Mineralocorticoid Receptor Signaling Research Articles

Mineralocorticoid receptor overexpression in embryonic stem cell-derived cardiomyocytes increases their beating frequency

Cardiac mineralocorticoid receptor (MR) activation triggers adverse cardiovascular events that could be efficiently prevented by mineralocorticoid antagonists. To gain insights into the pathophysiological role of MR function, we established embryonic stem (ES) cell lines from blastocysts of transgenic mice overexpressing the human MR driven by its proximal P1 or distal P2 promoter and presenting with cardiomyopathy, tachycardia, and arrhythmia. Cardiomyocyte differentiation allowed us to investigate the molecular mechanisms contributing to MR-mediated cardiac dysfunction. During cardiac differentiation, wild-type (WT) and recombinant ES cell cultures and excised beating patches expressed endogenous MR along with cardiac gene markers. The two-fold increase in MR protein detected in P1.hMR and P2.hMR cardiomyocytes led to a parallel increase in the spontaneous beating frequency of hMR-overexpressing cardiomyocytes compared with WT. The MR-mediated chronotropic effect was ligand-independent, could be partially repressed by spironolactone, and was accompanied by a significant two- to four-fold increase in mRNA and protein levels of the pacemaker channel HCN1, generating depolarizing If currents, thus revealing a potential new MR target. This was associated with modification in the expression of HCN4, the inward-rectifier potassium channel Kir2.1, and the L-type voltage-dependent calcium channel Cav1.2. We demonstrate that the amplification of MR signalling in ES-derived cardiomyocytes has a major impact on cardiomyocyte contractile properties through an important remodelling of ion channel expression, contributing to arrhythmias. Our results highlight the prominent role of MR function in cardiac physiology and support the benefit of MR antagonists in the management of cardiac dysfunctions.

Aldosterone: Role in the Cardiometabolic Syndrome and Resistant Hypertension

The prevalence of diabetes, hypertension, and cardiovascular disease (CVD) and chronic kidney disease (CKD) is increasing in concert with obesity. Insulin resistance, metabolic dyslipidemia, central obesity, albuminuria. and hypertension commonly cluster to comprise the cardiometabolic syndrome (CMS). Emerging evidence supports a shift in our understanding of the crucial role of elevated serum aldosterone in promoting insulin resistance and resistant hypertension. Aldosterone enhances tissue generation of oxygen free radicals and systemic inflammation. This increase in oxidative stress and inflammation, in turn, contributes to impaired insulin metabolic signaling, reduced endothelial-mediated vasorelaxation, and associated cardiovascular and renal structural and functional abnormalities. In this context, recent investigation indicates that hyperaldosteronism, which is often associated with obesity, contributes to impaired pancreatic β-cell function as well as diminished skeletal muscle insulin metabolic signaling. Accumulating evidence indicates that the cardiovascular and renal abnormalities associated with insulin resistance are mediated, in part, by aldosterone's nongenomic as well as genomic signaling through the mineralocorticoid receptor (MR). In the CMS, there are increased circulating levels of glucocorticoids, which can also activate MR signaling in cardiovascular, adipose, skeletal muscle, neuronal, and liver tissue. Furthermore, there is increasing evidence that fat tissue produces a lipid soluble factor that stimulates aldosterone production from the adrenal zona glomerulosa. Recently, we have learned that MR blockade improves pancreatic insulin release, insulin-mediated glucose utilization, and endothelium-dependent vasorelaxation as well as reduces the progression of CVD and CKD. In summary, aldosterone excess exerts detrimental metabolic effects that contribute to the development of the CMS and resistant hypertension as well as CVD and CKD.

