β2AR Activation Research Articles

A Thermodynamic Discrimination of Efficacy of GPCR Ligands using Absolute Binding Free Energy Calculations

G-protein-coupled receptors (GPCRs) play fundamental roles in most physiological processes by modulating diverse signaling pathways and thus have become one of the most important drug targets. Based on the fact that a variety of the extracellular signals are mediated in a ligand-specific manner such as inverse agonist, neutral antagonist and agonist, quantitative characterization of the ligand efficacy is essential for rational design of selective modulators for GPCR targets. As experimental methods used for this purpose are time-, cost- and labor-intensive, computational tools, if they were systematically able to predict the efficacy of GPCR ligands, would make a big impact on GPCR drug design. To tackle this issue, in this study we apply free energy perturbation molecular dynamics (FEP/MD) simulations to calculating absolute ligand binding free energy for β2-adrenergic receptor (β2AR)/ligand systems. Based on the characterization of binding free energy change with respect to simulation time and free energy decomposition, we present that computationally measured thermodynamic properties can be used as promising descriptors for discriminating the efficacy of GPCR ligands. In addition, the simulation results also provide further insights into β2AR activation dynamics and ligand binding affinity.

Dose-Effects of Aorta-Infused Clenbuterol on Spinal Cord Ischemia-Reperfusion Injury in Rabbits

BackgroundThe β2 adrenergic receptor (β2AR) plays an important role in ischemia-reperfusion (I/R) injury in various organs. Recently, a selective β2AR agonist clenbuterol was suggested to protect against cerebral I/R injury. This study was designed to investigate changes of β2ARs after spinal cord I/R injury and dose-effects of aorta-infused clenbuterol on spinal cord I/R injury in rabbits.MethodsSpinal cord ischemia was induced in New Zealand white rabbits by infrarenal abdominal aortic occlusion with a balloon catheter for 30 minutes except the sham group. During occlusion, nothing (I/R group), normal saline (NS group) or clenbuterol at different doses of 0.005, 0.01, 0.05, 0.1, 0.5, or 1 mg/kg (C0.005, C0.01, C0.05, C0.1, C0.5, and C1 groups) was infused into the occluded aortic segments. The hemodynamic data, blood glucose and serum electrolytes were measured during experimental period. Neurological function was assessed according to the modified Tarlov scales until 48 hours after reperfusion. After that, the lumbar spinal cord was harvested for β2AR immunohistochemistry and histopathologic evaluation in the anterior horns.ResultsThe β2AR expression in the anterior horns of the spinal cord was significantly higher in the I/R group than in the sham group. Tarlov scores and the number of viable α-motor neurons were higher in C0.01-C0.5 groups than in the NS group, C0.005 and C1 groups and were highest in the C0.1 group. Hypotension and hyperglycemia were found in the C1 group.Conclusionβ2ARs in the anterior horn were upregulated after spinal cord I/R injury. Aortic-infused clenbuterol (0.01–0.5 mg/kg) can attenuate spinal cord I/R injury dose-dependently during the ischemic period. The Optimal dosage was 0.1 mg/kg. Activation of β2AR could be a new therapeutic strategy for the treatment of spinal cord I/R injury.

Inducible cardiac arrhythmias caused by enhanced β1-adrenergic autoantibody expression in the rabbit

