Glucocorticoid Receptor Gene Expression Research Articles

Patterns of corticosterone exposure affect the subcellular localisation of mineralocorticoid and glucocorticoid receptor complexes and gene expression

Mineralocorticoid (MR) and glucocorticoid receptors (GR) act as transcription factors and major mediators of glucocorticoid signalling, with pivotal roles in regulating the stress response and hormonal signalling, mood, cognition and memory. The MR and GR and share many target genes, have a high degree of homology in their DNA binding (DBD) and ligand binding domain (LBD) but differ considerably in the N-terminal domain (NTD). Using Proximity Ligation Assay (PLA) we quantitatively assessed MR-GR complex subcellular localisation and transcriptional regulation in murine neuroblastoma (N2A) cells stimulated by constant or pulsatile corticosterone (CORT) patterns. We observe that continuous receptor activation by CORT caused localisation at the periphery of the cell nucleus. Truncation of the receptor Ligand Binding Domain (LBD) led to a stronger localisation of MR-GR complexes at the periphery of the cell nuclei. This was also observed for GR immunofluorescence (IF), while in cells expressing only MR or GR the mRNA response to pulsatile hormone treatment was substantially attenuated. However, there was no clearcut correlation between the spatial distribution of MR-GR complexes and the mRNA levels of target genes. Overall, our findings suggest that longer presence in the cell nucleus favors more peripheral nuclear localisation.

Heat stress affects milk yield, milk quality, and gene expression profiles in mammary cells of Girolando cows

Heat stress during lactation affects the physiological responses, hormonal release, health, and productivity of dairy cows. However, the adverse effects of heat stress on milk synthesis, immune response, and cellular apoptosis in mammary cells remains unknown in Bos indicus cows. This study aimed to understand the relationship between milk yield, milk quality, and the expression of genes related to milk synthesis, cell apoptosis, and immune response in mammary cells of Girolando cows. Twenty-four Girolando cows (3/4 Holstein and 1/4 Gir) were subjected to control (CT, with a temperature-humidity index ranging from 60 to 74, n = 12) or heat stress treatments (HS, with a temperature- humidity index ranging from 60 to 85, n = 12), from 111 to 120 d of lactation. Heat stress significantly increased the expression of heat shock proteins (HSPD1 and HSPD90AA1), insulin receptors (INSR), and prolactin receptors (PRLRsf) genes, and decreased the expression of glucocorticoid receptor (NR3C1) gene in mammary cells of the HS cows when compared with the CT cows. The HS cows exhibited significantly higher vaginal temperatures and cortisol release compared with the CT cows. Moreover, the HS cows had significantly lower dry matter intake and milk yield than CT cows. Although, HS cows showed higher percentage of lymphocytes in milk when compared with that from CT cows. There was no effect of heat stress on other leukocyte counts, somatic cell counts, bacterial counts in milk, or milk composition. Finally, this study demonstrated that Girolando cows are susceptible to heat stress, which decreases milk yield and affects the expression of genes linked to milk synthesis in the mammary cells.

Effects of DNA Methylation of HPA-Axis Genes of F1 Juvenile Induced by Maternal Density Stress on Behavior and Immune Traits in Root Voles (Microtus oeconomus)-A Field Experiment.

The literature shows that maternal stress can influence behavior and immune function in F1. Yet, most studies on these are from the laboratory, and replicated studies on the mechanisms by which maternal stress drives individual characteristics are still not fully understood in wild animals. We manipulated high- and low-density parental population density using large-scale field enclosures and examined behavior and immune traits. Within the field enclosures, we assessed anti-keyhole limpet hemocyanin immunoglobulin G (anti-KLH IgG) level, phytohemagglutinin (PHA) responses, hematology, cytokines, the depressive and anxiety-like behaviors and prevalence and intensity of coccidial infection. We then collected brain tissue from juvenile voles born at high or low density, quantified mRNA and protein expression of corticotropin-releasing hormone (CRH) and glucocorticoid receptor gene (NR3C1) and measured DNA methylation at CpG sites in a region that was highly conserved with the prairie vole CRH and NR3C1 promoter. At high density, we found that the F1 had a lower DNA methylation level of CRH and a higher DNA methylation level of NR3C1, which resulted in an increase in the expression levels of the CRH mRNA and protein expression and further reduced the expression levels of the NR3C1 mRNA and protein expression, and ultimately led to have delayed responses to acute immobilization stress. Juvenile voles born at high density also reduced anti-KLH IgG levels and PHA responses, increased cytokines, and depressive and anxiety-like behaviors, and the effects further led to higher coccidial infection. From the perspective of population density inducing the changes in behavior and immunity at the brain level, our results showed a physiological epigenetic mechanism for population self-regulation in voles. Our results indicate that altering the prenatal intrinsic stress environment can fundamentally impact behavior and immunity by DNA methylation of HPA-axis genes and can further drive population fluctuations in wild animals.

