Intravaginal sponges impregnated with the progesterone (P4) analogue fluorogestone acetate (FGA) induce synchronous oestrous behaviour and normal ovulatory cycle in goats. To explore alternatives using natural P4 from plants, we developed a method of ethanolic extraction and a specific enzyme immunoassay (EIA) to measure P4 in the different parts of the walnut tree Juglans regia. We found a very high concentration of P4, specifically in the leaves of the three most common French varieties (similar to 100 mg/kg of DM) but not in flowers, fruits, septa, husk, oil or cake. High concentrations of P4-and to a lesser extent its reduction metabolites and phytosterols-were also measured by Gas Chromatography-Mass Spectrometry/Mass Spectrometry in leaf extracts. P4 concentrations were five times higher in October than in June. P4 was detected in 182 varieties of Juglans regia ranging from 35 to 287 mg of P4 per kg of leaf DM. We collected large quantities of leaves over 6 years, which were used to manufacture feed pellets containing 32% of dry leaf for distribution to female goats. To determine their dietary acceptance and their efficacy in terms of P4 blood plasma concentration, three trials in ovariectomised goats and four trials in ovary-intact goats were performed (N = 83). The distribution of 600 g of pellets per day per ovary-intact goat over 3 days, 6 and 4 days before the introduction of males in April allowed us to achieve our objective of a significant increase of P4 plasma concentration to similar to 1.5 ng/mL measured by EIA from 24 to 72 h after the first distribution in the walnut pellet group (n = 13). The two control groups of goats (FGA, n = 12 and control, n = 10) showed no increase in plasma P4. However, despite this high P4 plasma concentration, goats of the walnut group had the same percentages of goats in oestrus at the first ovulation and of goats experiencing short ovulatory cycles after introduction of males (54 and 77%, respectively) as the group of control goats (80 and 90%), whereas the FGA goats showed very different percentages (100 and 0%, P < 0.01). It was concluded that whereas walnut leaves contain a high concentration of P4-and its reduction metabolites and phytosterols-the pellet feeding mode does not allow for restoration of oestrus behaviour and duration of the induced cycle consistently achieved with FGA-impregnated intravaginal sponges.

At temperate and polar latitudes, animals and humans experience seasonal changes that impact physiology and behavior. In these habitats, the prevalence and severity of certain psychiatric disorders fluctuate seasonally. Such patterns imply that an adaptive system fine-tunes brain physiology in response to annual environmental changes, and alterations to this system may adversely affect mental health. To date, the core neuronal circuitry of the seasonal control of brain functioning is still largely unknown. To address this question, we identified brain regions sensitive to seasonal changes, using neuroimaging in the domestic sheep (Ovis aries), an animal model commonly used to study seasonality. Here, we developed MRI neuroinformatics resources (templates and atlas) dedicated to the analysis of the sheep brain and revealed that seasons broadly modify grey matter organization and volume of both cortical and subcortical regions involved in the control of homeostasis, sensory processing, learning, memory, behavior control, and social cognition. Many of these regions were not previously known to be affected by seasonal variations, highlighting that the seasonal control of brain function involves plasticity mechanisms across multiple brain sites.

