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International Journal of Molecular... Aug 2020The orofacial pain pathway projects to the parabrachial and amygdala, and sex steroids have been shown to affect neuronal activity in these regions. GABA positive cells... (Comparative Study)
Comparative Study
The orofacial pain pathway projects to the parabrachial and amygdala, and sex steroids have been shown to affect neuronal activity in these regions. GABA positive cells in the amygdala are influenced by sex steroid metabolites to affect pain, and sex steroids have been shown to alter the expression of genes in the parabrachial, changing neuronal excitability. Mechanisms by which sex steroids affect amygdala and parabrachial signaling are unclear. The expression of genes in the parabrachial and amygdala in diestrus (low estradiol) and proestrus (high estradiol) female rats were evaluated in this study. First, varicella zoster virus was injected into the whisker pad of female rats to induce a pain response. Second, gene expression was quantitated using RNA-seq one week after injection. Genes that had the greatest change in expression and known to function in pain signaling were selected for the quantitation of protein content. Protein expression of four genes in the parabrachial and seven genes in the amygdala were quantitated by ELISA. In the parabrachial, neurexin 3 (Nrnx3) was elevated at proestrus. Nrnx3 has a role in AMPA receptor and GABA signaling. Neuronatin (Nnat) and protein phosphatase, Mg/Mn dependent 1E (Ppm1e) were elevated in the parabrachial of diestrus animals both genes having a role in pain signaling. Epoxide hydroxylase (Ephx2) was elevated in the parabrachial at proestrus and the vitamin D receptor (Vdr) was elevated in the amygdala. Ephx2 antagonists and vitamin D have been used to treat neuropathic pain. In conclusion, sex steroids regulate genes in the parabrachial and amygdala that might result in the greater pain response observed during diestrus.
Topics: Amygdala; Animals; Diestrus; Epoxide Hydrolases; Female; Gene Expression Regulation; Herpesvirus 3, Human; Injections; Nerve Tissue Proteins; Neuralgia; Proestrus; RNA, Messenger; Rats; Receptors, Calcitriol
PubMed: 32796585
DOI: 10.3390/ijms21165749 -
Animals : An Open Access Journal From... Jan 2023Myeloperoxidase (MPO), as a marker of neutrophil activation, has been associated with equine endometritis. However, in absence of inflammation, MPO is constantly...
Myeloperoxidase (MPO), as a marker of neutrophil activation, has been associated with equine endometritis. However, in absence of inflammation, MPO is constantly detected in the uterine lumen of estrous mares. The aim of this study was to characterize MPO in the uterus of mares under physiological conditions as a first step to better understand the role of this enzyme in equine reproduction. Total and active MPO concentrations were determined, by ELISA and SIEFED assay, respectively, in low-volume lavages from mares in estrus ( = 26), diestrus ( = 18) and anestrus ( = 8) in absence of endometritis. Immunohistochemical analysis was performed on 21 endometrial biopsies randomly selected: estrus ( = 11), diestrus ( = 6) and anestrus ( = 4). MPO, although mostly enzymatically inactive, was present in highly variable concentrations in uterine lavages in all studied phases, with elevated concentrations in estrus and anestrus, while in diestrus, concentrations were much lower. Intracytoplasmic immunoexpression of MPO was detected in the endometrial epithelial cells, neutrophils and glandular secretions. Maximal expression was observed during estrus in mid and basal glands with a predominant intracytoplasmic apical reinforcement. In diestrus, immunopositive glands were sporadic. In anestrus, only the luminal epithelium showed residual MPO immunostaining. These results confirm a constant presence of MPO in the uterine lumen of mares in absence of inflammation, probably as part of the uterine mucosal immune system, and suggest that endometrial cells are a source of uterine MPO under physiological cyclic conditions.
PubMed: 36766264
DOI: 10.3390/ani13030375 -
Biochemical Pharmacology Apr 2022Opioids, and numerous centrally active drugs, are metabolized by cytochrome P450 2D (CYP2D). There are sex and estrous cycle differences in brain oxycodone analgesia....