Alpha-synuclein-induced stress sensitivity renders the Parkinson’s disease brain susceptible to neurodegeneration

A link between chronic stress and Parkinson’s disease (PD) pathogenesis is emerging. Ample evidence demonstrates that the presynaptic neuronal protein alpha-synuclein (asyn) is closely tied to PD pathogenesis. However, it is not known whether stress system dysfunction is present in PD, if asyn is involved, and if, together, they contribute to neurodegeneration. To address these questions, we assess stress axis function in transgenic rats overexpressing full-length wildtype human asyn (asyn BAC rats) and perform multi-level stress and PD phenotyping following chronic corticosterone administration. Stress signaling, namely corticotropin-releasing factor, glucocorticoid and mineralocorticoid receptor gene expression, is also examined in post-mortem PD patient brains. Overexpression of human wildtype asyn leads to HPA axis dysregulation in rats, while chronic corticosterone administration significantly aggravates nigrostriatal degeneration, serine129 phosphorylated asyn (pS129) expression and neuroinflammation, leading to phenoconversion from a prodromal to an overt motor PD phenotype. Interestingly, chronic corticosterone in asyn BAC rats induces a robust, twofold increase in pS129 expression in the hypothalamus, the master regulator of the stress response, while the hippocampus, both a regulator and a target of the stress response, also demonstrates elevated pS129 asyn levels and altered markers of stress signalling. Finally, defective hippocampal stress signalling is mirrored in human PD brains and correlates with asyn expression levels. Taken together, our results link brain stress system dysregulation with asyn and provide evidence that elevated circulating glucocorticoids can contribute to asyn-induced neurodegeneration, ultimately triggering phenoconversion from prodromal to overt PD.

Prenatal auditory stimulation and impacts on physiological response to feed restriction in broiler chickens at market age

Feed restriction could induce physiological stress in broiler chickens, leading to welfare issues. Prenatal stimulation could improve stress-coping mechanisms in poultry. The present study aimed to elucidate the effects of subjecting developing embryos to auditory stimulation on physiological stress response to feed restriction in broiler chickens at market age. A total of 423 hatching eggs of Cobb 500 (Gallus domesticus) were subjected to the following auditory treatments: 1) no additional sound treatment other than the background sound of the incubator's compressors at 40 dB (CONTROL), 2) exposure to pre-recorded traffic noise at 90 dB (NOISE), and 3) exposure to Mozart's Sonata for Two Pianos in D Major, K 488 at 90 dB) (MUSIC). The NOISE and MUSIC treatments were for 20 min/h for 24 h (a total of 8 h/d), starting from embryonic days (ED) 12 to hatching. On d 42, an equal number of birds from each prenatal auditory stimulation (PAS) group were subjected to either ad libitum feeding (AL) or 30-h of feed restriction (FR) in a completely randomised design. The FR chickens exhibited significantly higher serum levels of corticosterone (CORT), and heat shock protein (HSP) 70 compared to those of AL. Prenatal auditory stimulation, particularly NOISE, led to lower serum levels of CORT and alpha-1-acid glycoprotein (AGP) levels compared to the CONTROL group. Additionally, NOISE significantly increased brain mRNA glucocorticoid receptor and HSP70 gene expression. The cecal population of E. coli and Lactobacillus spp. was not significantly affected by prenatal auditory stimulation. In conclusion, our findings suggest that prenatal auditory stimulation, particularly NOISE, positively impacts broiler chickens' ability to cope with feed restriction.

