Novel role for mineralocorticoid receptors in control of a neuronal phenotype.
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ABSTRACT:
SUBMITTER: McCann KE
PROVIDER: S-EPMC7234915 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Purpose: To compare the binding profiles of mineralocorticoid and glucocorticoid receptors after corticosterone treatment in neuroblastoma cells. Methods: Tagged glucocorticoid receptors (eGFP-rat GR) and mineralocorticoid receptors (mCherry-rat MR) were transiently transfected into Neuro2A mouse neuroblastoma cells and treated with (i) vehicle, (ii) corticosterone 100 nM for 20 mins and (iii) corticosterone 100 nM for 20 mins followed by media repacement twice and 40 mins further incubation in charcoal-stripped serum media. ChIP-nexus samples were prepared in triplicate for deep sequencing using Illumina NextSeq, as well as control samples taken from cell lysates before immunoprecipitation. Sequence data was analyzed and processed for the identification of enriched binding sites, using MACS2, and for the characteristic staggered ChIP-nexus borders, using the MACE pipeline (Wang et al., 2014). Results: Corticosterone-sensitive glucocorticoid receptor (GR) binding sites and mineralocorticoid receptor (MR) binding sites showed considerable overlap, by MACS2 analysis. GR and MR binding sites were highly similar showing recruitment to the same DNA sites, by MACE analysis (Wang et al., 2014). Further analyses showed GR could tether MR to the DNA by GR in a functional manner such that gene expression could be altered by the presence of tethered MR. Conclusions: MR can be recruited to DNA-binding sites by GR, even in the absence of MR DNA-binding. MR can interact in multiple modes at genomic DNA binding sites.
2019-03-15 | GSE115417 | GEO
Project description:Advances in synthetic chemistry, structural biology, molecular modelling and molecular cloning have enabled the systematic functional manipulation of transmembrane proteins. By combining genetically manipulated proteins with light-sensitive ligands, innately 'blind' neurobiological receptors can be converted into photoreceptors, which allows them to be photoregulated with high spatiotemporal precision. Here, we present the optochemical control of neuronal nicotinic acetylcholine receptors (nAChRs) with photoswitchable tethered agonists and antagonists. Using structure-based design, we produced heteromeric ?3?4 and ?4?2 nAChRs that can be activated or inhibited with deep-violet light, but respond normally to acetylcholine in the dark. The generation of these engineered receptors should facilitate investigation of the physiological and pathological functions of neuronal nAChRs and open a general pathway to photosensitizing pentameric ligand-gated ion channels.
| S-EPMC4977190 | biostudies-literature
Project description:The majority of glioblastoma (GBM) patients require the administration of dexamethasone (DEXA) to reduce brain inflammation. DEXA activates the glucocorticoid receptor (GR), which can consequently crosstalk with the mineralocorticoid receptor (MR). However, while GR signaling is well studied in GBM, little is known about the MR in brain tumors. We examined the implication of the MR in GBM considering its interplay with DEXA. Together with gene expression studies in patient cohorts, we used human GBM cell lines and patient-derived glioma stem cells (GSCs) to assess the impact of MR activation and inhibition on cell proliferation, response to radiotherapy, and self-renewal capacity. We show that in glioma patients, MR expression inversely correlates with tumor grade. Furthermore, low MR expression correlates with poorer survival in low grade glioma while in GBM the same applies to classical and mesenchymal subtypes, but not proneural tumors. MR activation by aldosterone suppresses the growth of some GBM cell lines and GSC self-renewal. In GBM cells, the MR antagonist spironolactone (SPI) can promote proliferation, radioprotection and cooperate with DEXA. In summary, we propose that MR signaling is anti-proliferative in GBM cells and blocks the self-renewal of GSCs. Contrary to previous evidence obtained in other cancer types, our results suggest that SPI has no compelling anti-neoplastic potential in GBM.
| S-EPMC8584062 | biostudies-literature
Project description: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 7days with corticosterone (100nM; 7.98 fold; P<0.01), 11-DHC (100nM; 7.14 fold; P<0.05) and dexamethasone (10nM; 7.16 fold; P<0.05), a synthetic glucocorticoid used as a positive control. Inhibition of 11β-HSD isoenzymes by 10μM 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 (10μM) had no effect on VSMC calcification in response to corticosterone or 11-DHC. In contrast, the mineralocorticoid receptor (MR) antagonist eplerenone (10μM) 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.
