Cinquante souris femelles prépubères ont reçu 5UI de PMSG et 48 heures plus tard l’un de ces cinq traitements : NaCl, du hCG ou du βNGF (0.1μg, 1μg, 10μg/souris). Les 3 doses de βNGF induisent l’ovulation de façon comparable à l’hCG (taux d’ovulation: hCG=80%, βNGF 0.1μg=80%, 1μg=80%, 10μg=100%, NaCl=10%). Pour vérifier si l’effet du βNGF implique l’activation des neurones à GnRH, nous avons réalisé un protocole d’induction de l’ovulation incluant des souris prétraitées ou non avec un antagoniste un récepteur de la GnRH : le Cétrorelix. Cent vingt-huit souris femelles prépubères ont été réparties dans cinq lots recevant un des cinq traitements suivants : βNGF (1μg), Cétrorelix (50ng) + βNGF (1μg), GnRH (50ng), Cétrorelix (50ng) + GnRH (50ng), NaCl. Le βNGF et le GnRH induisent l’ovulation et leurs effets sont supprimés par l’administration de Cétrorelix (Cétrorelix+βNGF vs βNGF : p<0.05). Ces résultats suggèrent que, chez la souris, le βNGF nécessite l’intervention des neurones à GnRH pour déclencher l’ovulation. Pour rechercher les sites d’action du βNGF au niveau de l’hypothalamus, nous avons utilisé une approche immunohistochimique pour localiser le récepteur p75NTR du βNGF sur des cerveaux de souris ovariectomisées et sous implant d’oestradiol. p75NTR a été retrouvé dans l’organe vasculaire de la lame terminale (OVLT), le noyau arqué (ARC), l’éminence médiane (ME). Parmi les cellules exprimant p75NTR, on trouve des neurones et des tanycytes.Parmi les neurones exprimant p75NTR, aucun n’exprime la GnRH ou la Kisspeptine. En conclusion, l’action de la GnRH est requise pour permettre au βNGF d’induire l’ovulation chez la souris, mais le βNGF n’agit pas directement sur les neurones à GnRH ni sur les neurones à kisspeptine. La poursuite de ce travail en utilisant des souris KO per la kisspeptine et son récepteur vise à établir si les systèmes βNGF agit en amont du système Kisspeptine

L’ovulation chez les mammifères

The triggering of ovulation in mammals can be done according to two modes: spontaneous and provoked. Spontaneous ovulation occurs during the estrous cycle as a result of internal endocrine factors. Induced ovulation is triggered by mating. In both cases, it is the increase in GnRH secretion that leads to an increase in LH that causes ovulation. The factors involved in the stimulation of GnRH secretion are different according to the two modalities, mainly kisspeptin for spontaneous ovulators and beta-NGF for provoked ovulators. Current protocols used to induce ovulation rely on a direct action on the ovary, through the use of heterologous gonadotropins, or by an action on the pituitary gland with GnRH agonists. These protocols present different disadvantages: loss of activity with time, supra-physiological stimulation, ethic questioning about the production of some of these products. New paradigms for triggering ovulation by targeting the hypothalamus, respectful of animal welfare and the environment are presented in this review. This review aims to introduce the reader to the cellular and molecular mechanisms involved in the regulation and triggering of ovulation as well as two new approaches that are being developed and that are respectful of the animal and of its environment.La maîtrise du moment de l’ovulation chez les mammifères d’élevage permet d’améliorer la fertilité, la gestion des mises bas, mais les méthodes actuelles sont basées sur l’utilisation de fortes doses d’hormones. Cette revue présente de nouveaux paradigmes pour maitriser le moment de l’ovulation, respectueux de la physiologie de l’animal, sans supplémentation hormonale basés sur la connaissance des mécanismes physiologiques

A customized long acting formulation of the kisspeptin analog C6 triggers ovulation in anestrus ewe

