Administration Of RU486 Research Articles

Effects of RU486 in Treatment of Traumatic Stress-Induced Glucocorticoid Dysregulation and Fear-Related Abnormalities: Early versus Late Intervention.

Central glucocorticoid receptor (GR) activity is enhanced following traumatic events, playing a key role in the stress-related cognitive abnormalities of posttraumatic stress disorder (PTSD). GR antagonists are expected to have potential as pharmacological agents to treat PTSD-related symptoms such as anxiety and fear memory disruption. However, an incubation period is usually required and stress-induced abnormalities do not develop immediately following the trauma; thus, the optimal intervention timing should be considered. Single prolonged stress (SPS) was employed as a rodent PTSD model to examine the effects of early or late (1–7 versus 8–14 days after the SPS) sub-chronic RU486 (a GR antagonist) administration. Behaviorally, fear conditioning and anxiety behavior were assessed using the fear-conditioning test and elevated T-maze (ETM), respectively. Neurochemically, the expressions of GR, FK506-binding proteins 4 and 5 (FKBP4 and FKBP5), and early growth response-1 (Egr-1) were assessed in the hippocampus, medial prefrontal cortex (mPFC), amygdala, and hypothalamus, together with the level of plasma corticosterone. Early RU486 administration could inhibit SPS-induced behavioral abnormalities and glucocorticoid system dysregulation by reversing the SPS-induced fear extinction deficit, and preventing SPS-reduced plasma corticosterone levels and SPS-induced Egr-1 overexpression in the hippocampus. Early RU486 administration following SPS also increased the FKBP5 level in the hippocampus and hypothalamus. Finally, both early and late RU486 administration inhibited the elevated hippocampal FKBP4 level and hypothalamus GR level in the SPS rats. Early intervention with a GR antagonist aids in the correction of traumatic stress-induced fear and anxiety dysregulation.

The Impact of Mouse Preterm Birth Induction by RU-486 on Microglial Activation and Subsequent Hypomyelination.

Preterm birth (PTB) represents 15 million births every year worldwide and is frequently associated with maternal/fetal infections and inflammation, inducing neuroinflammation. This neuroinflammation is mediated by microglial cells, which are brain-resident macrophages that release cytotoxic molecules that block oligodendrocyte differentiation, leading to hypomyelination. Some preterm survivors can face lifetime motor and/or cognitive disabilities linked to periventricular white matter injuries (PWMIs). There is currently no recommendation concerning the mode of delivery in the case of PTB and its impact on brain development. Many animal models of induced-PTB based on LPS injections exist, but with a low survival rate. There is a lack of information regarding clinically used pharmacological substances to induce PTB and their consequences on brain development. Mifepristone (RU-486) is a drug used clinically to induce preterm labor. This study aims to elaborate and characterize a new model of induced-PTB and PWMIs by the gestational injection of RU-486 and the perinatal injection of pups with IL-1beta. A RU-486 single subcutaneous (s.c.) injection at embryonic day (E)18.5 induced PTB at E19.5 in pregnant OF1 mice. All pups were born alive and were adopted directly after birth. IL-1beta was injected intraperitoneally from postnatal day (P)1 to P5. Animals exposed to both RU-486 and IL-1beta demonstrated microglial reactivity and subsequent PWMIs. In conclusion, the s.c. administration of RU-486 induced labor within 24 h with a high survival rate for pups. In the context of perinatal inflammation, RU-486 labor induction significantly decreases microglial reactivity in vivo but did not prevent subsequent PWMIs.

Effects of P4 Antagonist RU486 on VEGF and Its Receptors' Signaling during the In Vivo Transition from the Preovulatory to Periovulatory Phase of Ovarian Follicles.

The development of an adequate blood vessel network is crucial for the accomplishment of ovarian follicle growth and ovulation, which is necessary to support the proliferative and endocrine functions of the follicular cells. Although the Vascular Endothelial Growth Factor (VEGF) through gonadotropins guides ovarian angiogenesis, the role exerted by the switch on of Progesterone (P4) during the periovulatory phase remains to be clarified. The present research aimed to investigate in vivo VEGF-mediated mechanisms by inducing the development of periovulatory follicles using a pharmacologically validated synchronization treatment carried out in presence or absence of P4 receptor antagonist RU486. Spatio-temporal expression profiles of VEGF, FLT1, and FLK1 receptors and the two major MAPK/ERKs and PI3K/AKT downstream pathways were analyzed on granulosa and on theca compartment. For the first time, the results demonstrated that in vivo administration of P4 antagonist RU486 inhibits follicular VEGF receptors’ signaling mainly acting on the theca layer by downregulating the activation of ERKs and AKTs. Under the effect of RU486, periovulatory follicles’ microarchitecture did not move towards the periovulatory stage. The present evidence provides new insights on P4 in vivo biological effects in driving vascular and tissue remodeling during the periovulatory phase.