Early social isolation differentially affects the glucocorticoid receptor system and alcohol-seeking behavior in male and female Marchigian Sardinian alcohol-preferring rats

Adverse early life experiences during postnatal development can evoke long-lasting neurobiological changes in stress systems, thereby affecting subsequent behaviors including propensity to develop alcohol use disorder. Here, we exposed genetically selected male and female Marchigian Sardinian alcohol-preferring (msP) and Wistar rats to mild, repeated social deprivation from postnatal day 14 (PND14) to PND21 and investigated the effect of the early social isolation (ESI) on the glucocorticoid receptor (GR) system and on the propensity to drink and seek alcohol in adulthood. We found that ESI resulted in higher levels of GR gene and protein expression in the prefrontal cortex (PFC) in male but not female msP rats. In female Wistars, ESI resulted in significant downregulation of Nr3c1 mRNA levels and lower GR protein levels. In male and female msP rats, plasma corticosterone levels on PND35 were similar and unaffected by ESI. Wistar females exhibited higher levels of corticosterone compared with males, independently from ESI. In alcohol self-administration experiments we found that the pharmacological stressor yohimbine (0.0, 0.312, 0.625, and 1.25 mg/kg) increased alcohol self-administration in both rat lines, regardless of ESI. After extinction, 0.625 mg/kg yohimbine significantly reinstated alcohol seeking in female rats only. ESI enhanced reinstatement in female msP rats. Overall, the present results indicate that repeated social deprivation during the third week of postnatal life affects GR expression in a strain- and sex-dependent manner: such effect may contribute, at least partially, to the heightened sensitivity of female msP rats to the effects of yohimbine-induced alcohol seeking.

Effect of Acute and Cumulative Stress on Gene Expression in Mammary Tissue and Their Interactions with Physiological Responses and Milk Yield in Saanen Goats

This study addresses the hypothesis that different acute stressors can cumulatively decrease milk yield. In fact, in a time of global warming, the impact of environmental stress and farm management practices on milk production remains unclear. In this context, our objective was to investigate the effect of acute and cumulative stress on gene expression in mammary tissue and their interactions with physiological responses and milk yield in Saanen goats. Thirty lactating goats were subjected to two treatments: (1) control (CT), in which goats were maintained following a habitual routine under comfort conditions; (2) stress (ST), in which the goats were subjected to different types of environmental stress: heat stress, adrenocorticotropic hormone administration, hoof care management, and exposure to rain. These stressors were performed sequentially, with one stress per day on four consecutive lactation days, to evaluate their effect on milk quality and milk yield. Our results showed that compared to CT goats, cumulative stress increased the gene expression of glucocorticoid receptor (GR), interferon-gamma (IFN-γ), superoxide dismutase (SOD), and catalase (CAT) in mammary tissue, which are indicators of cortisol action, inflammatory response, and antioxidant enzymes. Furthermore, the acute challenges imposed on ST goats changed their rectal temperature and respiratory frequency and increased cortisol, glucose, cholesterol, triglycerides, and high-density lipoprotein release in plasma when compared to CT goats. Although these physiological and metabolic responses restore homeostasis, ST goats showed lower milk yield and higher somatic cell count in milk than CT goats. In conclusion, the results confirmed our initial hypothesis that different acute stressors cumulatively decrease the milk yield in Saanen goats.

From maternal glucocorticoid and thyroid hormones to epigenetic regulation of offspring gene expression: An experimental study in a wild bird species.

Offspring phenotype at birth is determined by its genotype and the prenatal environment including exposure to maternal hormones. Variation in both maternal glucocorticoids and thyroid hormones can affect offspring phenotype, but the underlying molecular mechanisms, especially those contributing to long-lasting effects, remain unclear. Epigenetic changes (such as DNA methylation) have been postulated as mediators of long-lasting effects of early-life environment. In this study, we determined the effects of elevated prenatal glucocorticoid and thyroid hormones on handling stress response (breath rate) as well as DNA methylation and gene expression of glucocorticoid receptor (GR) and thyroid hormone receptor (THR) in great tits (Parus major). Eggs were injected before incubation onset with corticosterone (the main avian glucocorticoid) and/or thyroid hormones (thyroxine and triiodothyronine) to simulate variation in maternal hormone deposition. Breath rate during handling and gene expression of GR and THR were evaluated 14 days after hatching. Methylation status of GR and THR genes was analyzed from the longitudinal blood cells sampled 7 and 14 days after hatching, as well as the following autumn. Elevated prenatal corticosterone level significantly increased the breath rate during handling, indicating an enhanced metabolic stress response. Prenatal corticosterone manipulation had CpG-site-specific effects on DNA methylation at the GR putative promoter region, while it did not significantly affect GR gene expression. GR expression was negatively associated with earlier hatching date and chick size. THR methylation or expression did not exhibit any significant relationship with the hormonal treatments or the examined covariates, suggesting that TH signaling may be more robust due to its crucial role in development. This study provides some support to the hypothesis suggesting that maternal corticosterone may influence offspring metabolic stress response via epigenetic alterations, yet their possible adaptive role in optimizing offspring phenotype to the prevailing conditions, context-dependency, and the underlying molecular interplay needs further research.