Seasonality of reproductive activity in rams and bucks is the major constraint in temperate and subtropical zones. Rapid alternation between 1 month of short days and 1 month of long days (LD) over three years in lightproof buildings eliminates this seasonality. We examined if this would also work in open barns, using only supplementary light. Over two years, one group of bucks (n = 7) was subjected to alternate 1 month of LD and 1 month of permanent light (LD-LL) and another group (n = 7) to alternate 1 month of LD and 1 month of natural light (LD-NL). A simultaneous control group, used for both experiments (CG1, n = 6; CG2, n = 6), remained under natural photoperiod. BW, testis weight (TW), plasma testosterone (T) and cortisol (C) were evaluated in all bucks. CG1 and CG2 bucks showed identical dramatic seasonal variations in BW (stable or decreasing in summer), TW (from 85 +/- 12 g in February to 127 +/- 7 g in July) and T (from 2.7 +/- 1.2 ng/mL in January-April to 24.3 +/- 3.2 ng/mL in June-October). By contrast, BW of LD-LL and LD-NL bucks increased regularly during the experiment. From 5 and 9 months after the experiment onset, LD-LL and LD-NL bucks, respectively, maintained constant TW of 115 +/- 5 g until the experiment end. After the first 3 months <5 ng/mL, T of LD-LL bucks remained constant (5-10 ng/mL) until the experiment end. By contrast, T of LD-NL bucks showed four periods of low (<5 ng/mL) and two periods of high concentrations (18.1 +/- 2.6 and 11.9 +/- 3.4 ng/mL). Plasma C remained low (5-8 ng/mL) and did not change with group or light treatment. These results show for the first time in any seasonal photoperiodic species that it is possible to maintain the sexual activity of males all year round in open buildings using alternating periods of LD and LL. By contrast, return to NL instead of LL every other month does not prevent seasonality in T concentration. These results raise interesting questions about the photoperiodic control of neuroendocrine regulation of seasonal sexual activity and suggest that these treatments can be used to manage males in open barns in farms and in artificial insemination centres. (Spanish and French versions of the full text are available as Supplementary Materials S1 and S2).(c) 2023 The Authors. Published by Elsevier B.V. on behalf of The Animal Consortium. This is an open access article under the CC BY license.

In mammals, especially rodents, social behaviours, such as parenting, territoriality or mate attraction, are largely based on olfactory communication through chemosignals. These behaviours are mediated by species-specific chemosignals, including small organic molecules and proteins that are secreted in the urine or in various fluids from exocrine glands. Chemosignal detection is mainly ensured by olfactory neurons in two specific sensory organs, the vomeronasal organ (VNO) and the main olfactory epithelium (MOE). This study aimed to characterise the olfactory communication in the fossorial ecotype of the water voles, Arvicola terrestris . We first measured the olfactory investigation of urine and lateral scent gland secretions from conspecifics. Our results showed that water voles can discriminate the sex of conspecifics based on the smell of urine, and that urinary male odour is attractive for female voles. Then, we demonstrated the ability of the VNO and MOE to detect volatile organic compounds (VOCs) found in water vole secretions using live-cell calcium imaging in dissociated cells. Finally, we evaluated the attractiveness of two mixtures of VOCs from urine or lateral scent glands in the field during a cyclical outbreak of vole populations.

Introduction Under temperate latitudes, reproduction in Ovis aries displays a marked seasonality, governed by the photoperiod. In natural conditions, the transition between sexual rest and sexual activity in both sexes is induced by the decrease of day light. Meanwhile, specific odors emitted by a sexually active male are able to reactivate the gonadotropic axis of anovulatory ewes. This physiological effect is called “male effect”, precisely ram effect in the ovine species. We have previously shown that the secreted proteins, namely Olfactory Binding Proteins (OBP), contained in the nasal mucus constitute the olfactory secretome (OS), the composition of which is determined by the status of oestrus cycle of females and differs between sexual rest and sexual activity periods. The objective of this study was to test the hypothesis that exposure to sexually active male can also modify the composition of ewes olfactory secretome during a male effect, as well as hormones produced by the reactivation of the oestrus cycle in sexual activity period under natural conditions. Methods We have set up a new non-invasive protocol of nasal mucus sampling and collected it from 12 ewes at different times during a ram effect. We analyzed the composition of their olfactory secretome by proteomics, mainly SDS-PAGE and MALDI-TOF mass spectrometry. As post-translational modifications of OBPs were a hallmark of ewes' sexual activity period, we were looking for glycosylation by western-blot and mass spectrometry. Results The efficiency of male effect was low in stimulated ewes as only 3 females displayed elevated progesterone levels in their blood. Besides, half of control ewes (non-stimulated ones) were cycled. We noticed a common OS profile in ewes in anoestrus, versus OS of cycled ones. A very clear and important result was the apparition of O -GlcNAcylation, previously detected only in sexual activity, after only 30 min of male introduction into the flock. Discussion This exploratory study paves the way for further experiments with larger flock to confirm and reinforce these results, and for eventually exploiting the nasal mucus as an indicator of females' receptivity to male odors.