Mineralocorticoid Receptors, Salt-Sensitive Hypertension, and Metabolic Syndrome

Obese persons with metabolic syndrome often have associated with salt-sensitive hypertension, microalbuminuria, and cardiac dysfunction, and the plasma aldosterone level in one-third of metabolic syndrome patients is clearly elevated. Hyperaldosteronism, which may be caused at least partially by certain adipocyte-derived factors, contributes to the development of proteinuria in obese hypertensive rats, and salt loading aggravates the proteinuria and induces cardiac diastolic dysfunction because of inadequate suppression of plasma aldosterone level. However, mineralocorticoid receptor (MR) antagonists prevent salt-induced renal and cardiac damage, suggesting that aldosterone excess and a high-salt diet exert an unfavorable synergistic action on the kidney and heart. In Dahl salt-sensitive rats, however, despite appropriate suppression of plasma aldosterone with a high-salt diet, salt loading paradoxically activated renal MR signaling, and the renal injury was markedly prevented by MR antagonists. Accordingly, we discovered an alternative pathway of MR activation in which Rac1, a small GTP-binding protein, activates MRs. Salt loading activates renal Rac1 in Dahl salt-sensitive rats, and Rac1 in turn induces MR activation, which results in renal injury, and the renal injury has been found to be prevented by Rac1 inhibitors. Moreover, several metabolic syndrome-related factors induce Rac1 activation, and one of them, hyperglycemia, activates MRs via Rac1 activation. Consistent with this, Rac1 inhibitors attenuated the proteinuria and renal injury in obese hypertensive animals. Thus, both salt and obesity activate Rac1 and cause MR activation. Abnormal activation of the aldosterone/MR pathway plays a key role in the development of salt-sensitive hypertension and renal injury in metabolic syndrome.

Corticosteroid regulation of Na +,K +-ATPase α1-isoform expression in Atlantic salmon gill during smolt development

The proposed mineralocorticoid-like signalling axis in teleost fish, consisting of the hormone 11-deoxycorticosterone (DOC) and a mineralocorticoid receptor (MR), has recently challenged our conception of cortisol being the only osmoregulatory corticosteroid in teleost fish. This paper aimed at comparing the osmoregulatory role of DOC with that of cortisol during the pre-adaptive development of SW-tolerance, smoltification, in Atlantic salmon. Using an in vitro gill block incubation system, the effect of DOC and cortisol in the gill was investigated from January to September, using Na +,K +-ATPase α-subunit isoforms α-1a and α-1b mRNA levels as targets for regulation by the hormones. Cortisol and DOC both conferred significant up-regulation of α-1a and α-1b mRNA levels at specific time-points during smoltification. However, the effect of cortisol and DOC on α-subunit isoforms varied seasonally between isoforms and hormones. The maximum induction of α-1a was 3- to 4-fold compared to controls whereas a 2-fold induction was observed for α-1b. The pattern and capacity of stimulation of α-1a through smoltification were similar for cortisol and DOC, whereas cortisol had an enhanced capacity to stimulate α-1b as compared to DOC. Even though there was no demonstrable change in cortisol or DOC sensitivity in the gill, the magnitude of the hormonal effects were seasonally dependent. This is the first report of DOC-induced effects on osmoregulatory targets in fish, thus indicating a role for DOC and MR signalling in osmoregulation.

Caveolin-1 Ablation Reduces the Adverse Cardiovascular Effects of N-ω-Nitro-l-Arginine Methyl Ester and Angiotensin II

Caveolae are the major cellular membrane structure through which extracellular mediators transmit information to intracellular signaling pathways. In vascular tissue (but not ventricular myocardium), caveolin-1 (cav-1) is the main component of caveolae; cav-1 modulates enzymes and receptors, such as the endothelial nitric oxide synthase and the angiotensin II (AngII) type 1 receptor. Evidence suggests that AngII and aldosterone (ALDO) are important mediators of ventricular injury. We have described a model of biventricular damage in rodents that relies on treatment with N-omega-nitro-l-arginine methyl ester (L-NAME (nitric oxide synthase inhibitor)) and AngII. This damage initiated at the vascular level and was observed only in the presence of ALDO and an activated mineralocorticoid receptor (MR). We hypothesize that cav-1 modulates the adverse cardiac effects mediated by ALDO in this animal model. To test this hypothesis, we assessed the ventricular damage and measures of inflammation, in wild-type (WT) and cav-1 knockout (KO) mice randomized to either placebo or L-NAME/AngII treatment. Despite displaying cardiac hypertrophy at baseline and higher blood pressure responses to L-NAME/AngII, cav-1 KO mice displayed, as compared with WT, decreased treatment-induced biventricular damage as well as decreased transcript levels of the proinflammatory marker plasminogen activator inhibitor-1. Additionally, L-NAME/AngII induced an increase in cardiac MR levels in WT but not cav-1-ablated mice. Moreover and despite similar circulating ALDO levels in both genotypes, the myocardial damage (as determined histologically and by plasminogen activator inhibitor-1 mRNA levels) was less sensitive to ALDO levels in cav-1 KO vs. WT mice, consistent with decreased MR signaling in the cav-1 KO. Thus, we conclude that the L-NAME/AngII-induced biventricular damage is mediated by a mechanism partially dependent on cav-1 and signaling via MR/ALDO.

