Characterizing the relationship between gonadotropin releasing hormone (GnRH), kisspeptin, and RFamide related peptide 3 (RFRP-3) neurons in the equine hypothalamus across the estrous cycle and in the anovulatory seasons.

To understand better the role that kisspeptin plays in regulating seasonal and estrous cycle changes in the mare, this study investigated the number, location and interactions between GnRH, kisspeptin and RFRP-3 neurons in the equine hypothalamus. Hypothalami were collected from mares during the non-breeding season, vernal transition and various stages of the breeding season. Fluorescent immunohistochemistry was used to label the neuropeptides of interest. GnRH cells were observed primarily in the arcuate nucleus (ARC), while very few labeled cells were identified in the pre-optic area (POA). Kisspeptin cells were identified primarily in the ARC, with a small number of cells observed dorsal to the ARC, surrounding the third ventricle (3V). The mean number of kisspeptin cells varied between animals and typically showed no pattern associated with season or stage of estrous cycle, but a seasonal difference was identified in the ARC population. Small numbers of RFRP-3 cells were observed in the ARC, ventromedial hypothalamus (VMH) and dorsomedial hypothalamus (DMH). The mean number of RFRP-3 cells appeared higher in pre-ovulatory animals compared to all other stages. The percentage of GnRH cell bodies with kisspeptin appositions did not change with season or stage of estrous cycle. The percentage of kisspeptin cells receiving inputs from RFRP-3 fibers did not vary with season or stage of estrous cycle. These interactions suggest the possibility of the presence of an ultra-short loop feedback system between these three peptides. The changes in RFRP-3 neurons suggest the possibility of a role in the regulation of reproduction in the horse, but it is unlikely to be as a gonadotropin inhibitory factor.

MC3R links nutritional state to childhood growth and the timing of puberty.

The state of somatic energy stores in metazoans is communicated to the brain, which regulates key aspects of behaviour, growth, nutrient partitioning and development1. The central melanocortin system acts through melanocortin 4 receptor (MC4R) to control appetite, food intake and energy expenditure2. Here we present evidence that MC3R regulates the timing of sexual maturation, the rate of linear growth and the accrual of lean mass, which are all energy-sensitive processes. We found that humans who carry loss-of-function mutations in MC3R, including a rare homozygote individual, have a later onset of puberty. Consistent with previous findings in mice, they also had reduced linear growth, lean mass and circulating levels of IGF1. Mice lacking Mc3r had delayed sexual maturation and an insensitivity of reproductive cycle length to nutritional perturbation. The expression of Mc3r is enriched in hypothalamic neurons that control reproduction and growth, and expression increases during postnatal development in a manner that is consistent with a role in the regulation of sexual maturation. These findings suggest a bifurcating model of nutrient sensing by the central melanocortin pathway with signalling through MC4R controlling the acquisition and retention of calories, whereas signalling through MC3R primarily regulates the disposition of calories into growth, lean mass and the timing of sexual maturation.

Cannabidiol has therapeutic potential for myofascial pain in female and male parkinsonian rats.

The musculoskeletal orofacial pain is a complex symptom of Parkinson's disease (PD) resulting in stomatognathic system dysfunctions aggravated by the disease rigidity and postural instability. We tested the effect of cannabidiol (CBD), a non-psychotomimetic constituent of Cannabis sativa, in PD-related myofascial pain. Wistar adult female and male rats orofacial allodynic and hyperalgesic responses were tested by Von Frey and formalin tests, before and 21 days past 6-OHDA lesion. Algesic response was tested after masseter muscle injection of CBD (10, 50, 100 µg in 10 µL) or vehicle. Males compared to females in all estrous cycles' phases presented reduced orofacial allodynia and hyperalgesia. According to the estrous cycle's phases, females presented distinct orofacial nociceptive responses, being the estrus phase well-chosen for nociceptive analysis after 6-OHDA lesion (phase with fewer hormone alterations and adequate length). Dopaminergic neuron lesion decreased mechanical and inflammatory nociceptive thresholds in females and males in a higher proportion in females. CBD local treatment reduced the increased orofacial allodynia and hyperalgesia, in males and females. The female rats were more sensitive to CBD effect considering allodynia, responding to the lowest dose. Although females and males respond to the effect of three doses of CBD in the formalin test, males showed a superior reduction in the hyperalgesic response. These results indicate that hemiparkinsonian female in the estrus phase and male answer differently to the different doses of CBD therapy and nociceptive tests. CBD therapy is effective for parkinsonism-induced orofacial nociception.

