Unraveling female communication through scent marks in the Norway rat.

Chemical communication by females remains poorly understood, with most attention focused on female advertisement of sexual receptivity to males or mother-offspring communication. However, in social species, scents are likely to be important for mediating competition and cooperation between females determining individual reproductive success. Here, we explore chemical signaling by female laboratory rats (Rattus norvegicus) to test i) whether females target their deployment of scent information differentially according to their sexual receptivity and the genetic identity of both female and male conspecifics signaling in the local environment and ii) whether females are attracted to gain the same or different information from female scents compared to males. Consistent with targeting of scent information to colony members of similar genetic background, female rats increased scent marking in response to scents from females of the same strain. Females also suppressed scent marking in response to male scent from a genetically foreign strain while sexually receptive. Proteomic analysis of female scent deposits revealed a complex protein profile, contributed from several sources but dominated by clitoral gland secretion. In particular, female scent marks contained a series of clitoral-derived hydrolases and proteolytically truncated major urinary proteins (MUPs). Manipulated blends of clitoral secretion and urine from estrus females were strongly attractive to both sexes, while voided urine alone stimulated no interest. Our study reveals that information about female receptive status is shared between females as well as with males, while clitoral secretions containing a complex set of truncated MUPs and other proteins play a key role in female communication.

The effects of diazepam on fear extinction in nulliparous and primiparous female rats.

Benzodiazepines undermine the success of exposure therapy in humans with anxiety disorders, and impair the long-term memory of fear extinction (the laboratory basis of exposure therapy) in rodents. However, most rodent studies on fear extinction and benzodiazepines have been conducted in male rodents. In female rodents, the estrous cycle influences the consolidation of fear extinction memories and sensitivity to benzodiazepines. In addition, pregnancy leads to long-term changes in the neurobiological, hormonal, and behavioural features of fear extinction, as well as the responsivity to benzodiazepines. Therefore, the present experiments examined the impact of benzodiazepines on fear extinction in female rats with and without reproductive experience. Age-matched nulliparous (no reproductive experience) and primiparous (one prior reproductive experience; tested one-month post-weaning) rats received fear conditioning to a discrete cue. The next day, rats were administered the benzodiazepine diazepam (2 mg/kg, s.c), or vehicle, prior to or immediately after extinction training. Rats were then tested the next day, drug free, for extinction retention. Similar to previous findings in males, diazepam impaired extinction retention in both nulliparous and primiparous rats when administered either pre- or post-extinction training. These findings may have potential clinical implications as they suggest that benzodiazepine use in conjunction with exposure therapy may undermine long-term treatment success in women with and without reproductive experience, although this remains to be tested in human populations. Moreover, these findings are theoretically important when considered in light of previous studies showing dissociable mechanisms of fear extinction in females pre- versus post-pregnancy.

Double perinatal stress reduces the sexual response of adult female Wistar rats.

BACKGROUND: Early-life stress affects physiological development and produces changes in various aspects of emotional behavior. AIM: We sought to examine the effects of double perinatal stress (DPS)-a combination of prenatal systemic hypoxic-ischemic (HI) insults and repeated early maternal separation-on the estrus cycle and sexual behavior of adult rats. METHODS: HI was induced by clamping the uterine arteries of pregnant rats for 45 minutes on the 18th day of gestation (HI group). Sham control animals received laparotomy and anesthesia only. Pups were born at term. Maternal separation was performed from postnatal day 1 (P1) (P0 = day of birth) to P15. At P90, the sexual response of females in estrus was evaluated. Statistical analysis was performed using 2-way analysis of variance followed by Tukey's test. OUTCOMES: We considered the estrous cycle and sexual behavior of female rats submitted to DPS, as well as the influence of female behavior on the sexual response of male rats. RESULTS: Rats submitted to DPS showed a reduction in the lordosis quotient and in the lordosis rate, suggesting a reduction in female sexual receptivity. DPS female rats showed a reduction in the number of hops and darts and in the genital exploration time rate, suggesting a reduction in sexual proceptivity. In addition, males that interacted with DPS females showed a reduction in the number of ejaculations and in copulatory efficiency. CLINICAL IMPLICATIONS: Developing a deeper understanding of perinatal factors that affect adult female sexual response will allow for more effective interventions to prevent and treat such changes. On the other hand, the analysis of the sexual response allows assessing the quality of life and the general state of health. STRENGTHS AND LIMITATIONS: The development of animal models to investigate the environmental factors that interfere in the female sexual response may allow researchers to propose and test new therapeutic strategies. On the other hand, care must be exercised when interpreting animal data and extrapolating these results to estimate the possible effects of perinatal stressors on the human sexual response. CONCLUSION: Our results revealed that females subjected to DPS showed long-term effects on sexual behavior. In conclusion, managing stressors in prenatal life and early postnatal life can prevent problems in adult sexual life and improve overall health.

