β-Lapachone attenuates cognitive impairment and neuroinflammation in beta-amyloid induced mouse model of Alzheimer's disease

Oxidative stress and neuroinflammation are critically involved in amyloid beta (Aβ) induced cognitive impairments. β-Lapachone (β-LAP) is a natural activator of NAD(P)H quinone oxidoreductase 1 (NQO1) which has antioxidant and anti-inflammatory properties. This study investigated the effect of β-LAP administration on Aβ-induced memory deficit, oxidative stress, neuroinflammation, and apoptosis cell death in the hippocampus. Forty BALB/c mice were allocated into control, sham, β-LAP (βL), Aβ, and Aβ + βL groups. Intracerebroventricular injection of Aβ_1-42 was used to induce Alzheimer’s disease (AD) model. Mice in the βL and Aβ + βL groups were treated with β-LAP (10 mg/kg, i.p) for 4 days. Results revealed that β-LAP attenuated memory impairment in the Aβ-received mice, as measured in the novel object recognition (NOR) and Barnes maze tests. Moreover, Aβ resulted in inflammasome activation evident by enhanced caspase-1 immunoreactivity and interleukin-1 beta (IL-1β) protein levels. However, β-LAP could markedly reduce reactive oxygen species (ROS) production and down-regulate mRNA expression of NLRP3 inflammasome and protein levels of cleaved caspase 1 and IL-1β. Additionally, β-LAP-treated mice showed increased SIRT1 levels and NAD⁺/NADH ratio in the hippocampus. These results were followed by fewer number of TUNEL-positive cell, reduced hippocampal atrophy and neuronal loss in the hippocampal dentate gyrus (DG). These results indicated that the protective effect of β-LAP against AD-associated cognitive deficits is partially through its strong antioxidant and anti-inflammatory actions.     

NecroX-5 alleviate lipopolysaccharide-induced acute respiratory distress syndrome by inhibiting TXNIP/NLRP3 and NF-κB

The activation of NLRP3 inflammasome and NF-κB pathway, associating with oxidative stress, have been implicated in the development of acute lung injury (ALI) and acute respiratory distress syndrome (ARDS). NecroX-5 has been reported to exhibit the effects of anti-oxidation and anti-stress in various diseases. However, the role of NecroX-5 in ALI has not been explicitly demonstrated. The aim of this study was to explore the therapeutic effects and potential mechanism action of NecroX-5 on ALI. Here, we found that NecroX-5 pretreatment dramatically diminished the levels of IL-1β, IL-18 and ROS in in RAW264.7 cells challenged with LPS and ATP. Furthermore, NecroX-5 suppressed the activation of NLRP3 inflammasome and NF-κB signal pathway. In addition, NecroX-5 also inhibited the thioredoxin-interacting protein (TXNIP) expression. In vivo, NecroX-5 reduced the LPS-induced lung histopathological injury, the number of TUNEL-positive cells, lung wet/dry (W/D) ratio, levels of total protein and inflammatory cytokines in the bronchoalveolar lavage fluid (BALF) in mice. Additionally, LPS-induced upregulation of myeloperoxidase (MPO), ROS production and malondialdehyde (MDA) were inhibited by NecroX-5 administration. Thus, our results demonstrate that NecroX-5 protects against LPS-induced ALI by inhibiting TXNIP/NLRP3 and NF-κB.     