Estrogen receptor 2 mediates intraspecific aggressive behaviors of the female Cricetulus barabensis in the estrous cycle.

The social behavior mechanisms have not been thoroughly reported in the solitary female striped dwarf hamster (Cricetulus barabensis). In this study, the handling bag test and neutral arena measurements were used to detect the changes of aggression in the face of rivals of different genders of wild striped dwarf hamsters. We found that female hamsters had the highest aggressive performance in proestrus, followed by estrus, and the lowest in metestrus and the dioestrus, and the increased aggression during the proestrus or estrus period was low-intensity aggression such as intimidation, shock, boxing and counterattack, or even ritualized non-harmful behaviors to drive away opponents. When confronted with male individuals, aggression in females decreased significantly during estrus. The concentration of plasma estradiol was the highest in estrus and the lowest in metestrus and dioestrus. In contrast, estrogen receptor 2 relative expression in the hypothalamus is the lowest in proestrus and highest in metestrus and dioestrus. Besides, both estradiol levels in plasma and estrogen receptor 2 mRNA in the hypothalamus were associated with aggression. These results will broaden our understanding of the molecular mechanism of how breeding phenotype is an essential driver in changing the social behavior of female Cricetulus barabensis.

Ovarian activity and antioxidant indices during estrous cycle of Barki ewes under effect of thyme, celery and salinomycin as feed additives.

This research aimed to examine the effects of thyme, celery and salinomycin on ovarian sex hormones, reproductive traits and antioxidant status during the estrous cycle. Seventy-five mature Barki ewes aged 2-3 years with an average weight of 40 ± 1.5 kg were assigned randomly into five groups (15 head/group). Group 1 was kept as the control; groups 2 and 3 received 20 g/head/day thyme (T) and celery (C) as dried herbs, respectively. Group 4 (T×C) received 10 g thyme + 10 g celery/head/day, and group 5 was treated with salinomycin 1 g/head/day. Blood samples were collected during follicular and luteal phases of the estrous cycle. Thyme and celery and the mixture of T×C increased (P < 0.01) estradiol-17ß (E2) during the follicular phase of the estrous cycle, while only the celery group showed a marked (P < 0.001) increase in progesterone (P4) during the luteal phase compared with the control. Salinomycin supplementation decreased (P < 0.05) E2 concentrations during the follicular and luteal phases of the estrous cycle. Supplementation with thyme and celery enhanced (P < 0.001) antioxidant capacity in the luteal phase compared with the follicular stage. The salinomycin group showed increased (P < 0.01) levels of reduced glutathione (GSH) and decreased malondialdehyde (MDA) levels compared with the control group throughout luteal phase. For the interaction between estrous phases and treatments, thyme, celery, and T×C supplementation revealed an increase (P < 0.05) in superoxide dismutase (SOD), GSH, and glutathione disulfide (GSSG) levels compared with the control group during the follicular and luteal phases. Thyme and celery supplementation improved the number of services per conception and fertilization from 1st and 2nd inseminations, respectively. In conclusion, the applied treatment had significant effects on reproductive performance and antioxidant status in ewes throughout the estrous cycle.

Neuroendocrine disruption is associated to infertility in chronically stressed female rats.