Anti-stress effects of combined block of glucocorticoid and mineralocorticoid receptors in the paraventricular nucleus of the hypothalamus.

Mineralocorticoid receptors (MRs), glucocorticoid receptors (GRs) and corticotropin-releasing factor (CRF) in the paraventricular nucleus of hypothalamus (PVN) are involved in the response to stress. The present study investigated the role of GRs and MRs in the PVN in regulating depressive and anxiety-like behaviours. To model chronic stress, rats were exposed to corticosterone treatment via drinking water for 21 days, and GR antagonist RU486 and MR antagonist spironolactone, alone and combined, were directly injected in the PVN daily for the last 7 days of corticosterone treatment. Behavioural tests were run on days 22 and 23. Depressive- and anxiety-like behaviours were evaluated in forced swim test, sucrose preference test, novelty-suppressed feeding test and social interaction test. The expression of GRs, MRs and CRF were detected by western blot. Rats exposed to corticosterone exhibited depressive- and anxiety-like behaviours. The expression of GRs and MRs decreased, and CRF levels increased in the PVN. The intra-PVN administration of RU486 increased the levels of GRs and CRF without influencing depressive- or anxiety-like behaviours. The spironolactone-treated group exhibited an increase in MRs without influencing GRs and CRF in the PVN and improved anxiety-like behaviours. Interestingly, the intra-PVN administration of RU486 and spironolactone combined restored expression of GRs, MRs and CRF and improved depressive- and anxiety-like behaviours. In this rat model of stress, the simultaneous restoration of GRs, MRs and CRF in the PVN might play an important role in the treatment of depression and anxiety.

Mifepristone (RU486) inhibits dietary lipid digestion by antagonizing the role of glucocorticoid receptor on lipase transcription

SummaryLipid digestion and absorption are tightly regulated to cope with metabolic demands among tissues. How these processes are coordinated is not well characterized. Here, we found that mifepristone (RU486) prevents lipid digestion both in flies and mice. In flies, RU486 administration suppresses lipid digestion by transcriptional downregulating Magro in guts. Similarly, intestinal lipid uptake in mice was also suppressed by RU486 through the glucocorticoid receptor (GR). Further studies showed that the pancreatic lipase Pnlip is a direct transcriptional target of GR in pancreas tissues. Glucocorticoid levels in mice fed a high fat diet (HFD) are significantly lower than those fed on a conventional diet, and RU486 administration inhibits HFD-induced obesity both in mice and flies. Our findings identified a novel mechanism of RU486 functions as a GR antagonist systematically regulating lipid metabolism, providing new insight on the role of Glucocorticoid/GR in Cushing disease, diabetes, and other related metabolic syndromes.

The ameliorating effects of tokishakuyakusan in a rat model of implantation failure involves endometrial gland leukemia inhibitory factor and decidualization