Opioids, and numerous centrally active drugs, are metabolized by cytochrome P450 2D (CYP2D). There are sex and estrous cycle differences in brain oxycodone analgesia. Here we investigated the mechanism examining the selective role of CYP2D in the brain on sex, estrous cycle, and hormonal regulation. Propranolol, CYP2D-specific mechanism-based inhibitor, or vehicle was delivered into cerebral ventricles 24 h before administering oxycodone (or oxymorphone, negative control) orally to male and female (in estrus and diestrus) rats. Ovariectomized and sham-operated females received no treatment, estradiol, progesterone or vehicle. Analgesia was measured using tail-flick latency, and brain drug and metabolite concentrations were measured by microdialysis. Data were analyzed by two-way or mixed ANOVA. Following propranolol (versus vehicle) inhibition and oral oxycodone, there were greater increases in brain oxycodone concentrations and analgesia, and greater decreases in brain oxymorphone/oxycodone ratios (an in vivo phenotype of CYP2D in brain) in males and females in estrus, compared to females in diestrus; with no impact on plasma drug concentrations. There was no impact of propranolol pre-treatment, sex, or cycle after oral oxymorphone (non-CYP2D substrate) on brain oxymorphone concentrations or analgesia. There was no impact of propranolol pre-treatment following ovariectomy on brain oxycodone concentrations or analgesia, which was restored in ovariectomized females following estradiol, but not progesterone, treatment. Sex, cycle, and estradiol regulation of CYP2D in brain in turn altered brain oxycodone concentration and response, which may contribute to the large inter-individual variation in response to the numerous centrally acting CYP2D substrate drugs, including opioids.
Topics: Analgesia; Analgesics, Opioid; Animals; Brain; Cytochrome P-450 Enzyme System; Estradiol; Estrous Cycle; Female; Male; Oxycodone; Oxymorphone; Pain; Progesterone; Propranolol; Rats; Rats, Wistar
PubMed: 35143755
DOI: 10.1016/j.bcp.2022.114949 -
Molecular Neurobiology Dec 2021Sex differences in opioid analgesia occur in rodents and humans, and could be due to differences in drug and metabolite levels. Thus, we investigated the sex and cycle...
Sex differences in opioid analgesia occur in rodents and humans, and could be due to differences in drug and metabolite levels. Thus, we investigated the sex and cycle differences in analgesia (nociception) from oxycodone in rats and related these to sex and cycle differences in brain and plasma oxycodone and metabolite levels. Since numerous opioids are CYP2D enzyme substrates and variation in CYP2D alters opioid drug levels and response, we also initiated studies to see if the sex and cycle differences observed might be due to differences in brain CYP2D activity. Across oxycodone doses, females in diestrus had higher analgesia (using tail flick latency) compared to males and females in estrus; we also demonstrated a direct effect of estrous cycle on analgesia within females. Consistent with the analgesia, females in diestrus had highest brain oxycodone levels (assessed using microdialysis) compared to males and females in estrus. Analgesia correlated with brain oxycodone, but not brain oxymorphone or noroxycodone levels, or plasma drug or metabolite levels. Propranolol (a CYP2D mechanism-based inhibitor), versus vehicle pre-treatments, increased brain oxycodone, and decreased brain oxymorphone/oxycodone drug level ratios (an in vivo CYP2D activity phenotype in the brain) in males and females in estrus, but not in females in diestrus. Brain oxymorphone/oxycodone inversely correlated with analgesia. Together, both sex and estrous cycle impact oxycodone analgesia and brain oxycodone levels, likely through regulation of brain CYP2D oxycodone metabolism. As CYP2D6 is expressed in human brain, perhaps similar sex and cycle influences also occur in humans.
Topics: Analgesia; Analgesics, Opioid; Animals; Brain; Estrous Cycle; Female; Male; Oxycodone; Pain Measurement; Rats; Rats, Wistar; Sex Characteristics
PubMed: 34581987
DOI: 10.1007/s12035-021-02560-1 -
Frontiers in Veterinary Science 2022The canine corpus luteum (CL) is able to synthetise, activate and deactivate 17b-estradiol (E2) and also expresses nuclear estrogen receptors in a time-dependent manner...
The canine corpus luteum (CL) is able to synthetise, activate and deactivate 17b-estradiol (E2) and also expresses nuclear estrogen receptors in a time-dependent manner during diestrus. Nevertheless, we are still missing a better comprehension of E2 functions in the canine CL, especially regarding the specific roles of estrogen receptor alpha (ERa) and ERb, encoded by and , respectively. For that purpose, we analyzed transcriptomic data of canine non-pregnant CL collected on days 10, 20, 30, 40, 50 and 60 of diestrus and searched for differentially expressed genes (DEG) containing predicted transcription factor binding sites (TFBS) for or . Based on biological functions of DEG presenting TFBS, expression of select transcripts and corresponding proteins was assessed. Additionally, luteal cells were collected across specific time points during diestrus and specificity of E2 responses was tested using ERa and/or ERb inhibitors. Bioinformatic analyses revealed 517 DEGs containing TFBS, from which 67 for both receptors. In general, abundance of predicted targets was greater in the beginning, while abundance of targets was greater in the end of diestrus. / ratio shifted from an increasing to a decreasing pattern from day 30 to 40 post ovulation. Specific receptor inhibition suggested an ERa-mediated positive regulation of CL function at the beginning of diestrus and an ERb-mediated effect contributing to luteal regression. In conclusion, our data points toward a broad spectrum of action of E2 and its nuclear receptors, which can also act as transcription factors for other genes regulating canine CL function.