Infertility is a growing worldwide public health problem, and stress is a main factor exerting detrimental effects on female reproduction. However, knowledge regarding the neuroendocrine changes caused by chronic stress in females is limited. Therefore, this study assessed the effects of stress on hormones that control female reproduction during the proestrus and diestrus stages of the estrous cycle, as well as its effects on fertility. Adult females were assigned to either a control or a stress group. Stress consisted of exposure, for 15 min, to cold-water immersion daily for 30 days. Estrous cyclicity, female sexual behavior, as well as hypothalamic kisspeptin, gonadotropin releasing hormone (GnRH) content, serum luteinizing hormone (LH), estradiol (E2), progesterone (P4), corticosterone (CORT) and fertility were assessed after chronic stress. The results show that chronically stressed females exhibited disrupted estrous cyclicity, decreased receptivity, low pregnancy rates and lower numbers of fetuses. The content of Kisspeptin and GnRH in the Anteroventral Periventricular/medial Preoptic Area decreased during proestrus, while Kisspeptin increased in the Arcuate nucleus in proestrus and diestrus. Serum LH decreased only during proestrus, whereas E2 and P4 concentrations decreased during proestrus and diestrus, with a concomitant increase in CORT levels in both stages. As a whole, these results indicate that chronic stress decreases Kisspeptin content in AVPV nucleus and GnRH in POA in females, and might induce disruption of the LH surge, consequently disrupting estrous cyclicity and fertility, leading to lower rates of pregnancy and number of fetuses.

Morphological plasticity of the tuberoinfundibular dopaminergic neurones in the rat during the oestrous cycle and lactation.

The hypothalamic tuberoinfundibular dopaminergic (TIDA) neurones are critical with respect to regulating prolactin secretion from the anterior pituitary. Under most physiological conditions, they are stimulated by prolactin to release dopamine into the median eminence which subsequently suppresses further prolactin secretion from the lactotrophs. During lactation, the TIDA neurones are known to undergo both electrophysiological and neurochemical changes that alleviate this negative-feedback, thus allowing circulating prolactin levels to rise. The present study aimed to determine whether TIDA neurone morphology, most notably spine density, is also modified during lactation. This was achieved by stereotaxically injecting the arcuate nucleus of female, tyrosine hydroxylase-promoter driven Cre-recombinase transgenic rats with Cre-dependent adeno-associated virus-expressing Brainbow. This resulted in the highly specifici transfection of between 10% and 30% of the TIDA neurones, thus allowing the morphologies on multiple individual neurones to be examined in a single hypothalamic slice. The transfected neurones exhibited a range of complex forms, including a diversity of soma and location of axonal origin. Neuronal spine counting showed that the density of somatic, but not dendritic, spines was significantly higher during lactation than at any other reproductive stage. There was also a significant fall in somatic spine density across the oestrous cycle from dioestrus to oestrus. Although the functional characteristics of the additional somatic spines have not been determined, if, as might be expected, they represent an increased excitatory input to the TIDA neurones, this could have important physiological implications by perhaps supporting altered neurotransmitter release at their neuroendocrine terminals. Enhanced excitatory input may, for example, favour the release of the opioid peptide enkephalin rather than dopamine, which is potentially significant because the expression of the peptide is known to increase in the TIDA neurones during lactation and, in contrast to dopamine, it stimulates rather than inhibits prolactin secretion from the pituitary.

Regulation of the hypothalamic GnRH-GnIH system by putrescine in adult female rats and GT1-7 neuronal cell line.

The gonadotropin-releasing hormone-gonadotropin inhibitor (GnRH-GnIH) system in the hypothalamus of mammals is the key factor that controls the entire reproductive system. The aim of this study was to immunolocalize GnIH (RFRP-3) in the hypothalamus during the estrous cycle and to study the effect of putrescine on the expression of GnRH-I and GnIH through both in vivo and in vitro (GT1-7 cells) approach and the circulatory levels of GnRH-I, GnIH, and gonadotropins were also investigated. The study also aims in analyzing all the immunofluorescence images by measuring the relative pixel count of an image. This study showed the effect of putrescine on the morphology of ovary, uterus, and the expression of the steroidogenic acute regulatory protein in the ovary. This study showed GnIH expression was intense during the diestrus and moderate during proestrus and estrus, whereas mild staining during the metestrus. The study further showed that putrescine supplementation to adult female rats increased both GnRH-I expression in the hypothalamus as well as the GnRH-I levels in circulation. The study, for the first time, also showed that putrescine supplementation decreased the expression and release of GnIH. These effects of upregulating GnRH-I expression and downregulating GnIH expression were confirmed by in vitro experiments using GT1-7 cells. Putrescine supplementation also increased the gonadotropin levels in the serum. To summarize, putrescine can regulate the hypothalamic-pituitary-gonadal axis by increasing the GnRH-I, luteinizing hormone, and follicle-stimulating hormone levels and suppressing GnIH levels. This is the first report showing the simultaneous effects of putrescine on the regulation of both GnRH-I and GnIH in the hypothalamus.