Ethnopharmacological relevanceTokishakuyakusan (TSS) is a Kampo medicine that is prescribed for the treatment of infertility in Japan. However, its precise mechanism of action remains unclear. Aim of the studyLeukemia inhibitory factor (LIF) in the endometrium plays an indispensable role in embryo implantation and is linked to infertility or implantation failure. Previously, we demonstrated that TSS ameliorated implantation failure induced by mifepristone (RU-486), an antagonist of progesterone, in rats. Herein, we aimed to clarify whether the ameliorating effect of TSS on implantation failure in the rat model involves endometrial LIF. Additionally, we determined whether decidualization, the dysfunction of which is linked to infertility or implantation failure similar to LIF, progesterone, and other implantation-related factors, are involved in the effect of TSS. Materials and methodsThe implantation failure rat model was developed via the subcutaneous administration of RU-486 (7 mg/kg) on day 3 post-coitus. Sesame oil was administered as the vehicle control. Rats were fed a diet containing 1% or 3% TSS or a control diet from day 13 pre-coitus. Subsequently, the implantation sites were assessed, and plasma progesterone levels were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) on day 8 post-coitus. The LIF mRNA of the endometrial gland, which was segmented via laser-microdissection from the endometrial tissue, was measured, and endometrial LIF immunostaining was carried out on day 5. The gene expression of different factors related to implantation, including decidualization and progesterone-responsiveness on days 5 and 6, were measured. The human endometrial Ishikawa cell line derived from human adenocarcinoma was treated with TSS (30–300 μg/mL) for 24 h, and the LIF concentrations in the cell culture supernatants were measured. ResultsRU-486 decreased the number of implantation sites in the uterus of rats; however, the decrease was significantly alleviated by TSS (3%-diet), which tended to increase plasma progesterone. In rats with RU-486-induced implantation failure, endometrial gland LIF mRNA and endometrial LIF protein were markedly decreased while the gene expression of both decidualization-related factors such as interleukin-11, insulin-like growth factor binding protein-1, and cyclooxygenase-2, and progesterone responsive-related factors such as FK506 binding protein 5, were significantly decreased. These changes in the uterus of rats with implantation failure were significantly alleviated by TSS (3%-diet). Additionally, TSS significantly enhanced LIF protein production and LIF mRNA in Ishikawa cells. ConclusionsThe mechanism whereby TSS ameliorates RU-486-induced implantation failure in rats may involve the alleviation of decreased LIF production derived from the endometrial gland, and a dysfunction of decidualization, including lower progesterone responsiveness in the model. These findings may partly contribute to the interpretation of the beneficial effects of TSS on infertility.

Glucocorticoid receptor antagonism prevents microglia-mediated neuronal remodeling and behavioral despair following chronic unpredictable stress

Synaptic deficits and neuronal dystrophy in the prefrontal cortex (PFC) are linked to behavioral and cognitive symptoms in depressed individuals. Preclinical studies indicate that chronic stress causes synaptic deficits on pyramidal neurons in the PFC that contribute to behavioral and cognitive impairments. Our recent work shows that chronic stress provokes microglia-mediated neuronal remodeling via neuronal colony stimulating factor (CSF)-1 signaling, leading to synaptic deficits and depressive-like behaviors. Other reports indicate that elevated corticosterone causes pyramidal neuron atrophy and microglia activation in the medial PFC, implicating glucocorticoid signaling in microglia-mediated neuronal remodeling following chronic stress. In this study, male mice were exposed to chronic unpredictable stress (CUS) and received daily administration of glucocorticoid receptor antagonist RU486 (25 mg/kg, i.p.). As expected, CUS exposure caused adrenal hypertrophy and elevated plasma corticosterone levels. Glucocorticoid receptor blockade prevented behavioral despair and cognitive impairments following CUS. Moreover, RU486 administration diminished CUS-induced CSF1 signaling in the PFC and reduced markers of phagocytosis on purified microglia. Confocal imaging in Thy1-GFP(M) mice showed that CUS increased microglia-mediated neuronal remodeling, and RU486 administration attenuated microglial engulfment of neuronal elements and prevented dendritic spine density deficits on pyramidal neurons following CUS. These results demonstrate that chronic stress-induced glucocorticoid signaling promotes CSF1 signaling and microglia-mediated neuronal remodeling in the medial PFC, which contributes to development of behavioral despair and cognitive impairments. This study presents primary evidence that neuroendocrine responses engage neuron-microglia interactions in the PFC; further implicating microglia in stress-induced neuronal remodeling, PFC dysfunction, and associated behavioral consequences.

Abstract # 2066 Chronic stress-induced glucocorticoid signaling drives microglia-mediated neuronal remodeling and depressive-like behaviors