| S-EPMC5111541 | biostudies-literature
Project description:The mineralocorticoid receptor (MR) is an emerging target in the field of alcohol research. The MR is a steroid receptor in the same family as the glucocorticoid receptor, with which it shares the ligand corticosterone in addition to the MR selective ligand aldosterone. Recent studies have shown correlations between central amygdala (CeA) MR expression and alcohol drinking in rats and macaques, as well as correlations between aldosterone and alcohol craving in individuals with alcohol use disorder (AUD). Additionally, our previous work demonstrated that systemic treatment with the MR antagonist spironolactone reduced alcohol self-administration and response persistence in both male and female rats. This study examined if reductions in self-administration following MR antagonist treatment were related to dysregulation of MR-mediated corticosterone negative feedback. Female rats treated with spironolactone (50 mg/kg; IP) showed increased plasma corticosterone following self-administration, which correlated with reduced alcohol self-administration. Next, local microinjection of the MR-selective antagonist eplerenone was used to identify the brain-regional locus of MR action on alcohol self-administration. Eplerenone infusion produced dose-dependent reductions in alcohol self-administration in the CeA, but had no effect in the dorsal hippocampus. Finally, to assay the functional role of CeA MR expression in alcohol self-administration, CeA MR was knocked down by antisense oligonucleotide (ASO) infusion prior to alcohol self-administration. Rats showed a transient reduction in alcohol self-administration 1 day after ASO infusion. Together these studies demonstrate a functional role of CeA MR in modulating alcohol self-administration and make a case for studying MR antagonists as a novel treatment for AUD.
| S-EPMC7657087 | biostudies-literature
Project description:The type 2 ryanodine receptor/calcium release channel (RyR2), required for excitation-contraction coupling in the heart, is abundant in the brain. Chronic stress induces catecholamine biosynthesis and release, stimulating ?-adrenergic receptors and activating cAMP signaling pathways in neurons. In a murine chronic restraint stress model, neuronal RyR2 were phosphorylated by protein kinase A (PKA), oxidized, and nitrosylated, resulting in depletion of the stabilizing subunit calstabin2 (FKBP12.6) from the channel complex and intracellular calcium leak. Stress-induced cognitive dysfunction, including deficits in learning and memory, and reduced long-term potentiation (LTP) at the hippocampal CA3-CA1 connection were rescued by oral administration of S107, a compound developed in our laboratory that stabilizes RyR2-calstabin2 interaction, or by genetic ablation of the RyR2 PKA phosphorylation site at serine 2808. Thus, neuronal RyR2 remodeling contributes to stress-induced cognitive dysfunction. Leaky RyR2 could be a therapeutic target for treatment of stress-induced cognitive dysfunction.
| S-EPMC3690518 | biostudies-literature
Project description:Chronic aldosterone administration increases glomerular filtration rate, whereas inhibition of mineralocorticoid receptors (MRs) markedly attenuates glomerular hyperfiltration and hypertension associated with primary aldosteronism or obesity. However, the mechanisms by which aldosterone alters glomerular filtration rate regulation are poorly understood. In the present study, we hypothesized that aldosterone suppresses tubuloglomerular feedback (TGF) via activation of macula densa MR. First, we observed the expression of MR in macula densa cells isolated by laser capture microdissection and by immunofluorescence in rat kidneys. Second, to investigate the effects of aldosterone on TGF in vitro, we microdissected the juxtaglomerular apparatus from rabbit kidneys and perfused the afferent arteriole and distal tubule simultaneously. Under control conditions, TGF was 2.8±0.2 ?m. In the presence of aldosterone (10(-8) mol/L), TGF was reduced by 50%. The effect of aldosterone to attenuate TGF was blocked by the MR antagonist eplerenone (10(-5) mol/L). Third, to investigate the effect of aldosterone on TGF in vivo, we performed micropuncture, and TGF was determined by maximal changes in stop-flow pressure P(sf) when tubular perfusion rate was increased from 0 to 40 nL/min. Aldosterone (10(-7) mol/L) decreased ?P(sf) from 10.1±1.4 to 7.7±1.2 mm Hg. In the presence of l-NG-monomethyl arginine citrate (10(-3) mol/L), this effect was blocked. We conclude that MRs are expressed in macula densa cells and can be activated by aldosterone, which increases nitric oxide production in the macula densa and blunts the TGF response.
