Glucocorticoid Receptor Content Research Articles

Multi-Residue Detection of Eight Glucocorticoids by Nano-Au/Fluticasone Propionate Electrochemical Immunosensor.

Based on an indirect competitive method, a novel nano-Au/fluticasone propionate electrochemical immunosensor was successfully fabricated by combining the nanoscale effect, superior conductivity of nano-Au, stable Au-S chemical bond as well as strong interaction between glucocorticoid and the receptor, which was used to simultaneously detect eight kinds of glucocorticoids. The modified immunosensors' electrochemical properties were explored by means of a cyclic voltammetry (CV) method and electrochemical impedance spectroscopy (EIS) measurements. Two factors (glucocorticoid receptor concentration, incubation time) were studied in order to obtain the optimal results. The immunosensor presents attractive electrochemical performance with a wide linear range (between 0.1 and 1500 ng⋅mL-1) and low detection limit (between 0.057 and 0.357 ng⋅mL-1), realizing the rapid multi-residue detection of a large class of glucocorticoids. Two glucocorticoids (hydrocortisone, triamcinolone) were detected in actual skincare samples, which obtained satisfactory detection results.

Resistance Training Modulates Hippocampal Neuroinflammation and Protects Anxiety-Depression-like Dyad Induced by an Emotional Single Prolonged Stress Model.

Stress is a triggering factor for anxious and depressive phenotypes. Exercise is known for its action on the central nervous system. This study aimed to evaluate the role of resistance exercise in an anxiety-depression-like dyad in a model of stress. Male Swiss mice (35-day-old) were exercised, three times a week for 4 weeks on nonconsecutive days. The resistance exercise consisted of climbing a 1-m-high ladder 15 times. After mice were subjected to an emotional single prolonged stress (Esps) protocol. Seven days later, they were subjected to anxiety and depression predictive behavioral tests. The results showed that exercised mice gain less weight than sedentary from weeks 3 to 5. Resistance exercise was effective against an increase in immobility time in the forced swim test and tail suspension test and a decrease in grooming time of mice subjected to Esps. Resistance exercise protected against the decrease in the percentage of open arms time and open arm entries, and the increase in the anxiety index in Esps mice. Four-week resistance exercise did not elicit an antidepressant/anxiolytic phenotype in non-stressed mice. Esps did not alter plasma corticosterone levels but increased the hippocampal glucocorticoid receptor content in mice. Resistance exercise protected against the decrease in hippocampal levels of tropomyosin kinase B (TRκB), the p-Akt/Akt, and the p-mTOR/mTOR ratios of Esps mice. Resistance exercise proved to be effective in decreasing hippocampal neuroinflammation in Esps mice. Resistance exercise protected against the increase in the hippocampal Akt/mTOR pathway and neuroinflammation, and anxiety/depression-like dyad in Esps exposed mice.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12035-022-03069-x.

Social-single prolonged stress as an ether-free candidate animal model of post-traumatic stress disorder: Female and male outcomings

BackgroundSingle Prolonged Stress (SPS) is a valid animal model that reflects the core of post-traumatic stress disorder (PTSD) phenotypes. Although SPS has been a pivotal tool, it can bring ethics approval difficulties due to the use of ether as a stressor. The present study evaluated if changing a chemical (ether) with a social stressor resembles the PTSD hallmark symptoms. MethodsFemale and male young adult rats were distributed in Sham and Social-SPS groups. Rats in Social-SPS groups were subjected to stress, whereas those in Sham groups remained undisturbed. One set of animals performed the behavioral tests, elevated plus-maze (EPM) and Y-maze. Plasma corticosterone levels and cortical and hippocampal molecular protein contents were analyzed. Another set of animals performed the dexamethasone suppression test. ResultsA significant decrease in the percentage of time spent and the number of entries in open arms and an increase in anxiety index in the EPM were observed in rats of the social-SPS groups. In the Social-SPS groups, rats reduced the spontaneous alternations in Y-maze. The Social-SPS exposure enhanced the HPA-axis feedback and increased glucocorticoid receptor contents in the cerebral cortex and hippocampus of rats. A decrease in the content of synaptic integrity-related proteins, synaptophysin, and PSD-95, were found in the cortex and hippocampus of rats subjected to social-SPS. There were no statistical differences between males and females in any parameter analyzed. LimitationsThe absence of a task to recap criterion E ‘arousal’ and predictive validity experiments. ConclusionsThis study reveals that social-SPS recapitulated the main clusters required for a candidate animal model of PTSD.

