Transcriptome analysis during 4-vinylcyclohexene diepoxide exposure-induced premature ovarian insufficiency in mice.

The occupational chemical 4-Vinylcyclohexene diepoxide (VCD) is a reproductively toxic environmental pollutant that causes follicular failure, leading to premature ovarian insufficiency (POI), which significantly impacts a woman's physical health and fertility. Investigating VCD's pathogenic mechanisms can offer insights for the prevention of ovarian impairment and the treatment of POI. This study established a mouse model of POI through intraperitoneal injection of VCD into female C57BL/6 mice for 15 days. The results were then compared with those of the control group, including a comparison of phenotypic characteristics and transcriptome differences, at two time points: day 15 and day 30. Through a comprehensive analysis of differentially expressed genes (DEGs), key genes were identified and validated some using RT-PCR. The results revealed significant impacts on sex hormone levels, follicle number, and the estrous cycle in VCD-induced POI mice on both day 15 and day 30. The DEGs and enrichment results obtained on day 15 were not as significant as those obtained on day 30. The results of this study provide a preliminary indication that steroid hormone synthesis, DNA damage repair, and impaired oocyte mitosis are pivotal in VCD-mediated ovarian dysfunction. This dysfunction may have been caused by VCD damage to the primordial follicular pool, impairing follicular development and aggravating ovarian damage over time, making it gradually difficult for the ovaries to perform their normal functions.

Daily GnRH agonist treatment effectively delayed puberty in female rats without long-term effects on sexual behavior or estrous cyclicity.

The present study examined the long-term effects of suppressing puberty with a GnRH agonist on reproductive physiology and behavior in female rats. We have recently reported that administration of the GnRH agonist leuprolide acetate (25 µg/kg) daily between postnatal day (PD) 25-50 delayed puberty and disrupted the development of copulatory behavior and sexual motivation in male rats. However, pilot data from our lab suggest that this low dose of leuprolide acetate (25 µg/kg) was not high enough to significantly delay puberty in female rats. Therefore, we injected female Long-Evans rats with leuprolide acetate at a higher dose (50 µg/kg) or 0.9% sterile saline, daily , starting on PD 25 and ending on PD 50. Vaginal opening was monitored daily starting on PD 30 for signs of pubertal onset and first estrous cycle. In addition, we measured estrous cyclicity starting approximately 2 weeks after the last injection of leuprolide (∼PD 64). Immediately after monitoring estrous cyclicity, the female rats were mated on their first day in behavioral estrus using the partner-preference paradigm, with and without physical contact (PD 95-110). We found that this dose of leuprolide (50 µg/kg) significantly delayed puberty; however, neither estrous cyclicity nor sexual motivation was significantly affected by periadolescent exposure to leuprolide. Together with our findings in male rats, these results add to our understanding of the developmental effects of chemically suppressing puberty in rats.

Alginic acid induces oxidative stress-mediated hormone secretion disorder, apoptosis and autophagy in mouse granulosa cells and ovaries.

Alginic acid (AA) is a kind of polysaccharide extracted from brown seaweeds and has been widely used in food industry. Certain positive effects of AA, such as anti-inflammation and anti-allergy, have been reported. Nevertheless, as a potential chemical contaminant of the environment, its impact on female reproductive system remains to be investigated. The purpose of this study is to explore the impact of AA on ovary and to investigate the further cellular mechanism. Primarily, in vitro cultured mouse ovary granulosa cells (GCs) were treated with AA at a concentration of 10µM for 24 h. The cells and supernatant were collected and subjected to further measures. The results demonstrated that after being treated with 10µM AA for 24 h the levels of estradiol and progesterone in supernatant were down-regulated. And excessive reactive oxygen species (ROS) and declined antioxidant capacity were also determined. Additionally, a large number of apoptotic bodies and autophagic vesicles were found in the experimental cells, and the mitochondria-mediated apoptotic pathway was demonstrated to play a main role in GCs apoptosis. To further investigate the effect of AA on ovary, the female ICR mice were administered with AA (10 mg/ kg bodyweight) intraperitoneally for successive 35 days, and the estrus phase was recorded simultaneously. After exposure, the ovaries and blood samples were collected for further analysis. The results revealed that the estrus period of the mice was shortened and the interestrus period was extended after being treated with AA for 35 days. At the organismal level, the numbers of antral follicles and atresia follicles increased and the levels of pro-apoptosis and autophagy-related proteins were detected upregulated after AA treatment. Taken together, both in vivo and in vitro data suggested that AA has toxicity on female reproduction by disrupting estrogen production and inducing oxidative stress, mitochondria-mediated apoptosis and autophagy. Our results provide new scientific basis and the concern for controlling the increasing use of AA.

