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Glucocorticoids are powerful modulators of brain function. They act via mineralocorticoid and glucocorticoid receptors (MR and GR). These are best understood as transcription factors. Although many glucocorticoid effects depend on the modulation of gene transcription, it is a major challenge to link gene expression to function given the large-scale, apparently pleiotropic genomic responses. The extensive sets of MR and GR target genes are highly specific per cell type, and the brain contains many different (neuronal and non-neuronal) cell types. Next to the set "trait" of cellular context, the "state" of other active signaling pathways will affect MR and GR transcriptional activity. Here, we discuss receptor specificity and contextual factors that determine the transcriptional outcome of MR/GR signaling, experimental possibilities offered by single-cell transcriptomics approaches, and reflect on how to make sense of lists of target genes in relation to understanding the functional effects of steroid receptor activation., (© 2022 The Authors. Journal of Neuroendocrinology published by John Wiley & Sons Ltd on behalf of British Society for Neuroendocrinology.)

The glucocorticoid stress hormones affect brain function via high-affinity mineralocorticoid receptors (MRs) and lower-affinity glucocorticoid receptors (GRs). MR and GR not only differ in affinity for ligands, but also have distinct, sometimes opposite, actions on neuronal excitability and other cellular and higher-order parameters related to cerebral function. GR and MR messenger RNA (mRNA) levels are often used as a proxy for the responsiveness to glucocorticoids, assuming proportionality between mRNA and protein levels. This may be especially relevant for the MR, which because of its high affinity is already largely occupied at low basal (trough) hormone levels. Here we explore how GR and MR mRNA levels are associated with the expression of a shared target gene, glucocorticoid-induced leucine zipper (GILZ, coded by Tsc22d3 ) with basal and elevated levels of corticosterone in male mice, using in situ hybridization. Depending on the hippocampal subfield and the corticosterone levels, mRNA levels of MR rather than GR mostly correlated with GILZ mRNA in the hippocampus and hypothalamus at the bulk tissue level. At the individual cell level, these correlations were much weaker. Using publicly available single-cell RNA sequencing data, we again observed that MR and GR mRNA levels were only weakly correlated with target gene expression in glutamatergic and GABAergic neurons. We conclude that MR mRNA levels can be limiting for receptor action, but many other cell-specific and region-specific factors ultimately determine corticosteroid receptor action. Altogether, our results argue for caution while interpreting the consequences of changed receptor expression for the response to glucocorticoids., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Endocrine Society.)

Synthetic glucocorticoids are clinically used to treat auto-immune and inflammatory disease. Despite the high efficacy, glucocorticoid treatments causes side effects such as obesity and insulin resistance in many patients. Via their pharmacological target, the glucocorticoid receptor (GR), glucocorticoids suppress endogenous glucocorticoid secretion. Endogenous, but not synthetic, glucocorticoids activate the mineralocorticoid receptor (MR) and side effects of synthetic glucocorticoids may thus not only result from GR hyperactivation but also from MR hypoactivation. Here, we tested the hypothesis that reactivation of MR with corticosterone add-on treatment can attenuate the metabolic effects of the synthetic glucocorticoid dexamethasone. Male 8-week-old C57Bl/6J mice received a high-fat diet supplemented with dexamethasone or vehicle, and were subcutaneously implanted with low-dose corticosterone- or vehicle-containing pellets. Dexamethasone strongly reduced body weight and fat mass gain, while corticosterone add-on partially normalized this. Dexamethasone-induced hyperglycemia and hyperinsulinemia were exacerbated by corticosterone add-on, which was prevented by MR antagonism. In subcutaneous white adipose tissue, corticosterone add-on prevented the dexamethasone-induced expression of intracellular lipolysis genes. In brown adipose tissue, dexamethasone also upregulated gene expression of brown adipose tissue identity markers, lipid transporters and lipolysis enzymes, which was prevented by corticosterone add-on. In conclusion, corticosterone add-on treatment prevents several, while exacerbating other metabolic effects of dexamethasone. While the exact role of MR remains elusive, this study suggests that corticosterone suppression by dexamethasone contributes to its effects in mice., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Koorneef, van der Meulen, Kooijman, Sánchez-López, Scheerstra, Voorhoeve, Ramesh, Rensen, Giera, Kroon and Meijer.)