Antioxidant Activity Of Rat Liver With A Low Resistance To Hypoxia After Systemic Ischemia Reperfusion

Objective — To assess the antioxidant activity of rat liver after systemic ischemia reperfusion (IRP). Material and Methods — The study was conducted on 70 male rats. For all animals of the treatment group (n=35) under ether anesthesia, we were stopping stopping systemic circulation for five minutes. After that, the animals were given an external cardiac massage and artificial lung ventilation. We did not perform circulatory arrest after ether anesthesia in animals of the control group (n=35). In all animals, we were measuring the levels of serum hormones (corticosterone, aldosterone), the content of glucocorticoid and mineralocorticoid receptors in liver homogenates, and the activity of enzymes of the antioxidant system (superoxide dismutase and catalase). We were making control measurements on days 1, 3, 5, 7, 14, 21, and 35 after the simulated IRP. Results — On day 1 after simulation of IRP development, the levels of cortisol and aldosterone in the serum of treatment group rats were significantly higher, by 14.3% and 33.5%, respectively, compared with the control group. In response to stress (IRP), we observed the highest concentration of cortisol in the blood of treatment group rats on day 3 (p=0.0002), which decreased afterwards. On day 1 after IRP, there was a reduction in the activity of superoxide dismutase and catalase in treatment group rats, by 50.3% and by 29%, respectively (p<0.0001). The lowest antioxidant activity in the rat liver after IRP was observed on days 3-7. Conclusion — Systemic IRP is associated with pronounced changes in the dynamics of corticosteroid receptors in the liver, which leads to a reduction in the activity of key antioxidant enzymes.

Sex-Dependent Actions of Prenatal Stress on the Activity of the Hypothalamo-Hypophyseal-Adrenocortical System in Rats: The Role of Corticosteroid Receptors in the Brain

Prenatal stress is regarded as a risk factor for the development of diseases such as mental, cardiovascular, and metabolic disorders in later life. Dysregulation of the activity of the hypothalamo-hypophyseal-adrenocortical system (HHAS) is one of the mechanisms producing these disorders. Regulation of the HHAS is mediated by two types of receptor – mineralocorticoid (MR) and glucocorticoid receptors (GR) – in the corticolimbic structures of the brain. The link between prenatal stress, sex, and HHAS activity, as well as the role of central MR and GR in this interaction, has received insufficient study. We have investigated the effects of prenatal stress on HHAS activity in adult male and female rats and the expression of GR and MR in the hippocampus and medial prefrontal cortex (mPFC). Prenatal stress was modeled by subjecting pregnant female Wistar rats to 1-h restraint from days 15 to 19 of pregnancy. Basal HHAS activity and stress corticosterone levels in response to 30-min restraint were studied in their adult offspring. GR and MR expression was assessed by western blotting and also, in the hippocampus, by an immunohistochemical method. Prenatally stressed males displayed prolonged stress responses, while females showed increases in basal and stress reactivity, along with increases in the sensitivity of the HHAS to feedback signals. In males, changes in HHAS activity were accompanied by decreases in GR and MR content in the hippocampus and mPFC with an unaltered GR:MR ratio. Prenatally stressed females showed increased expression of GR protein and decreased quantities of MR protein in the hippocampus and an increased GR:MR ratio in both areas of the brain. The hippocampus of male and female animals showed changes in GR and MR content in field CA3 and the dentate gyrus. These data indicate that sex-dependent changes in GR and MR protein expression in the hippocampus and mPFC contribute to modifications of HHAS activity due to prenatal stress.

The effect of free radical stress correction on corticoid signaling in the kidney of rat with different resistance to hypoxia after systemic circulation arrest

Aim. To assess the influence of the pathogenetic action of the succinate-containing drug on corticosteroid regulation in the kidney of rats with different resistance to hypoxia during recovery after systemic circulation arrest.
 Methods. The object of the study was male non-inbred white rats weighing 200220 g. A week after testing for resistance to hypoxia, a 5-minute systemic circulation arrest was simulated by intrathoracic clamping of the vascular bundle of the heart, followed by resuscitation. In the postresuscitation period, the experimental rats were once daily injected with a solution containing inosine + nicotinamide + riboflavin + succinic acid, and the control rats 0.9% Sodium Chloride solution. The observation period was 35 days. We studied the content of corticosterone, aldosterone in blood plasma, gluco- and mineralocorticoid receptors, carbonylated proteins, dityrosine, and products that react with thiobarbituric acid in homogenates of the kidneys. Statistical data were presented as mean and standard deviation M. Nonparametric KruskalWallis (N) and MannWhitney (U) tests followed by Dunn test, Spearman correlation analysis were used. Differences were considered statistically significant at p 0.05.
 Results. The use of succinate-containing preparation reduced the intensity of free radical processes in both groups of animals. Against this background, in low-resistance rats, on the 1st day, the concentration of glucocorticoid receptors statistically significantly increased to 117% (p 0.05), and then was comparable to the control; the greatest statistically significant changes in the level of mineralocorticoid receptors occurred on the 1st day (increase by 25%, p 0.001) and at 2135th days (decrease by 2230%, p 0.001). In highly resistant rats, the correction led to a shift in the maximum content of glucocorticoid receptors from the last day (134% of the control level, p 0.01 without therapy) to the 1st (123%, p 0.05 with succinate-containing therapy) and maintaining the receptors level comparable to the initial, in the future. The level of mineralocorticoid receptors in highly resistant rats was lower than in low resistant rats, both in the group without correction and with correction.
 Conclusion. Correction of the course of the postresuscitation period with a succinate-containing drug in animals with a low resistance to hypoxia against the background of a decrease in the intensity of carbonyl stress and restoration of feedback mechanisms causes stabilization of glucocorticoid receptors level and a decrease in mineralocorticoid receptors to control values by the end of the experiment; in organisms highly resistant to hypoxia, against the background of correction, the activity of lipid peroxidation decreases and the level of both types of receptors are restored.