Decamethylcyclopentasiloxane affects estradiol production in female rats but not H295R cells.

Sex hormones, such as androgens and estrogens, are predominantly produced in the gonads (ovaries and testes) and adrenal cortex. Endocrine-disrupting chemicals (EDCs) are substances that mimic, block, or interfere with hormones in the endocrine systems of humans and organisms. EDCs mainly act via nuclear receptors and steroidogenesis-related enzymes. In the OECD conceptual framework for testing and assessment of EDCs, several well-known assays are used to identify the potential disruption of nuclear receptors both in vivo and in vitro, whereas the H295R steroidogenesis assay is the only assay that detects the disruption of steroidogenesis. Forskolin and prochloraz are often used as positive controls in the H295R steroidogenesis assay. Decamethylcyclopentasiloxane (D5) was suspected one of EDCs, but the effects of D5 on steroidogenesis remain unclear. To establish a short-term in vivo screening method that detects the disruption of steroidogenesis, rats in the present study were fed a diet containing forskolin, prochloraz, or D5 for 14 days. Forskolin increased plasma levels of 17ß-estradiol (E2) and testosterone as well as the mRNA level of Cyp19 in both the adrenal glands and ovaries. Prochloraz induced the loss of cyclicity in the sexual cycle and decreased plasma levels of E2 and testosterone. D5 increased E2 levels and shortened the estrous cycle in a dose-dependent manner; however, potential endocrine disruption was not detected in the H295R steroidogenesis assay. These results demonstrate the importance of comprehensively assessing the endocrine-disrupting effects of chemicals on steroidogenesis in vivo.

Molecular characterization of physis tissue and hormonal profiles of female rats neonatally exposed to low-dose bisphenol A.

Physis is a complex cartilaginous structure that is critical for longitudinal bone growth. As one of the endocrine-disrupting chemicals, bisphenol A (BPA) can interfere with the physis by deranging the complex networks of nutritional, cellular, paracrine, and endocrine factors, and this affects longitudinal bone growth, leading to different growth outcomes. However, the exact mechanisms underlying these phenomena remain unclear. Therefore, understanding the molecular pathways involved in the physis after neonatal exposure to low-dose BPA may permit the identification of research targets for therapeutics, which may aid in modulating the process of growth plate closure. In the present study, female Sprague-Dawley rats were exposed to 0.05 mg·kg-1·day-1 of BPA and corn oil vehicle from postnatal day 1 (PND1) to 15 via subcutaneous injection. Next-generation RNA sequencing was performed for the mRNA isolated from the physis. The levels of osteocalcin (OC), growth hormone (GH), and insulin-like growth factor 1 (IGF-1) were measured on PND30 (BPA0.05mg vs. Vehicle; n = 5 for each group). We observed statistically significant enrichment of gene sets in the BPA0.05mg tissues compared with the Vehicle tissues. Further analysis of the differentially expressed genes (DEGs) identified BPA0.05mg-specific networks of secreted proteins (LEP, NPY, AGT, ACE2, C4B, and C4BPA) as well as those of local matrix and protease proteins (FBN2, LAMC2, ADAMTS16, and ADAMTS19). Furthermore, the levels of OC and GH were affected by BPA exposure. Our results revealed the specific molecular characteristics of physis contaminated with BPA and may provide new clues for physis maturation and supervision of industrial products and occupational exposure.

Comparison of Steroidogenic and Ovulation-Inducing Effects of Orthosteric and Allosteric Agonists of Luteinizing Hormone/Chorionic Gonadotropin Receptor in Immature Female Rats.