Sex-specific behavioral and physiological changes during single parenting in a biparental species, Columba livia

Many species exhibit biparental care to maximize fitness. When a partner is lost, the surviving partner may alter their behavior to compensate offspring. Whether both sexes use the same physiological mechanisms to manifest their change in behavior remains elusive. We investigated behaviors and mechanisms associated with the alteration of parental care post-partner removal in a biparental avian species, the rock dove (Columba livia). We hypothesized that rock dove single parents experience sex-biased changes in neural genomic transcription and reproductive behaviors, and these changes are related to chick development. We manipulated parental partner presence and measured parental attendance, offspring growth, gene expression of glucocorticoid receptors (GR) and mineralocorticoid receptors (MR) in the pituitary, and GR, MR, and estrogen receptor beta (ER-β) in the hypothalamus. We also measured circulating plasma concentrations of the stress-associated hormone corticosterone and the parental care-associated hormone prolactin. We also quantified prolactin gene (PRL) expression changes in the pituitary, as well as prolactin receptor (PRLR) expression in the hypothalamus and pituitary. We found that single mothers and fathers maintained similar provisioning levels as paired parents, but spent less cumulative time brooding chicks. Chicks of single parents were smaller than paired-parented chicks after three days post-hatch. Mothers in both treatment groups experienced higher expression of hypothalamic GR as compared to fathers. Single parents experienced lower PRL gene expression in the pituitary as compared to paired parents. No significant differences were found for the circulating hormones or other genes listed.

The potential role of GSK-3β signaling pathway for amelioration actions of ketamine on the PTSD rodent model

RationalePost-traumatic stress disorder (PTSD) is a complex, chronic psychiatric disorder typically triggered by life-threatening events and, as yet, lacks a specialized pharmacological treatment. The potential therapeutic role of ketamine, an N-methyl-D-aspartate receptor antagonist, in mitigating PTSD has been the subject of investigation. ObjectiveThe aim of this study was to elucidate alterations in the glycogen synthase kinase-3β (GSK-3β) signaling pathway in response to ketamine intervention, using the single prolonged stress (SPS) model of PTSD at a molecular level. MethodsPTSD-like symptoms were simulated using the SPS model. Ketamine (10 mg/kg) and GSK-3β antagonist SB216763 (5 mg/kg) were then administered intraperitoneally. Stress-related behavior was evaluated through the open field test (OFT) and the elevated plus maze test (EMPT). Additionally, brain activity was analyzed using quantitative electroencephalography (qEEG). Changes in protein and mRNA expressions of glucocorticoid receptor (GR), brain-derived neurotrophic factor (BDNF), GSK-3β, phosphorylated ser-9 GSK-3β (p-GSK-3β), FK506 binding protein 5 (FKBP5), and corticotropin-releasing hormone (CRH) were assessed in the hypothalamus via western blot and qPCR. ResultsSPS-exposed rats exhibited reduced distance and time spent in the center of the open arms, a pattern divergent from control rats. qEEG readings revealed SPS-induced increases in alpha power, low gamma and high gamma power. Furthermore, SPS triggered an upregulation in the protein and gene expression of GSK-3β, GR, BDNF, p-GSK-3β, and FKBP5, and downregulated CRH expression in the hypothalamus. Ketamine administration following the SPS procedure counteracted these changes by increasing the time spent in the center of the OFT, the distance traversed in the open arms of the EMPT, and mitigating SPS-induced alterations in cerebral cortex oscillations. Moreover, ketamine reduced the protein levels of GSK-3β, GR, p-GSK-3β, and altered the ratio of p-GSK-3β to GSK-3β. Gene expression of GSK-3β, GR, BDNF, and FKBP5 decreased in the SPS-Ket group compared to the SPS-Sal group. ConclusionsKetamine appeared to remediate the abnormal GSK-3β signaling pathway induced by SPS. These findings collectively suggest that ketamine could be a promising therapeutic agent for PTSD symptoms, working through the modulation of the GSK-3β signaling pathway.

Is chronic stress a causal mechanism for small mammal population cycles? Reconciling the evidence.