Previous studies demonstrated burst pacing and intravenous infusion of ACh induced sustained atrial tachycardia when rabbits were immunized to produce β2-adrenergic receptor (β2AR)-activating autoantibodies. The objective of this study was to examine the arrhythmogenic effect of β1-adrenergic receptor (β1AR)-activating autoantibodies in the rabbit. Eight New Zealand white rabbits were immunized with a β1AR second extracellular loop peptide to raise β1AR antibody titers. A catheter-based electrophysiological study was performed on anesthetized rabbits before and after immunization. Arrhythmia occurrence was determined in response to burst pacing before and after ACh infusion in incremental concentrations of 10 μM, 100 μM, and 1 mM. The baseline sinus heart rate before and after immunization averaged 149 ± 17 per min and 169 ± 16 per min, respectively (P < 0.05). In the preimmune studies, there were five sustained (≥10 s) arrhythmias in 32 induction attempts, which occurred in only four of eight rabbits. In the postimmune studies, there were 22 sustained arrhythmias in 32 induction attempts, which occurred in all eight rabbits (P < 0.0001 for the independent effect of immunization). Of the 22 sustained arrhythmias postimmunization, 15 were sinus tachycardia compared with only two before immunization (P < 0.01 for the independent effect of immunization). Postimmune (but not preimmune) rabbit sera demonstrated specific binding to β1AR and induced significant β1AR activation in transfected cells in vitro. No cross-reactivity with β2AR was observed. In conclusion, in contrast with rabbits with β2AR-activating autoantibodies that demonstrate predominantly atrial tachycardias, enhanced autoantibody activation of β1AR in the rabbit leads to tachyarrhythmias mainly in the form of sustained sinus tachycardia.

Developmental α2-adrenergic regulation of noradrenergic synaptic facilitation at cerebellar GABAergic synapses

In the central nervous system, the normal development of neuronal circuits requires adequate temporal activation of receptors for individual neurotransmitters. Previous studies have demonstrated that α2-adrenoceptor (α2-AR) activation eliminates spontaneous action potentials of interneurons in the cerebellar molecular layer (MLIs) and subsequently reduces the frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) in Purkinje cells (PCs) after the second postnatal week. The magnitude of the α2-adrenergic reduction in sIPSC frequency is enhanced during the third postnatal week because of an increase in firing-derived sIPSCs. However, little is known about the effects of α2-AR activation by noradrenaline (NA) on cerebellar GABAergic synaptic transmission that is accompanied by the activation of other AR subtypes, α1- and β-ARs. Here, we developmentally examined the roles of α2-AR activation in the noradrenergic facilitation of sIPSCs in cerebellar PCs. Until the second postnatal week, when substantial inhibitory effects of α2-ARs are absent, NA potentiated sIPSCs and maintained the increased sIPSC frequency, suggesting that NA causes long-lasting facilitation of GABAergic synaptic transmission through α1- and β-AR activation. After the second postnatal week, NA transiently increased the sIPSC frequency, whereas blocking α2-ARs sustained the noradrenergic sIPSC facilitation and increase in the firing rate of MLIs, suggesting that α2-AR activation suppresses the noradrenergic facilitation of GABAergic synaptic transmission. The simultaneous activation of α1- and β-ARs by their specific agonists mimicked the persistent facilitation of sIPSC frequency, which required extracellular signal-regulated kinase 1/2 activation. These findings indicate that NA acts as a neurotrophic factor that strengthens GABAergic synaptic transmission in the developing cerebellar cortex and that α2-ARs temporally restrain the noradrenergic facilitation of sIPSCs after GABAergic synaptogenesis.

β 1 -Adrenergic Receptor and Sphingosine-1-Phosphate Receptor 1 (S1PR1) Reciprocal Downregulation Influences Cardiac Hypertrophic Response and Progression to Heart Failure

Background— The sphingosine-1-phosphate receptor 1 (S1PR1) and β 1 -adrenergic receptor (β1AR) are G-protein–coupled receptors expressed in the heart. These 2 receptors have opposing actions on adenylyl cyclase because of differential G-protein coupling. Importantly, both of these receptors can be regulated by the actions of G-protein–coupled receptor kinase-2, which triggers desensitization and downregulation processes. Although classic signaling paradigms suggest that simultaneous activation of β1ARs and S1PR1s in a myocyte would simply result in opposing action on cAMP production, in this report we have uncovered a direct interaction between these 2 receptors, with regulatory involvement of G-protein–coupled receptor kinase-2. Methods and Results— In HEK (human embryonic kidney) 293 cells overexpressing both β1AR and S1PR1, we demonstrated that β1AR downregulation can occur after stimulation with sphingosine-1-phosphate (an S1PR1 agonist), whereas S1PR1 downregulation can be triggered by isoproterenol (a β-adrenergic receptor agonist) treatment. This cross talk between these 2 distinct G-protein–coupled receptors appears to have physiological significance, because they interact and show reciprocal regulation in mouse hearts undergoing chronic β-adrenergic receptor stimulation and in a rat model of postischemic heart failure. Conclusions— We demonstrate that restoration of cardiac plasma membrane levels of S1PR1 produces beneficial effects that counterbalance the deleterious β1AR overstimulation in heart failure.