Dietary salt restriction activates mineralocorticoid receptor signaling in volume-overloaded heart failure

Whether a high plasma aldosterone concentration induced by strict salt restriction promotes cardiac remodeling remains controversial. Male Sprague–Dawley rats at 10 weeks of age were given normal salt (NS) (1.5% NaCl) or low salt (LS) (0.05% NaCl) diets. Each animal underwent aortocaval fistula creation for volume-overloaded heart failure or sham surgery. All rats with a fistula received either vehicle or a non-hypotensive dose of spironolactone (200 mg/kg/day) by gavage. Two weeks later, the LS diet significantly increased the plasma aldosterone level in the sham-operated and fistula-created rats (2677 ± 662 pg/ml and 2406 ± 422 pg/ml) compared with that in rats given the NS diet (518 ± 18 pg/ml and 362 ± 45 pg/ml, respectively). In sham-operated rats, the difference in plasma aldosterone level did not affect the extent of myocardial fibrosis (1.8 ± 0.1% with LS diet vs. 1.5 ± 0.3% with NS diet). However, the increase in myocardial fibrosis in fistula-created rats was more prominent with the LS diet than with the NS diet (4.7 ± 0.3% vs. 3.4 ± 0.1%). In addition, the fistula-created rats on the LS diet expressed significantly increased oxidative stress and transforming growth factor-beta compared with those on the NS diets ( P < 0.05). These increases in the fistula-created rats on the LS diet were significantly suppressed by the non-hypotensive dose of spironolactone ( P < 0.05). These results suggest that increased plasma aldosterone level with strict salt restriction activated the mineralocorticoid receptor signaling in volume-overloaded condition, resulting in increased myocardial fibrosis.

Persistent Angiotensin II-independent Activation of Mineralocorticoid Receptor Signaling is Associated With the Cardiac Dysfunction Induced by Long-term Diabetes Mellitus

Persistent Angiotensin II-independent Activation of Mineralocorticoid Receptor Signaling is Associated With the Cardiac Dysfunction Induced by Long-term Diabetes Mellitus

Deletion of Mineralocorticoid Receptors From Macrophages Protects Against Deoxycorticosterone/Salt-Induced Cardiac Fibrosis and Increased Blood Pressure

Increased mineralocorticoid levels plus high salt promote vascular inflammation and cardiac tissue remodeling. Mineralocorticoid receptors are expressed in many cell types of the cardiovascular system, including monocytes/macrophages and other inflammatory cell types. Although mineralocorticoid receptors are expressed in monocytes/macrophages, their role in regulating macrophage function to date has not been investigated. We, thus, used the Cre/LoxP-recombination system to selectively delete mineralocorticoid receptors from monocytes/macrophages with the lysozyme M promoter used to drive Cre expression (MR(flox/flox)/LysM(Cre/-) mice). Male mice from each genotype (MR(flox/flox) or wild-type and MR(flox/flox)/LysM(Cre/-) mice) were uninephrectomized, given 0.9% NaCl solution to drink, and treated for 8 days or 8 weeks with either vehicle (n=10) or deoxycorticosterone (n=10). Equivalent tissue macrophage numbers were seen for deoxycorticosterone treatment of each genotype at 8 days; in contrast, plasminogen activator inhibitor type 1 and NAD(P)H oxidase subunit 2 levels were increased in wild-type but not in MR(flox/flox)/LysM(Cre/-) mice given deoxycorticosterone. Baseline expression of other inflammatory genes was reduced in MR(flox/flox)/LysM(Cre/-) mice compared with wild-type mice. At 8 weeks, deoxycorticosterone-induced macrophage recruitment and connective tissue growth factor and plasminogen activator inhibitor type 1 mRNA levels were similar for each genotype; in contrast, MR(flox/flox)/LysM(Cre/-) mice showed no increase in cardiac fibrosis or blood pressure, as was seen in wild-type mice at 8 weeks. These data demonstrate the following points: (1) mineralocorticoid receptor signaling regulates basal monocyte/macrophage function; (2) macrophage recruitment is not altered by loss of mineralocorticoid receptor signaling in these cells; and (3) a novel and significant role is seen for macrophage signaling in the regulation of cardiac remodeling and systolic blood pressure in the deoxycorticosterone/salt model.