Quercetin attenuates NLRP3 inflammasome activation and apoptosis to protect INH-induced liver injury via regulating SIRT1 pathway

Severe hepatotoxicity greatly limits the clinical application of the first-line anti-tuberculosis drug isoniazid(INH). Quercetin(Que) has multiple pharmacological properties, and is regarded as a potential protective agent against a variety of organ injuries. However, the exact effect of quercetin on INH-induced hepatotoxicity and the underlying mechanisms are not yet completely understood. In this study, liver injury models were established in rats and L02 cells to reveal the protective effect of Que on INH-induced hepatotoxicity and the relevant mechanism. The in vivo results indicated that Que pretreatment reduced the level of ALT/AST, improved the liver histopathological changes and substantially mitigated apoptosis in rats. In vitro, it evidently relieved INH-induced cell viability loss and apoptosis in L02 cells. Furthermore, the studies on mechanisms elucidated that Que remarkably elevated the expression of SIRT1 and suppressed NLRP3 inflammasome activation. Meanwhile, Que significantly inhibited the level of tumor suppressor P53, Bax, cleaved-cas3 expressionl and increased Bcl-2 expression to reduce apoptosis in vivo and in vitro. However, SIRT1 inhibitor EX527 reversed the suppression of Que on NLRP3 inflammasome activation and the protection of Que on rat liver injury and cell apoptosis. In short, our findings showed that Que exhibited protective effects against INH-induced liver damage via inhibiting the activation of NLRP3 inflammasome and apoptosis in a SIRT-dependent manner.     

Reversal of the deleterious effects of chronic dietary HFCS-55 intake by PPAR-δ agonism correlates with impaired NLRP3 inflammasome activation

Although high-fructose corn syrup (HFCS-55) is the major sweetener in foods and soft-drinks, its potential role in the pathophysiology of diabetes and obesity (“diabesity”) remains unclear. Peroxisome-proliferator activated receptor (PPAR)-δ agonists have never been tested in models of sugar-induced metabolic abnormalities. This study was designed to evaluate (i) the metabolic and renal consequences of HFCS-55 administration (15% wt/vol in drinking water) for 30 weeks on male C57Bl6/J mice and (ii) the effects of the selective PPAR-δ agonist GW0742 (1 mg/kg/day for 16 weeks) in this condition. HFCS-55 caused (i) hyperlipidemia, (ii) insulin resistance, and (iii) renal injury/inflammation. In the liver, HFCS-55 enhanced the expression of fructokinase resulting in hyperuricemia and caused abnormalities in known insulin-driven signaling events. In the kidney, HFCS-55 enhanced the expression of the NLRP3 (nucleotide-binding domain and leucine-rich-repeat-protein 3) inflammasome complex, resulting in caspase-1 activation and interleukin-1β production. All of the above effects of HFCS-55 were attenuated by the specific PPAR-δ agonist GW0742. Thus, we demonstrate for the first time that the specific PPAR-δ agonist GW0742 attenuates the metabolic abnormalities and the renal dysfunction/inflammation caused by chronic HFCS-55 exposure by preventing upregulation of fructokinase (liver) and activation of the NLRP3 inflammasome (kidney).     

Dexmedetomidine promotes liver regeneration in mice after 70% partial hepatectomy by suppressing NLRP3 inflammasome not TLR4/NFκB

Inflammasome activation is mediated by NOD-like receptors (NLRs) that play important role in cellular proliferation. NLRP3 senses the widest array of stimuli. But its role in the liver regeneration after partial hepatectomy (PHx) is still unknown. Dexmedetomidine (Dex) has been documented to protect the liver against ischemia-reperfusion injury via the suppression of the TLR4/NF-κB pathway, which is important for NLRP3 inflammasome activation and liver regeneration. We tested whether Dex contributes to liver regeneration, and investigated its consequent effect on inflammasome activation. In vitro, L02 human liver cells were treated with Dex at different concentrations. The 70% PHx was performed in C57 BL/6 mice as PHx group, and sham-operated animals as Sham group, Dex-treated animals were assigned into two groups: Dex + PHx, which received single intraperitoneal injections of Dex (25 μg/kg) before PHx 30 mins; Dex + PHx + Dex, which received additional Dex (25 μg/kg) after PHx for 3 days. Dex significantly inhibited the proliferation of Lo2 cells in vitro and decreased the expression of TLR4/NFκB. In vivo, Dex + PHx exhibited promoted effect on liver regeneration and liver function recovery via inhibiting NLRP3 inflammasome activation. Dex + PH + Dex inhibited the liver regeneration, which may be associated with suppressed expression levels of TLR4/NFκB pathway. Though Dex pretreatment contributed to liver regeneration and function recovery via inflammation suppression, excessive inflammation suppression accompanied with TLR4 suppression could be related to the diminished liver regeneration, suggesting that TLR4/NFκB played important role in liver regeneration and Dex + PHx might be a useful therapeutic strategy to promote liver regeneration in clinical.     