Cyantraniliprole impairs reproductive parameters by inducing oxidative stress in adult female wistar rats.

Cyantraniliprole is a synthetic insecticide used to control pests of up to 23 different types of crops. It is able to modulate ryanodine-like calcium channels, which are widely found in the organism of insects and mammals. The objective of this research was to verify the possible reproductive effects of adult female Wistar rats exposure to cyantraniliprole. Animals (67 days old) were exposed to the chemical at doses of 10 or 150 mg/kg/day, orally, for 28 consecutive days (control animals received only the vehicle). Vaginal secretions were collected during the exposure period to assess the regularity of the estrous cycle; the liver, kidneys, pituitary gland, adrenal gland, uterus, and ovaries were collected and weighed; reproductive organs were assessed for histopathological evaluation and for biochemical markers of oxidative stress and progesterone plasma level was measured. Both doses caused negative changes in the morphology and redox system of the uterus and ovaries. Animals exposed to 10 mg/kg also exhibited higher level of plasma progesterone, elevated levels of lipid peroxidation in reproductive organs, increased superoxide dismutase activity in the uterus and glutathione peroxidase activity on the ovary, while the 150 mg/kg group exhibited an increment in estrous cycle length and diminished uterine glandular epithelium. Based on these results, we conclude that cyantraniliprole may have acted as an endocrine disruptor, and its effects are mediated by oxidative stress.

Combinational exposure to Bisphenol A and a high-fat diet causes trans-generational Malfunction of the female reproductive system in mice.

Bisphenol A (BPA) is a common endocrine disruptor and a high-fat diet (HFD) also affects fertility. However, little is known about the long-term consequences of simultaneous exposure to BPA and a HFD on reproductive health. Herein, we assessed the effects of maternal exposure to BPA in combination with a HFD on reproductive function in subsequent generations of female mice and evaluated its effects on the hypothalamic-pituitary-gonadal axis. We found that the combination of maternal exposure to BPA and a HFD led to increased urine BPA levels, precocious puberty, altered estrous cyclicity, decreased follicle numbers, and altered hypothalamic Kiss1 methylation status in F1 and F2 mice. Therefore, we demonstrated that maternal exposure to BPA in combination with a HFD exerts a trans-generational effect on female reproduction.

Long-term treatment of Nicotinamide mononucleotide improved age-related diminished ovary reserve through enhancing the mitophagy level of granulosa cells in mice.

Ovarian aging affects the reproductive health of elderly women due to decline in oocyte quality, which is closely related to mitochondrial dysfunction. Nicotinamide mononucleotide (NMN), as a precursor of NAD+, effectively regulate mitochondria metabolism in mice. However, roles of NMN in improving age-related diminished ovary reserve remain to be determined. In present study, 4, 8, 12, 24, 40-week old female ICR mice were collected and a 20-week-long administration of NMN was conducted to 40-week-old mice (60WN), meanwhile the control group is given water (60WC). First, we found that 20-week-long administration of NMN to 40-week-old mice exhibited anti-aging and anti-inflammatory effects on organ structures, along with the improvement of estrus cycle condition and endocrine function. The number of primordial, primary, secondary, antral follicles and corpora luteum of ovaries in 60WN group was significantly increased compared with those in 60WC group. Additionally, the protein and gene expressions of P16 of ovaries were significantly reduced in 60WN group than in 60WC group. the mitochondria biogenesis, autophagy level, and proteases activity enhanced in granulosa cells after 20-week-administration of NMN. Present results indicate that NMN has the potential to save diminished ovary reserve by long-term treatment, providing a basis for exploring the role of NMN in anti-ovarian aging by enhancing the mitophagy level of granulosa cells.