The mineralocorticoid receptor and its coregulators

The mineralocorticoid receptor (MR) is a member of the nuclear receptor superfamily and is essential for controlling sodium transport in epithelial tissues such as the kidney and colon. Moreover, it is also present in other non-epithelial tissues and is capable of activation by both mineralocorticoids and glucocorticoids. A challenge in understanding transcriptional regulation by the MR and other nuclear receptors is to determine how tissue- and ligand-specificity is achieved. Over the past decade, it has become clear that a heterogeneous group of non-receptor proteins termed as coregulators are required to either enhance or repress nuclear receptor-mediated transactivation of target genes. A subset of these coregulators may be expected to confer specificity to MR-mediated responses by virtue of their variable tissue expression and selectivity for different ligands. Specific coregulator-MR interactions may be a suitable target in the rational design of tissue-specific MR modulators as has been described for other steroid receptors. However, the number of coregulators identified to date for the MR is very limited compared with other nuclear receptors. Understanding the full complement of MR coregulators is essential for unraveling the complexity of MR signaling pathways and will facilitate the development of selective MR modulators.

Mineralocorticoid Receptor Overexpression in Basolateral Amygdala Reduces Corticosterone Secretion and Anxiety

The amygdala plays a critical role in the development of anxiety and the regulation of stress hormone secretion. Reciprocally, stress and stress hormones can induce amygdala hypertrophy, a phenomenon related to enhanced anxiety. As such, modulating amygdaloid function can potentially reduce maladaptive features of the stress response. The amygdala contains two kind of receptor for corticosteroids, the adrenal steroid hormone released during stress: glucocorticoid receptors (GRs) and mineralocorticoid receptors (MRs). Although high-affinity MRs are heavily occupied during basal conditions, low-affinity GRs are heavily occupied only by stress levels of glucocorticoids. Prolonged and heavy occupancy of GRs tends to mediate the deleterious effects of glucocorticoids on neurons, whereas MR occupancy tends to mediate beneficial effects. In this report, we overexpress MR in neurons of adult rat basolateral amygdala, with a herpes simplex viral vector coding for two copies of MR. Such overexpression reduced anxiety, as measured on an elevated plus-maze, and reduced the magnitude of glucocorticoid secretion after an acute stressor. Thus, increasing MR signaling in basolateral amygdala could be valuable in management of stress disorders.

S1-1 The Role and Regulation of Mineralocorticoid Receptor Activation in Heart Failure

Mineralocorticoid receptor (MR) activation by either administration of exogenous mineralocorticoids, or by endogenous glucocorticoids, promotes oxidative stress, vascular inflammation and cardiac fibrosis in experimental animals. These studies suggest potential mechanisms for the benefits observed in recent large scale clinical trials investigating the cardioprotective effects of MR antagonists given in conjunction with current best practice. Given that few patients showed elevated plasma aldosterone, novel mechanisms involved in activating the MR in the failing heart are now being investigated. We have investigated the contribution of specific cell types to MR-mediated cardiac pathology using Cre-Lox technology to create tissue specific MR knockouts mice. Our data suggests that MR signalling in macrophages contributes to the regulation of proinflammatory and profibrotic genes associated with the onset of cardiac fibrosis. Importantly, macrophage MR null mice given DOC/salt for 8 weeks show no increase in cardiac fibrosis and systolic blood pressure, in contrast to wild type littermates. Given that macrophage recruitment is equivalent in MR macrophage knockout mice and wild type mice these data suggest that macrophage MR may play a distinct role in tissue remodelling but that MR activation elsewhere in the cardiovascular system is also important. Selective modulators of the MR that are specifically directed towards non-epithelial cardiovascular tissue would increase tissue protection without the side effects associated with renal MR blockade. Moreover, activation of the MR by other ligands would argue for the use of MR antagonists in renal and cardiovascular protection even when plasma aldosterone is normal.