Abstract A103: The neurotransmitter norepinephrine and β2-AR activation of MB-231 breast cancer cells alters fibrillar collagen structure

Abstract The impact of psychosocial stress on breast cancer pathogenesis has become an area of intense research in the past several years. Sympathetic nervous system (SNS) activation and release of the catecholamine norepinephrine (NE) is a major stress pathway that promotes breast cancer pathogenesis. We have evidence that SNS activation can regulate breast tumor growth and metastasis by altering the tumor extracellular matrix (ECM), specifically fibrillar collagen. Alterations in fibrillar collagen microstructure can be detected using second harmonic generation multiphoton laser scanning microscopy (SHG MPLSM). This non-linear optical technique takes advantage of endogenous light scattering by non-centrosymmetric structures, including collagen. Stress-induced changes to the tumor collagen microstructure can be quantified by measuring the ratio of forward- to backward-scattered light (F/B ratio), as well as the directional coherence of resident collagen fibers. Here, we demonstrate that NE can directly stimulate breast cancer cells and tumor stromal cells to elicit changes in collagen microstructure. In these experiments, cells were pretreated with NE, washed to remove NE, and placed in 3D-type I collagen matrices. Pretreatment of MDA-MB-231 metastatic breast cancer cells with NE significantly lowered the F/B ratio and decreased fiber directional coherence of type I collagen gels. Treatment of MB-231 cells with the β2-adrenergic receptor (β2-AR) selective agonist, terbutaline, mimicked the effects of NE. Furthermore, NE pretreatment of the β-AR-negative metastatic breast cancer line, 4T1, did not elicit alterations in SHG F/B ratio or fiber coherence in collagen gels, compared to control cells. These results suggest that NE acts through tumor cell β2-AR to alter collagen microstructure. Future experiments will elucidate the molecular mediators of this collagen reorganization and determine the impact of such changes on tumor metastatic capacity. These experiments will provide new insights into mechanisms underlying stress-induced tumor pathogenesis that may reveal new therapeutic avenues for the treatment of breast cancer. Citation Format: Ryan P. Dawes, Kathleen A. Burke, Petr Stastka, Edward B. Brown, Kelley S. Madden. The neurotransmitter norepinephrine and β2-AR activation of MB-231 breast cancer cells alters fibrillar collagen structure. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research: Genetics, Biology, and Clinical Applications; Oct 3-6, 2013; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2013;11(10 Suppl):Abstract nr A103.

Mucosa of human detrusor impairs contraction and β‐adrenoceptor‐mediated relaxation

To elucidate the impact of the mucosa on detrusor muscle function by investigating force of contraction under various stimulatory conditions and during subsequent relaxation using catecholamines. Detrusor tissue was obtained from patients who had undergone cystectomy for bladder cancer and strips of intact or mucosa-denuded muscle were set up for force measurement. Preparations were precontracted with KCl, carbachol or electric-field stimulation (EFS). Precontracted strips were relaxed using increasing concentrations of catecholamines in the absence and presence of the subtype-selective β-adrenoceptor (AR) blockers CGP 20712A (β1-ARs), ICI 118,551 (β2-ARs), and L-748,337 (β3-ARs). Force development in response to KCl (40 mM), carbachol (1 μM) or EFS was larger in the absence of mucosa than in intact muscle strips. The force of contraction of mucosa-denuded strips with detached urothelium incubated in the same chamber was as low as in intact strips. Noradrenaline relaxed precontracted detrusor strips to a significantly larger extent and at lower concentrations in denuded than in intact strips. CGP 20712A did not affect noradrenaline-induced relaxation of denuded and intact strips, and ICI 118,551 did not influence noradrenaline-induced relaxation in denuded strips, but abolished the difference between denuded and intact strips. The antagonism of the relaxant effects of noradrenaline by L-748,337 was slightly smaller in intact than denuded strips. The mucosa of human detrusor strips impairs force development when stimulated with KCl, carbachol or EFS. The mucosa also blunts the relaxing effects of catecholamines. The latter effect does not involve the activation of β1-ARs but only of β2-ARs, whereas β3-ARs mediate the relaxation of human detrusor.