ACUTE ANDROGEN AND GLUCOCORTICOID RECEPTOR RESPONSE FOLLOWING A MODERATE INTENSITY RESISTANCE EXERCISE BOUT

PURPOSE: The purpose of this study was to examine the acute androgen receptor (AR) and glucocorticoid receptor (GR) response to a moderate intensity resistance exercise bout in resistance trained (RT) and untrained men (UT). METHODS: RT men (n = 10; X ± SD, age = 21.3 ± 1.7 yrs, height = 175.8 ± 6.8 cm, body mass = 84.5 ± 13.5 kg, back squat 1RM = 154.3 ± 19.3 kg, training history = 5.4 ± 2.0 yrs) and UT men (n = 9; X ± SD, age = 20.8 ± 3.1 yrs, height = 178.7 ± 8.9 cm, body mass = 81.0 ± 14.0 kg, squat 1RM = 108.1 ± 13.7 kg, training history = 0.7 ± 1.7 yrs) volunteered for this study. Prior to the RE bout, subjects were strength tested for back squat (BS) and leg extension (LE). Subjects returned 4-7 days later between 10am-2pm, and completed a RE bout consisting of 6 sets of 10 repetitions of BS at 75% 1RM, immediately followed by 4 sets of 10 repetitions of LE at 75% 1RM with 1.5 min rest between all sets. Muscle samples were collected from the vastus lateralis prior to exercise (PRE) and 10 min (10P), 30 min (30P), 60 min (60P), and 180 min (180P) post exercise. Total AR and GR expression was determined via western blotting. Receptor data were not normally distributed, thus all receptor data were analyzed using Mann-Whitney U test, Friedman test, and Wilcoxon signed-rank test. RESULTS: For total AR expression, there were no differences between time points within the RT group (p > .05); however, there were differences between time points within the UT group (p = .016). In the UT group, total AR expression significantly decreased at 30P (-19%∆, z = -2.192, p = .027) and 60P (-11%∆, z = -2.192, p = .027) post exercise, but returned to baseline values by 180P (z = -.178, p > .05). For Total GR content, there were no differences between time points within the RT or UT groups (p > .05). Total GR content was significantly greater in the RT group compared to the UT group at 10P (Mann-Whitney U = 19, z = -2.123, p = .035). CONCLUSIONS: While no changes were observed for AR expression in the RT group, the UT subjects experienced a significant decrease in AR expression at 30P and 60P suggesting acute AR responses vary depending on training status. No differences were seen across time for the GR in either group; however, RT and UT subjects were different from each other at 10P. It is unclear if these responses are related to the acute hormonal response; therefore, future research will address this.

Gastrodiae Rhizoma Water Extract Ameliorates Hypothalamic-Pituitary-Adrenal Axis Hyperactivity and Inflammation Induced by Chronic Unpredictable Mild Stress in Rats.

Gastrodiae Rhizoma is a highly valuable traditional herbal medicine commonly used to treat neurological disorders. The present study is designed to determine the antidepressant-like effect of the Gastrodiae Rhizoma water extract (GRWE) on a depression model and the potential mechanisms. The chronic unpredictable mild stress (CUMS) rat model was used to induce depression. The sucrose preference test, open field test, forced swimming test, and tail suspension test were performed to assess the depressive-like behaviors, respectively. Hypothalamic–pituitary–adrenal (HPA) function was measured via plasma corticosterone (CORT), adrenocorticotrophic hormone (ACTH), hypothalamic corticotropin-releasing factor (CRF), and glucocorticoid receptor (GR) concentrations. Plasma concentrations of proinflammatory cytokines including interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) were also evaluated. The results showed that GRWE significantly attenuates the behavioral abnormalities in CUMS rats, as shown by elevated sucrose consumption, raised locomotor activity, and reduced immobility duration. Moreover, GRWE treatment reduced CORT, ACTH, CRF, and GR levels and decreased the plasma IL-1β, IL-6, and TNF-α concentrations. These findings indicate that GRWE improves depressive behaviors in a chronic stress model of rats; its effect may be ascribed to the modulation of the HPA axis activity and inflammatory response.

Features of the dynamics of corticosteroid receptors in the myocardium of animals with different resistance to hypoxia in the post resuscitation period