PubMed: 35982918
DOI: 10.3389/fvets.2022.885257 -
Frontiers in Veterinary Science 2020The vaginal microbiota plays an important role in the health of dairy cattle, and it could be manipulated for the prevention and treatment of reproduction-related...
The vaginal microbiota plays an important role in the health of dairy cattle, and it could be manipulated for the prevention and treatment of reproduction-related infections. The present study profiles and compares the vaginal microbiota of healthy dairy heifers during the estrous cycle focusing the results in follicular (estrus) and luteal (diestrus) phases using 16S rRNA sequencing of the V3-V4 hypervariable region. Twenty 13-16-months-old virgin dairy heifers from a single farm were included in this study. Vaginal swabs and blood samples were obtained during estrus (6-8 h before artificial insemination) and diestrus (14 days after insemination). Estrus was evaluated by an activity monitoring system and confirmed with plasma progesterone immunoassay. Results showed that the taxonomic composition of the vaginal microbiota was different during the follicular and luteal phases. At the phylum level, the most abundant bacterial phyla were Tenericutes, Firmicutes, and Bacteroidetes which comprised more than 75% of the vaginal microbiota composition. The next more abundant phyla, in order of decreasing abundance, were Proteobacteria, Actinobacteria, Fusobacteria, Epsilonbacteraeota, and Patescibacteria. Together with Tenericutes, Firmicutes, and Bacteroidetes represented more than 96% of the bacterial composition. , were the most abundant genera or families. The results also showed that the vaginal microbiota of dairy heifers was non-lactobacillus dominant. The genus was always found at a low relative abundance during the estrous cycle being more abundant in the follicular than in the luteal phase. Despite more research is needed to explore the potential use of native vaginal microbiota members as probiotics in dairy heifers, this study represents an important step forward. Understanding how the microbiota behaves in healthy heifers will help to identify vaginal dysbiosis related to disease.
PubMed: 32719814
DOI: 10.3389/fvets.2020.00371 -
Animal Reproduction Science Aug 2023Phoenixin-14 (PNX-14) is a regulatory neuropeptide encoded by the SMIM20 gene, which has been implicated in the reproductive cycle by modulating the...
Phoenixin-14 (PNX-14) is a regulatory neuropeptide encoded by the SMIM20 gene, which has been implicated in the reproductive cycle by modulating the hypothalamic-pituitary-gonadal (HPG) axis. Recently, we showed that PNX-14 is downregulated in bitches with cystic endometrial hyperplasia and pyometra. The objective of this study was to determine the expression of Smim20, PNX-14, and its putative receptor GRP173 in the canine ovary (both healthy and those with ovarian cysts), periovarian adipose tissue (PAT) and in the endometrium during the oestrous cycle. The expression was analysed by RT-qPCR and Western blot. In tissue sections, peptides were localised by immunofluorescent assays, and blood plasma concentrations of PNX-14 were detected by EIA. The results demonstrated increased levels of PNX in bitches in the anestrus groups compared to diestrus animals. The expression of GPR173 increased in PAT during the diestrus phase and endometrial tissue in late diestrus bitches. In the ovary, strong signals of PNX-14 and GPR173 were detected in the luteal and follicular cells. Furthermore, bitches with cystic ovaries were characterised by elevated circulating PNX levels and a significantly higher expression of PNX and GPR173 in gonadal tissues, when compared with healthy animals. Moreover, a positive correlation between PNX and progesterone in the blood of healthy bitches was noted, which changed to a negative correlation in females affected by cystic ovaries. These studies expand the knowledge regarding the expression and localization of the PNX/GRP173 system in canine reproductive organs during physiological and pathological conditions.
Topics: Female; Animals; Dogs; Neuropeptides; Peptides; Endometrial Hyperplasia; Endometrium; Adipose Tissue; Dog Diseases
PubMed: 37356348
DOI: 10.1016/j.anireprosci.2023.107282 -
Heliyon Dec 2022The medial preoptic (MPN) and the ventromedial hypothalamic nuclei (VMN) modulate the estrogen receptor (ER)-dependent female sexual behavior, a response that is...