Chronic stress has long been hypothesized to play a role in driving population cycles. Christian (1950) hypothesized that high population density results in chronic stress and mass "die-offs" in small mammal populations. Updated variations of this hypothesis propose that chronic stress at high population density may reduce fitness, reproduction, or program aspects of phenotype, driving population declines. We tested the effect of density on the stress axis in meadow voles (Microtus pennsylvanicus) by manipulating population density in field enclosures over three years. Using fecal corticosterone metabolites as a non-invasive measure of glucocorticoid (GC) concentrations, we found that density alone was not associated with GC differences. However, we found that the seasonal relationship of GC levels differed by density treatment, with high-density populations having elevated GC levels early in the breeding season and decreasing towards late summer. We additionally tested hippocampal glucocorticoid receptor and mineralocorticoid receptor gene expression in juvenile voles born at different densities, with the hypothesis that high density may reduce receptor expression, altering negative feedback of the stress axis. We found that females had marginally higher glucocorticoid receptor expression at high density, no effect in males, and no detectable effect of density on mineralocorticoid receptor expression in either sex. Hence, we found no evidence that high density directly impairs negative feedback in the hippocampus, butrather female offspring may be better equipped for negative feedback. We compare our findings with prior studies to attempt to disentangle the complicated relationship between density, seasonality, sex, reproduction and the stress axis.

Covariation between glucocorticoid levels and receptor expression modulates embryo development and postnatal phenotypes in gulls

The hypothalamic-pituitary-adrenocortical axis can translate, through glucocorticoid secretion, the prenatal environment to development to produce phenotypes that match prevailing environmental conditions. However, whether developmental plasticity is modulated by the interaction between circulating glucocorticoids and receptor expression remains unclear. Here, we tested whether covariation between plasma corticosterone (CORT) and glucocorticoid receptor gene (Nr3c1) expression in blood underlies embryonic developmental programming in yellow-legged gulls (Larus michahellis). We examined variations in circulating levels of CORT and the expression and DNA methylation patterns of Nr3c1 in response to two ecologically relevant prenatal factors: adult alarm calls (a cue of predator presence) and changes in prenatal light environment (a cue of competitive disadvantage). We then determined whether embryonic development and postnatal phenotypes were associated with CORT levels and Nr3c1 expression, and explored direct and indirect relationships between the prenatal environment, hormone-receptor covariation, and postnatal phenotypes. Prenatal exposure to alarm calls increased CORT levels and up-regulated Nr3c1 expression in gull chicks, while exposure to light cues reduced both hormone levels and receptor expression. Chicks prenatally exposed to alarm calls showed altered DNA methylation profiles in the Nr3c1 regulatory region, but patterns varied throughout the breeding season and between years. Moreover, our results suggest a negative relationship between DNA methylation and expression in Nr3c1 , at least at specific CpG sites. The interplay between circulating CORT and Nr3c1 expression affected embryo developmental timing and vocalizations, as well as hatchling mass and fitness-relevant behaviours. These findings provide a link between prenatal inputs, glucocorticoid function and phenotypic outcomes, suggesting that hormone-receptor interaction may underlie developmental programming in free-living animals.

Intergenerational Perioperative Neurocognitive Disorder in Young Adult Male Rats with Traumatic Brain Injury.

The authors tested the hypothesis that the effects of traumatic brain injury, surgery, and sevoflurane interact to induce neurobehavioral abnormalities in adult male rats and in their offspring (an animal model of intergenerational perioperative neurocognitive disorder). Sprague-Dawley male rats (assigned generation F0) underwent a traumatic brain injury on postnatal day 60 that involved craniectomy (surgery) under 3% sevoflurane for 40 min followed by 2.1% sevoflurane for 3 h on postnatal days 62, 64, and 66 (injury group). The surgery group had craniectomy without traumatic brain injury, whereas the sevoflurane group had sevoflurane only. On postnatal day 90, F0 males and control females were mated to generate offspring (assigned generation F1). Acutely, F0 injury rats exhibited the greatest increases in serum corticosterone and interleukin-1β and -6, and activation of the hippocampal microglia. Long-term, compared to controls, F0 injury rats had the most exacerbated corticosterone levels at rest (mean ± SD, 2.21 ± 0.64 vs. 7.28 ± 1.95 ng/ml, n = 7 - 8; P < 0.001) and 10 min after restraint (133.12 ± 33.98 vs. 232.83 ± 40.71 ng/ml, n = 7 - 8; P < 0.001), increased interleukin-1β and -6, and reduced expression of hippocampal glucocorticoid receptor (Nr3c1; 0.53 ± 0.08 fold change relative to control, P < 0.001, n = 6) and brain-derived neurotrophic factor genes. They also exhibited greater behavioral deficiencies. Similar abnormalities were evident in their male offspring, whereas F1 females were not affected. The reduced Nr3c1 expression in F1 male, but not female, hippocampus was accompanied by corresponding Nr3c1 promoter hypermethylated CpG sites in F0 spermatozoa and F1 male, but not female, hippocampus. These findings in rats suggest that young adult males with traumatic brain injury are at an increased risk of developing perioperative neurocognitive disorder, as are their unexposed male but not female offspring.