Mapping the estrous cycle to context-specific extinction memory.

In Pavlovian renewal paradigms, intact female rats have previously failed to exhibit renewal of appetitive behavior after extinction. However, when treated with exogenous estradiol, female rats exhibit robust renewal behavior. The current study aims to investigate whether the estrous cycle can influence renewal of appetitive behaviors and activity in brain areas known to support the renewal effect. We further aimed to examine whether the estrous cycle would similarly affect renewal of two different types of appetitive behaviors. We first establish that rats in the proestrous stage of the estrous cycle during extinction exhibit elevated renewal behavior compared with rats in either metestrous/diestrous stages, and only rats in proestrus during extinction training (but not during the renewal test) exhibit elevated renewal behavior. Furthermore, we show that this estrous cycle dependent effect on renewal only applies to the conditioned approach behavior toward the food delivery site but not the conditioned approach behavior toward the light cue associated with food delivery. Finally, we examined FOS activity within the prelimbic and infralimbic areas of the medial prefrontal cortex, the dorsal and ventral hippocampal formation, the paraventricular nucleus of the thalamus, the nucleus accumbens, and areas of the amygdala. Particularly in the hippocampus and amygdala, FOS expression which corresponded to the behavioral differences between groups was observed. Results from this study suggest that context information processing may vary as a function of endogenous female hormones across the gonadal hormone cycle and that encoding and retrieval of this information is accomplished in a state-specific manner. (PsycINFO Database Record (c) 2019 APA, all rights reserved).

Environmentally relevant exposure to dibutyl phthalate disrupts DNA damage repair gene expression in the mouse ovary†.

Phthalates have a history of reproductive toxicity in animal models and associations with adverse reproductive outcomes in women. Human exposure to dibutyl phthalate (DBP) occurs via consumer products (7-10 µg/kg/day) and medications (1-233 µg/kg/day). Most DBP toxicity studies have focused on high supraphysiological exposure levels; thus, very little is known about exposures occurring at environmentally relevant levels. CD-1 female mice (80 days old) were treated with tocopherol-stripped corn oil (vehicle control) or DBP dissolved in oil at environmentally relevant (10 and 100 µg/kg/day) or higher (1000 µg/kg/day) levels for 30 days to evaluate effects on DNA damage response (DDR) pathway genes and folliculogenesis. DBP exposure caused dose-dependent effects on folliculogenesis and gene expression. Specifically, animals exposed to the high dose of DBP had more atretic follicles in their ovaries, while in those treated with environmentally relevant doses, follicle numbers were no different from vehicle-treated controls. DBP exposure significantly reduced the expression of DDR genes including those involved in homologous recombination (Atm, Brca1, Mre11a, Rad50), mismatch repair (Msh3, Msh6), and nucleotide excision repair (Xpc, Pcna) in a dose-specific manner. Interestingly, staining for the DNA damage marker, γH2AX, was similar between treatments. DBP exposure did not result in differential DNA methylation in the Brca1 promoter but significantly reduced transcript levels for the maintenance DNA methyltransferase, Dnmt1, in the ovary. Collectively, these findings show that oral exposure to environmentally relevant levels of DBP for 30 days does not significantly impact folliculogenesis in adult mice but leads to aberrant ovarian expression of DDR genes.

Sex differences in central nervous system plasticity and pain in experimental autoimmune encephalomyelitis.