GW24-e1132 β3-Adrenoceptor mediates metabolic remodelling in the rabbit model of tachypacing-induced atrial fibrillation

ObjectivesBeta 3-adrenoceptor (β3-AR) is closely associated with the energy metabolism. This study aimed to explore the role of β3-AR in energy remodelling in rabbit model of pacing-induced atrial fibrillation (AF).MethodsRabbits...

β3-Adrenoceptor activation attenuates atherosclerotic plaque formation in ApoE−/− mice through lowering blood lipids and glucose

To examine the effects of β3-adrenoceptor (β3-AR) activation on atherosclerotic plaque development in ApoE(-/-) mice. Thirty six week-old male ApoE(-/-) mice on a high-fat diet were treated with atorvastatin (10 mg·kg(-1)·d(-1), po), BRL37344 (β3-AR agonist, 1.65 or 3.30 μg/kg, ip, twice a week) or SR52390A (β3-AR antagonist, 50 μg/kg, ip, twice a week) for 12 weeks. Wild-type C57BL/6J mice receiving a normal diet were taken as healthy controls. At the end of the treatments, serum levels of triglycerides (TG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), non-high density lipoprotein cholesterol (nHDL-C), glucose and insulin were measured. The thoracic aortas were dissected out, the area of atherosclerotic plaques and extent of fibrosis in the plaques were examined using HE and Masson's trichome staining, respectively. Compared to wild-type mice, ApoE(-/-) mice fed on a high-fat diet exhibited prominent hyperlipidemia and insulin resistance, associated with large area of atherosclerotic plaques and great extent of fibrosis in aortas. Atorvastatin significantly decreased the serum levels of TC and nHDL-C, and reduced the plaque area and collagen content in aortas. BRL37344 significantly decreased the serum levels of TG, TC, nHDL-C, glucose and insulin, and increased HDL-C and the insulin sensitivity, and dose-dependently reduced the plaque area and collagen content in aortas. SR52390A treatment did not affect any parameters studied. The β3-AR agonist impedes the progression of atherosclerosis in ApoE(-/-) mice, through improvement of the lipid and glucose profiles.

Circulation : Clinical Summaries

<i>Circulation</i> : Clinical Summaries

Catecholamine-Induced β2-Adrenergic Receptor Activation Mediates Desensitization of Gastric Cancer Cells to Trastuzumab by Upregulating MUC4 Expression

Trastuzumab is currently used for patients with Her2(+) advanced gastric cancer. However, the response rate to trastuzumab among the patients is low. The molecular mechanisms underlying trastuzumab resistance in gastric cancer are unknown. Our in vitro data show that activation of β2-adrenergic receptor (β2-AR) triggered by catecholamine caused "targeting failure" of trastuzumab in gastric cancer cells. The antitumor activities of trastuzumab were significantly impeded by chronic catecholamine stimulation in gastric cancer cells and in the mice bearing human gastric cancer xenografts. Mechanistically, catecholamine induced upregulation of the MUC4 expression at both transcription and protein levels via activating STAT3 and ERK. The effects of catecholamine could be effectively blocked by β2-AR antagonist ICI-118,551, indicating that β2-AR-mediated signaling pathway plays a key role in upregulation of MUC4, which was previously demonstrated to interfere with the recognition and physical binding of trastuzumab to Her2 molecules. Moreover, a significant elevation of the MUC4 level was observed in the xenograft tissues in nude mice chronically treated with isoproterenol. Knockdown of MUC4 restored the binding activities of trastuzumab to Her2-overexpressing gastric cancer cells. In addition, coexpression of β2-AR and MUC4 were observed in gastric cancer tissues. Our data indicated a novel trastuzumab resistance mechanism, by which catecholamine-induced β2-AR activation mediates desensitization of gastric cancer cells to trastuzumab through upregulating the MUC4 expression.