Dexamethasone alleviate allergic airway inflammation in mice by inhibiting the activation of NLRP3 inflammasome

Dexamethasone (DEX) is the mainstay treatment for asthma, which is a common chronic airway inflammation disease. However, the mechanism of DEX resolute symptoms of asthma is not completely clear. Here, we aimed to analyze the effect of DEX on airway inflammation in OVA-induced mice and whether this effect is related to the inhibition of the activation of NLRP3 inflammasome. Female (C57BL/6) mice were used to establish the allergic airway inflammation model by inhalation OVA. The number of inflammatory cells in the bronchi alveolar lavage fluid (BALF) was counted by Swiss-Giemsa staining, and the contents of IL-1β, IL-18, IL-5 and IL-17 were detected by ELISA. The degree of inflammatory cells infiltration and mucous cells proliferation in lung tissue were separately observed by H&E and PAS staining. The proteins expression of NLRP3, pro-caspase-1, caspase-1, IL-1β, IL-6 and IL-17 in lung tissue were detected by Western blotting. We found that DEX significantly inhibited OVA-induced inflammatory cells infiltration, airway mucus secretion and goblet cell proliferation in mice. The total and classified numbers of inflammatory cells and the levels of IL-1β, IL-18, IL-5 and IL-17 in the BALF of the experimental group were significantly lower than those of the model group after DEX treatment. DEX also significantly inhibited the activity of NLRP3 inflammasome and reduced the protein contents of Pro-Caspase-1, Caspase-1, Capase-1/Pro-Caspase-1, IL-1β, IL-6 and IL-17 in lung tissues. Our study suggested that DEX alleviates allergic airway inflammation by inhibiting the activity of NLRP3 inflammasome and the levels of IL-1β and IL-18.     

Evaluation of cellular response to perfluorooctane sulfonate in human umbilical vein endothelial cells

Perfluorooctane sulfonate (PFOS), a stable fluorosurfactant and global pollutant, can be bioaccumulated. Recently, the possible adverse effects of PFOS on human health have raised concern. In this study, we examined the effects of PFOS on the expression of three important receptors related to endocrine-disrupting chemicals, six inflammatory-related genes, the production of reactive oxygen species (ROS) and the ability of cell adhesion in human umbilical vein endothelial cells (HUVECs). The results demonstrated that the expression of PPARγ and ERα were up-regulated after exposure to PFOS. After a high dose exposure (100 mg/L) for 48 h, the expression of IL-1β, IL-6, COX-2, NOS3, ICAM-1 and P-Selectin were all notably up-regulated, whereas all of them did not show any significant changes after a low dose exposure (50 mg/L) for 24 h. Moreover, the expression of IL-1β, COX-2 and NOS3 were significantly up-regulated, when cells were exposed to 100 mg/L PFOS for 24 h. Meanwhile, the amount of ROS induced by the exposure to high-dose PFOS was significantly increased with increasing incubation times. Furthermore, the adhesion of THP-1 cells onto HUVECs was significantly increased after exposure to 100 mg/L PFOS for 48 h, as observed for the expression of ICAM-1 and P-Selectin. In conclusion, our data suggest that PFOS exposure may play an important role in the vascular inflammatory disorders and endothelial dysfunctions.     