Multiple sclerosis (MS) is a neurodegenerative autoimmune disease with many known structural and functional changes in the central nervous system. A well-recognized, but poorly understood, complication of MS is chronic pain. Little is known regarding the influence of sex on the development and maintenance of MS-related pain. This is important to consider, as MS is a predominantly female disease. Using the experimental autoimmune encephalomyelitis (EAE) mouse model of MS, we demonstrate sex differences in measures of spinal cord inflammation and plasticity that accompany tactile hypersensitivity. Although we observed substantial inflammatory activity in both sexes, only male EAE mice exhibit robust staining of axonal injury markers and increased dendritic arborisation in morphology of deep dorsal horn neurons. We propose that tactile hypersensitivity in female EAE mice may be more immune-driven, whereas pain in male mice with EAE may rely more heavily on neurodegenerative and plasticity-related mechanisms. Morphological and inflammatory differences in the spinal cord associated with pain early in EAE progression supports the idea of differentially regulated pain pathways between the sexes. Results from this study may indicate future sex-specific targets that are worth investigating for their functional role in pain circuitry.

Effects of plane of nutrition and arginine on ovarian follicles in non-pregnant sheep: Cell proliferation, and expression of endothelial nitric oxide and its receptor.

The aim of this study was to investigate the role of the nitric oxide (NO) system in ovarian function, by determining if arginine (Arg) supplementation impacts follicle number, cell proliferation, and expression of the NO system members in nutritionally compromised ewes. Ewes were randomly assigned into maintenance (C, 100% requirements), excess (O; 2xC), or restricted (U; 0.6xC) diets 8 weeks prior to Arg treatment. Ewes were individually fed twice daily with pelleted diets. Ewes from each nutritional group were randomly assigned to one of two treatments: saline or Arg, which was initiated on day 0 of the estrous cycle and administered 3 times per day. Ovaries were collected at the early-luteal, mid-luteal and late-luteal/follicular phases of the estrous cycle to determine 1) the number of surface follicles, 2) follicle cell proliferation marked by Ki67 protein expression, and 3) expression of endothelial nitric oxide (eNOS; NOS3) and soluble guanylyl cyclase beta (sGC; GUCY1B3) protein and mRNA in granulosa (G) and theca (T) layers using immunohistochemistry followed by image analysis and qPCR, respectively. During nutritional treatment, C maintained body weight, O gained 6±1.2 kg, and U lost 14±1.3 kg. Our data show that: 1) Ki67 was expressed in all ovarian compartments, eNOS protein was detected in blood vessels of T and stroma, and sGC protein was detected in T cells, and blood vessels of T layer and other ovarian compartments; 2) plane of nutrition affected the number of surface follicles, and thus folliculogenesis, cell proliferation in the T layer, eNOS and sGC protein expression in T, and NOS3 and GUCY1B3 mRNA expression in G; 3) Arg treatment affected cell proliferation in G and T, eNOS and sGC protein expression in T, mRNA expression of NOS3 in T in all groups, and GUCY1B3 in G depending on the stage of the estrous cycle; and 4) G and T cell proliferation, and expression of eNOS and sGC protein in T was affected by the stage of the estrous cycle. Our data demonstrated that plane of nutrition and Arg are involved in the regulation of follicular functions in non-pregnant sheep.

The Effect of Diet-induced Obesity on Toxicological Parameters in the Polygenic Sprague-Dawley Rat Model.

The obese rodent serves as an indispensable tool for proof-of-concept efficacy and mode-of-action pharmacology studies. Yet the utility of this disease model as an adjunct to the conventional healthy animal in the nonclinical safety evaluation of anti-obesity pharmacotherapies has not been elucidated. Regulatory authorities have recommended employing disease models in toxicology studies when necessary. Our study investigated standard and exploratory toxicology parameters in the high-fat diet (HFD)-induced obese, polygenic Sprague-Dawley rat model in comparison to chow diet (CD)-fed controls. We sought to establish feasibility of the model for safety testing and relevance to human obesity pathophysiology. We report that both sexes fed a 45% kcal HFD for 29 weeks developed obesity and metabolic derangements that mimics to a certain extent, common human obesity. Minor clinical pathologies were observed in both sexes and considered related to CD versus HFD differences. Histopathologically, both sexes exhibited mild obesity-associated findings in brown and subcutaneous white fat, bone, kidneys, liver, lung, pancreas, salivary parotid glands, and skeletal muscle. We conclude that chronic HFD feeding in both sexes led to the development of an obese but otherwise healthy rat. Therefore, the diet-induced obese Sprague-Dawley rat may serve as a suitable model for evaluating toxicity findings encountered with anti-obesity compounds.