Mineralocorticoid receptor mRNA expression is increased in human hippocampus following brief cerebral ischaemia

We have previously reported that neuronal endangerment in vitro and hypothermic transient global ischaemia in vivo each result in increased mineralocorticoid receptor (MR) expression. In both models MR induction is associated with increased neuronal survival, and blocking MR signalling reduces neuronal survival. Furthermore, transgenic overexpression of human MR promotes neuronal survival both in vitro and in vivo. Here we have assessed whether brief periods of cerebral ischaemia in human subjects, such as occurs in cardiac arrest from which successful resuscitation is achieved, are associated with a sustained increase in hippocampal MR mRNA expression. Human post-mortem brain sections from patients who had died in the weeks following cardiac arrest were analysed for MR mRNA expression by in situ hybridization. Sustained upregulation of MR mRNA expression was observed in the dentate gyrus region of human hippocampus following a brief episode of cerebral ischaemia. This confirms that MR mRNA expression is regulated following neuronal injury in human brain, and suggests that the benefits of increased MR expression seen in animal models of ischaemia may also be observed in humans.

Injury-induced mineralocorticoid receptor expression involves differential promoter usage: A novel role for the rat MRβ variant

Neuronal injury results in increased mineralocorticoid receptor (MR) expression and is associated with increased neuronal survival, suggesting that enhancing MR signalling may have therapeutic implications. MR has a complex gene structure with at least three untranslated exons (α, β, γ) each with unique promoters and a common coding region. We examined whether distinct cellular stressors differentially regulate exon-specific MR transcripts. MRβ transcript was specifically upregulated in rat primary cortical cultures undergoing hypothermic oxygen–glucose deprivation (OGD/H) through activation of its own promoter. This effect was mediated in part by ERK signalling as blockade with PD98059 inhibited OGD/H-induced MRβ promoter activity. A specific increase in MRβ transcript expression was also found in vivo in hypothermic anoxic neonatal rat hippocampus. These results demonstrate a novel key role for the MRβ transcript in response to injury and suggest that some of the known neuroprotective effects of hypothermia may be mediated through increased MR expression.

Corticosteroid receptors, macrophages and cardiovascular disease

The mineralocorticoid receptor (MR) and glucocorticoid receptor are ligand-activated transcription factors that have important physiological and pathophysiological actions in a broad range of cell types including monocytes and macrophages. While the glucocorticoids cortisol and corticosterone have well-described anti-inflammatory actions on both recruited and tissue resident macrophages, a role for the mineralocorticoid aldosterone in these cells is largely undefined. Emerging evidence, however, suggests that MR signalling may promote pro-inflammatory effects. This review will discuss the current understanding of the role of corticosteroid receptors in macrophages and their effect on diseases involving inflammation, with a particular focus on cardiovascular disease.

The Mineralocorticoid Receptor in Endothelial Physiology and Disease: Novel Concepts in the Understanding of Erectile Dysfunction

Aldosterone is a steroid hormone that controls blood pressure by binding to the mineralocorticoid receptor (MR), a ligand-activated transcription factor, and regulating genes that play a role in salt and water homeostasis in the kidney. Dysregulation of the mineralocorticoid system reveals its crucial role in various human diseases including hypertension, atherosclerosis, cardiac failure, mineralocorticoid resistance, and disorders of the nervous system. Recently, experimental animal models of mineralocorticoid/salt-induced hypertension and atherosclerosis have revealed an epithelial, pro-inflammatory role for MR activation. Extensive investigation has begun to elucidate the mechanisms underlying the vascular effects of MR activation which involve its direct role in cardiomyocytes, vascular smooth muscle cells, and endothelial cells. More specifically, in patients with cardiovascular risk factors and disease, including diabetes, hypertension, and/or congestive heart failure, an excess of MR activation has been shown to have a negative impact on endothelial function hence disrupting the physiological balance between vasoconstriction and vasodilation. Such a mechanism may play a role in the pathogenesis of erectile dysfunction (ED), a condition that occurs frequently in patients with increased cardiovascular risk and involves endothelial dysregulation of vascular relaxation. The aim of this review is to summarize the latest concepts in MR signaling, with particular attention to the endothelium, and to discuss the potential benefits of tissue-selective MR blockade in treating subsets of ED patients, such as those with congestive heart failure and hypertension, in which the MR system may be over activated.