Glucocorticoids enhance memory consolidation of emotionally arousing events via largely unknown molecular mechanisms. This glucocorticoid effect on the consolidation process also requires central noradrenergic neurotransmission. The intracellular pathways of these two stress mediators converge on two transcription factors: the glucocorticoid receptor (GR) and phosphorylated cAMP response element-binding protein (pCREB). We therefore investigated, in male rats, whether glucocorticoid effects on memory are associated with genomic interactions between the GR and pCREB in the hippocampus. In a two-by-two design, object exploration training or no training was combined with post-training administration of a memory-enhancing dose of corticosterone or vehicle. Genomic effects were studied by chromatin immunoprecipitation followed by sequencing (ChIP-seq) of GR and pCREB 45 min after training and transcriptome analysis after 3 hr. Corticosterone administration induced differential GR DNA-binding and regulation of target genes within the hippocampus, largely independent of training. Training alone did not result in long-term memory nor did it affect GR or pCREB DNA-binding and gene expression. No strong evidence was found for an interaction between GR and pCREB. Combination of the GR DNA-binding and transcriptome data identified a set of novel, likely direct, GR target genes that are candidate mediators of corticosterone effects on memory consolidation. Cell-specific expression of the identified target genes using single-cell expression data suggests that the effects of corticosterone reflect in part non-neuronal cells. Together, our data identified new GR targets associated with memory consolidation that reflect effects in both neuronal and non-neuronal cells., (© 2021 The Authors. European Journal of Neuroscience published by Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Glucocorticoids regulate numerous processes in human physiology, but deregulated or excessive glucocorticoid receptor (GR) signaling contributes to the development of various pathologies including metabolic syndrome. For this reason, GR antagonists have considerable therapeutic value. Yet, the only GR antagonist that is clinically approved to date - mifepristone - exhibits cross-reactivity with other nuclear steroid receptors like the progesterone receptor. In this study, we set out to identify novel selective GR antagonists by combining rational chemical design with an unbiased in vitro and in vivo screening approach. Using this pipeline, we were able to identify CORT125329 as the compound with the best overall profile from our octahydro series of novel GR antagonists, and demonstrated that CORT125329 does not exhibit cross-reactivity with the progesterone receptor. Further in vivo testing showed beneficial activities of CORT125329 in models for excessive corticosterone exposure and short- and long-term high-fat diet-induced metabolic complications. Upon CORT125329 treatment, most metabolic parameters that deteriorated upon high-fat diet feeding were similarly improved in male and female mice, confirming activity in both sexes. However, some sexually dimorphic effects were observed including male-specific antagonism of GR activity in brown adipose tissue and female-specific lipid lowering activities after short-term CORT125329 treatment. Remarkably, CORT125329 exhibits beneficial metabolic effects despite its lack of GR antagonism in white adipose tissue. Rather, we propose that CORT125329 treatment restores metabolic activity in brown adipose tissue by stimulating lipolysis, mitochondrial activity and thermogenic capacity. In summary, we have identified CORT125329 as a selective GR antagonist with strong beneficial activities in metabolic disease models, paving the way for further clinical investigation., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Objective: Brown adipose tissue (BAT) displays a strong circadian rhythm in metabolic activity, but it is unclear how this rhythm is regulated. As circulating levels of corticosterone coincide with the rhythm of triglyceride-derived fatty acid (FA) uptake by BAT, we investigated whether corticosterone regulates BAT circadian rhythm., Methods: Corticosterone levels were flattened by implanting mice with subcutaneous corticosterone-releasing pellets, resulting in constant circulating corticosterone levels., Results: Flattened corticosterone rhythm caused a complete loss of circadian rhythm in triglyceride-derived fatty acid uptake by BAT. This effect was independent of glucocorticoid receptor expression in (brown) adipocytes and was not caused by deregulation of clock gene expression or overexposure to glucocorticoids, but rather seemed mediated by reduced sympathetic innervation of BAT. In a mouse model of hyperlipidemia and metabolic syndrome, long-term experimental flattening of corticosterone - and thus rhythm in BAT function - resulted in adiposity., Conclusions: This study highlights that a physiological rhythm in glucocorticoids is an important regulator of BAT function and essential for the maintenance of metabolic health., (Copyright © 2021 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

Glucocorticoids mediate numerous essential processes in the human body via binding to the glucocorticoid receptor (GR). Excessive GR signaling can cause disease, and GR antagonists can be used to treat many symptoms of glucocorticoid-induced pathology. The purpose of this study was to characterize the tissue-specific properties of the selective GR antagonist CORT125281. We evaluated the antagonistic effects of CORT125281 upon acute and subchronic corticosterone exposure in mice. In the acute corticosterone setting, hypothalamus-pituitary-adrenal-axis activity was investigated by measurement of basal- and stress-induced corticosterone levels, adrenocorticotropic hormone levels and pituitary proopiomelanocortin expression. GR signaling was evaluated by RT-PCR analysis of GR-responsive transcripts in liver, muscle, brown adipose tissue (BAT), white adipose tissue (WAT) and hippocampus. Pretreatment with a high dose of CORT125281 antagonized GR activity in a tissue-dependent manner. We observed complete inhibition of GR-induced target gene expression in the liver, partial blockade in muscle and BAT and no antagonism in WAT and hippocampus. Tissue distribution only partially explained the lack of effective antagonism. CORT125281 treatment did not disinhibit the hypothalamus-pituitary-adrenal neuroendocrine axis. In the subchronic corticosterone setting, CORT125281 partially prevented corticosterone-induced hyperinsulinemia, but not hyperlipidemia and immune suppression. In conclusion, CORT125281 antagonizes GR transcriptional activity in a tissue-dependent manner and improves corticosterone-induced hyperinsulinemia. Tailored dosing of CORT125281 may allow tissue-specific inhibition of GR transcriptional activity.