In a large part of western Europe, water voles have a dynamic reproductive activity across the year, leading to regular multiyear outbreaks, which cause significant damage in mid-mountain grassland ecosystems. Water vole populations are currently managed with rodenticides. But their toxicity to the ecosystem imposes to find means of population control that are more environment-friendly. In this context, we began to explore here aspects of water voles’ olfactory communication and tested the attractivity of several species-specific chemosignals identified in their urine or lateral scent glands’ secretion. One of these odor compounds, namely, 3-ethyl-2,5-dimethyl-pyrazine, induced a strong attraction of female water voles, in terms of longer olfactory investigation. By contrast, this compound seems to be rather repulsive to males. The potential use of 2,5-dimethyl-pyrazine for the control of water vole populations in relation to outbreaks needs to be further evaluated, especially by field-trapping tests performed under different environmental conditions.

Most small rodent species display cyclic fluctuations in their population density. The mechanisms behind these cyclical variations are not yet clearly understood. Density-dependent effects on reproductive function could affect these population variations. The fossorial water vole ecotype, Arvicola terrestris, exhibits multi-year cyclical dynamics with outbreak peaks. Here, we monitored different water vole populations over 3 years, in spring and autumn, to evaluate whether population density is related to male reproductive physiology. Our results show an effect of season and inter-annual factors on testis mass, plasmatic testosterone level, and androgen-dependent seminal vesicle mass. By contrast, population density does not affect any of these parameters, suggesting a lack of modulation of population dynamics by population density.

Mammals living at temperate latitudes typically display annual cyclicity in their reproductive activity: births are synchronized when environmental conditions are most favorable. In a majority of these species, day length is the main proximate factor used to anticipate seasonal changes and to adapt physiology. The brain integrates this photoperiodic signal through key hypothalamic structures, which regulate the reproductive axis. In this context, our study aimed to characterize regulations that occur along the hypothalamo-pituitary-gonadal (HPG) axis in male fossorial water voles (Arvicola terrestris, also known as Arvicola amphibius) throughout the year and to further probe the implication of photoperiod in these seasonal regulations. Our monthly field monitoring showed dramatic seasonal changes in the morphology and activity of reproductive organs, as well as in the androgen-dependent lateral scent glands. Moreover, our data uncovered seasonal variations at the hypothalamic level. During the breeding season, kisspeptin expression in the arcuate nucleus (ARC) decreases, while RFRP3 expression in the dorsomedial hypothalamic nucleus (DMH) increases. Our follow-up laboratory study revealed activation of the reproductive axis and confirmed a decrease in kisspeptin expression in males exposed to a long photoperiod (summer condition) compared with those maintained under a short photoperiod (winter condition) that retain all features reminiscent of sexual inhibition. Altogether, our study characterizes neuroendocrine and anatomical markers of seasonal reproductive rhythmicity in male water voles and further suggests that these seasonal changes are strongly impacted by photoperiod.

Background and Aim: Goats are widely distributed in southwest Algeria. The Saharan goat is perfectly adapted to the harsh conditions of arid areas, and it is characterized by resistance to long photoperiod and reduced metabolic needs, allowing the survival of its offspring by maintaining lactation. Several studies have demonstrated that parturition and lactation are critical periods that induce hormone, energy, and lipid status changes in mammals. However, the relationship between the blood biochemical parameters of parturition control and lactation functions in the Algerian Saharan goat has not been thoroughly documented. Therefore, this study assesses hormone and metabolite levels during parturition and early lactation in Saharan goats reared in arid areas. Materials and Methods: Experiments were performed on 14 multiparous female goats, and blood samples were collected during parturition, 4 days postpartum (D1PP-D4PP), and during the first 12 weeks of lactation (W1-W12) to analyze prolactin, cortisol, glucose (GLU), total proteins (TP), cholesterol (CHO), triglycerides (TGs), total lipids (TL), low-density lipoproteins (LDLs), high-density lipoproteins (HDLs), and very LDLs (VLDLs). Results: Statistical data analysis revealed a significant (p<0.05) increase in plasma prolactin concentrations at W1 after parturition, reaching maximum values at W3 and W9, and remained high until W12 of lactation. Plasma cortisol levels were high at parturition, reaching two peaks at W3 and W9, and then decreased at W5, W7, and W12 of lactation. No significant changes were found in serum GLU levels during the first 7 weeks of lactation compared with parturition day; then, the levels became significantly (p<0.05) lower at W8, W11, and W12 of lactation. Plasma TP increased significantly (p<0.05) at D3PP, W1, and W4, then decreased significantly (p<0.05) at W8. In addition, this decrease coincided with that of GLU production. Serum CHO, TGs, TL, LDLs, and VLDLs, were low at parturition and high at D4PP and during the first 3 months of lactation. Furthermore, HDL levels were low at D3PP, 1st, and 3rd months and high at the 2nd month of lactation. Conclusion: This study emphasized the impact of parturition and the 1st weeks of lactation on endocrine and metabolic changes in indigenous goats living in the Algerian Sahara Desert. These results can be used to monitor and improve farming management and understand physiological adaptive strategies, mainly lactation function sustainability, of this goat living in marginal zones.