Modification of mineralocorticoid receptor function by Rac1 GTPase: implication in proteinuric kidney disease

Blockade of mineralocorticoid receptor has been shown to improve the clinical outcomes of proteinuric kidney diseases. However, little is known about the regulation of mineralocorticoid receptor-dependent transcriptional activity in renal disease. Here we identify a new role for Rac1, a member of the Rho family GTPases, as a potent activator of mineralocorticoid receptor signal transduction both in vitro and in vivo. Transient transfection assays in HEK 293 cells revealed that constitutively active Rac1 (CA-Rac1) enhanced mineralocorticoid receptor-dependent reporter activity, which was accompanied by increased nuclear translocation of mineralocorticoid receptor. CA-Rac1 facilitated mineralocorticoid receptor nuclear accumulation also in podocytes via p21-activated kinase phosphorylation. In mice lacking Rho GDP-dissociation inhibitor-alpha (Arhgdia(-/-) mice), renal abnormalities, including heavy albuminuria and podocyte damage, were associated with increased Rac1 (but not RhoA) and mineralocorticoid receptor signaling in the kidney, without alteration in systemic aldosterone status. Pharmacological intervention with a Rac-specific small-molecule inhibitor diminished mineralocorticoid receptor overactivity and renal damage in this model. Furthermore, albuminuria and histological changes in Arhgdia(-/-) mice were suppressed by mineralocorticoid receptor blockade, confirming the pathological role of Rac1-mineralocorticoid receptor interaction. Our results provide evidence that signaling cross-talk between Rac1 and mineralocorticoid receptor modulates mineralocorticoid receptor activity and identify Rac1 as a therapeutic target for chronic kidney disease.

Glucocorticoid signaling and exercise-induced downregulation of the mineralocorticoid receptor in the induction of adult mouse dentate neurogenesis by treadmill running

Physical exercise is known to promote adult neurogenesis, although the underlying mechanisms remain unclear. Glucocorticoid (corticosterone in rodents) is a factor that is known to affect neurogenesis. As physical exercise modulates corticosterone secretion, we hypothesized that corticosterone signaling is involved in exercise-induced adult neurogenesis. We chose treadmill running (TR) to accurately define the intensity and duration of exercise. Our results showed that 5 weeks of TR increased the doublecortin (DCX)-positive neuronal progenitor cells (NPCs) in adult hippocampus and transiently increased the serum corticosterone level at the end of the TR protocol. This protocol reduced the levels of hippocampal mineralocorticoid receptor (MR); however, glucocorticoid receptor levels were unaltered. We then investigated whether reducing corticosterone levels by bilateral adrenalectomy (ADX) attenuated the TR-enhanced adult neurogenesis. Our results showed that ADX not only blocked the TR-induced downregulation of MR, but also reduced the number of TR-enhanced NPCs. In order to examine the role of MR downregulation in TR-induced adult neurogenesis, animals were treated repeatedly with a selective MR antagonist, spironolactone, for 3 weeks. The results revealed that spironolactone increased the number of spontaneously occurring and TR-induced NPC in the dentate area. Further analysis revealed that spironolactone treatment did not alter precursor cell proliferation, but increased the number of DCX-positive NPCs, suggesting that blockage of MR signaling either facilitates the differentiation of progenitor cells towards neurons and/or enhances the survival of NPCs. Taken together, the data indicated that induction of NPCs in the dentate area of adult hippocampus by TR is partly due to the downregulation of glucocorticoid/MR signaling, which subsequently enhances differentiation along a neuronal lineage and/or NPC survival.