International audienceThe modulation of the kisspeptin system holds promise as a treatment for human reproductive disorders and for managing livestock breeding. The design of analogs has overcome some unfavorable properties of the endogenous ligands. However, for applications requiring a prolongation of drug activity, such as ovulation induction in the ewe during the non-breeding season, additional improvement is required. To this aim, we designed and tested three formulations containing the kisspeptin analog C6. Two were based on polymeric nanoparticles (NP1 and NP2) and the third was based on hydrogels composed of a mixture of cyclodextrin polymers and dextran grafted with alkyl side chains (MD/pCD). Only the MD/pCD formulation prolonged C6 activity, as shown by monitoring luteinizing hormone (LH) plasma concentration (elevation duration 23.4 +/- 6.1, 13.7 +/- 4.7 and 12.0 +/- 2.4 h for MD/pCD, NP1 and NP2, respectively). When compared with the free C6 (15 nmol/ewe), the formulated (MD/pCD) doses of 10, 15 and 30 nmol/ewe, but not the 90 nmol/ewe dose, provided a more gradual release of C6 as shown by an attenuated LH release during the first 6 h post-treatment. When tested during the non-breeding season without progestogen priming, only, the formulated 30 nmol/ewe dose triggered ovulation (50% of ewes). Hence, we showed that a formulation with an adapted action time would improve the efficacy of C6 with respect to inducing ovulation during the non-breeding season. This result suggests that formulations containing a kisspeptin analog might find applications in the management of livestock reproduction but also point to the possibility of their use for the treatment of some human reproductive pathologies

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

International audienceThe 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 and compared it with human recombinant β-NGF (hβ-NGF). We showed that the β-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 β-NGF induced ovulation through an increase in GnRH secretion provoked by a glial dependent P75 mediated mechanism

Treating mares with the long-acting kisspeptin analog C6 increases circulating gonadotropins but does not trigger ovulation

International audienceThe role of the neuropeptide kisspeptin (Kp) in mammalian reproduction is well established. Nevertheless, species-specific differences exist. In the horse, administration of the shortest endogenous Kp isoform, Kp10, is unable to trigger ovulation even though it increases plasma gonadotropins concentrations. To check if this issue would be dependent on Kp10 short half-life, we tested two degradation-resistant Kp analogs. The first analog was based on the equine Kp10 sequence (eC6), the second on the ovine Kp10 sequence (oC6). During the non-breeding season, a dose of 150 nmol/mare of either molecule had no significant effect on LH concentration, while oC6 provided a better stimulation of FSH than eC6 (p = 0.01). Furthermore, oC6 was more effective when injected intravenously than intramuscularly. Due to its best pharmacodynamics profile, oC6 (150 nmol/mare) was probed for ovulation induction during the breeding season. The molecule was injected during the preovulatory phase when the follicle diameter ranged from 34 to 37 mm and a uterine oedema was observed. oC6 consistently increased the total amount of gonadotropins released (FSH, p = 0.01 and LH, p = 0.02). However, as shown by transrectal ultrasonography and plasma progesterone levels, oC6 did not anticipate ovulation compared to the control group. Our results provide further evidence of the peculiar reproductive endocrinology of the mare but leave open questions regarding the exact role of Kp in the control of ovulation and breeding in the mare, which we attempt to identify and discuss

Highly-Sensitive In Vitro Bioassays for FSH, TSH, PTH, Kp, and OT in Addition to LH in Mouse Leydig Tumor Cell

We demonstrate here that highly sensitive in vitro bioassays for FSH, TSH, and PTH can be set up in mouse Leydig Tumor Cells (mLTC), in addition to the normal LH/CG bioassay, after they were transfected with expression vectors encoding the corresponding Gs Protein-Coupled Receptors (GsPCR), such as FSHR, TSHR, or PTHR. Although the β2 adrenergic receptor is also a GsPCR, its expression in mLTC led to a significant but very low cAMP response compared to those observed with FSH, TSH, or PTH. Similarly, after transfection of the GiPCR MT1 melatonin receptor, we did not observe any inhibitory effect by melatonin of the LH or hCG stimulation. Interestingly, after transfection of mLTC with the human kisspeptin receptor (hKpR), which is a GqPCR, we observed a dose-dependent synergy of 10−12–10−7 M kisspeptin variants with a fixed concentration of 0.3 nM LH or hCG. Without any exogenous receptor transfection, a 2 h preincubation with OT or AVP led to a dose-dependent cAMP response to a fixed dose of LH or hCG. Therefore, highly sensitive in vitro bioassays for various hormones and other GPCR ligands can be set up in mLTC to measure circulating concentrations in only 3–10 µL of blood or other body fluids. Nevertheless, the development of an LHRKO mLTC cell line will be mandatory to obtain strict specificity for these bioassays to eliminate potential cross-reaction with LH or CG