Melatonin (MLT) is a biological modulator of circadian and seasonal rhythms and reproduction. The photoperiodic information is detected by retinal photoreceptors and transmitted through nerve transmissions to the pineal gland, where MLT is synthesized and secreted at night into the blood. MLT interacts with two G protein-coupled receptors, MT1 and MT2. The aim of our work was to provide evidence for the presence of MLT receptors in the ovine pineal gland and define their involvement on melatonin secretion. For the first time, we identified the expression of MLT receptors with the specific 2-[125I]-MLT agonistic radioligand in ovin pinealocytes. The values of Kd and Bmax are 2.24 ± 1.1 nM and 20 ± 6.8 fmol/mg. MLT receptors are functional and inhibit cAMP production and activate ERK1/2 through pertussis toxin-sensitive Gi/o proteins. The MLT receptor antagonist/ inverse agonist luzindole increased cAMP production (189 ± 30%) and MLT secretion (866 ± 13%). The effect of luzindole on MLT secretion was additive with the effect of well-described activators of this pathway such as the β-adrenergic agonist isoproterenol and the α-adrenergic agonist phenylephrine. Co-incubation of all three compounds increased MLT secretion by 1236 ± 199%. These results suggest that MLT receptors are involved in the negative regulation of the synthesis of its own ligand in pinealocytes. While adrenergic receptors promote MLT secretion, MLT receptors mitigate this effect to limit the quantity of MLT secreted by the pineal gland.

Seasonally breeding mammals display timely physiological switches between reproductive activity and sexual rest, which ensure synchronisation of births at the most favourable time of the year. These switches correlate with seasonal changes along the hypothalamo-pituitary-gonadal axis, but they are primarily orchestrated at the hypothalamic level through environmental control of KISS1-dependent GnRH release. Our field study shows that births of fossorial water voles, Arvicola terrestris, are concentrated between March and October, which indicates the existence of an annual reproductive cycle in this species. Monthly field monitoring for over a year further reveals dramatic seasonal changes in the morphology of the ovary, uterus and lateral scent glands, which correlate with the reproductive status. Finally, we demonstrate seasonal variation in kisspeptin expression within the hypothalamic arcuate nucleus. Altogether, this study demonstrates a marked rhythm of seasonal breeding in the water vole and we speculate that this is governed by seasonal changes in photoperiod.

Goats are seasonal breeders with the main cue controlling the timing of breeding season being photoperiod. Hence, the season of birth impacts puberty onset: spring-born goats reach puberty in autumn, at 7 mo of age, whereas autumn-born goats reach puberty at 1 yr during the next reproductive season. The aim of this study was to determine whether exposure of autumn-born young females to sexually active males could counteract the delay in puberty onset observed in autumn-born goats. Females exposed to sexually active males (n = 8) reached puberty earlier than isolated females (n = 8), with exposed females ovulating at a mean age of 3.5 mo. To our knowledge, such precocious puberty onset obtained through social stimulation has never been described in the literature. Moreover, those exposed females exhibited estrus behavior for most ovulations. Our results indicate that in goats born out of season, exposure to sexually active bucks is a really efficient approach to induce early puberty, suggesting that social interactions could have a crucial impact on the regulation of pubertal transition.