Converging evidence indicates that chronic stress elicits neurobiological responses, including pyramidal neuron remodeling and synaptic loss in the medial prefrontal cortex (PFC), that contribute to development of affective disorders. Our recent studies show that chronic stress provokes microglia-mediated neuronal remodeling via increased colony stimulating factor (CSF)-1 signaling in the medial PFC, leading to synaptic deficits and development of depressive-like behaviors. Separate studies show that elevated glucocorticoids cause neuronal remodeling and microglia activation in the medial PFC, implicating glucocorticoid receptor activation in stress-induced CSF1 signaling and microglia-mediated neuronal remodeling in the medial PFC. In the present studies C57BL/6 male mice were exposed to chronic unpredictable stress (CUS) and received daily administration of glucocorticoid receptor antagonist RU486 (25 mg/kg, i.p.). Glucocorticoid receptor blockade prevented depressive-like behaviors and cognitive impairments following CUS. Moreover, RU486 administration dampened CUS-induced CSF1 levels in the PFC and reduced CSF1 receptor along with markers of phagocytosis on sorted microglia. Confocal imaging in Thy1-GFP(M) mice revealed that CUS increased microglia-mediated neuronal remodeling and reduced spine density on apical dendrites of pyramidal neurons in the medial PFC. Notably, RU486 administration reduced microglia engulfment of neuronal elements and prevented dendritic spine density deficits on pyramidal neurons after CUS. These findings demonstrate that chronic stress-induced glucocorticoid signaling drives CSF1 signaling in the medial PFC, which provokes microglia-mediated neuronal remodeling and subsequent depressive-like behaviors and cognitive impairments.

Cortisol promotes staining-type hypermelanosis in juvenile Japanese flounder

Hypermelanosis of the blind side of Japanese flounder frequently occurs in post-metamorphic juveniles reared in hatcheries. Due to the morphological similarity between darkened blind-side skin and normal ocular-side skin, hypermelanosis is regarded as a transition from normal blind-side skin to that of the ocular side. Hypermelanosis appears only in rearing tanks without sand on the bottom, and its incidence tends to increase under stressful conditions; however, the specific mechanisms responsible for its occurrence have not been identified to date. In teleosts, the hypothalamus-pituitary-interrenal axis (HPI axis) is activated under stress, which then accelerates cortisol secretion. Therefore, in this study, we orally administered cortisol to Japanese flounder to examine its effect on hypermelanosis. The results of our study indicated that cortisol promoted hypermelanosis in a dose-dependent manner. In addition, the effect of cortisol was canceled out in juveniles reared in tanks with bottom sand, suggesting that the well-documented inhibitory effect of bottom sand is stronger than the stimulatory effect of cortisol (i.e., stress) on hypermelanosis. However, administration of the cortisol antagonist RU486 failed to inhibit hypermelanosis, suggesting the contribution of other stimulatory factors to this process. Therefore, our findings strongly suggest that stress-induced cortisol release has the adverse effect of promoting hypermelanosis in juvenile flounders reared without bottom sand.

Early-life Photoperiod Influences Depression-like Behavior, Prepulse Inhibition of the Acoustic Startle Response, and Hippocampal Astrogenesis in Mice

Environmental factors during early life stages affect behavioral and physiological phenotypes in adulthood. We examined the effect of photoperiods during development on neurogenesis and affective behaviors during adolescence/adulthood using C57BL/6J mice. Mice were born and raised until weaning under long-day conditions (LDs) or short-day conditions (SDs), followed by a 12L12D cycle until adulthood. Adult mice born under SD showed a shorter latency to first immobility in the forced swim test when compared with the mice born under LD. The mice born under SD also exhibited significantly lower prepulse inhibition, which is a characteristic of schizophrenia. However, the mice exposed to SD and LD during the prenatal period only did not show differences in prepulse inhibition. At 4 weeks of age, there were less 5-bromo-2′-deoxyuridine (BrdU)-positive cells in the dentate gyrus (DG) of the hippocampus of mice born under SD when compared with mice born under LD. Double immunostaining showed that the mice born under SD showed less BrdU/glial fibrillary acidic protein (GFAP, an astrocyte marker) cells when compared with mice born under LD. Furthermore, expression of the glucocorticoid receptor in the DG was higher in mice born under SD, and the photoperiod-dependent changes in the number of BrdU-positive cells in the DG were abolished by administration of RU486, a glucocorticoid receptor antagonist. These results suggest that the photoperiod in early life alters astrogenesis in the hippocampus via the hypothalamic–pituitary-adrenal axis and may relate to affective behaviors in adulthood.