Dynamic and Sex-Specific Changes in Gonadotropin-Releasing Hormone Neuron Activity and Excitability in a Mouse Model of Temporal Lobe Epilepsy.

Reproductive endocrine disorders are prominent comorbidities of temporal lobe epilepsy (TLE) in both men and women. The neural mechanisms underlying these comorbidities remain unclear, but hypothalamic gonadotropin-releasing hormone (GnRH) neurons may be involved. Here, we report the first direct demonstrations of aberrant GnRH neuron function in an animal model of epilepsy. Recordings of GnRH neuron firing and excitability were made in acute mouse brain slices prepared two months after intrahippocampal injection of kainate (KA) or control saline, a well-established TLE model in which most females develop comorbid estrous cycle disruption. GnRH neurons from control females showed elevated firing and excitability on estrus compared with diestrus. By contrast, cells from KA-injected females that developed prolonged, disrupted estrous cycles (KA-long) showed the reverse pattern. Firing rates of cells from KA-injected females that maintained regular cycles (KA-regular) were not different from controls on diestrus, but were reduced on estrus. In KA-injected males, only GnRH neurons in the medial septum displayed elevated firing. In contrast to the diestrus versus estrus and sex-specific changes in firing, GnRH neuron intrinsic excitability was elevated in all KA-injected groups, indicating a role for afferent synaptic and neuromodulatory inputs in shaping overall changes in firing activity. Furthermore, KA-injected females showed cycle-stage-specific changes in circulating sex steroids on diestrus and estrus that also differed between KA-long and KA-regular groups. Together, these findings reveal that the effects of epilepsy on the neural control of reproduction are dynamic across the estrous cycle, distinct in association with comorbid estrous cycle disruption severity, and sex-specific.

Molecular cloning and epigenetic change detection of Kiss1 during seasonal reproduction in Chinese indigenous sheep.

Like most seasonal domesticated species, sheep are short-day breeders, which means that the reproduction axis is activated by short days. The annual photoperiodic cycle affects the amount of daylength information that is transmitted to the hypothalamic-pituitary-gonadal (HPG) axis by regulating pulsatile secretion of gonadotrophin-releasing hormone from the hypothalamus. Kisspeptin, which is encoded by Kiss1, plays a major role in reproductive seasonality. Based on results from our previous Solexa sequencing data obtained from Tan (T) and Small Tail Han (STH) sheep during anoestrus and the breeding season, full-length mRNA information for ovine Kiss1 was obtained; 894bp in T sheep and 1145bp in STH sheep. Both encode 135 amino acids. Additionally, T and STH sheep have different transcription start sites of Kiss1. Kiss1 expression during oestrus was significantly higher than that during dioestrus, both in T and STH sheep (P<0.01). We also found a strong relationship between Kiss1 mRNA levels and histone H3 acetylation status in the 5' promoter region of ovine Kiss1. These data indicated that epigenetic modification occurs during reproduction in sheep, and this is the first report that histone H3 deacetylation occurs in the hypothalamus of seasonal sheep breeders during the transition from dioestrus to oestrus.

Food restriction-induced changes in motivation differ with stages of the estrous cycle and are closely linked to RFamide-related peptide-3 but not kisspeptin in Syrian hamsters.