| S-EPMC3299000 | biostudies-literature
Project description:Mineralocorticoid and glucocorticoid receptors (MRs and GRs) constitute a functionally important dual receptor system detecting and transmitting circulating corticosteroid signals. High expression of MRs and GRs occurs in the same cells in the limbic system, the primary site of glucocorticoid action on cognition, behavior, and mood; however, modes of interaction between the receptors are poorly characterized. We used chromatin immunoprecipitation with nucleotide resolution using exonuclease digestion, unique barcode, and single ligation (ChIP-nexus) for high-resolution genome-wide characterization of MR and GR DNA binding profiles in neuroblastoma cells and demonstrate recruitment to highly similar DNA binding sites. Expressed MR or GR showed differential regulation of endogenous gene targets, including Syt2 and Ddc, whereas coexpression produced augmented transcriptional responses even when MRs were unable to bind DNA (MR-XDBD). ChIP confirmed that MR-XDBD could be tethered to chromatin by GR. Our data demonstrate that MR can interact at individual genomic DNA sites in multiple modes and suggest a role for MR in increasing the transcriptional response to glucocorticoids.
| S-EPMC6462215 | biostudies-literature
Project description:Abnormalities that characterize pulmonary arterial hypertension include impairment in the structure and function of pulmonary vascular endothelial and smooth muscle cells. Aldosterone levels are elevated in human pulmonary arterial hypertension and in experimental pulmonary hypertension, while inhibition of the aldosterone-binding mineralocorticoid receptor attenuates pulmonary hypertension in multiple animal models. We explored the role of mineralocorticoid receptor in endothelial and smooth muscle cells in using cell-specific mineralocorticoid receptor knockout mice exposed to sugen/hypoxia-induced pulmonary hypertension. Treatment with the mineralocorticoid receptor inhibitor spironolactone significantly reduced right ventricular systolic pressure. However, this is not reproduced by selective mineralocorticoid receptor deletion in smooth muscle cells or endothelial cells. Similarly, spironolactone attenuated the increase in right ventricular cardiomyocyte area independent of vascular mineralocorticoid receptor with no effect on right ventricular weight or interstitial fibrosis. Right ventricular perivascular fibrosis was significantly decreased by spironolactone and this was reproduced by specific deletion of mineralocorticoid receptor from endothelial cells. Endothelial cell-mineralocorticoid receptor deletion attenuated the sugen/hypoxia-induced increase in the leukocyte-adhesion molecule, E-selectin, and collagen IIIA1 in the right ventricle. Spironolactone also significantly reduced pulmonary arteriolar muscularization, independent of endothelial cell-mineralocorticoid receptor or smooth muscle cell-mineralocorticoid receptor. Finally, the degree of pulmonary perivascular inflammation was attenuated by mineralocorticoid receptor antagonism and was fully reproduced by smooth muscle cell-specific mineralocorticoid receptor deletion. These studies demonstrate that in the sugen/hypoxia pulmonary hypertension model, systemic-mineralocorticoid receptor blockade significantly attenuates the disease and that mineralocorticoid receptor has cell-specific effects, with endothelial cell-mineralocorticoid receptor contributing to right ventricular perivascular fibrosis and smooth muscle cell-mineralocorticoid receptor participating in pulmonary vascular inflammation. As mineralocorticoid receptor antagonists are being investigated to treat pulmonary arterial hypertension, these findings support novel mechanisms and potential mineralocorticoid receptor targets that mediate therapeutic benefits in patients.
| S-EPMC8216367 | biostudies-literature
Project description:Cannabinoid CB2 receptor (CB2) agonists are potential analgesics void of psychotropic effects. Peripheral immune cells, neurons and glia express CB2; however, the involvement of CB2 from these cells in neuropathic pain remains unresolved. We explored spontaneous neuropathic pain through on-demand self-administration of the selective CB2 agonist JWH133 in wild-type and knockout mice lacking CB2 in neurons, monocytes or constitutively. Operant self-administration reflected drug-taking to alleviate spontaneous pain, nociceptive and affective manifestations. While constitutive deletion of CB2 disrupted JWH133-taking behavior, this behavior was not modified in monocyte-specific CB2 knockouts and was increased in mice defective in neuronal CB2 knockouts suggestive of increased spontaneous pain. Interestingly, CB2-positive lymphocytes infiltrated the injured nerve and possible CB2transfer from immune cells to neurons was found. Lymphocyte CB2depletion also exacerbated JWH133 self-administration and inhibited antinociception. This work identifies a simultaneous activity of neuronal and lymphoid CB2that protects against spontaneous and evoked neuropathic pain.
| S-EPMC7384863 | biostudies-literature