Effects of heat-sensitive moxibustion on HPA axis in rats with irritable bowel syndrome

To observe the effects of heat-sensitive moxibustion on corticotropin releasing hormone (CRH), adrenocorticotrophic hormone (ACTH) and corticosterone (CORT) in rats with irritable bowel syndrome (IBS), and to explore the possible mechanism of heat-sensitive moxibustion on IBS. According to random number table, 56 SD male rats were randomly divided into a blank group (n=8), a model group (n=8), a moxibustion group (n=32), and a mifepristone group (RU-486 group, n=8). The rats in the blank group were treated with normal feeding; the rats in the model group, RU-486 group and moxibustion group were treated with chronic non-predictable stimulation for 21 days to establish IBS model. After model establishment, the rats in the moxibustion group were treated with moxibustion at "Mingmen" (GV 4) for 40 min, once a day for 14 days; the tail temperature was recorded every 5 min; according to the change of tail temperature, the rats were divided into a heat-sensitive moxibustion group and a non-heat-sensitive moxibustion group, and 8 rats were randomly selected in the two groups. The rats in the RU-486 group were treated with gastric administration of RU-486 for 14 days, while the rats in the blank group, model group and moxibustion groups were treated with identical volume of 0.9% NaCl. The rat general condition, body mass, behavioristics, intestinal propulsive rate and visceral sensitivity were observed in each group; ELISA method was used to detect serum CRH, ACTH and CORT; optical microscope was applied to observe the morphological changes of colon. (1) After model establishment, rats were in rest state, fatigued, with withered hair and dim ear; the stool was dry or watery; the body mass were slowly increased; the number of crossed grid and standing frequency were significantly reduced; visceral sensitivity was increased and intestinal propulsion rate was decreased; no obvious inflammatory cell infiltration was observed under microscope. (2) After intervention, compared with the blank group, the body mass and visceral sensitivity in the RU-486 group were not significantly different (both P>0.05), but the intestinal propulsion rate was decreased significantly (P<0.01). Compared with the blank group, the body mass of heat-sensitive moxibustion group and non-heat-sensitive moxibustion group was lower (both P<0.01), but the visceral sensitivity and intestinal propulsion rate were similar (both P>0.05). Compared with the model group, the body mass and visceral sensitivity were improved in the RU-486 group (P<0.05, P<0.01), but the intestinal propulsion rate was similar (P>0.05). The body mass, visceral sensitivity and intestinal propulsion rate of the heat-sensitive moxibustion group and the non-heat-sensitive moxibustion group were superior to those of the model group (P<0.05, P<0.01), and the body mass and intestinal propulsion rate of heat-sensitive moxibustion group were superior to those of non-heat-sensitive moxibustion group (both P<0.05). (3) After intervention, compared with the blank group, the contents of CRH, ACTH and CORT in the model group were significantly increased (P<0.05, P<0.01). Compared with the model group, the contents of CRH, ACTH and CORT of the heat-sensitive moxibustion group were statistically reduced (P<0.05, P<0.01), and the contents of CRH and ACTH in the non-heat-sensitive moxibustion group were statistically reduced (P<0.05, P<0.01); the content of CRH in the RU-486 group was reduced (P<0.05), but the contents of ACTH and CORT were increased (P<0.05, P<0.01). Compared with the non-heat-sensitive moxibustion group, the heat-sensitive moxibustion group was better in the improvement of CRH (P<0.05), but there was no significant difference of ACTH and CORT between the two groups (both P>0.05). Heat-sensitive moxibustion could reduce the contents of CRH, ACTH and CORT through the HPA axis, and improve the function of gastrointestinal motility to treat IBS.

Prostaglandin-Endoperoxide Synthase 1 Mediates the Timing of Parturition in Mice Despite Unhindered Uterine Contractility.

Cyclooxygenase (COX)-derived prostaglandins stimulate uterine contractions and prepare the cervix for parturition. Prior reports suggest Cox-1 knockout (KO) mice exhibit delayed parturition due to impaired luteolysis, yet the mechanism for late-onset delivery remains unclear. Here, we examined key factors for normal onset of parturition to determine whether any could account for the delayed parturition phenotype. Pregnant Cox-1KO mice did not display altered timing of embryo implantation or postimplantation growth. Although messenger RNAs of contraction-associated proteins (CAPs) were differentially expressed between Cox-1KO and wild-type (WT) myometrium, there were no differences in CAP agonist-induced intracellular calcium release, spontaneous or oxytocin (OT)-induced ex vivo uterine contractility, or in vivo uterine contractile pressure. Delayed parturition in Cox-1KO mice persisted despite exogenous OT treatment. Progesterone (P4) withdrawal, by ovariectomy or administration of the P4-antagonist RU486, diminished the delayed parturition phenotype of Cox-1KO mice. Because antepartum P4 levels do not decline in Cox-1KO females, P4-treated WT mice were examined for the effect of this hormone on in vivo uterine contractility and ex vivo cervical dilation. P4-treated WT mice had delayed parturition but normal uterine contractility. Cervical distensibility was decreased in Cox-1KO mice on the day of expected delivery and reduced in WT mice with long-term P4 treatment. Collectively, these findings show that delayed parturition in Cox-1KO mice is the result of impaired luteolysis and cervical dilation, despite the presence of strong uterine contractions.