Aim. To characterize the peculiarities of the dynamics of the level of corticosteroid receptors in the myocardium of animals with different resistance to hypoxia in the post-resuscitation period.
 Methods. Experimental studies were carried out on male non-inbred white rats, divided into 2 groups by hypoxia resistance. A 5-minute arrest of the systemic circulation was modeled under ether anesthesia with intrathoracic clamping of the cardiovascular cluster with subsequent resuscitation. The observation period was 35 days. The content of corticosterone and aldosterone was determined in the blood plasma, the concentration of glucocorticoid and mineralocorticoid receptors in myocard homogenates was determined as well.
 Results. On days 1 to 3 of the post-resuscitation period in rats highly resistant to hypoxia, the dynamics of plasma corticosterone concentration and the content of corticosteroid receptors was unidirectional. Starting from the 5th day, against the background of a statistically significant decrease in the level of plasma corticosterone, a gradual increase in the density of corticosteroid receptors, mostly glucocorticoid, was observed, most pronounced on the 14th day and remaining until the end of the observation. In animals with low resistance to hypoxia, the dynamics of corticosteroid receptors was characterized by a predominance of mineralocorticoid content in almost all periods of observation. On days 13 of post-resuscitation period on the background of high concentrations of corticosteroid hormones, the minimum content of glucocorticoid receptors was noted. A decrease in the mineralocorticoid receptor level was recorded only on the first day, and in all subsequent periods of the experiment, the control indicators were significantly higher by 1.41.6 times. Strengthened mineralocorticoid signaling in the myocardium, characteristic of animals with low resistance to hypoxia, may be associated with the development of hypertrophy and fibrosis, inflammation, impaired electrical function. An increase in glucocorticoid receptors, characteristic of animals with a high resistance to hypoxia, has an adaptive effect, limiting the inflammatory response, the potential mechanism may be associated with increased expression of type 11-hydroxysteroid dehydrogenase.
 Conclusion. The identified features can have a significant influence on the course of the post-resuscitation period and determine the long-term forecast.

Application of natural compounds–based gold nanoparticles for the treatment of hemolytic anemia in an anemic mouse model: Formulation of a novel drug from relationship between the nanotechnology and hematology sciences

From time immemorial, people have tried to treat several diseases using natural compounds, including plants. Recently, researchers proposed that plants and herbal nanoparticles possess many remedial potentials. The results of this study confirmed the ability of an aqueous extract of Allium eriophyllum Boiss leaf grown under in vitro conditions for the biosynthesis of gold nanoparticles (AuNPs) and also revealed the anti‐hemolytic anemia activity of AuNPs in an anemic rodent model. These nanoparticles were characterized using Fourier‐transform infrared spectroscopy (FT‐IR) spectroscopy, ultraviolet–visible spectroscopy, X‐ray diffraction, field‐emission scanning electron microscopy (FE‐SEM), and transmission electron microscopy (TEM). TEM and FE‐SEM images showed the biosynthesized nanoparticles as having a uniform spherical morphology and diameters in the range of 5–30 nm. In vivo design, the induction of hemolytic anemia, was done using phenylhydrazine in 40 mice. Then, the mice were randomly divided into five groups: HAuCl4, A. eriophyllum, AuNPs, untreated, and control. AuNPs significantly (p ≤ 0.01) decreased the concentration of glucocorticoid receptors in the serum, liver, and spleen, and also alkaline phosphatase, aspartate aminotransferase, alanine aminotransferase, gamma‐glutamyl transferase, total and conjugated bilirubin, cholesterol, triglyceride, low‐density lipoprotein, urea, creatinine, ferrous, ferritin, and erythropoietin in the serum increased the concentrations of superoxide dismutase, catalase, and glutathione peroxidase (GPx) in the serum, liver, and spleen and also total protein, high‐density lipoprotein, and albumin in serum in the control mice as compared to the anemic mice. Also, AuNPs significantly (p ≤ 0.01) increased the body weight; anti‐inflammatory cytokine (IL4, IL5, IL10, IL13, and IFNα) concentration; and the total platelet, white blood cells, lymphocyte, neutrophil, monocyte, eosinophil, and basophil counts and red blood cell parameters but decreased the weight and volume of the liver and spleen and their subcompartments and decreased the concentration of pro‐inflammatory cytokines (IL1, IL6, IL12, IL18, IFNY, and TNFα) compared to the untreated mice. In vitro design, using 2,2‐diphenyl‐1‐picrylhydrazyl test, revealed similar antioxidant potentials for A. eriophyllum, AuNPs, and butylated hydroxytoluene. In addition, AuNPs were similar to A. eriophyllum and had a high cell viability dose dependently against the human umbilical vein endothelial cell line. In conclusion, the results of the chemical characterization confirm that the leaves of A. eriophyllum can be used to produce AuNPs with a remarkable amount of anti‐hemolytic anemia property.

Особенности свободнорадикального окисления и динамики содержания кортикостероидных рецепторов в легких у животных с разной чувствительностью к гипоксии в постреанимационном периоде