Theaflavin 3, 3'-Digallate Delays Ovarian Aging by Improving Oocyte Quality and Regulating Granulosa Cell Function.

Ovarian aging refers to the gradual decline of ovarian function with increasing physiological age, manifested as decreased ovarian reserve, elevated aging-related markers, and reduced oocyte quality. With a declining female fertility and a growing aging population, it is urgent to delay ovarian aging to maintain fertility and improve the life quality of women. Theaflavin 3, 3'-digallate (TF3) is a naturally bioactive polyphenol compound extracted from black tea, and its antioxidant properties play an important role in maintaining human health and delaying aging; however, the effects of TF3 on female reproduction and ovarian function are not yet clear. Here, we show that TF3 can preserve primordial follicle pool, partially restore the estrous cycle, and increase the offspring number of aged mice. Meanwhile, TF3 gavage increased the number of oocytes retrieved, decreased the level of reactive oxygen species, increased the level of glutathione, and decreased the abnormal rate of oocyte spindle after ovulation induction. Moreover, TF3 inhibited human granulosa cell apoptosis and improved their antioxidative stress ability. High-throughput sequencing and small-molecule-targeted pharmacological prediction show that TF3 affects multiple pathways and gene expression levels, mainly involved in reproductive and developmental processes. It may also affect cellular function by targeting mTOR to regulate the autophagic pathway, thereby delaying the process of ovarian aging. This study shows that TF3 can be used as a potential dietary supplement to protect ovary function from aging and thereby improving the life quality of advanced-age women.

The effectiveness of resveratrol in treatment of PCOS on the basis of experimental model in rats.

PURPOSE OF THE STUDY: Polycystic ovary syndrome (PCOS) is a widespread endocrine disorder in women of reproductive age. Further research is required to justify new directions of effective targeted therapy of this condition. Resveratrol possesses anti-inflammatory, antioxidant and antidiabetic properties. The purpose of this study was to evaluate the potential effectiveness of resveratrol in PCOS based on the created model of this disease in Wistar rats. MATERIALS AND METHODS: The PCOS model was created by oral administration of letrozole to female Wistar rats.. The animals received resveratrol at a dosage of 20 mg/kg and 30 mg/kg for the next 30 days. Then ovariectomy was performed for histological confirmation of the effectiveness of resveratrol in the treatment of PCOS. Regularity of estrous cycle, animal's body mass and the level of soluble receptors for advanced glycation end products (sRAGE) in the blood of rats were also evaluated in dynamics. RESULTS: The study revealed that administration of resveratrol leads to dose-dependent restoration of normal morphology of ovarian tissue, normalizes regularity of estrous cycle and decreases body weight of rats with PCOS. CONCLUSION: The results obtained in rats suggest that resveratrol may be a promising agent for the treatment of PCOS in women.

El consumo semicrónico moderado de alcohol altera la foliculogénesis y la calidad núcleo-citoplasmática oocitaria, en el ratón / Moderate semi-chronic alcohol consumption alters folliculogenesis and oocyte nuclear-cytoplasmic quality in mice

Resumen El consumo crónico de alcohol es un problema de salud mundial que afecta particularmente a la población femenina. Sin embargo, los efectos de la ingesta semicrónica en cantidades moderadas a bajas en el ovario y el oocito son poco conocidos. En un modelo murino, se administró etanol al 10% en agua de bebida (hembras tratadas) o agua (hembras control) por 15 días, y luego de la superovulación o no (ovulación espontánea), se analizó el ciclo estral y la calidad ovárico-gamética. En las hembras tratadas, la frecuencia y duración del diestro aumentó, y las frecuencias de folículos y cuerpos lúteos disminuyeron vs hembras controles, valores que se restauraron luego de la superovulación. Sin embargo, en las hembras tratadas, la tasa de proliferación celular folicular y el desbalance de la expresión ovárica de VEGF (factor de crecimiento endotelial) persistieron luego de la superovulación. El número de ovocitos ovulados con metafase II anormal, fragmentados y activados partenogenéticamente fue mayor en las hembras tratadas respecto las controles. En conclusión, el consumo semicrónico moderado de alcohol produce anestro, ciclo estral irregular, foliculogénesis deficiente y anomalías núcleo-citoplasmáticas en los oocitos ovulados. Estas alteraciones podrían constituirse en un factor etiológico de pérdida gestacional temprana y desarrollo embrionario anormal luego del consumo de alcohol.