Aims/hypothesis: Brown adipose tissue (BAT) improves energy metabolism by combusting glucose and lipids into heat. Agonism of the glucagon-like peptide-1 receptor (GLP-1R) within the central nervous system activates BAT in mice. Moreover, in patients with type 2 diabetes, GLP-1R agonism lowers body weight and improves glucose and lipid levels, possibly involving BAT activation. Interestingly, people from South Asian descent are prone to develop cardiometabolic disease. We studied the effect of GLP-1R agonism on BAT in humans, specifically in South Asians and Europids without obesity or type 2 diabetes., Methods: Twelve Dutch South Asian and 12 age- and BMI-matched Europid nondiabetic men received 12 weeks extended-release exenatide (Bydureon) in this single-arm prospective study. Before and after treatment, BAT was visualized by a cold-induced [ 18 F]FDG-PET/CT scan and a thermoneutral MRI scan, and resting energy expenditure (REE), substrate oxidation, body composition and fasting plasma glucose and serum lipids were determined. Appetite was rated using a visual analogue scale., Results: Since the effect of exenatide on metabolic parameters did not evidently differ between ethnicities, data of all participants were pooled. Exenatide decreased body weight (-1.5 ± 0.4 kg, p < 0.01), without affecting REE or substrate oxidation, and transiently decreased appetite ratings during the first weeks. Exenatide also lowered triglycerides (-15%, p < 0.05) and total cholesterol (-5%, p < 0.05), and tended to lower glucose levels. Notably, exenatide increased BAT metabolic volume (+28%, p < 0.05) and mean standardized uptake value (+11%, p < 0.05) ([ 18 F]FDG-PET/CT), without affecting supraclavicular adipose tissue fat fraction (MRI)., Conclusions/interpretation: We show for the first time that GLP-1R agonism increases [ 18 F]FDG uptake by BAT in South Asian and Europid men without obesity or type 2 diabetes., Trial Registry: Clinicaltrials.gov NCT03002675., Competing Interests: Declaration of competing interest The statements in this manuscript represent the opinions of the authors and not per se those of AstraZeneca. AstraZeneca reviewed the report prior to publication. The authors declare no conflict of interest., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Glucocorticoid hormones have important effects on brain function in the context of acute and chronic stress. Many of these are mediated by the glucocorticoid receptor (GR). GR has transcriptional activity which is highly context-specific and differs between tissues and even between cell types. The outcome of GR-mediated transcription depends on the interactome of associated coregulators. Selective GR modulators (SGRMs) are a class of GR ligands that can be used to activate only a subset of GR-coregulator interactions, thereby giving the possibility to induce a unique combination of agonistic and antagonistic GR properties. We describe SGRM action in animal models of brain function and pathology, and argue for their utility as molecular filters, to characterize context-specific GR interactome and transcriptional activity that are responsible for particular glucocorticoid-driven effects in cognitive processes such as memory consolidation. The ultimate objective of this approach is to identify molecular processes that are responsible for adaptive and maladaptive effects of glucocorticoids in the brain., (© 2019 The Author(s)Published by S. Karger AG, Basel.)

Medication for nonalcoholic fatty liver disease (NAFLD) is an unmet need. Glucocorticoid (GC) stress hormones drive fat metabolism in the liver, but both full blockade and full stimulation of GC signaling aggravate NAFLD pathology. We investigated the efficacy of selective glucocorticoid receptor (GR) modulator CORT118335, which recapitulates only a subset of GC actions, in reducing liver lipid accumulation in mice. Male C57BL/6J mice received a low-fat diet or high-fat diet mixed with vehicle or CORT118335. Livers were analyzed histologically and for genome-wide mRNA expression. Functionally, hepatic long-chain fatty acid (LCFA) composition was determined by gas chromatography. We determined very-low-density lipoprotein (VLDL) production by treatment with a lipoprotein lipase inhibitor after which blood was collected to isolate radiolabeled VLDL particles and apoB proteins. CORT118335 strongly prevented and reversed hepatic lipid accumulation. Liver transcriptome analysis showed increased expression of GR target genes involved in VLDL production. Accordingly, CORT118335 led to increased lipidation of VLDL particles, mimicking physiological GC action. Independent pathway analysis revealed that CORT118335 lacked induction of GC-responsive genes involved in cholesterol synthesis and LCFA uptake, which was indeed reflected in unaltered hepatic LCFA uptake in vivo. Our data thus reveal that the robust hepatic lipid-lowering effect of CORT118335 is due to a unique combination of GR-dependent stimulation of lipid (VLDL) efflux from the liver, with a lack of stimulation of GR-dependent hepatic fatty acid uptake. Our findings firmly demonstrate the potential use of CORT118335 in the treatment of NAFLD and underscore the potential of selective GR modulation in metabolic disease.