HIV-1 Viral Protein R Activates NLRP3 Inflammasome in Microglia: implications for HIV-1 Associated Neuroinflammation

Human Immunodeficiency virus (HIV) enters the brain soon after seroconversion and induces chronic neuroinflammation by infecting and activating brain macrophages. Inflammasomes are cytosolic protein complexes that mediate caspase-1 activation and ensuing cleavage and release of IL-1β and ?18 by macrophages. Our group recently showed that HIV-1 infection of human microglia induced inflammasome activation in NLRP3-dependent manner. The HIV-1 viral protein R (Vpr) is an accessory protein that is released from HIV-infected cells, although its effects on neuroinflammation are undefined. Infection of human microglia with Vpr-deficient HIV-1 resulted in reduced caspase-1 activation and IL-1β production, compared to cells infected with a Vpr-encoding HIV-1 virus. Vpr was detected at low nanomolar concentrations in cerebrospinal fluid from HIV-infected patients and in supernatants from HIV-infected primary human microglia. Exposure of human macrophages to Vpr caused caspase-1 cleavage and IL-1β release with reduced cell viability, which was dependent on NLRP3 expression. Increased NLRP3, caspase-1, and IL-1β expression was evident in HIV-1 Vpr transgenic mice compared to wild-type littermates, following systemic immune stimulation. Treatment with the caspase-1 inhibitor, VX-765, suppressed NLRP3 expression with reduced IL-1β expression and associated neuroinflammation. Neurobehavioral deficits showed improvement in Vpr transgenic animals treated with VX-765. Thus, Vpr-induced NLRP3 inflammasome activation, which contributed to neuroinflammation and was abrogated by caspase-1 inhibition. This study provides a new therapeutic perspective for HIV-associated neuropsychiatric disease.     

Inhibiting NLRP3 inflammasome with MCC950 ameliorates perioperative neurocognitive disorders,suppressing neuroinflammation in the hippocampus in aged mice

Perioperative neurocognitive disorders (PND) are characterized by deficits in cognitive functions in the elderly following anesthesia and surgery. Effective clinical interventions for preventing this disease are limited. Growing evidence demonstrates that activation of NOD-like receptor protein3 (NLRP3) inflammasome is involved in neurodegenerative diseases. We therefore hypothesized that activation of NLRP3 inflammasome is linked to neuroinflammation and the subsequent cognitive impairments that occurred in an animal model of PND. In this study, 18-month-old C57BL/6 mice were subjected to an exploratory laparotomy under isoflurane anesthesia to mimic clinical human abdominal surgery. For interventional studies, mice received NLRP3 specific inhibitor MCC950 (10 mg/kg) or the vehicle only intraperitoneally. Behavioral studies were performed at 6 and 7 d after surgery using open field and fear conditioning tests, respectively. Interleukin-1β (IL-1β), interleukin-18 (IL-18), tumor necrosis factor-α (TNF-α), ionized calcium-binding adaptor molecule-1 (IBA1) positive cells, glial fibrillary acidic protein (GFAP) positive cells, NLRP3, apoptosis-associated speck-like protein containing a CARD (ASC), and cleaved caspase-1 were measured at 3 days post-surgery. Brain-derived neurotrophic factor (BDNF) and postsynaptic density protein 95 (PSD95) were measured at 7 days post-surgery. Our data indicates that surgery-induced cognitive impairments were associated with significant increases in IL-1β, IL-18, TNF-α, NLRP3, ASC, cleaved caspase-1, IBA1-positive cells and GFAP-positive cells, and decreases in BDNF and PSD95 expression in the hippocampus. Notably, administration with MCC950 attenuated inflammatory changes and rescued surgery-induced cognitive impairments. Our study suggests that surgery induces neuroinflammation and cognitive deficits that are partly attributed to the activation of NLRP3 inflammasome in the hippocampus of aged mice.     