Цель исследования - изучение особенностей динамики показателей свободнорадикального окисления и уровня кортикостероидных рецепторов в легких у животных с разной чувствительностью к гипоксии в постреанимационном периоде. Методика. Эксперимент выполнен на самцах беспородных белых крыс, разделенных по устойчивости к гипоксии на 2 группы: неустойчивые и высокоустойчивые. Остановку системного кровообращения продолжительностью 5 мин моделировали под общим эфирным наркозом (через 1 нед после тестирования на устойчивость к гипоксии) интраторакальным пережатием сосудистого пучка сердца с последующей реанимацией. Через 1, 3, 5, 7, 14, 21 и 35 сут животных под эфирным наркозом декапитировали. В плазме крови определяли содержание кортикостерона и альдостерона, в гомогенатах легких - концентрацию глюкокортикоидных и минералокортикоидных рецепторов, уровни продуктов, реагирующих с тиобарбитуровой кислотой, карбонилированных белков, железозависимое образование битирозина, активность супероксиддисмутазы, каталазы, содержание восстановленного глутатиона. Результаты. Установлено, что для неустойчивых к гипоксии животных характерна высокая интенсивность свободнорадикального окисления, проявляющаяся развитием карбонильного стресса на фоне снижения активности ключевых антиоксидантов и сопряженная с выраженными изменениями динамики кортикостероидных рецепторов: в первые 3 сут постреанимационного периода наблюдалось снижение уровня кортикостероидных рецепторов, а к концу 35-суточного мониторинга значительное нарастание концентрации минералокортикоидных рецепторов. Высокую степень устойчивости к гипоксии отличает относительно низкая интенсивность свободнорадикального окисления, характеризующаяся только усилением липопероксидации за счет адекватной емкости и сохранности водорастворимых антиоксидантных систем, экранирующих белки. В раннем восстановительном периоде это сопровождается сохранением уровня кортикостероидных рецепторов, в позднем - существенным ростом концентрации глюкокортикоидных рецепторов. Заключение. Увеличение содержания глюкокортикоидных рецепторов у высокоустойчивых к гипоксии животных в позднем восстановительном периоде (14-е - 35-е сут) играет важную роль, предположительно путем модулирования течения воспалительного процесса, ограничения неконтролируемых иммунных реакции и повреждение тканей. Следствием значительного нарастания концентрации минералокортикоидных рецепторов у животных с низкой устойчивостью к гипоксии может быть усиление сосудистого ремоделирования и развитие фиброза в легких. Aim. The aim of the study was to characterize changes in free radical oxidation and corticosteroid receptor density in the lungs of animals with different sensitivity to hypoxia in the postresuscitation period. Methods. Experiments were performed on mongrel male white rats divided into two groups based on their hypoxia resistance, non-resistant and highly resistant. One week after testing the rats for resistance to hypoxia, a 5-min arrest of systemic circulation was modeled under ether anesthesia by intrathoracic compression of the vascular bundle of the heart with subsequent resuscitation. The follow-up period was 35 days. At 1, 3, 5, 7, 14, 21, and 35 days, the animals were sacrificed by decapitation under ether anesthesia, and blood and tissue samples were collected. Concentrations of corticosterone and aldosterone were measured in plasma; concentrations of glucocorticoid and mineralocorticoid receptors, thiobarbituric acid reactive substances, and carbonylated proteins, iron-dependent formation of dityrosine, activities of superoxide dismutase and catalase, and concentration of reduced glutathione were measured in lung homogenates. Results. Hypoxia non-resistant rats had a high intensity of free radical oxidation as evident from the development of carbonyl stress associated with decreased activities of key antioxidants and pronounced changes in the dynamics of corticosteroid receptors. A reduced level of corticosteroid receptors was observed in the first three days of resuscitation period and followed by a significant increase in mineralocorticoid receptors at the end of 35-day monitoring. High resistance to hypoxia was characterized by a relatively low intensity of free radical oxidation, evident only from increased lipid peroxidation, due to an adequate capacity and preservation of water-soluble antioxidants. During the early recovery period, high resistance to hypoxia was associated with preserved level of corticosteroid receptors whereas during late recovery, the high hypoxia resistance was associated with a significant increase in the concentration of glucocorticoid receptors. Conclusion. The increased density of glucocorticoid receptors in hypoxia high-resistant animals during the late recovery period (days 14-35) plays an important role, presumably by modulating the inflammatory process and restricting uncontrolled immune responses and tissue damage. The significant increase in mineralocorticoid receptors in hypoxia low-resistance animals may result in stimulation of vascular remodeling and development of pulmonary fibrosis.

Glucocorticoids regulate adipose tissue protein concentration in a depot- and sex-specific manner

Preclinical and clinical findings indicate that glucocorticoids (GC) induce lipid accumulation in visceral depots, while inhibiting lipid stores from subcutaneous depots. Whereas some suggest that this is due to adipose depot specific concentration of glucocorticoid receptors (GR) or 11beta-hydroxysteroid dehydrogenase 1 (11β-HSD1), others demonstrate these events emerge from increases in interleukin-1 beta (IL-1β) from macrophages within distinct depots. Regardless of the mechanisms, most of these studies occur in males and thus lack evaluation of sex differences. Here, we examined the impact of 2-week corticosterone (CORT) (3 mg/kg/day) or saline treatment on GR, 11β-HSD1 and IL-1β protein concentration in intra-abdominal (epididymal/parametrial, and visceral) and subcutaneous (inguinal) depots in male and female Sprague Dawley rats. The objective was to examine if factors that regulate GC-induced adipose depot metabolism and distribution, differ between males and females. CORT inhibited, but did not decrease, body weight gain in both sexes. 11β-HSD1 was similar between the sexes in all adipose depots. CORT increased IL-1β in both sexes only in gonadal adipose tissue. Overall, males had greater GR protein concentration in all adipose depots, whereas females had more IL-1β in intra-abdominal adipose depots. Given the male-biased increase in intra-abdominal GR protein concentration, the data suggest that males may be more prone to CORT-induced increases in visceral obesity, which may have implications for increased risk for metabolic diseases. Overall, the data suggest that the effects of GC signaling in adipose tissue are multifaceted, dependent on sex, and the inherent adipocyte characteristics.Lay summaryResearch supports that glucocorticoids (GC) induce visceral adipose tissue accumulation, however few studies have examined if these GC-mediated outcomes are similar between males and females. This study investigates if female rats differentially respond to corticosterone treatment. Results indicate that male rats may have an increased susceptibility to CORT-induced accumulation of visceral adipose tissue compared with females, which may have implication for sex-specific risk for metabolic diseases.