Overexpression of mineralocorticoid and transdominant glucocorticoid receptor blocks the impairing effects of glucocorticoids on memory

It is well established that mineralocorticoid receptors (MRs) and glucocorticoid receptors (GRs) influence hippocampal-dependent spatial memory. MRs are saturated in the presence of low corticosterone (CORT) levels; consequently receptor protein levels play a rate-limiting role in regulating the positive effects of MR-mediated gene transcription. In this study, viral vector-mediated transgene expression was used to simultaneously manipulate both MR and GR signaling. This approach allowed us to investigate the effects of spatially restricted overexpression of MR and a negative transdominant GR (TD) in the dentate gyrus (DG) subfield of the hippocampus, on short term and long term spatial memory in animals overexpressing one copy of MR or TD, two copies of MR ("MR/MR"), or one copy of each ("MR/TD"). Expression of transgenes did not influence the acquisition (learning) phase of the Morris water maze task. However, we found an overall enhancing effect of MR/MR expression on short term memory performance. In addition, rats expressing TD and MR/TD blocked the high CORT-induced impairments on long term spatial memory retrieval. These findings illustrate the potential beneficial effects of increasing MR signaling or decreasing GR signaling to enhance specific aspects of cognitive function.

Glucocorticoid-Related Signaling Effects in Vascular Smooth Muscle Cells

Mineralocorticoid receptor blockade protects from angiotensin II-induced target-organ damage. 11beta-Hydroxysteroid dehydrogenase type 2 protects the mineralocorticoid receptor from activation by glucocorticoids; however, high glucocorticoid concentrations and absent 11beta-hydroxysteroid dehydrogenase type 2 in some tissues make glucocorticoids highly relevant mineralocorticoid receptor ligands. We investigated the effects of corticosterone (10(-6) to 10(-12) mol/L) on early vascular mineralocorticoid receptor signaling by Western blotting, confocal microscopy, and myography. Corticosterone initiated extracellular signal-regulated kinase 1/2 phosphorylation in rat vascular smooth muscle cells at > or =10(-11) mol/L doses. Protein synthesis inhibitors had no effect, indicating a nongenomic action. Corticosterone also stimulated c-Jun N-terminal kinase, p38, Src, and Akt phosphorylation at 15 minutes and enhanced angiotensin II-induced signaling at 5 minutes. A specific epidermal growth factor receptor blocker, AG1478, as well as the Src inhibitor PP2, markedly reduced corticosterone-induced extracellular signal-regulated kinase 1/2 phosphorylation, as did preincubation of cells with the mineralocorticoid receptor antagonist spironolactone. Silencing mineralocorticoid receptor with small interfering RNA abolished corticosterone-induced effects. Corticosterone (10(-9) mol/L) enhanced phenylephrine-induced contraction of intact aortic rings. These effects were dependent on the intact endothelium, mineralocorticoid receptor, and mitogen-activated protein kinase kinase 1/extracellular signal-regulated kinase signaling. We conclude that corticosterone induces rapid mineralocorticoid receptor signaling in vascular smooth muscle cells that involves mitogen-activated protein kinase kinase/extracellular signal-regulated kinase-dependent pathways. These new mineralocorticoid receptor-dependent signaling pathways suggest that glucocorticoids may contribute to vascular disease via mineralocorticoid receptor signaling, independent of circulating aldosterone.

EF Domains Are Sufficient for Nongenomic Mineralocorticoid Receptor Actions

The mineralocorticoid receptor (MR) is important for salt homeostasis and reno-cardiovascular pathophysiology. Signaling mechanisms include, besides classical genomic pathways, nongenomic pathways with putative pathophysiological relevance involving the mitogen-activated protein kinases ERK1/2. We determined the MR domains required for nongenomic signaling and their potential to elicit pathophysiological effects in cultured cells under defined conditions. The expression of full-length human MR or truncated MR consisting of the domains CDEF (MR CDEF), DEF (MR DEF), or EF (MR EF) renders cells responsive for the MR ligand aldosterone with respect to nongenomic ERK1/2 phosphorylation, whereas only full-length MR and MR CDEF conferred genomic responsiveness. ERK1/2 phosphorylation depends on the EGF receptor and cSRC kinase. MR EF expression is sufficient to evoke the aldosterone-induced increase of collagen III levels, similar to full-length MR expression. Our data suggest that nongenomic MR signaling is mediated by the EF domains and present the first proof of principle showing that nongenomic signaling can be sufficient for some pathophysiological effects. The minimum amino acid motif required for nongenomic MR signaling and its importance in various effects have yet to be determined.