Evaluation of EpiDerm full thickness-300 (EFT-300) as an in vitro model for skin irritation: Studies on aliphatic hydrocarbons

The aim of this study was to understand the skin irritation effects of saturated aliphatic hydrocarbons (HCs), C9–C16, found jet fuels using in vitro 3-dimensional EpiDerm full thickness-300 (EFT-300) skin cultures. The EFT-300 cultures were treated with 2.5 μl of HCs and the culture medium and skin samples were collected at 24 and 48 h to measure the release of various inflammatory biomarkers (IL-1α, IL-6 and IL-8). To validate the in vitro results, in vivo skin irritation studies were carried out in hairless rats by measuring trans epidermal water loss (TEWL) and erythema following un-occlusive dermal exposure of HCs for 72 h. The MTT tissue viability assay results with the EFT-300 tissue show that 2.5 μl/tissue (≈4.1 μl/cm²) of the HCs did not induce any significant changes in the tissue viability for exposure times up to 48 h of exposure. Microscopic observation of the EFT-300 cross-sections indicated that there were no obvious changes in the tissue morphology of the samples at 24 h, but after 48 h of exposure, tridecane, tetradecane and hexadecane produced a slight thickening and disruption of stratum corneum. Dermal exposures of C12–C16 HCs for 24 h significantly increased the expression of IL-1α in the skin as well as in the culture medium. Similarly, dermal exposure of all HCs for 24 h significantly increased the expression of interleukin-6 (IL-6) and IL-8 in the skin as well as in the culture medium in proportion to the HC chain length. As the exposure time increased to 48 h, IL-6 concentrations increased 2-fold compared to the IL-6 values at 24 h. The in vivo skin irritation data also showed that both TEWL and erythema scores increased with increased HCs chain length (C9–C16). In conclusion, the EFT-300 showed that the skin irritation profile of HCs was in the order of C9 ⩽ C10 ⩽ C11 ⩽ C12 < C13 ≈ C14 ≈ C16 and that the tissue was an excellent in vitro model to predict in vivo irritation and to understand the structural activity relationship of HCs.     

PFOS facilitates liver inflammation and steatosis: An involvement of NLRP3 inflammasome-mediated hepatocyte pyroptosis

Perfluorooctane sulfonate (PFOS) is a fluorinated organic pollutant with substantial accumulation in mammalian liver tissues. However, the impact of chronic PFOS exposure on liver disease progression and the underlying molecular mechanisms remain elusive. Herein, we for the first time revealed that micromolar range of PFOS exposure initiates the activation of NLR pyrin domain containing 3 (NLRP3) inflammasome to drive hepatocyte pyroptosis. We showed that 5 mg/kg/day PFOS exposure may exacerbated liver inflammation and steatosis in high-fat diet (HFD)-fed mice with concurrently elevated expression of NLRP3 and caspase-1. PFOS exposure resulted in viability impairment and LDH release in BRL-3A rat liver cells. 25–100 μM concentrations of PFOS exposure activated the NLRP3 inflammasome, leading to consequent GSDMD cleavage, IL-1β release and the initiation of pyroptosis in a dose-dependent manner, whereas treatment with 10 μM NLRP3 inhibitor MCC950 abrogated this effect. Moreover, pretreatment of 5 mM ROS scavenger N-acetyl-L-cysteine (NAC) ameliorated PFOS-induced NLRP3 inflammasome activation and pyroptosis. Collectively, our data highlight a pivotal role of pyroptotic death in PFOS-mediated liver inflammation and metabolic disorder.     

Mechanistic studies of the toxicity of zinc gluconate in the olfactory neuronal cell line Odora

Zinc is both an essential and potentially toxic metal. It is widely believed that oral zinc supplementation can reduce the effects of the common cold; however, there is strong clinical evidence that intranasal (IN) zinc gluconate (ZG) gel treatment for this purpose causes anosmia, or the loss of the sense of smell, in humans. Using the rat olfactory neuron cell line, Odora, we investigated the molecular mechanism by which zinc exposure exerts its toxic effects on olfactory neurons. Following treatment of Odora cells with 100 and 200 μM ZG for 0–24 h, RNA-seq and in silico analyses revealed up-regulation of pathways associated with zinc metal response, oxidative stress, and ATP production. We observed that Odora cells recovered from zinc-induced oxidative stress, but ATP depletion persisted with longer exposure to ZG. ZG exposure increased levels of NLRP3 and IL-1β protein levels in a time-dependent manner, suggesting that zinc exposure may cause an inflammasome-mediated cell death, pyroptosis, in olfactory neurons.     