Abstract Chronic alcohol consumption is a global health problem that particularly affects the female population. However, the ef-fects of semi-chronic ethanol intake in low-moderate amounts on the ovary and oocyte are poorly understood. In a mouse model, 10% ethanol was administered in drinking water (treated females) or water (control females) for 15 days, and after superovulation or not (spontaneous ovulation), the estrous cycle and ovarian-gametic quality were analyzed. In treated females, the frequency and duration of the diestrus increased, and the frequencies of follicles and corpus luteum decreased vs control females, values that restored after superovulation. However, in treated females, the follicular cell proliferation rate and the imbalance in ovarian expression of VEGF (endothelial growth factor) persisted after superovulation. The number of ovulated oocytes with abnormal metaphase II, fragmented and parthenogenetically activated was higher in treated females than in control ones. In conclusion, moderate semi-chronic alcohol consumption produces anestrum, irregular estrous cycle, poor folliculogenesis, and nuclear-cytoplasmic abnormalities in ovulated oocytes. These alterations could constitute an etiological factor of early gestational loss and abnormal embryonic development after alcohol consumption.

Effects of chronic exposure to bisphenol A in adult female mice on social behavior, vasopressin system, and estrogen membrane receptor (GPER1).

Bisphenol A (BPA), an organic synthetic compound found in some plastics and epoxy resins, is classified as an endocrine disrupting chemical. Exposure to BPA is especially dangerous if it occurs during specific "critical periods" of life, when organisms are more sensitive to hormonal changes (i.e., intrauterine, perinatal, juvenile or puberty periods). In this study, we focused on the effects of chronic exposure to BPA in adult female mice starting during pregnancy. Three months old C57BL/6J females were orally exposed to BPA or to vehicle (corn oil). The treatment (4 µg/kg body weight/day) started the day 0 of pregnancy and continued throughout pregnancy, lactation, and lasted for a total of 20 weeks. BPA-treated dams did not show differences in body weight or food intake, but they showed an altered estrous cycle compared to the controls. In order to evidence alterations in social and sociosexual behaviors, we performed the Three-Chamber test for sociability, and analyzed two hypothalamic circuits (well-known targets of endocrine disruption) particularly involved in the control of social behavior: the vasopressin and the oxytocin systems. The test revealed some alterations in the displaying of social behavior: BPA-treated dams have higher locomotor activity compared to the control dams, probably a signal of high level of anxiety. In addition, BPA-treated dams spent more time interacting with no-tester females than with no-tester males. In brain sections, we observed a decrease of vasopressin immunoreactivity (only in the paraventricular and suprachiasmatic nuclei) of BPA-treated females, while we did not find any alteration of the oxytocin system. In parallel, we have also observed, in the same hypothalamic nuclei, a significant reduction of the membrane estrogen receptor GPER1 expression.

Regulation of uterine function during estrous cycle, anestrus phase and pregnancy by steroids in red deer (Cervus elaphus L.).

Steroid synthesis and production in ruminant uterus is not obvious, especially in seasonally reproduced. We compared steroid production by investigating enzymes involved in red deer uterine steroid metabolism in reproductive seasons. Blood and uteri (endometrium and myometrium) were collected post mortem from hinds on 4th day (N = 8), 13th day of the cycle (N = 8), anestrus (N = 8) and pregnancy (N = 8). The expression of cytochrome P450 aromatase (P450), 3 -beta-hydroxysteroid dehydrogenase (3ß-HSD), 17 -beta-hydroxysteroid dehydrogenase (17ß-HSD), aldo-keto reductase family 1 C1 (AKR1C1), estrogen receptor alpha (ERα), and progesterone receptors (PRs), were analyzed using real-time-PCR and Western Blotting. Plasma samples were assayed for 17-beta-estradiol (E2), progesterone (P4), luteinizing hormone (LH), follicle-stimulating hormone (FSH), and testosterone (T4) concentrations by EIA. Hinds at the beginning of the estrous cycle, mainly in endometrium, were characterized by a high mRNA expression of 3ß-HSD, AKR1C1, PRs and ERα, contrary to the expression in myometrium during pregnancy (P < 0.05). For P4, E2, and FSH, concentration was the highest during the 13th day of the estrous cycle (P < 0.05). Uterine steroid production and output in hinds as a representative seasonally reproduced ruminant occurred mainly during the estrous cycle and sustained in anestrus.