Signaling pathways involved in the rapid biphasic effect of aldosterone on Na+/H+ exchanger in rat proximal tubule cells

The receptors and signaling pathways for nongenomic effects of aldosterone (Aldo) on the proximal Na+/H+ exchanger are still unknown; therefore, the aim of this study was to investigate the mineralocorticoid receptor (MR) and/or glucocorticoid receptor (GR) participation in rapid Aldo effects on NHE1 (basolateral Na+/H+ exchanger isoform) and cytosolic calcium concentration ([Ca2+]i). In addition, phospholipase C (PLC), protein kinase C (PKC), and mitogen-activated protein kinase kinase (MEK) involvement in signaling pathways of such effects was evaluated, using immortalized proximal tubule cells of rat (IRPTC) as an experimental model. MR and GR expression was investigated using reverse transcription polymerase chain reaction and immunoblotting. The intracellular pH recovery rate (after acid loading) and [Ca2+]i were determined by the probes BCECF-AM and FURA 2-AM, respectively. Aldo (10−12 M) promoted a moderate increase in [Ca2+]i and stimulation of NHE1, whereas Aldo (10−6 M) greatly increased the [Ca2+]i, but inhibited the NHE1. BAPTA-AM (a calcium chelator), GR antagonism and inhibition of PLC, PKC and MEK pathway abolished the biphasic and dose-dependent effect of Aldo on NHE1 and decreased the [Ca2+]i; whereas MR do not appear to participate in this rapid signaling in IRPTC cells. The reduction of GR content, by gene silencing, abolished the Aldo effect on NHE1, in low concentration, confirming the importance of this receptor in the rapid modulation of proximal sodium and hydrogen transports.

Chronic Stress Causes Sex-Specific and Structure-Specific Alterations in Mitochondrial Respiratory Chain Activity in Rat Brain.

Chronic restraint stress (CRS) induces a variety of changes in brain function, some of which are mediated by glucocorticoids. The response to stress occurs in a sex-specific way, and may include mitochondrial and synaptic alterations. The synapse is highly dependent on mitochondrial energy supply, and when mitochondria become dysfunctional, they orchestrate cell death. This study aimed to investigate the CRS effects on mitochondrial respiratory chain activity, as well as mitochondrial potential and mass in cell body and synapses using hippocampus, cortex and striatum of male and female rats. Rats were divided into non-stressed (control) and stressed group (CRS during 40days). Results showed that CRS increased complex I-III activity in hippocampus. We also observed an interaction between CRS and sex in the striatal complex II activity, since CRS induced a reduction in complex II activity in males, while in females this activity was increased. Also an interaction was observed between stress and sex in cortical complex IV activity, since CRS induced increased activity in females, while it was reduced in males. Glucocorticoid receptor (GR) content in cortex and hippocampus was sexually dimorphic, with female rats presenting higher levels compared to males. No changes were observed in GR content, mitochondrial potential or mass of animals submitted to CRS. It was concluded that CRS induced changes in respiratory chain complex activities, and some of these changes are sex-dependent: these activities are increased in the striatal mitochondria by CRS protocol mainly in females, while in males it is decreased.

Vitamin D interferes with glucocorticoid responsivenessin human peripheral blood mononuclear target cells.