In male goats, being in permanent visual contact with females in estrus does not prevent seasonal variation in certain endocrine hormone levels and sexual activities. In this study, we tested whether continuous and full contact with females in estrus prevented seasonal endocrinological variation in bucks. In 1 experiment (Exp. 1), we verified that the sudden introduction of goats in estrus increased the plasma concentrations of androgen in bucks during the nonbreeding season under our experimental conditions. In another experiment (Exp. 2), we tested the ability of estrous goats to prevent seasonal inhibition of LH and androgen secretions in bucks kept in permanent and full contact with them. In Exp. 1, 3 groups of bucks (n = 5 in each group) were isolated from females from the months of July to January. On January 27, one group continued being isolated from females; a second group was exposed to ovariectomized, untreated goats; and a third group was exposed to ovariectomized goats with induced estrus. Plasma androgen concentrations were determined every 2 h from 8 h before to 8 h after the introduction of females. The introduction of estrus-induced goats significantly increased androgen concentrations, which were higher than in the isolated bucks, as well as in those exposed to untreated goats (P < 0.05). In Exp. 2 (n = 5 per group), one group of bucks was isolated from females from October to July, whereas two other groups remained in contact with ovariectomized goats, either untreated or regularly induced to estrus. In the three groups of bucks, plasma concentrations of LH were determined once during the months of October, February, March, and June, whereas androgen concentrations were determined weekly from October to July. The mean plasma LH and androgen concentrations were low and did not differ among the groups of bucks during the normal seasonal period of sexual inactivity (P > 0.05). We conclude that full contact and sexual interactions with estrus-induced goats failed to stop the seasonality of LH and androgen plasma concentrations of bucks, although bucks could respond to the introduction of females by acute increases in plasma LH and androgen.

In mammals, melatonin is responsible for the synchronisation of seasonal cycles to the solar year. Melatonin is secreted by the pineal gland with a profile reflecting the duration of the night and acts via the pituitary pars tuberalis (PT), which in turn modulates hypothalamic thyroid hormone status via seasonal changes in the production of locally‐acting thyrotrophin. Recently, we demonstrated that, in the Soay sheep, photoperiodic induction of Tshb expression and consequent downstream hypothalamic changes occur over a narrow range of photoperiods between 12 and 14 hours in duration. In the present study, we aimed to extend our molecular characterisation of this pathway, based on transcriptomic analysis of photoperiodic changes in the pituitary and hypothalamus of ovariectomised, oestradiol‐implanted Ile‐de‐France ewes. We demonstrate that photoperiodic treatments applied before the winter solstice elicit two distinctive modes of accelerated reproductive switch off compared to ewes held on a simulated natural photoperiod, with shut‐down occurring markedly faster on photoperiods of 13 hours or more than on photoperiods of 12 hours and less. This pattern of response was reflected in gene expression profiles of photoperiodically sensitive markers, both in the PT (Tshb, Fam150b, Vmo1, Ezh2 and Suv39H2) and in tanycytes (Tmem252 and Dct). Unexpectedly, the expression of Dio2 in tanycytes did not show any noticeable increase in expression with lengthening photoperiods. Finally, the expression of Kiss1, the key activator of gonadotrophin‐releasing hormone release, was proportionately decreased by lengthening photoperiods, in a pattern that correlated strongly with gonadotrophin suppression. These data show that stepwise increases in photoperiod lead to graded molecular responses at the level of the PT, a progressive suppression of Kiss1 in the hypothalamic arcuate nucleus and luteinising hormone/follicle‐stimulating hormone release by the pituitary, despite apparently unchanged Dio2 expression in tanycytes. We hypothesise that this apparent discontinuity in the seasonal neuroendocrine response illustrates the transient nature of the thyroid hormone‐mediated response to long days in the control of circannual timing.

The water vole Arvicola terrestris is endemic to Europe where its outbreak generates severe economic losses for farmers. Our project aimed at characterising putative chemical signals used by this species, to develop new sustainable methods for population control that could also be used for this species protection in Great Britain. The water vole, as well as other rodents, uses specific urination sites as territorial and sex pheromone markers, still unidentified. Lateral scent glands and urine samples were collected from wild males and females caught in the field, at different periods of the year. Their volatile composition was analysed for each individual and not on pooled samples, revealing a specific profile of flank glands in October and a specific profile of urinary volatiles in July. The urinary protein content appeared more contrasted as males secrete higher levels of a lipocalin than females, whenever the trapping period. We named this protein arvicolin. Male and female liver transcript sequencing did not identify any expression of other odorant-binding protein sequence. This work demonstrates that even in absence of genome, identification of chemical signals from wild animals is possible and could be helpful in strategies of species control and protection.