Effects of Radix Polygalae on Cognitive Decline and Depression in Estradiol Depletion Mouse Model of Menopause.

Postmenopausal syndrome refers to symptoms caused by the gradual decrease in female hormones after mid-40 years. As a target organ of estrogen, decrease in estrogen causes various changes in brain function such as a decrease in choline acetyltransferase and brain-derived neurotrophic factor; thus, postmenopausal women experience cognitive decline and more depressive symptoms than age-matched men. Radix Polygalae has been used for memory boosting and as a mood stabilizer and its components have shown neuroprotective, antidepressant, and stress relief properties. In a mouse model of estrogen depletion induced by 4-vinylcyclohexene diepoxide, Radix Polygalae was orally administered for 3 weeks. In these animals, cognitive and depression-related behaviors and molecular changes related to these behaviors were measured in the prefrontal cortex and hippocampus. Radix Polygalae improved working memory and contextual memory and despair-related behaviors in 4-vinylcyclohexene diepoxide-treated mice without increasing serum estradiol levels in this model. In relation to these behaviors, choline acetyltransferase and brain-derived neurotrophic factor in the prefrontal cortex and hippocampus and bcl-2-associated athanogene expression increased in the hippocampus. These results implicate the possible benefit of Radix Polygalae in use as a supplement of estrogen to prevent conditions such as postmenopausal depression and cognitive decline.

Effect of urolithins A and B on ectopic endometrial growth in a murine model of endometriosis.

Endometriosis is an often painful disease in reproductive-aged women, in which endometrial-like tissue grows outside the uterine cavity. Since the limited current therapeutic alternatives fail in alleviating the symptoms and based on our previous research in in vitro models using the same compounds as the ones used in the present study, we aimed to evaluate the effects of urolithins A (UA) and B (UB) on the growth and survival of endometriotic-like lesions in a murine model of endometriosis. Female BALB/C mice were surgically induced with endometriosis and treated with 2.5 mg kg-1 day-1 intraperitoneal UA or UB. The mice were monitored daily and weighed and the estrous stage was determined. After 28 days of treatment, lesions were counted, measured, excised, and fixed. Both urolithins proved not to affect the estrous cycle or body weight of the mice. UA completely prevented endometriotic-like lesions, while UB diminished the implant volume (p < 0.05). Treatment also reduced epithelial and stromal cell proliferation within the implants (p < 0.001 and p < 0.01, respectively) and apoptosis was enhanced (p < 0.05 and p < 0.01, respectively). These results are promising and reveal that urolithins A and B, separately, have a beneficial effect on the overall endometriotic growth without affecting the body weight or estrous cycle.

The damage effect of heat stress and psychological stress combined exposure on uterus in female rats.

AIMS: Explore the effects of heat stress and psychological stress combined exposure on the uterus and its underlying mechanisms. MAIN METHODS: Sixty female Sprague-Dawley rats were randomly assigned to four groups: control group, psychological stress group, high ambient temperature group, and high ambient temperature combined with psychological stress group. All treatments were administered for two weeks. During this period, the estrous cycle, body weights and rectal temperature were measured regularly. Then, ovarian weight coefficient, serum estradiol (E2) and progesterone (P) concentration, uterine histomorphological alterations, levels of tumor necrosis factor alpha (TNF-α), malondialdehyde (MDA) and superoxide dismutase (SOD), and the expressions of ovarian hormone receptors, leukemia inhibitory factor (LIF) and its receptor, homeobox gene A10 (HoxA10), Wnt5a, Wnt7a, ß-catenin, and P-ß-cateninY142 in the uterus and endometrium were detected. KEY FINDINGS: High temperature combined with psychological stress lead to body weight, body temperature, ovarian hormones and estrus cycle disorder, uterine gland ducts expansion and endometrial thickness reduction, and the decreased expression of endometrial receptivity markers (LIF and HoxA10). Further, disturbed expression of E2 and P receptors in endometrium, elevated MDA and TNF-α levels, and decreased Wnt5a, Wnt7a and P-ß-cateninY142 content were found. Our data suggested that co-exposure to high temperature and psychological stress could aggravate uterine damage probably by inducing ovarian hormonal disorder and the subsequent oxidative stress and inflammation, and reduce the endometrial function through suppressing Wnt signaling. SIGNIFICANCE: This will provide the scientific basis for improving female reproductive health, and preventing and treating reproductive disorders.