Gonadotropins, including human chorionic gonadotropin (hCG), are used to induce ovulation, but they have a number of side effects, including ovarian hyperstimulation syndrome (OHSS). A possible alternative is allosteric luteinizing hormone (LH)/hCG receptor agonists, including the compound TP4/2 we developed, which remains active when administered orally. The aim was to study the effectiveness of TP4/2 (orally, 40 mg/kg) as an ovulation inducer in FSH-stimulated immature female rats, compared with hCG (s.c., 15 IU/rat). TP4/2 stimulated progesterone production and corpus luteum formation; time-dependently increased the ovarian expression of steroidogenic genes (Star, Cyp11a1, Cyp17a1) and genes involved in ovulation regulation (Adamts-1, Cox-2, Egr-1, Mt-1); and increased the content of metalloproteinase ADAMTS-1 in the ovaries. These effects were similar to those of hCG, although in some cases they were less pronounced. TP4/2, in contrast to hCG, maintained normal LH levels and increased the ovarian expression of the LH/hCG receptor gene, indicating preservation of ovarian sensitivity to LH, and did not cause a sustained increase in expression of vascular endothelial growth factor-A involved in OHSS. Thus, TP4/2 is an effective ovulation inducer that, unlike hCG, has a lower risk of OHSS and ovarian LH resistance due to its moderate stimulating effect on steroidogenesis.

Reduced sexual motivation of diabetic female rats: Restoration with insulin.

The aim of this study was to evaluate female rat sexual motivation in a model of diabetes mellitus type 1. Severe hyperglycemia was induced in ovariectomized Wistar rats by injecting streptozotocin [STZ, 100 mg/kg, i.p.]. Ten days later, females received estradiol benzoate (10 µg/rat, s.c.) plus progesterone (3 mg/rat, s.c.). A group of STZ-treated animals was administered with insulin (2-4 U) every 12 h for 10 days, which normalized glucose levels. In the partner preference (PP) and sexual incentive motivation (SIM) tests, control females spent more time close to a sexually experienced male (SE) than with a castrated male (CM). STZ-treated females stayed the same amount of time with both stimuli, that is, they lost their sexual preference. We also evaluated the sense of smell using two behavioral tests, one related to sexual odors (SO) and another one to food odors (FO). In the SO test, control females spent more time sniffing the sawdust coming from cages that contained SE males; hyperglycemic females remained the same amount of time sniffing the sawdust of both stimuli: SE and CM. In the FO test, no differences were found between control and STZ-treated groups. Insulin treatment reverted the changes observed in hyperglycemic females in the PP, SIM and SO tests. These data suggest that severe hyperglycemia decreases sexual motivation and that insulin recovers such diminution.

Increase in sexual motivation throughout adolescence in the cycling female rat.

Sexual behavior in the female rat is a highly motivated behavior first displayed during adolescence, a developmental period when neural circuits underlying motivation are not mature. This study characterizes the natural development of sexual motivation and behavior of female rats. We compared the incentive value of the male for mid-adolescent (PNDs:39-43), late adolescent (PNDs:49-53), and adult (PNDs:90-115) cycling females, using a male-female preference task and an ultrasonic vocalization emission test following exposure to a male or female stimulus animal. Furthermore, display of sexual and social behaviors during an interaction with a male or a nonreceptive female was assessed. Mid-adolescent rats exhibited a reduced preference for the male than adults and performed less attempts to access the male. Unlike late adolescent and adult females, mid-adolescent rats did not increase their ultrasonic vocalization emission after interacting with a male relative to a female. Although most of the sexual behavior did not differ between groups, mid-adolescent females showed lower lordosis magnitude and higher levels of play and social investigation during a sexual interaction, giving rise to a unique behavioral profile. Present results indicate that the sexual behavior repertoire is fully displayed by mid-adolescence, but sexual motivation is low and increases into late adolescence.

Effect of genistein on the gene and protein expressions of CXCL-12 and EGR-1 in the rat ovary.