Glucocorticoids (GCs) are widely used in the treatment of inflammatory and autoimmune diseases; however, patients are often resistant to GC effects. Current studies indicate that vitamin D reduces the risk or modifies the course of autoimmune diseases posing vitamin D supplementation as a prevention or therapeutic option. Herein, we investigated whether vitamin D can modify the response to GCs at the molecular level. To this end, peripheral blood mononuclear cells (PBMCs) were isolated from healthy vitamin D-deficient women and incubated with either the active metabolite 1,25(OH)2D3 (VitD) for 11days or dexamethasone (Dex) for the last 2days in the presence or absence of VitD. Ex vivo GC sensitivity was assessed by the expression of the glucocorticoid receptor (GR) responsive gene GILZ with RT-PCR. Long-term incubation of PBMCs with VitD significantly decreased the Dex-induced augmentation of GILZ expression. Since the intracellular concentration of GR and the GR nuclear translocation are critical determinants of GC sensitivity, we next evaluated the effect of VitD on these factors. RT-PCR and western-blot analysis revealed that VitD reduced the expression of GR. This effect was abolished by the HDAC-specific inhibitor trichostatin A, implying that HDAC was implicated in this effect. Moreover, NCoR1 mRNA was significantly decreased upon treatment with VitD either alone or as pre-treatment to Dex, suggesting that a possible increase in expression of this co-repressor was not involved. In addition, immunofluorescence analysis showed that VitD hindered the Dex-induced GRα nuclear translocation, an effect verified by subcellular fractionation and western-blot experiments. To further explore the underpinning mechanism, we examined the potential of VitD to: (1) strengthen the FK506-binding protein 5 (FKBP5) negative feedback loop and (2) modify the phosphorylation status of GR. Remarkably, VitD decreased FKBP5 expression and decreased phosphorylation at Ser211, while enhancing phosphorylation of GR at Ser203. Overall, VitD decreases the ex vivo GC sensitivity and this effect is, at least in part, attributed both to decrease of GR expression owing to a mechanism that engages HDAC and inhibition of GR translocation to nucleus via differential modulation of the phosphorylation state of GR. Our study provides, for the first time, evidence that long-term action of VitD induces GC resistance in PBMCs from healthy volunteers and offers a possible mechanistic basis for VitD-triggered attenuation of GC effects.

RELATIONSHIP BETWEEN THE CORTICAL THICKNESS AND SERUM CORTISOL LEVELS IN DRUG-NAÏVE, FIRST-EPISODE PATIENTS WITH MAJOR DEPRESSIVE DISORDER: A SURFACE-BASED MORPHOMETRIC STUDY.

In major depressive disorder (MDD) patients, higher morning cortisol levels due to a hyperactive hypothalamic-pituitary-adrenal (HPA) axis have been reported. The aim of the present study was to evaluate the relationship between cortical thinning and the serum cortisol levels during the first depressive episode in drug-naïve MDD patients using an automated surface-based morphometry (SBM) method. The institutional review board approved this prospective study. MR imaging data were obtained using a 3T scanner by a three-dimensional fast-spoiled gradient recalled acquisition with steady state (3D-FSPGR). Thirty drug-naïve patients with MDD and 41 age- and gender-matched healthy subjects (controls) were enrolled. We then used the SBM method (Freesurfer) to generate cortical thickness maps, and measured the cortical thickness in each subject. Morning blood samples were drawn from all participants for cortisol measurements. We found the serum cortisol levels were significantly higher in the MDD patients than in the controls. The MDD patients manifested significant thinning of the left lateral orbitofrontal cortex compared with the controls. There was a significant negative linear correlation between the thickness of the left lateral orbitofrontal cortex and the serum cortisol levels in the MDD patients. In the early stage of MDD, the thickness of the lateral orbitofrontal cortex was significantly reduced, and also showed a significant inverse correlation with the serum cortisol levels. Since the lateral orbitofrontal cortex contains a high concentration of glucocorticoid receptor, glucocorticoid receptor-mediated signaling transductions could contribute to neurotoxicity, which might occur when there are high cortisol levels in patients with MDD.

Rapid Glucocorticoid Feedback Inhibition of ACTH Secretion Involves Ligand-Dependent Membrane Association of Glucocorticoid Receptors.

The hypothesis that rapid glucocorticoid inhibition of pituitary ACTH secretion mediates a feedforward/feedback mechanism responsible for the hourly glucocorticoid pulsatility was tested in cultured pituitary cells. Perifusion with 30 pM CRH caused sustained the elevation of ACTH secretion. Superimposed corticosterone pulses inhibited CRH-stimulated ACTH release, depending on prior glucocorticoid clearance. When CRH perifusion started after 2 hours of glucocorticoid-free medium, corticosterone levels in the stress range (1 μM) caused a delayed (25 min) and prolonged inhibition of CRH-stimulated ACTH secretion, up to 60 minutes after corticosterone withdrawal. In contrast, after 6 hours of glucocorticoid-free medium, basal corticosterone levels inhibited CRH-stimulated ACTH within 5 minutes, after rapid recovery 5 minutes after corticosterone withdrawal. The latter effect was insensitive to actinomycin D but was prevented by the glucocorticoid receptor antagonist, RU486, suggesting nongenomic effects of the classical glucocorticoid receptor. In hypothalamic-derived 4B cells, 10 nM corticosterone increased immunoreactive glucocorticoid receptor content in membrane fractions, with association and clearance rates paralleling the effects on ACTH secretion from corticotrophs. Corticosterone did not affect CRH-stimulated calcium influx, but in AtT-20 cells, it had biphasic effects on CRH-stimulated Src phosphorylation, with early inhibition and late stimulation, suggesting a role for Src phosphorylation on the rapid glucocorticoid feedback. The data suggest that the nongenomic/membrane effects of classical GR mediate rapid and reversible glucocorticoid feedback inhibition at the pituitary corticotrophs downstream of calcium influx. The sensitivity and kinetics of these effects is consistent with the hypothesis that pituitary glucocorticoid feedback is part of the mechanism for adrenocortical ultradian pulse generation.

Modulation of central glucocorticoid receptors in short- and long-term experimental hyperthyroidism.