Metabolic status impacts on the emotional brain to induce behavior that maintains energy balance. While hunger suppresses the fear circuitry to promote explorative food-seeking behavior, satiety or obesity may increase fear to prevent unnecessary risk-taking. Here we aimed to unravel which metabolic factors, that transfer information about the acute and the chronic metabolic status, are of primary importance to regulate fear, and to identify their sites of action within fear-related brain regions. We performed a de novo analysis of central and peripheral metabolic factors that can penetrate the blood-brain barrier using genome-wide expression data across the mouse brain from the Allen Brain Atlas (ABA). The central fear circuitry, as defined by subnuclei of the amygdala, the afferent hippocampus, the medial prefrontal cortex and the efferent periaqueductal gray, was enriched with metabolic receptors. Some of their corresponding ligands were known to modulate fear (e.g., estrogen and thyroid hormones) while others had not been associated with fear before (e.g., glucagon, ACTH). Additionally, several of these enriched metabolic receptors were coexpressed with well-described fear-modulating genes ( Crh, Crhr1 , or Crhr2 ). Co-expression analysis of monoamine markers and metabolic receptors suggested that monoaminergic nuclei have differential sensitivity to metabolic alterations. Serotonergic neurons expressed a large number of metabolic receptors (e.g., estrogen receptors, fatty acid receptors), suggesting a wide responsivity to metabolic changes. The noradrenergic system seemed to be specifically sensitive to hypocretin/orexin modulation. Taken together, we identified a number of novel metabolic factors (glucagon, ACTH) that have the potential to modulate the fear response. We additionally propose novel cerebral targets for metabolic factors (e.g., thyroid hormones) that modulate fear, but of which the sites of action are (largely) unknown.

The glucocorticoid hormone cortisol acts throughout the body to support circadian processes and adaptation to stress. The glucocorticoid receptor is the target of cortisol and of synthetic glucocorticoids, which are used widely in the clinic. Both agonism and antagonism of the glucocorticoid receptor may be beneficial in disease, but given the wide expression of the receptor and involvement in various processes, beneficial effects are often accompanied by unwanted side effects. Selective glucocorticoid receptor modulators are ligands that induce a receptor conformation that allows activation of only a subset of downstream signaling pathways. Such molecules thereby combine agonistic and antagonistic properties. Here we discuss the mechanisms underlying selective receptor modulation and their promise in treating diseases in several organ systems where cortisol signaling plays a role., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Glucocorticoids influence a wide range of metabolic processes in the human body, and excessive glucocorticoid exposure is known to contribute to the development of metabolic disease. We evaluated the utility of the novel glucocorticoid receptor (GR) antagonist CORT125281 for its potential to overcome adiposity, glucose intolerance, and dyslipidemia and compared this head-to-head with the classic GR antagonist RU486 (mifepristone). We show that, although RU486 displays cross-reactivity to the progesterone and androgen receptor, CORT125281 selectively inhibits GR transcriptional activity. In a mouse model for diet-induced obesity, rhythmicity of circulating corticosterone levels was disturbed. CORT125281 restored this disturbed rhythmicity, in contrast to RU486, which further inhibited endogenous corticosterone levels and suppressed adrenal weight. Both CORT125281 and RU486 reduced body weight gain and fat mass. In addition, CORT125281, but not RU486, lowered plasma levels of triglycerides, cholesterol, and free fatty acids and strongly stimulated triglyceride-derived fatty acid uptake by brown adipose tissue depots. In combination with reduced lipid content in brown adipocytes, this indicates that CORT125281 enhances metabolic activity of brown adipose tissue depots. CORT125281 was also found to increase liver lipid accumulation. Taken together, CORT125281 displayed a wide range of beneficial metabolic activities that are in part distinct from RU486, but clinical utility may be limited due to liver lipid accumulation. This warrants further evaluation of GR antagonists or selective modulators that are not accompanied by liver lipid accumulation while preserving their beneficial metabolic activities., (Copyright © 2018 Endocrine Society.)