Characterization of a differential reinforcement of low rates of responding task in non-deprived male and female rats: Role of Sigma-1 receptors.

Impulsive action can be defined as the inability to withhold a response and represents one of the dimensions of the broad construct impulsivity. Here, we characterized a modified differential reinforcement of low rates of responding (DRL) task developed in our laboratory, in which impulsive action is measured in ad libitum fed/watered subjects. Specifically, we first determined the effects of both sex and estrous cycle on impulsive action by systematically comparing male and estrous-synchronized female subjects. In addition, we evaluated the convergent validity of this modified DRL task by testing the effects of the D2R/5HT2AR antagonist, aripiprazole, and the noncompetitive NMDAR antagonist, MK-801. Finally, we tested the effects of the selective antagonist BD-1063 and agonist PRE-084 of Sigma-1 receptor (Sig-1R) on impulsive action using this modified DRL task. We found that female rats showed and increased inability to withhold a response when compared to males, and this effect was driven by the metestrus/diestrus phase of the estrous cycle. In addition, aripiprazole and MK-801 fully retained their capability to reduce and increase impulsive action, respectively. Finally, the selective Sig-1R antagonist, BD-1063 dose-dependently reduced the inability to withhold a response in both sexes, though more potently in female rats. In summary, we show that impulsive action, as measured in a modified DRL task which minimizes energy-homeostatic influences, is a function of both sex and estrous cycle. Furthermore, we validate the convergent validity of the task and provide evidence that Sig-1R antagonism may represent a novel pharmacological strategy to reduce impulsive action.

Metformin and exenatide upregulate hepatocyte nuclear factor-4α, sex hormone binding globulin levels and improve hepatic triglyceride deposition in polycystic ovary syndrome with insulin resistance rats.

OBJECTIVE: To explore the efficacy and underlying mechanisms of metformin and exenatide in reversing reproductive and metabolic disturbances in letrozole combined with high-fat diet-induced polycystic ovary syndrome (PCOS) rats. METHODS: Rats with PCOS and insulin resistance (IR) were induced by intra-gastric instillation of letrozole combined with a high-fat diet and verified by histological screening of vaginal exfoliated cells. After metformin and exenatide supplementation, body weight, chow intake and ovarian morphology were observed. Serum biochemical profiles were analyzed using ELISA, while the levels of key anabolism-related proteins, including sex hormone binding globulin (SHBG), hepatocyte nuclear factor-4α (HNF-4α), PI3K, and AKT, were determined using western blotting. RESULTS: The estrus cycle and ovarian morphology of rats with PCOS and IR were significantly recovered following metformin and exenatide treatment, with decreased body weight and chow intake. Furthermore, PCOS-induced changes in metabolic disorders including IR and hepatic triglyceride (TG) deposition, and hyperandrogenemia were reversed by treatment with both drugs. Specifically, the levels of HNF-4α and SHBG in liver tissue of rats with PCOS and IR were upregulated significantly. CONCLUSIONS: Both metformin and exenatide could recover the estrous cycle and ovarian morphology, reduce body weight and high-fat chow intake, and improve glycolipid metabolism disorders and hyperandrogenemia in PCOS with IR rat models. Interestingly, our findings also highlight the potential of both therapeutic agents for improving IR by regulating the liver PI3K/AKT pathway, reducing the deposition of hepatic TG, as well as upregulating the levels of SHBG and HNF-4α in PCOS with IR rat liver tissue.