Glucocorticoid Receptor Gene (NR3C1) Expression in the Pathogenesis of Depression in Cancer.

This study aimed to compare the NR3C1 expression among cancer patients with major depressive disorder (cancer depression), cancer patients without major depressive disorder (cancer non-depression), and major depressive disorder patients without cancer (general depression), as a preliminary investigation of epigenetic changes in the glucocorticoid receptor gene. From May 2019 to November 2019, patients were recruited from the Department of Psychiatry, Cancer Center in Busan, Korea. For gene expression studies, primers were designed using the Primer3 web tool (http://frodo.wi.mit.edu/primer3), and amplification reactions were performed. Expression levels of NR3C1 were lower in cancer depression and general depression than in cancer non-depression group. Given that we observed downregulation of the NR3C1 gene expression in depressive patients regardless of cancer status, it appears that methylation changes in NR3C1 may contribute to the pathophysiology of depression. The results of this study imply that the expression of NR3C1 may be decreased in major depressive disorder.

Striatal miR-183-5p inhibits methamphetamine-induced locomotion by regulating glucocorticoid receptor signaling.

MicroRNA (miRNA)-mediated striatal gene regulation may play an important role in methamphetamine (METH) addiction. This study aimed to identify changes in novel miRNAs and their target genes during METH self-administration and investigate their roles in METH-induced locomotion. RNA sequencing analysis revealed that mir-183-5p was upregulated in the striatum of METH self-administered rats, and target gene prediction revealed that the glucocorticoid receptor (GR) gene, Nr3c1, was a potential target gene for mir-183-5p. We confirmed that single and repeated METH administrations increased METH-induced locomotion and plasma corticosterone levels in rats. Additionally, increased miR-185-5p expression and decreased GR gene expression were observed only in the repeated-METH-injection group but not in the single-injection group. We then investigated the effects of miR-183-5p on METH-induced locomotion using a miR-183-5p mimic and inhibitor. Injection of a mir-183-5p mimic in the striatum of rats attenuated METH-induced locomotion, whereas injection of a miR-183-5p inhibitor enhanced the locomotor activity in METH-administered rats. Furthermore, the miR-183-5p mimic reduced the phosphorylation of tyrosine hydroxylase (TH) whereas the inhibitor increased it. Taken together, these results indicate that repeated METH injections increase striatal miR-183-5p expression and regulate METH-induced locomotion by regulating GR expression in rats, thereby suggesting a potential role of miR-183-5p as a novel regulator of METH-induced locomotion.

Harm of circadian misalignment to the hearts of the adolescent wistar rats

PurposeThe purpose of this study was to observe the harm of circadian misalignment (CM), caused by an inverted photoperiod (IP), on the hearts of the adolescent Wistar rats, and to explore the mechanisms leading to harm.MethodsAn IP was used to create a CM model. A total of 174 Wistar rats were randomly divided into circadian alignment (CA) and CM groups (87 rats per group). The different activity rhythms of the two groups of rats were adjusted through different light/dark cycles for 90 days. We recorded the rhythmic activity trajectory and sleep time of the rats. After 90 days of modeling, we performed various analyses (i.e., blood pressure, weight, cardiac ultrasound tests, serological tests, cardiac tissue immunofluorescence, immunohistochemistry, transmission electron microscopy on myocardial mitochondria, western blotting, and quantitative polymerase chain reactions).Results(1) The IP protocol caused CM in rats. (2) CM rats showed significantly higher blood pressure during the day (resting phase). They also showed significantly higher serum levels of angiotensin II and epinephrine during the day compared to the CA rats. (3) CM caused up-regulation of gene expression of adrenergic receptors α1 (α1-AR) and β1 (β1-AR) and down-regulation of the glucocorticoid receptor (Gr) gene expression in rat hearts. It also caused downregulation of Bmal1 expression. In addition, the changes in Bmal1 and Per2 correlated with the changes in β1-AR and α1-AR. (4) CM had adverse effects on multiple molecular proteins of the heart. (5) CM increased the collagen fibers in the rat heart and increased the destruction of mitochondria. (6) Eventually, CM caused a decrease in the pumping function of the heart and decreased the coronary blood flow rate.Conclusions(1) CM significantly affected the cardiac structure and function in the adolescent rats through a variety of mechanisms. (2) CM can regulate the expression of myocardial clock genes, and it is likely to have an impact on the heart through this pathway.