Dexamethasone reduces side population fraction through downregulation of ABCG2 transporter in MCF-7 breast cancer cells.

Side population (SP) cells represent a rare population among breast cancer cells. SP cells have been reported to act as cancer stem‑like cells, and to participate in the development of multidrug resistance via modulating the expression of ATP-binding cassette subfamily G member 2 (ABCG2). Dexamethasone is a corticosteroid drug that has been used as an adjuvant treatment to enhance the efficacy of chemotherapeutic agents; however, its effects in breast cancer have yet to be thoroughly investigated. In the present study, the effects of dexamethasone were investigated using the human MCF‑7 breast cancer cell line, and SPs were examined in detail. Cellular proliferation, SP fractions and ABCG2 expression were examined following treatment of MCF‑7 cells with dexamethasone. Dexamethasone was revealed to cause a dose‑ and time‑dependent decrease in cancer cell proliferation, and it also decreased the size of the SP fraction of MCF‑7 cells and the expression of the ABCG2 transporter. The effects of dexamethasone on cellular proliferation, SP fraction and ABCG2 expression were abolished following the administration of the glucocorticoid antagonist RU486. These results suggested that dexamethasone may target breast cancer cell SPs and thus increase the sensitivity of tumor cells to chemotherapy. Therefore, it may be hypothesized that dexamethasone can be used as a chemosensitizer in the adjuvant treatment of patients with breast cancer.

Testicular germ cells apoptosis following exposure to chronic stress in rats

ObjectiveTo evaluate the effects of chronic stress on testosterone hormone level and germ cells apoptosis in testes and the inhibitory role of glucocorticoid receptor antagonist, RU486. MethodsAdult male Sprague–Dawley rats were randomly assigned to four groups (n = 6): control; stress; RU486; stress/RU486. Animals in RU486 and stress/RU486 subjected to subcutaneous injections of 2.5 mg RU486/kg 1 h before stress session. Rats were submitted to chronic restraint stress (1 h daily for 12 consecutive days) whereas control animals were not subjected to stress. Serum testosterone assay was performed and the occurrence of DNA fragmentation in the testis sections was examined using TUNEL staining. ResultsChronic restraint stress significantly induced a decrease in serum testosterone level with increase in apoptosis in spermatogonia. Systemic administration of RU486 significantly restored serum testosterone levels and attenuates stress-induced apoptosis in spermatogonia. ConclusionThe restraint stress-induced change in serum testosterone levels and seminiferous tubules apoptosis closely associated with the glucocorticoid receptor.

Glucocorticoid receptor mediated the propofol self-administration by dopamine D1 receptor in nucleus accumbens

Propofol, a widely used anesthetic, can cause addictive behaviors in both human and experimental animals. In the present study, we examined the involvement of glucocorticoid receptor (GR) signaling in the molecular process by which propofol may cause addiction. The propofol self-administration model was established by a fixed ratio 1 (FR1) schedule of reinforced dosing over successive 14days in rats. On day 15, the rats were treated with dexamethasone, a GR agonist (10–100μg/kg), or RU486, a GR antagonist (10–100μg/kg) at 1h prior to the last training. The animal behaviors were recorded automatically by the computer. The expression of dopamine D1 receptor in the nucleus accumbens (NAc) was examined by Western blot and the concentrations of plasma corticosterone were measured by enzyme-linked immunosorbent assay (ELISA). To further examine the specificity of GR in the process, mineralocorticoid receptor (MR) antagonist, spironolactone, and dexamethasone plus MR agonist, aldosterone, were also tested. Administration of dexamethasone (100μg/kg) or RU486 (⩾10mg/kg) significantly attenuated the rate of propofol maintained active nose-poke responses and infusions, which were accompanied by reductions in both plasma corticosterone level and the expression of D1 receptor in the NAc. Neither spironolactone alone nor dexamethasone combined with aldosterone affected the propofol-maintaining self-administrative behavior, indicating GR, but not MR, modulates the propofol reward in rats. In addition, neither the food-maintaining sucrose responses under FR1 schedule nor the locomotor activity was affected by any doses of dexamethasone or RU486 tested. These findings provide evidence that GR signaling may play an important role in propofol reward.