Hyperthyroidism is associated with a significant increase in circulating glucocorticoid levels and hyperactivity of the hypothalamic-pituitary-adrenal (HPA) axis. The aim of this study was to examine whether the HPA axis hyperactivity observed in hyperthyroidism may be explained by a disturbed feedback inhibition of endogenous glucocorticoids through two specific intracellular receptors in the brain: the high affinity mineralocorticoid receptor (MR) and the lower affinity glucocorticoid receptor (GR). Cytosolic receptor binding and gene expression was assessed in rats with short (7 days) and long standing (60 days) eu- and hyperthyroidism. Glucocorticoid receptor number and binding affinity (Kd) in the hippocampus were measured using [(3)H2]-dexamethasone radioreceptor assay. In situ hybridization was employed to examine the effects of hyperthyroidism on the GR and MR mRNA levels in the hippocampus and the pituitary. Both short- and long-term hyperthyroid rats showed pronounced reduction in the concentration of cytosolic GR in the hippocampus, without changes in binding affinity or changes in GR expression. In contrast, GR mRNA in the pituitary increased after 7 days and decreased after 60 days of thyroxin treatment. MR mRNA was moderately affected. Hyperthyroidism is associated with significant decreases in hippocampal GR levels supporting the hypothesis that hyperactivity of the HPA axis observed in experimentally induced hyperthyroidism may be attributed, at least in part, to decreased negative feedback at the level of the hippocampus. These findings further support the notion that a central locus is principally responsible for the hyperactivity of the HPA axis observed in hyperthyroidism.

Are receptor concentrations correlated across tissues within individuals? A case study examining glucocorticoid and mineralocorticoid receptor binding.

Hormone receptors are a necessary (although not sufficient) part of the process through which hormones like corticosterone create physiological responses. However, it is currently unknown to what extent receptor concentrations across different target tissues may be correlated within individual animals. In this study, we examined this question using a large dataset of radioligand binding data for glucocorticoid receptors (GRs) and mineralocorticoid receptors (MRs) in 13 different tissues in the house sparrow (Passer domesticus) (n=72). Our data revealed that individual house sparrows tended to exhibit higher or lower receptor binding across all tissues, which could be part of what creates the physiological and behavioral syndromes associated with different hormonal profiles. However, although statistically significant, the correlations between tissues were very weak. Thus, when each tissue was independently regressed on receptor concentrations in the other tissues, multivariate analysis revealed significant relationships only for sc fat (for GR) and whole brain, hippocampus, kidney, omental fat, and sc fat (for MR). We also found significant pairwise correlations only between receptor concentrations in brain and hippocampus, and brain and kidney (both for MR). This research reveals that although there are generalized individual consistencies in GR and MR concentrations, possibly due to such factors as hormonal regulation and genetic effects, the ability of 2 different tissues to respond to the same hormonal signal appears to be affected by additional factors that remain to be identified.

Effects of colostrum versus formula feeding on hepatic glucocorticoid and α1- and β2-adrenergic receptors in neonatal calves and their effect on glucose and lipid metabolism

Neonatal energy metabolism in calves has to adapt to extrauterine life and depends on colostrum feeding. The adrenergic and glucocorticoid systems are involved in postnatal maturation of pathways related to energy metabolism and calves show elevated plasma concentrations of cortisol and catecholamines during perinatal life. We tested the hypothesis that hepatic glucocorticoid receptors (GR) and α1- and β2-adrenergic receptors (AR) in neonatal calves are involved in adaptation of postnatal energy metabolism and that respective binding capacities depend on colostrum feeding. Calves were fed colostrum (CF; n=7) or a milk-based formula (FF; n=7) with similar nutrient content up to d 4 of life. Blood samples were taken daily before feeding and 2h after feeding on d 4 of life to measure metabolites and hormones related to energy metabolism in blood plasma. Liver tissue was obtained 2h after feeding on d 4 to measure hepatic fat content and binding capacity of AR and GR. Maximal binding capacity and binding affinity were calculated by saturation binding assays using [3H]-prazosin and [3H]-CGP-12177 for determination of α1- and β2-AR and [3H]-dexamethasone for determination of GR in liver. Additional liver samples were taken to measure mRNA abundance of AR and GR, and of key enzymes related to hepatic glucose and lipid metabolism. Plasma concentrations of albumin, triacylglycerides, insulin-like growth factor I, leptin, and thyroid hormones changed until d 4 and all these variables except leptin and thyroid hormones responded to feed intake on d 4. Diet effects were determined for albumin, insulin-like growth factor I, leptin, and thyroid hormones. Binding capacity for GR was greater and for α1-AR tended to be greater in CF than in FF calves. Binding affinities were in the same range for each receptor type. Gene expression of α1-AR (ADRA1) tended to be lower in CF than FF calves. Binding capacity of GR was related to parameters of glucose and lipid metabolism, whereas β2-AR binding capacity was negatively associated with glucose metabolism. In conclusion, our results indicate a dependence of GR and α1-AR on milk feeding immediately after birth and point to an involvement of hepatic GR and AR in postnatal adaptation of glucose and lipid metabolism in calves.