The effect of genistein (GEN) on the gene expression level of stromal cell-derived factor-1/CXCL-12 and early growth response gene-1 was studied in ovarian tissue of young and initially ageing (early stages in the ageing process) female rats. Forty, young female Sprague Dawley (SD) rats of 2-3 months old (200 ±20 g) and forty, initially ageing female SD rats of 10-12 months (490 ± 20 g) old were selected. According to the weight, rats were divided into control group, low-dose group (L), medium-dose group (M) and a high-dose group (H) and were given 15, 30 and 60 mg/kg GEN respectively. The positive control (Oestrogen) group was given 0.5 mg/kg diethylstilbestrol. The treatment lasted for 30 days. The mRNA expression of C-X-C motif chemokine ligand 12 (CXCL-12) and early growth response factor-1 (EGR-1) was measured by real-time PCR, and protein expression of EGR-1 was detected by Western blot. When compared to the negative control group (NC), the ovary/body weight ratio in the young rats decreased in the GEN group, but the difference was not significant. Similarly, compared with NC, the ovary/body weight ratio in the initially ageing rats also decreased with the increase in GEN concentration, but the decrease was significant in M and H groups (p < .01). The administration of GEN enhanced both the gene and protein expression levels of CXCL-12 and EGR-1 in the ovary. Pearson's correlation analysis showed a synergistic effect between CXCL-12 and EGR-1. Thus, we conclude that the effect of GEN on CXCL-12 and EGR-1 in the initially ageing group was obvious than that in the younger group.

[Effects of chronic stress during pregnancy on composition and diversity of intestinal microbiota in female rats and offspring].

Objective: To investigate the effect of chronic stress of pregnant rats on the gut microbiota of female rats and offspring, and explore the role of intestinal microbiota in chronic stress during pregnancy. Methods: In November 2019, SPF-grade healthy adult SD rats were selected. 16 female rats were randomly divided into control group and model group, with 8 in each group; 12 male rats were randomly divided into model mating group (8) and control mating group (4) . A model of chronic unpredictable mild stress (CUMS) during pregnancy was established. Blood samples were collected from the iliac vein of the female rats 1 day before and 1, 7, and 14 days after the CUMS protocol, and measured for plasma corticosterone content by radioimmunoassay. After the stress was completed, fresh feces of the female rats were collected for testing. The offspring's fresh stool samples were collected on postnatal day 20 (PND20) , and they were divided into control offspring group and model offspring group samples. The sequence of 16S rRNAV3-V4 regions of microorganisms in the feces of offspring was determined by Illumina MiSeq technique; and the interaction between microbial community structure and diversity were analyzed. Results: The content of plasma corticosterone in the model group was higher than that in the control group on the 7th and 14th day of stress (P<0.05) . Compared with the control group, the Sobs index, Chao index, ACE index and Shannon index of the model group were decreased (P<0.05) . The number of unique species abundance (OTU) in the control group was 130, and 91 in the model group. The relative abundance of female Firmicutes in the control group (64.87%) was higher than that in the model group, and the relative abundance of Bacteroides (31.72%) was lower than that of the model group (46.35%) . The Sobs index, Chao index, ACE index, Simpson index and Shannon index of the control offspring group were higher than those of the model offspring group (P<0.05) . The number of unique OTUs in the model offspring group was 75, and 93 in the control offspring group. The relative abundance of Firmicutes (60.24%) in the control offspring group was higher than that of the model offspring group (52.95%) . Conclusion: Chronic stress during pregnancy can not only lead to the disorder of intestinal flora in female rats, but also lead to the change of intrauterine environment, thus affecting the diversity of intestinal flora in offspring.

Neonatal exposure to bisphenol A advances pubertal development in female rats.

Neonatal exposure to bisphenol A (BPA) is hypothesized to advance pubertal development. However, the effects of neonatal BPA exposure on pubertal development has not been described. In this study, female Sprague-Dawley rats were exposed to 0.05, 0.5, 5, or 10 mg·kg-1 ·day-1 BPA, or corn oil vehicle alone from postnatal day 1 (PND1) to PND10 via subcutaneous injection. We evaluated day of vaginal opening (DVO), ovarian morphology, serum hormone concentrations, and hypothalamic expression of Gnrh1 and Kiss1 in female rats at PND35. DVO was significantly advanced in rats exposed to 5 and 10 mg·kg-1 ·day-1 BPA. Serum hormone concentrations increased as BPA dose increased. Additionally, hypothalamic Gnrh1 and Kiss1 expression were increased with BPA exposure; rats exposed to 10 mg·kg-1 ·day-1 BPA had significantly upregulated hypothalamic Gnrh1 and Kiss1 expressions in terms of both messenger RNA and protein levels. Our results suggest that exposure to a 10 mg·kg-1 ·day-1 dose of BPA might advance pubertal development significantly. In addition, within the range of 0 to 10 mg·kg-1 ·day-1 , neonatal exposure to BPA may affect pubertal development in a dose-dependent manner.