RU486 facilitates or disrupts the sensitization of sexual behaviors by estradiol in the ovariectomized Long–Evans rat: Effect of timecourse

An acute injection of estradiol benzoate (EB) to the ovariectomized (OVX) rat activates low levels of lordosis, and subsequent progesterone (P) administration augments lordosis and recruits a complete pattern of sexual behavior including appetitive behaviors (e.g., hops/darts and solicitations). However, repeated injections of 5μg or 10μg EB (but not 2μg EB), administered every 4days to sexually-experienced OVX rats results in a behavioral sensitization, such that lordosis quotients (LQs) and appetitive behaviors progressively increase. We have shown that adrenal P does not play a critical role because behavioral sensitization to EB is not prevented by adrenalectomy. Here we tested whether P receptors play a role by examining the effect of chronic administration of the P receptor antagonist RU486 at a dose that reliably inhibits sexual behavior in fully primed OVX rats. Females were treated with EB (5 or 10μg), and 5mg RU486 dissolved in 0.4mL vehicle (VEH; 80% sesame oil, 15% benzyl benzoate, 5% benzyl alcohol) 48h and 5h prior to each of 7 tests, respectively, occurring at 4-day intervals in unilevel 4-hole pacing chambers. Control animals were treated with 2, 5, or 10μg EB+VEH. As expected, sensitization did not occur in females treated with 2μg EB+VEH, and those females received fewer intromissions and ejaculations than all other groups. RU486 did not prevent the sensitization of LQ, moderate and high lordosis magnitudes (LM2 and LM3) or appetitive sexual behaviors on early tests, and in fact potentiated appetitive behaviors, LQ, LM2 and LM3, consistent with its facilitative actions in females treated with EB-alone, as we and others have reported previously. However, despite the initial facilitation, blocking P receptors by chronic administration of RU486 inhibited the maintenance of behavioral sensitization to EB.

Blockade of glucocorticoid receptors with RU486 attenuates cardiac damage and adipose tissue inflammation in a rat model of metabolic syndrome.

Glucocorticoids are stress hormones that modulate metabolic, inflammatory and cardiovascular processes. We recently characterized DahlS.Z-Lepr(fa)/Lepr(fa) (DS/obese) rats, derived from a cross between Dahl salt-sensitive (DS) and Zucker rats, as a new animal model of metabolic syndrome (MetS). We have now investigated the effects of glucocorticoid receptor (GR) blockade on cardiac and adipose tissue pathology and gene expression, as well as on glucose metabolism in this model. DS/obese rats were treated with the GR blocker RU486 (2 mg kg(-1) per day, subcutaneous) for 4 weeks beginning at 9 weeks of age. Age-matched homozygous lean (DahlS.Z-Lepr(+)/Lepr(+), or DS/lean) littermates of DS/obese rats served as controls. Treatment of DS/obese rats with RU486 attenuated left ventricular (LV) fibrosis and diastolic dysfunction, as well as cardiac oxidative stress and inflammation, without affecting hypertension or LV hypertrophy. Administration of RU486 to DS/obese rats also inhibited the upregulation of GR and 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) expression at the mRNA and protein levels in the heart; it attenuated adiposity and adipose tissue inflammation, as well as the upregulation of GR and 11β-HSD1 mRNA and protein expression in adipose tissue; it ameliorated fasting hyperinsulinemia as well as insulin resistance and glucose intolerance. Our results thus implicate the glucocorticoid-GR axis in the pathophysiology of MetS, and they suggest that GR blockade has therapeutic potential for the treatment of this condition.