The role of maternal exercise on placental, behavioral and genetic alterations induced by prenatal stress

The present study aimed to evaluate the effects of treadmill maternal exercise on alterations induced by prenatal stress in neonatal mice. Female and male Balb/c mice were divided into five groups: control (CON), prenatal restraint stress (PNS), prenatal restraint stress and physical exercise before pregnancy (PNS + EX1), prenatal restraint stress and physical exercise during pregnancy (PNS + EX2), and prenatal restraint stress and physical exercise before and during pregnancy (PNS + EX3). Exercise was performed using a treadmill, at a speed of 10 m/min, for 60 min, 5 days a week. Maternal behavior was assessed on days 3, 4 and 5 postpartum (PPD). Placental gene expression of glucocorticoid receptor (GR), 11-β-hydroxysteroid dehydrogenase 2 (11β-HSD2), 5-hydroxytryptamine receptor 1A (5HT1AR), and corticotropin releasing hormone receptor 1 (CRHR1) were analyzed. In neonatal mice, the gene expression of GR, mineralocorticoid receptor (MR), CRHR1, 5HTr1, oxytocin Receptor 1 (OXTr1), tropomyosin related kinase B (TRκB), brain-derived neurotrophic factor exon I (BDNF I), and BDNF IV was analyzed in the brain (PND0) and hippocampus (PND10). Maternal exercise improved (p < 0.05) maternal care. In the placenta, maternal exercise prevented (p < 0.01) the increase in GR expression caused by PNS. In the brain from PND0, exercise before pregnancy prevented (p = 0.002) the decreased CRHR1 expression promoted by PNS. In the hippocampus of PND10 males, PNS decreased (p = 0.0005) GR expression, and exercise before pregnancy prevented (p = 0.003) this effect. In PND10 females, maternal exercise prevented (p < 0.05) the PNS-induced increase in MR expression. PNS + EX2 males showed increased (p < 0.01) BDNF I gene expression and PNS + EX1 females demonstrated increased (p = 0.03) BDNF IV expression. In conclusion, maternal physical exercise may play a role in modulating maternal-fetal health and may contribute to preventing neurodevelopmental changes induced by prenatal stress.

Corticosterone Levels and Glucocorticoid Receptor Gene Expression in High Drinking in the Dark Mice and Their Heterogeneous Stock (HS/NPT) Founder Line

The High Drinking in the Dark (HDID-1) line of mice has been selectively bred for achieving high blood alcohol levels (BALs) in the Drinking in the Dark task, a model of binge-like drinking. Recently, we determined that glucocorticoid receptor (GR) antagonism with either mifepristone or CORT113176 (a selective GR antagonist) reduced binge-like ethanol intake in the HDID-1 mice, but not in their founder line, HS/NPT. Here, we examined whether the selection process may have altered glucocorticoid functioning by measuring (1) plasma corticosterone levels and (2) expression of the genes encoding GR (Nr3c1) and two of its chaperone proteins FKBP51 and FKBP52 (Fkbp5 and Fkbp4) in the brains (nucleus accumbens, NAc) of HDID-1 and HS/NPT mice. We observed no genotype differences in baseline circulating corticosterone levels. However, HDID-1 mice exhibited a greater stimulated peak corticosterone response to an IP injection (of either ethanol or saline) relative to their founder line. We further observed reduced basal expression of Fkbp4 and Nr3c1 in the NAc of HDID-1 mice relative to HS/NPT mice. Finally, HDID-1 mice exhibited reduced Fkbp5 expression in the NAc relative to HS/NPT mice following an injection of 2 g/kg ethanol. Together, these data suggest that selective breeding for high BALs may have altered stress signaling in the HDID-1 mice, which may contribute to the observed selective efficacy of GR antagonism in reducing binge-like ethanol intake in HDID-1, but not HS/NPT mice. These data have important implications for the role that stress signaling plays in the genetic risk for binge drinking.