Expression and Regulation of CD73 during the Estrous Cycle in Mouse Uterus.

Cluster of differentiation 73 (CD73, also known as ecto-5'-nucleotidase) is an enzyme that converts AMP into adenosine. CD73 is a surface enzyme bound to the outside of the plasma membrane expressed in several cells and regulates immunity and inflammation. In particular, it is known to inhibit T cell-mediated immune responses. However, the regulation of CD73 expression by hormones in the uterus is not yet clearly known. In this study, we investigated the expression of CD73 in ovariectomized mice treated with estrogen or progesterone and its regulation in the mouse uterus during the estrous cycle. The level of CD73 expression was dynamically regulated in the uterus during the estrous cycle. CD73 protein expression was high in proestrus, estrus, and diestrus, whereas it was relatively low in the metestrus stage. Immunofluorescence revealed that CD73 was predominantly expressed in the cytoplasm of the luminal and glandular epithelium and the stroma of the endometrium. The expression of CD73 in ovariectomized mice was gradually increased by progesterone treatment. However, estrogen injection did not affect its expression. Moreover, CD73 expression was increased when estrogen and progesterone were co-administered and was inhibited by the pretreatment of the progesterone receptor antagonist RU486. These findings suggest that the expression of CD73 is dynamically regulated by estrogen and progesterone in the uterine environment, and that there may be a synergistic effect of estrogen and progesterone.

Chronic exposure to low concentrations of chlorpyrifos affects normal cyclicity and histology of the uterus in female rats.

Chlorpyrifos (CPF), the most used insecticide in Argentina, can act as an endocrine disruptor at low doses. We previously demonstrated that chronic exposure to CPF induces hormonal imbalance in vivo. The aim of this work was to study the effects of low concentrations of CPF (0.01 and 1 mg/kg/day) on the reproductive system of virgin adult rats. In the ovary, we studied the effects of CPF on steroidogenesis by determining steroid hormone content by RIA and CYP11 and CYP19 enzyme expression by qRT-PCR. The estrous cycle was evaluated by microscopic observation of vaginal smear, as well as by changes in uterine histology. In endometrium, we determined the fractal dimension and expression of PCNA, ERα and PR by IHC. Our results showed that chronic exposure to CPF affects ovarian steroid synthesis, causing alterations in the normal cyclicity of animals. In addition, CPF induced proliferative changes in the uterus, suggesting that it could affect reproduction or act as a risk factor in the development of uterine proliferative pathologies.

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.

Estrous cycle modulates the anxiogenic effects of caffeine in the elevated plus maze and light/dark box in female rats.

Caffeine is a commonly used stimulant of the central nervous system that reduces fatigue, increases alertness, and exerts positive effects on emotion through actions on various brain structures. High doses of caffeine can cause headaches, heart palpitations, hyperactivity, and anxiety symptoms. Consequently, reducing the consumption of stimulant substances, such as sugar and caffeine, is proposed to ameliorate symptoms of premenstrual syndrome in women. The administration of steroid hormones has been suggested to modulate the effects of caffeine, but unknown is whether endogenous hormone variations during the estrous cycle modulate the pharmacological effects of caffeine. The present study evaluated the effects of caffeine (10, 20, and 40 mg/kg) during metestrus-diestrus and proestrus-estrus of the ovarian cycle in rats on anxiety-like behavior using the elevated plus maze and light/dark box. During metestrus-diestrus, all doses of caffeine increased anxiety-like behavior, indicated by the main variables in both behavioral tests (i.e., higher Anxiety Index and lower percent time spent on the open arms in the elevated plus maze and less time spent in the light compartment in the light/dark box). During proestrus-estrus, only 20 and 40 mg/kg caffeine increased these parameters of anxiety-like behavior, albeit only slightly. In conclusion, caffeine increased anxiety-like behaviors in metestrus-diestrus, with an attenuation of these effects of lower doses of caffeine in proestrus-estrus. These effects that were observed in metestrus-diestrus and proestrus-estrus may be associated with low and high concentrations of steroid hormones, respectively, that naturally occur during these phases of the ovarian cycle.