Placental claudin expression and its regulation by endogenous sex steroid hormones

Tight junctions (TJs) form continuous intercellular contacts controlling the paracellular transportation across the cell-to-cell junction. TJ components include the peripheral protein zonula occludens-1 (ZO-1), junctional adhesion molecules (JAMs), and integral proteins such as occludin and claudins. Among the junction proteins, claudins play a major role in regulation of paracellular electrolyte transportation. This study explores the expression and distribution of tight junctions and their regulation during pregnancy. To study the regulation of claudin family, we examined expression of mouse placental tight junction proteins, including claudin-1 to -24, with real-time PCR and Western blotting and distribution of tight junction proteins with immunohistochemistry. Pregnant C57/BL6 mice were used in this study. The pregnant mice were divided into three groups depending on pregnant day (on days 12, 16, and 20 of gestation). Regarding the transcription levels, claudin-1, claudin-2, claudin-4, and claudin-5 expression levels were relatively high compared to other claudin family in all periods of pregnancy. Claudin-4 and 5 expressions, which reduce ion permeability, were increased over a period of time. However, claudin-2 expression, that is the responsive protein for a decrease in paracellular conductance, was decreased. Following this modulation of expression during mid-term pregnancy, we identified endogenous hormonal modulation of claudin family using estrogen receptor antagonist ICI 182,780 and progesterone receptor antagonist RU-486. After administration of ICI and RU-486, expression of claudin-4 mRNA and protein was increased. In addition, immunohistochemistry was performed to identify their localization for inferring permeability in placenta. Due to the function of claudins as effectors of ion transport at the end of regulatory pathways, they must be transducing proteins that modulate the function of claudins and thus link the physiologic inputs to the final effectors. This study will provide the claudin expressions and their localization in the mouse placenta, and their regulation by endogenous hormones. Taken together, the results of this study may contribute to assuming the roles and regulatory mechanism of these tight junction genes regarding maternal-fetal ion transportation in the placenta.

Dexamethasone potentiates in vitro blood-brain barrier recovery after primary blast injury by glucocorticoid receptor-mediated upregulation of ZO-1 tight junction protein.

Owing to the frequent incidence of blast-induced traumatic brain injury (bTBI) in recent military conflicts, there is an urgent need to develop effective therapies for bTBI-related pathologies. Blood-brain barrier (BBB) breakdown has been reported to occur after primary blast exposure, making restoration of BBB function and integrity a promising therapeutic target. We tested the hypothesis that treatment with dexamethasone (DEX) after primary blast injury potentiates recovery of an in vitro BBB model consisting of mouse brain endothelial cells (bEnd.3). DEX treatment resulted in complete recovery of transendothelial electrical resistance and hydraulic conductivity 1 day after injury, compared with 3 days for vehicle-treated injured cultures. Administration of RU486 (mifepristone) inhibited effects of DEX, confirming that barrier restoration was mediated by glucocorticoid receptor signaling. Potentiated recovery with DEX treatment was accompanied by stronger zonula occludens (ZO)-1 tight junction immunostaining and expression, suggesting that increased ZO-1 expression was a structural correlate to BBB recovery after blast. Interestingly, augmented ZO-1 protein expression was associated with specific upregulation of the α(+) isoform but not the α(-) isoform. This is the first study to provide a mechanistic basis for potentiated functional recovery of an in vitro BBB model because of glucocorticoid treatment after primary blast injury.

Enhanced corticosteroid signaling alters synaptic plasticity in the dentate gyrus in mice lacking the fragile X mental retardation protein

The fragile X mental retardation protein (FMRP) is an important regulator of protein translation, and a lack of FMRP expression leads to a cognitive disorder known as fragile X syndrome (FXS). Clinical symptoms characterizing FXS include learning impairments and heightened anxiety in response to stressful situations. Here, we report that, in response to acute stress, mice lacking FMRP show a faster elevation of corticosterone and a more immediate impairment in N-methyl-d-aspartate receptor (NMDAR) dependent long-term potentiation (LTP) in the dentate gyrus (DG). These stress-induced LTP impairments were rescued by administering the glucocorticoid receptor (GR) antagonist RU38486. Administration of RU38486 also enhanced LTP in Fmr1−/y mice in the absence of acute stress to wild-type levels, and this enhancement was blocked by application of the NMDAR antagonist 2-amino-5-phosphonopentanoic acid. These results suggest that a loss of FMPR results in enhanced GR signaling that may adversely affect NMDAR dependent synaptic plasticity in the DG.