Maternal deprivation effect on morphine-induced CPP is related to changes in opioid receptors in selected rat brain regions (hippocampus, prefrontal cortex, and nucleus accumbens)

Early-life environmental conditions affect offspring's development. Maternal deprivation (MD) can induce persistent changes that give rise to neuropsychiatric diseases including substance abuse disorders. However, long-lasting mechanisms that determine vulnerability to drug addiction remain unknown. We hypothesized that MD could induce changes in Opioid system, HPA (hypothalamic-pituitary-adrenal) axis, and BDNF (brain-derived neurotrophic factor), so may be involved in the drug abuse in later life. Male offspring of Wistar rats (n = 8 per group) were subjected to 3 h of daily MD during postnatal days 1–14. In adulthood, morphine-induced CPP (conditioned place preference) was investigated using two doses of morphine (3 and 5 mg/kg). Serum corticosterone level was measured by ELISA method. The expression level of genes in selected brain regions (hippocampus, prefrontal cortex, and nucleus accumbens) was determined by qPCR (quantitative PCR). A greater morphine-induced CPP was observed in MD rats with 3 and 5 mg/kg morphine compared to controls. MD group had a higher corticosterone level. A significant decrease was observed in the expression of BDNF gene (in all of the selected brain regions) and GR (glucocorticoid receptor) gene (in the hippocampus and nucleus accumbens) in MD rats. Also, a significant increase in the expression of μ Opioid receptor (in all of the selected brain regions) and κ Opioid receptor (in the prefrontal cortex and nucleus accumbens) was observed in MD rats. Our results suggest that MD induces alterations in the HPA axis function, BDNF level, and Opioid receptors system that enhance vulnerability to morphine at adulthood.

The Effect of 12 Hz Extremely Low-frequency Electromagnetic Field on Visual Memory of Male Macaque Monkeys.

Today, humans live in a world surrounded by electromagnetic fields. Numerous studies have been conducted to discover the biological, physiological, and behavioral effects of electromagnetic fields on humans and animals. Given the biological similarities between monkeys and humans, The present research aimed to examine Visual Memory (VM), hormonal, genomic, and anatomic changes, in the male rhesus macaques exposed to an Extremely Low-Frequency Magnetic Field (ELF-MF). Four male rhesus macaques (Macaca mulatta) were used. For the behavioral tests, the animals should be fasting for 17 hours. For the tests such as visual memory, the animal's cooperation was necessary. Using the radiation protocol, we exposed two monkeys to a 12-Hz electromagnetic field with a magnitude of 0.7 μT (electromagnetic radiation) four hours a day for a month. Before and after the exposure, a visual memory test was conducted using a coated device (visible reward) on a movable stand. Ten milliliters of blood was obtained from the femoral artery of each monkey, and half of it was used to examine cortisol serum levels using the MyBioSource kit (made in the USA). The other half of the blood was used to extract lymphocytes for assaying expressions of Glucocorticoid Receptor (GR) genes before and after radiation using the PCR method. Anatomic studies of the amygdala were carried out based on pre- and post-radiation Magnetic Resonance Imaging (MRI). Research results indicated that visual memory in male primates increased significantly after exposure to the 12-Hz frequency. Hormonal analysis at the 12-Hz frequency showed a decrease in cortisol serum levels. However, visual memory and serum cortisol levels did not change considerably in male primates in the control group. There was no considerable amygdala volumetric difference after exposure to the 12-Hz frequency. The expression of the GR genes decreased in the 12-Hz group compared to the control group. In short, these results indicated that ELF might benefit memory enhancement because exposure to the 12-HZ ELF can enhance visual memory. This outcome may be due to a decrease in plasma cortisol and or expression of GR genes. Moreover, direct amygdala involvement in this regard cannot be recommended. The effects of Extremely Low-Frequency Electromagnetic Fields (ELF-EMF) of 12 Hz on monkeys were studied.The results showed a reduction in the serum cortisol levels and the expression of GR genes.The amygdala anatomical area changes were not significant in the experimental group.In the experimental group, visual memory (delay of 30- and 60-s evaluation) improved after exposure to a frequency of 12 Hz. Extremely low-frequency electromagnetic fields are among the most important factors affecting humans. This study aimed to determine the fields of 12-Hz frequency on the visual memory changes of male monkeys. The importance of research is due to the cognitive similarity of monkeys to humans. The findings of the research can be attributed to humans. Behavioral, hormonal, genetic, and anatomical studies indicated improvement in visual memory (test monkeys versus control monkeys). This study demonstrates the effect of the 12-Hz frequency on the monkey's visual memory. Researchers can study 12-Hz frequency in other cognitive indices.