We tested the hypothesis that the effects of food restriction on behavioral motivation are mediated by one or both of the RFamide peptides, RFamide-related peptide-3 (RFRP-3) and kisspeptin (Kp) in female Syrian hamsters (Mesocricetus auratus). Female hamsters fed ad libitum and given a choice between food and adult male hamsters are highly motivated to visit males instead of food on all four days of the estrous cycle, but after 8days of mild food restriction (75% of ad libitum intake) they shift their preference toward food every day of the estrous cycle until the day of estrus, when they shift their preference back toward the males. In support of a role for RFRP-3 in these behavioral changes, the preference for food and the activation of RFRP-3-immunoreactive (Ir) cells in the dorsomedial hypothalamus (DMH) showed the same estrous cycle pattern in food-restricted females, but no association was observed between behavior and the activation of Kp cells in the hypothalamic arcuate nucleus or preoptic area. Next, we tested the hypothesis that food-restriction-induced activation of RFRP-3-Ir cells is modulated by high levels of ovarian steroids at the time of estrus. In support of this idea, on nonestrous days, mild food restriction increased activation of RFRP-3-Ir cells, but failed to do so on the day of estrus even though this level of food restriction did not significantly decrease circulating concentrations of estradiol or progesterone. Furthermore, in ovariectomized females, food-restriction-induced increases in activation of RFRP-3-Ir cells were blocked by systemic treatment with progesterone alone, estradiol plus progesterone, but not estradiol alone. Central infusion with RFRP-3 in ad libitum-fed females significantly decreased sexual motivation and produced significant increases in 90-minute food hoarding, in support of the hypothesis that elevated central levels of RFRP-3 are sufficient to create the shift in behavioral motivation in females fed ad libitum. Together, these results are consistent with the hypothesis that high levels of ingestive motivation are promoted during the nonfertile phase of the estrous cycle by elevated activation of RFRP-3-Ir cells, and RFRP-3-Ir cellular activation is modulated by ovarian steroids around the time of estrus, thereby diverting attention away from food and increasing sexual motivation.

Effect of adding different levels of rumen protected choline to the diet on productive and reproductive performance of female goats and growth of their kids from birthing to weaning.

Forty female goats in the third parity were randomly divided into four similar groups. The experiment was started 20 days before mating and lasted until the end of the suckling period for 60 days and weaning their kids. The first group were fed diet without supplementation and kept as control while in the second, third and fourth groups, each doe was fed diets with rumen protected choline (RPC) at the rate of 10, 20 and 40 g/day, respectively. Results showed that number of doe kidding twins and triplets and litter weight of kids born per group increased with increasing the level of RPC in the diet of goats and viability rate of born kids during the suckling period improved due to RPC supplementation in the diets of their mothers. Duration of estrous, days from weaning to estrous, days from kidding to estrous and kidding interval decreased significantly, while conception rate increased due to adding RPC. Milk choline concentrations and total choline secretion though milk were progressively increased significantly with increasing the level of RPC supplementation. Live body weight and daily body gain of their suckling male and female kids at weaning increased significantly with increasing RPC levels in the diets of their mothers.

Ovarian hormones modify anxiety behavior and glucocorticoid receptors after chronic social isolation stress.

Chronic social isolation could lead to a disruption in the Hypothalamic-Pituitary-Adrenal (HPA) axis, resulting in anxiety and depressive-like behaviors but cycling estrogens could modify these behaviors. The aim of this study was to determine if changes in ovarian hormones during the normal cycle could interact with social isolation to alter anxiety and depressive-like behaviors. In parallel, we examined the expression of glucocorticoid receptor (GR) and synaptic vesicle protein synaptophysin in the hippocampus and hypothalamus of Sprague Dawley normal cycling female rats. We assigned rats to either isolated or paired housing for 8 weeks. To assess anxiety and depressive-like behaviors, we used the open field test and forced swim test, respectively. Female rats were tested at either diestrus, estrus, or proestrus stage of the estrous cycle. After behaviors, rats were perfused and brains collected. Brain sections containing hippocampus and hypothalamus were analyzed using immunohistochemistry for synaptophysin and glucocorticoid receptor (GR) levels. We found an increase in depressive-like behaviors for isolated animals compared to paired housed rats, regardless of the estrous cycle stage. Interestingly, we found a decrease in anxiety behaviors in females in the estrus stage accompanied by a decrease in GR expression in hippocampal DG and CA3. However, no changes in synaptophysin were observed in any of the areas of studied. Our results support the beneficial effects of circulating ovarian hormones in anxiety, possibly by decreasing GR expression.