Alzheimer’s Disease: New Concepts on the Role of Autoimmunity and NLRP3 Inflammasome in the Pathogenesis of the Disease

Alzheimer’s disease (AD), recognized as the most common neurodegenerative disorder, is clinically characterized by the presence of extracellular beta-amyloid (Aβ) plaques and by intracellular neurofibrillary tau tangles, accompanied by glial activation and neuroinflammation. Increasing evidence suggests that self-misfolded proteins stimulate an immune response mediated by glial cells, inducing the release of inflammatory mediators and the recruitment of peripheral macrophages into the brain, which in turn aggravate AD pathology.The present review aims to update the current knowledge on the role of autoimmunity and neuroinflammation in the pathogenesis of the disease, indicating a new target for therapeutic intervention. We mainly focused on the NLRP3 microglial inflammasome as a critical factor in stimulating innate immune responses, thus sustaining chronic inflammation. Additionally, we discussed the involvement of the NLRP3 inflammasome in the gut-brain axis. Direct targeting of the NLRP3 inflammasome and the associated receptors could be a potential pharmacological strategy since its inhibition would selectively reduce AD neuroinflammation.     

2-Deoxy-d-glucose attenuates sevoflurane-induced neuroinflammation through nuclear factor-kappa B pathway in vitro

ObjectSevoflurane, one of the most commonly used anesthetics in clinic, induced neuroinflammation and caused cognitive impairment. 2-deoxy-d-glucose (2-DG) is a synthetic analogue of glucose and is clinically used in medical imaging safely.MethodsWe examined the effect of 2-DG on sevoflurane-induced neuroinflammation in the mouse primary microglia cells. Mouse microglia cells were treated with 4.1% sevoflurane for 6 h to examine the expression of interleukin (IL)-6 and tumor necrosis factor (TNF-α) and activation of nuclear factor-kappa B (NF-κB). Pyrrolidine dithiocarbamate (PDTC) or 2-DG was used 1 h before sevoflurane treatment.ResultsIn the present study, we found that sevoflurane increased level of IL-6 and TNF-α through activating NF-κB signaling, and that 2-DG reduced sevoflurane-induced increase in IL-6 and TNF-α and nuclear NF-κB in microglia cells.ConclusionOur data suggests that NF-κB signaling pathway could be a target for sevoflurane-induced neuroinflammation and 2-DG might be a potential therapy to prevent or treat sevoflurane-induced neuroinflammation.     

A novel role for the mineralocorticoid receptor in glucocorticoid driven vascular calcification