Mirtazapine reduces the expression of cocaine-induced locomotor sensitization in male and female Wistar rats.

BACKGROUND: Epidemiological studies have described that women are more vulnerable to the reinforcing effects of cocaine. In animals, the findings are similar: female rats show higher levels of cocaine self-administration and increased cocaine-induced locomotor activity. In contrast, women with depression respond better to treatment with antidepressants, however their therapeutic response to tetracyclic antidepressants is lower. Several studies have shown that mirtazapine-a tetracyclic antidepressant-decreases the behavioral effects of cocaine in male rats. The objective of this study was to evaluate the efficacy of daily dosing of mirtazapine on cocaine-induced locomotor activity and sensitization in naive female rats compared to male rats. METHODS: Male and female Wistar rats were daily dosed with 10 mg/kg of cocaine. During extinction, cocaine was withdrawn and the groups received daily mirtazapine (30 mg/kg, i.p.) or saline. Tamoxifen was administered during the antagonism phase. After each administration, locomotor activity for each animal was recorded for 30 min in transparent Plexiglass activity chambers. RESULTS: In this study, a higher cocaine locomotor response was found in females than in males and the mirtazapine was equally effective in decreasing cocaine-induced locomotor activity and the expression of locomotor sensitization in male and female rats. In addition, co-administration of mirtazapine and tamoxifen enhanced the efficacy of mirtazapine in female rats. CONCLUSION: The results suggest that mirtazapine may be considered an effective therapeutic option for the treatment of cocaine use disorder in men and women.

The mycotoxin deoxynivalenol activates GABAergic neurons in the reward system and inhibits feeding and maternal behaviours.

Deoxynivalenol (DON) or vomitoxin, is a trichothecene mycotoxin produced mainly by Fusarium graminearum and culmorum. Mycotoxins or secondary metabolic products of mold fungi are micro-pollutants, which may affect human and animal health. The neuronal and behavioural actions of DON were analysed in the present study. To address, which neurons can be affected by DON, the neuronal activation pattern following intraperitoneal injection of DON (1 mg/kg) was investigated in adult male rats and the results were confirmed in mice, too. DON-induced neuronal activation was assessed by c-Fos immunohistochemistry. DON injection resulted in profound c-Fos activation in only the elements of the reward system, such as the accumbens nucleus, the medial prefrontal cortex, and the ventral tegmental area. Further double labelling studies suggested that GABAergic neurons were activated by DON treatment. To study the behavioural relevance of this activation, we examined the effect of DON on feed intake as an example of reward-driven behaviours. Following DON injection, feed consumption was markedly reduced but returned to normal the following day suggesting an inhibitory action of DON on feed intake without forming taste-aversion. To further test how general the effect of DON on goal-directed behaviours is, its actions on maternal behaviour was also examined. Pup retrieval latencies were markedly increased by DON administration, and DON-treated mother rats spent less time with nursing suggesting reduced maternal motivation. In a supplementary control experiment, DON did not induce conditioned place preference arguing against its addictive or aversive actions. The results imply that acute uptake of the mycotoxin DON can influence the reward circuit of the brain and exert inhibitory actions on goal-directed, reward-driven behaviours. In addition, the results also suggest that DON exposure of mothers may have specific implications.

GPR30 receptor promotes preoperative anxiety-induced postoperative hyperalgesia by up-regulating GABAA-α4ß1δ subunits in periaqueductal gray in female rats.