Expression of ODC1, SPD, SPM and AZIN1 in the hypothalamus, ovary and uterus during rat estrous cycle.

The aim of the present study was to investigate variation in the expression pattern of ornithine decarboxylase (ODC1), spermine (SPM), spermidine (SPD) and antizyme inhibitor (AZIN1) in hypothalamus, ovary and uterus during the estrous cycle of rats. Further, to understand any correlation between polyamines and GnRH I expression in hypothalamus; effect of putrescine treatment on GnRH I expression in hypothalamus and progesterone and estradiol levels in serum were investigated. The study also aims in quantifying all the immunohistochemistry images obtained based on pixel counting algorithm to yield the relative pixel count. This algorithm uses a red green blue (RGB) colour thresholding approach to quantify the intensity of the chromogen present. The result of the present study demonstrates almost similar expression pattern of polyamine and polyamine related factors, ODC1, SPD, SPM and AZIN1, with that of hypothalamic GnRH I, all of which mainly localized in the medial preoptic area (MPA) of the hypothalamus, during the proestrus, estrus and diestrus. This suggest that hypothalamic GnRH I expression is under regulation of polyamines. The study showed significant increase in hypothalamic GnRH I expression for both the doses of putrescine treatment to adult female rats. Further, it was shown that in ovary expression pattern of ODC1, SPM, SPD and AZIN1 were similar with that of steroidogenic factor, StAR during the estrous cycle, and putrescine supplementation increased significantly estradiol and progesterone levels in serum, all suggesting ovarian polyamines are involved in regulation of ovarian steroidogenesis. Localization of these factors in the theca and granulosa cells suggest involvement of polyamines in the process of folliculogenesis and luteinization; and ODC1, SPD, SPM and AZIN1 in oocyte further suggests polyamine role in maintenance of oocyte physiology. Finally, in uterus SPM and AZIN1 were localized throughout the estrous cycle, being comparatively more during the metestrus phase. There was intense immunostaining of SPD in the luminal and glandular epithelium during the metestrus and diestrus phases of the estrous cycle suggesting these all the three polyamines as such play important role in regulation of uterine physiology.

AgRP to Kiss1 neuron signaling links nutritional state and fertility.

Mammalian reproductive function depends upon a neuroendocrine circuit that evokes the pulsatile release of gonadotropin hormones (luteinizing hormone and follicle-stimulating hormone) from the pituitary. This reproductive circuit is sensitive to metabolic perturbations. When challenged with starvation, insufficient energy reserves attenuate gonadotropin release, leading to infertility. The reproductive neuroendocrine circuit is well established, composed of two populations of kisspeptin-expressing neurons (located in the anteroventral periventricular hypothalamus, Kiss1AVPV, and arcuate hypothalamus, Kiss1ARH), which drive the pulsatile activity of gonadotropin-releasing hormone (GnRH) neurons. The reproductive axis is primarily regulated by gonadal steroid and circadian cues, but the starvation-sensitive input that inhibits this circuit during negative energy balance remains controversial. Agouti-related peptide (AgRP)-expressing neurons are activated during starvation and have been implicated in leptin-associated infertility. To test whether these neurons relay information to the reproductive circuit, we used AgRP-neuron ablation and optogenetics to explore connectivity in acute slice preparations. Stimulation of AgRP fibers revealed direct, inhibitory synaptic connections with Kiss1ARH and Kiss1AVPV neurons. In agreement with this finding, Kiss1ARH neurons received less presynaptic inhibition in the absence of AgRP neurons (neonatal toxin-induced ablation). To determine whether enhancing the activity of AgRP neurons is sufficient to attenuate fertility in vivo, we artificially activated them over a sustained period and monitored fertility. Chemogenetic activation with clozapine N-oxide resulted in delayed estrous cycles and decreased fertility. These findings are consistent with the idea that, during metabolic deficiency, AgRP signaling contributes to infertility by inhibiting Kiss1 neurons.