Vascular calcification, which is common in the elderly and in patients with atherosclerosis, diabetes and chronic renal disease, increases the risk of cardiovascular morbidity and mortality. It is a complex, active and highly regulated cellular process that resembles physiological bone formation. It has previously been established that pharmacological doses of glucocorticoids facilitate arterial calcification. However, the consequences for vascular calcification of endogenous glucocorticoid elevation have yet to be established. Glucocorticoids (cortisol, corticosterone) are released from the adrenal gland, but can also be generated within cells from 11-keto metabolites of glucocorticoids (cortisone, 11-dehydrocorticosterone [11-DHC]) by the enzyme, 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1). In the current study we hypothesized that endogenous glucocorticoids facilitate vascular smooth muscle cell (VSMC) calcification and investigated the receptor-mediated mechanism underpinning this process.In vitro studies revealed increased phosphate-induced calcification in mouse VSMCs following treatment for 7 days with corticosterone (100 nM; 7.98 fold; P < 0.01), 11-DHC (100 nM; 7.14 fold; P < 0.05) and dexamethasone (10 nM; 7.16 fold; P < 0.05), a synthetic glucocorticoid used as a positive control. Inhibition of 11β-HSD isoenzymes by 10 μM carbenoxolone reduced the calcification induced by 11-DHC (0.37 fold compared to treatment with 11-DHC alone; P < 0.05). The glucocorticoid receptor (GR) antagonist mifepristone (10 μM) had no effect on VSMC calcification in response to corticosterone or 11-DHC. In contrast, the mineralocorticoid receptor (MR) antagonist eplerenone (10 μM) significantly decreased corticosterone- (0.81 fold compared to treatment with corticosterone alone; P < 0.01) and 11-DHC-driven (0.64 fold compared to treatment with 11-DHC alone; P < 0.01) VSMC calcification, suggesting this glucocorticoid effect is MR-driven and not GR-driven. Neither corticosterone nor 11-DHC altered the mRNA levels of the osteogenic markers PiT-1, Osx and Bmp2. However, DAPI staining of pyknotic nuclei and flow cytometry analysis of surface Annexin V expression showed that corticosterone induced apoptosis in VSMCs.This study suggests that in mouse VSMCs, corticosterone acts through the MR to induce pro-calcification effects, and identifies 11β-HSD-inhibition as a novel potential treatment for vascular calcification.     

Ethanol and nitric oxide modulate expression of glucocorticoid receptor in the rat adrenal cortex

BackgroundThis study was performed to investigate expression and distribution of glucocorticoid receptor (GR) in the rat adrenal cortex, acute effect of ethanol on its expression and possible role of endogenous nitric oxide (NO) in this phenomenon.MethodsAdult female Wistar rats showing diestrus day 1 were treated with: a) ethanol (2 or 4 g/kg body weight (b.w.), ip), b) N^ω-nitro-L-argmine methyl ester (L-NAME), well-known competitive inhibitor of all isoforms of NO synthase (NOS), (30 mg/kg b.w., sc) followed by ethanol (4 g/kg, ip) 3 h later and c) L-NAME (30 mg/kg b.w., sc) followed by saline (ip) 3 h later. Untreated rats were used as controls. Adrenocortical expression of GR was estimated by immunohistochemistry.ResultsStrong nuclear GR staining was observed throughout the cortex of control rats. Acute ethanol treatment significantly decreased the expression of GR in the zona fasciculata and zona reticularis. Blockade of NO formation had no influence on this effect of ethanol, whereas L-NAME itself induced significant decline in GR immunoreactivity.ConclusionsObtained findings are the first to demonstrate localization and distribution of the GR throughout the rat adrenal cortex and to suggest that ethanol as well as endogenous NO may modulate adrenocortical expression of this steroid receptor.     

The impact of neonatal exposure to 17alpha-ethynylestradiol on the development of kisspeptin neurons in female rats

Neonatal exposure to 17alpha-ethynylestradiol (EE) at relatively low doses leads to delayed effects characterized by the early onset of age-related anovulation. Kisspeptin neurons in the anteroventral periventricular nucleus (AVPV), located at the anterior hypothalamus, are proposed to play key roles in appearance of these delayed effects after maturation. To understand the initial changes, we investigated Kiss1 mRNA expression in the anterior and posterior hypothalamus before weaning in female rats that received neonatal exposure to EE at various doses (0.002–2000 μg/kg). The level of Kiss1 mRNA in the anterior hypothalamus was decreased from 0.002 μg/kg which did not induce delayed effects. In the posterior hypothalamus, Kiss1 mRNA expression did not differ among the groups except 2000 μg/kg group. These results suggest that neonatal exposure to EE affects the development of kisspeptin neurons and kisspeptin neurons in the AVPV are highly susceptible to neonatal EE treatment.