BACKGROUND: G-protein coupled estrogen receptor 30 (GPR30) was proved the specific estrogen receptor relating to mechanical hyperalgesia. Studies have shown that the GABAA receptor subunits α4, ß1, and δ in the periaqueductal gray (PAG) neurons promote the descending facilitation system. This study inquired into whether and how GPR30 and GABAA-α4ß1δ in the PAG promote preoperative anxiety-induced postoperative hyperalgesia in female rats. METHODS: All the female rats were subjected to the single prolonged stress (SPS) to stimulate preoperative anxiety. Subsequently, mechanical allodynia was evaluated before and after the incision, based on the paw withdrawal mechanical threshold (PWMT). The selective GPR30 agonist G1 and antagonist G15 were locally microinjected into the PAG. The expression of GPR30, protein kinase A (PKA), and GABAA receptor subunits α4, ß1, and δ in the PAG neurons were detected using western blotting and immunofluorescence. RESULTS: Behavioral testing revealed that Group S and Group I decreased the nociceptive threshold levels of PWMT in female rats. PWMT in Group S + I decreased more than that of Group S and Group I. Further, results of western blotting showed the expression of GPR30, PKA, and GABAA α4, ß1, and δ subunits significantly up-regulated in Group S + I, and immunofluorescence indicated that the neurons of PAG in Group S + I appeared simultaneously immunopositive for GPR30 and GABAA α4, ß1, and δ receptors. After microinjection of G1 into the PAG, female rats with plantar incision continued to exhibit significant hyperalgesia until postoperative 48 h. On the other hand, microinjection of G15 with SPS and plantar incision procedure relieved postoperative hyperalgesia in female rats. Western blotting demonstrated that intra-PAG injection of G15 markedly decreased the GPR30, PKA, and GABAA α4, ß1, and δ levels in Group G15 + I. CONCLUSIONS: Our results indicate that the GPR30-PKA-GABAAα4ß1δ pathway in the PAG promotes preoperative anxiety-induced postoperative hyperalgesia in female rats. This mechanism might be a potential novel therapeutic target for hyperalgesia in females.

Does Chronic Hyperglycemia Affect Female Rat Sexual Behavior? Differences in Paced and Non-Paced Mating.

INTRODUCTION: Diabetes mellitus has been associated with sexual dysfunction; however, in women this relationship is controversial. A study using a model of type 2 diabetes mellitus (DM2) failed to find a reduced receptivity in the non-paced mating (NPM), but the appetitive aspects of female sexual behavior have not been evaluated, for example, in the paced mating (PM) paradigm. AIM: To evaluate all components of female sexual behavior (in NPM and PM) in a model of DM2 using ovariectomized (OVX) (treated with steroids) or intact female rats (non-OVX) in natural proestrus. METHODS: Neonatal females (3-4 days) were administered streptozotocin (STZ, 70 mg/kg, intraperitoneally) or citrate buffer. At week 8, a glucose tolerance test was performed. At week 10, half of the females were OVX, and in the other half (non-OVX) the estrous cycle was monitored. At the twelfth week, the sexual behavior tests were conducted; OVX females were treated with estradiol benzoate (10 µg, -24 hours) and progesterone (3 mg, -4 hours), whereas the non-OVX were evaluated on vaginal proestrus. MAIN OUTCOME MEASURES: We registered in NPM and PM receptivity (lordosis quotient and intensity), as well as the number of proceptive and aggressive behaviors. Additionally, in PM we calculated the percentage of exits and the return latencies after receiving stimulation and the time the female remained in the male's compartment. RESULTS: The STZ-treated females presented glucose intolerance and were hyperglycemic. Neonatal STZ treatment provoked changes in the females' sexual behavior depending on the paradigm and the hormonal condition. In the NPM, STZ-OVX females had decreased lordosis quotient and intensity and increased aggression, whereas, in the STZ-non-OVX females, there was a decrease in proceptivity; such changes were not observed in PM. Regardless of whether the STZ-treated females were OVX, they failed to perform the pacing behavior. CLINICAL IMPLICATION: These data support the idea that chronic mild hyperglycemia, like that observed in DM2 (which represents 90% of the clinical cases), provokes marginal changes in most aspects of female sexual behavior. STRENGTHS & LIMITATIONS: The main strength of this work is the evaluation of consummatory and motivational aspects of female sexual behavior in a model of DM2. The main limitation is the duration of the experimental design that does not resemble the course of the disease in humans. No histologic or biochemical analyses were performed. CONCLUSION: These results suggest that chronic hyperglycemia produces decreases in sexual behavior. Hernández-Munive AK, Rebolledo-Solleiro D, Fernández-Guasti A. Does Chronic Hyperglycemia Affect Female Rat Sexual Behavior? Differences in Paced and Non-Paced Mating. J Sex Med 2019;16:1130-1142.