The medial preoptic (MPN) and the ventromedial hypothalamic nuclei (VMN) modulate the estrogen receptor (ER)-dependent female sexual behavior, a response that is inhibited by tamoxifen (TAM), a modulator of the steroid receptor activation. With the objective to assess TAM action in the brain areas involved in the modulation sexual cues, an animal model on long-term TAM therapy to intact female rats, was used to mimic the 5-year prophylactic TAM therapy offered to women at higher risk of breast cancer. After three months treatment, female sexual behavior with a stud male rat was evaluated. Upon sacrifice, the brains were removed and the MPN and the ventrolateral division of the VMN were screened for the effects of TAM in the expression of ERα, ERβ and progesterone receptor. Results show that TAM inhibited the receptive component of the female sexual behavior. Even though TAM decreased estrogen and progesterone levels to values similar to the ones of estrous and diestrus rats, the biochemical data failed to demonstrate such possible causation for the behavioral response. In fact, TAM administration induced a constant low level of ovarian hormones that changed the pattern of ER and PR expression as well as receptor co-expression in the brain areas regulating the behavioral response, dissimilar to the ones seen in the cycle phases with the same low hormone levels. Nevertheless, present data suggests that by affecting ER- and/or PR-dependent mechanisms, TAM may modulate the hypothalamus, a region known to participate in several social behaviors.
PubMed: 36593822
DOI: 10.1016/j.heliyon.2022.e12362 -
Reproduction in Domestic Animals =... Oct 2022In this study, uterine blood flow area (BFA) has been evaluated for the first time using power Doppler ultrasound (PD) as a marker of endometritis in mares and jennies....
In this study, uterine blood flow area (BFA) has been evaluated for the first time using power Doppler ultrasound (PD) as a marker of endometritis in mares and jennies. The uterine BFA in healthy mares was greater in oestrus than in diestrus (p < .001). However, differences in endometrial blood flow between oestrus and diestrus were not observed in mares with endometritis. The uterine blood flow in healthy jennies is not affected by the oestrus cycle. Both species showed an increase in endometrial BFA in pathological uterine conditions compared to controls. BFA was a good marker of endometritis with an area under curve (AUC) (estrus:0.94 (p < .001) diestrus:0.98 (p < .001) in mares and AUC (0.91 (p < .0001) in jennies. The results of this preliminary study suggest that PD ultrasound in combination with computerized image analysis has the potential to be a very useful tool in the diagnosis of endometritis.
Topics: Animals; Endometritis; Endometrium; Equidae; Female; Horse Diseases; Horses; Uterus
PubMed: 35467047
DOI: 10.1111/rda.14132 -
Biology of Reproduction Jul 2021Zearalenone (ZEN), a nonsteroidal estrogenic mycotoxin, is detrimental to female reproduction. Altered chemical biotransformation, depleted primordial follicles and a...
Zearalenone (ZEN), a nonsteroidal estrogenic mycotoxin, is detrimental to female reproduction. Altered chemical biotransformation, depleted primordial follicles and a blunted genotoxicant response have been discovered in obese female ovaries, thus, this study investigated the hypothesis that obesity would enhance ovarian sensitivity to ZEN exposure. Seven-week-old female wild-type nonagouti KK.Cg-a/a mice (lean) and agouti lethal yellow KK.Cg-Ay/J mice (obese) received food and water ad libitum, and either saline or ZEN (40 μg/kg) per os for 15 days. Body and organ weights, and estrous cyclicity were recorded, and ovaries collected posteuthanasia for protein analysis. Body and liver weights were increased (P < 0.05) in the obese mice, but obesity did not affect (P > 0.05) heart, kidney, spleen, uterus, or ovary weight and there was no impact (P > 0.05) of ZEN exposure on body or organ weight in lean or obese mice. Obese mice had shorter proestrus (P < 0.05) and a tendency (P = 0.055) for longer metestrus/diestrus. ZEN exposure in obese mice increased estrus but shortened metestrus/diestrus length. Neither obesity nor ZEN exposure impacted (P > 0.05) circulating progesterone, or ovarian abundance of EPHX1, GSTP1, CYP2E1, ATM, BRCA1, DNMT1, HDAC1, H4K16ac, or H3K9me3. Lean mice exposed to ZEN had a minor increase in γH2AX abundance (P < 0.05). In lean and obese mice, LC-MS/MS identified alterations to proteins involved in chemical metabolism, DNA repair and reproduction. These data identify ZEN-induced adverse ovarian modes of action and suggest that obesity is additive to ZEN-induced ovotoxicity.
Topics: Animals; Estrogens, Non-Steroidal; Female; Mice; Mycotoxins; Ovary; Proteome; Zearalenone
PubMed: 33855340
DOI: 10.1093/biolre/ioab069