To survive in temperate latitudes, species rely on the photoperiod to synchronize their physiological functions, including reproduction, with the predictable changes in the environment. In sheep, exposure to decreasing day length reactivates the hypothalamo-pituitary-gonadal axis, while during increasing day length, animals enter a period of sexual rest. Neural stem cells have been detected in the sheep hypothalamus and hypothalamic neurogenesis was found to respond to the photoperiod. However, the physiological relevance of this seasonal adult neurogenesis is still unexplored. This longitudinal study, therefore aimed to thoroughly characterize photoperiod-stimulated neurogenesis and to investigate whether the hypothalamic adult born-cells were involved in the seasonal timing of reproduction. Results showed that time course of cell proliferation reached a peak in the middle of the period of sexual activity, corresponding to decreasing day length period. This enhancement was suppressed when animals were deprived of seasonal time cues by pinealectomy, suggesting a role of melatonin in the seasonal regulation of cell proliferation. Furthermore, when the mitotic blocker cytosine-b-D-arabinofuranoside was administered centrally, the timing of seasonal reproduction was affected. Overall, our findings link the cyclic increase in hypothalamic neurogenesis to seasonal reproduction and suggest that photoperiod-regulated hypothalamic neurogenesis plays a substantial role in seasonal reproductive physiology.

Chromosomal abnormalities are notable causes of infertility in horses. Mares show various degrees of estrous behavior, and ultrasound examination often reveals an underdeveloped genital tract. This article reports investigations on fertility in a Haflinger sibship with a healthy, normally developed, fertile mare with at least three healthy offspring. Chromosomal analysis performed incidentally and blinded for this mare revealed 63,X/64,XX/65,XXX mosaicism. Two closely related mares were also mosaics (63,X/64,XX), and one of them was a carrier of a marker chromosome. Repeated examinations of the mare and seven relatives (four mares and three stallions) did not provide evidence for sub- or in-fertility. They had no developmental abnormalities or conspicuous body conditions. Peripheral blood samples were collected for analysis of the karyotype and molecular analyses. Chromosomes were Giemsa stained and 4′,6-diamidino-2-phenylindole banded to identify numerical or structural aberrations of chromosomes and identification of sex chromosomes, respectively. Fluorescence in situ hybridization was performed with an equine Y-chromosome painting probe to identify and count the sex chromosomes, and polymerase chain reaction analysis was used to test for the presence of the SRY gene and investigating chimerism. The present article demonstrates the necessity of further studies analyzing chromosomal X0 mosaics to improve the predictive value of chromosomal aberrations on fertility.

In rodents, early exposure to adult male is well known to induce an early puberty in females (Vandenbergh effect). This phenomenon has been less studied in other mammals. In goats, despite our extensive knowledge about the "male-effect" phenomenon in adults (i.e. ovulation induced by the introduction of the male during the anestrous), there are few data on the consequences of an early exposure of females to males. Here, we evaluated the puberty onset of young alpine goats when raised since weaning with intact bucks (INT), with castrated bucks (CAS) or isolated from bucks (ISOL). The INT group had the first ovulation 1.5 month before the two other groups. Despite the earlier puberty the INT group of females had normal and regular ovarian cycles. Morphological study of the genital tract showed that at 6 months, uterus of INT goats was 40% heavier than CAS and ISOL goats. Moreover, INT females had a myometrium significantly thicker and INT was the only group having corpora lutea. In our study, INT females were pubescent in the month following the entry of bucks into the breeding season, suggesting that only sexually active bucks provide the signal responsible for puberty acceleration. By removing direct contact with the bucks, we showed that somatosensory interactions were dispensable for an early puberty induction. Finally, no difference in the GnRH network (fiber density and number of synaptic appositions) can be detected between pubescent and non-pubescent females, suggesting that the male stimulations triggering puberty onset act probably on upstream neuronal networks, potentially on kisspeptin neurons.