Effect of genistein on IGF-1 and IGFBP-1 in young and aged female rat ovary.

The objective of this study was to assess the effects of genistein (GEN) on expression of insulin-like growth factor 1 (IGF-1) and insulin-like growth factor binding protein 1 (IGFBP-1) in young and aged rat ovary. Forty young female Sprague Dawley (SD) rats (200 ± 20 g) and forty aged female SD rats (490 ± 20 g) were selected and according to weight, they were divided into the following five groups with eight animals in each: negative control group (NC), low-dose group (L), middle-dose group (M), high-dose group (H) and positive control group (PC). GEN group received GEN of 15, 30, 60 mg/kg respectively. It lasted 30 days. Concentrations of serum hormones, IGF-1 and IGFBP-1 were determined by enzyme-linked immunosorbent assay (ELISA). Gene and protein expressions of IGF-1 and IGFBP-1 were determined by real-time PCR and Western blot respectively. Compared with NC, GEN significantly increased oestradiol-17ß(E2 ) level in aged rat, reduced luteinizing hormone (LH) level in young and aged rat. Serum levels of IGFBP-1 in young rats were significantly higher in GEN groups (p < 0.05). mRNA and protein expression levels of IGF-1 and IGFBP-1 were positively correlated with GEN dose. GEN could significantly reduce the ratio of IGF-1/IGFBP-1 of aged rats. Multivariate Cox regression analysis result showed IGF-1 and IGFBP-1 levels significantly correlated with GEN dose. We speculate that there is an association between the addition of GEN and expression of IGF-1 and IGFBP-1, and the relationship between them is different in young and aged rat.

Mechanistic effect of human umbilical cord blood derived mesenchymal stem cells on the submandibular salivary gland in ovariectomized rats.

We performed this study to understand the effect of human umbilical cord blood derived mesenchymal stem cells (hUCB-MSCs) on the submandibular gland after bilateral ovariectomy. For this, 21 adult female rats were distributed equally among 3 groups: the sham-operated group (SHAM); the ovariectomized group (OVX); and the OVX group that received repeated intravenous injections of the hUCB-MSCs (OVX + hUCB-MSCs). We used reverse transcription - PCR to analyze for the gene expression of AQPs 3, 4, 5, and BMP-6. The cellular localization and expression of human CD105, human CD34, proliferating nuclear antigen (PCNA), single-stranded DNA (ss-DNA), caspase 3, AQP1, and α smooth muscle actin (α-SMA) were determined immunohistochemically. In the OVX group, a significant decrease in the gene expression of AQP3, AQP4, and BMP6, as well as the acinar area % was detected, while area % of granular convoluted tubules (GCTs) showed a significant increase. A significant decrease in area % staining positively for AQP1 and α-SMA was noted. An obvious improvement in the structure of the submandibular gland was demonstrated in the group injected with hUCB-MSCs, as well as a significant increase in the gene expression of AQP3, AQP4, and BMP6. The acinar and GCT area %, as well as the different measured markers, were relatively normal. This demonstrates that E2-deficiency induces structural changes to the submandibular gland. Moreover, a definite amelioration of the structure and function of the submandibular gland was detected after the administration of hUCB-MSCs.

Western diet triggers Toll-like receptor 4 signaling-induced endothelial dysfunction in female Wistar rats.

Overconsumption of a diet rich in fat and carbohydrates, called the Western diet, is a major contributor to the global epidemic of cardiovascular disease. Despite previously documented cardiovascular protection exhibited in female rats, this safeguard may be lost under certain metabolic stressors. We hypothesized that female Wistar rats challenged by a Western diet composed of 21% fat and 50% carbohydrate (34.1% sucrose) for 17 wk would develop endothelial dysfunction via endothelial Toll-like receptor 4 (TLR4) signaling. Western diet-fed female rats exhibited dysregulation of metabolism, revealing increased body weight and abdominal fat, decreased expression of adiponectin in white adipose tissue, glucose intolerance, and impaired insulin sensitivity. Western diet exposure increased hepatic triglycerides and cholesterol alongside hepatic steatosis, categorizing nonalcoholic fatty liver disease. Moreover, a Western diet negatively affected vascular function, revealing hypertension, impaired endothelium-dependent vasorelaxation, aortic remodeling, and increased reactive oxygen species (ROS) production. Aortic protein expression of TLR4 and its downstream proteins were markedly increased in the Western diet-fed group in association with elevated serum levels of free fatty acids. In vitro experiments were conducted to test whether free fatty acids contribute to vascular ROS overproduction via the TLR4 signaling pathway. Cultured endothelial cells were stimulated with palmitate in the presence of TAK-242, a TLR4 signaling inhibitor. Palmitate-induced overgeneration of ROS in endothelial cells was abolished in the presence of TAK-242. Our data show that a Western diet induced endothelial dysfunction in female rats and suggest that endothelial TLR4 signaling may play a key role in abolishing female cardiovascular protection. NEW & NOTEWORTHY A Western diet induced elevated levels of free fatty acids, produced nonalcoholic fatty liver disease, and provoked endothelial dysfunction in female rats in association with Toll-like receptor 4 signaling-mediated vascular reactive oxygen species production. Limited consumption of a Western diet in premenopausal women may decrease their risk of cardiovascular complications.

Zinc Alleviates Lipopolysaccharide Interference with Both Body Temperature and Sickness Behavior in Virgin Female Rats.

OBJECTIVES: Previous studies from our group showed that lipopolysaccharide (LPS) exposure induces several signs of sickness behavior, including a decrease in food consumption, body weight gain, adipsia, and a biphasic effect in tympanic temperature with a first phase of hypothermia, followed by an increased tympanic temperature. LPS can activate a chain of nonspecific host responses, including the immune response, and decreased zinc levels. In addition, there are differences in the immune response between males and females, particularly fever, with sex hormones interfering with body temperature. This study aims to characterize the effects of zinc treatment on tympanic temperature, body weight gain, food and water consumption, and general activity in open field of virgin female rats exposed to a dose of LPS that was previously reported to induce sickness behavior. METHODS: Virgin female Wistar rats were treated with either saline (S) or LPS. One hour later, the S group received another injection of saline (S + S group), half of the LPS group received saline (LPS + S group) and the other half received zinc (LPS + Zn group). Tympanic temperature, body weight, and water and food consumption were measured for 96 h. Measurements and observations started 2 h after LPS administration. RESULTS: Treatment with zinc attenuated LPS-increased temperature, decreased the body weight gain and food consumption, and water consumption was increased. CONCLUSION: Zinc treatment is beneficial as it reduces the increased tympanic temperature induced by LPS, but it does not influence other sickness behavior caused by exposure to LPS.

Effects of 900-MHz electromagnetic fields exposure throughout middle/late adolescence on the kidney morphology and biochemistry of the female rat.

The effect of the electromagnetic field (EMF) established when cell phones are in use on human health, and particularly the head, has been the subject of major scientific research. Phones are usually carried near the lumbar region when not in use, and the kidneys will also inevitably be affected by such fields. We investigated the effects on the kidneys of female rats exposed to a continuous 900-megahertz (MHz) EMF for 1 h daily in mid-late adolescence. Control, sham, and EMF groups were established. The EMF was applied to the application group rats daily on postnatal days 35-59. A pseudo-megahertz effect was applied to sham group rats. All animals were euthanized on postnatal day 60. Right kidney tissues were subjected to routine procedures. Malondialdehyde, total antioxidant status, and total oxidant status (TOS) were investigated in left kidneys, and the oxidative stress index (OSI) was also calculated from these. Histopathological analysis revealed no pathology in either the control or sham groups. However, findings including hemorrhage in glomerulus, vacuolization and irregularity in the proximal and distal tubular epithelium, diffuse glomerular degeneration and edema, occasional degeneration in Bowman capsules, hemorrhage in the medullary region, disturbed nucleus location and morphology, and tubular edema in the cortex were observed in the EMF groups. TOS and OSI values were lower in the EMF group (9.4316 ± 1.0211 and 0.8461 ± 0.0826, respectively) and the sham group (8.2171 ± 0.6437 and 0.7358 ± 0.0545, respectively) than in the control group (11.1522 ± 1.3389 and 1.0085 ± 0.1174, respectively) ( p < 0.05). In conclusion, exposure to a continuous 900-MHz EMF for 1 h daily during middle and late adolescence may cause various changes in the female rat kidney at postnatal day 60.