Yohimbine Promotes Cardiac NE Release and Prevents LPS-Induced Cardiac Dysfunction via Blockade of Presynaptic α2A-Adrenergic Receptor

Myocardial depression is an important contributor to mortality in sepsis. We have recently demonstrated that α2-adrenoceptor (AR) antagonist, yohimbine (YHB), attenuates lipopolysaccharide (LPS)-induced myocardial depression. However, the mechanisms for this action of YHB are unclear. Here, we demonstrated that YHB decreased nitric oxide (NO) and tumor necrosis factor-alpha (TNF-α) levels in the myocardium and plasma, attenuated cardiac and hepatic dysfunction, but not kidney and lung injuries in endotoxemic mice. Immunohistochemical analysis revealed that cardiac α2A-AR was mostly located in sympathetic nerve presynaptic membrane; YHB decreased cardiac α2A-AR level and promoted cardiac norepinephrine (NE) release in endotoxemic mice. Reserpine that exhausted cardiac NE without markedly decreasing plasma NE level abrogated the inhibitory effects of YHB on cardiac TNF-α and iNOS expression as well as cardiac dysfunction, but not the suppressive effects of YHB on plasma TNF-α and NO elevation in LPS-challenged mice. Furthermore, both reserpine and YHB significantly inhibited LPS-induced myocardial apoptosis. α1-AR, β2-AR, but not β1-AR antagonists reversed the inhibitory effect of YHB on LPS-stimulated myocardial apoptosis. However, β1-AR antagonist attenuated LPS-caused cardiomyocyte apoptosis, partly abolished the protective effect of YHB on the left ventricular ejection fraction in endotoxemic mice. Altogether, these findings indicate that YHB attenuates LPS-induced cardiac dysfunction, at least in part, through blocking presynaptic α2A-AR and thus increasing cardiac NE release. YHB-elevated cardiac NE improves cardiac function via suppressing cardiac iNOS and TNF-α expression, activating β1-AR and inhibiting cardiomyocyte apoptosis through α1- and β2-AR in endotoxemic mice. However, cardiac β1-AR activation promotes LPS-induced cardiomyocyte apoptosis.

Activation of β2-adrenoceptor enhances synaptic potentiation and behavioral memory via cAMP-PKA signaling in the medial prefrontal cortex of rats

The prefrontal cortex (PFC) plays a critical role in cognitive functions, including working memory, attention regulation, behavioral inhibition, as well as memory storage. The functions of PFC are very sensitive to norepinephrine (NE), and even low levels of endogenously released NE exert a dramatic influence on the functioning of the PFC. Activation of β-adrenoceptors (β-ARs) facilitates synaptic potentiation and enhances memory in the hippocampus. However, little is known regarding these processes in the PFC. In the present study, we investigate the role of β2-AR in synaptic plasticity and behavioral memory. Our results show that β2-AR selective agonist clenbuterol facilitates spike-timing-dependent long-term potentiation (tLTP) under the physiological conditions with intact GABAergic inhibition, and such facilitation is prevented by co-application with the cAMP inhibitor Rp-cAMPS. Loading postsynaptic pyramidal cells with Rp-cAMPS, the PKA inhibitor PKI(5-24), or the G protein inhibitor GDP-β-S significantly decreases, but does not eliminate, the effect of clenbuterol. Clenbuterol suppresses the GABAergic transmission, while blocking GABAergic transmission by the GABA(A) receptor blocker partially mimics the effect of clenbuterol. In behavioral tests, a post-training infusion of clenbuterol into mPFC enhances 24-h trace fear memory. In summary, we observed that prefrontal cortical β2-AR activation by clenbuterol facilitates tLTP and enhances trace fear memory. The mechanism underlying tLTP facilitation involves stimulating postsynaptic cAMP-PKA signaling cascades and suppressing GABAergic circuit activities.

Activation of the G(s) heterotrimer monitored in living cells with novel G(s) biosensors

Our understanding of conformational changes associated with receptor activation has been advanced profoundly by the crystal structure of agonist‐bound β2 adrenergic receptor (β2AR) coupled to G(s). However, there is uncertainty regarding the extent to which conformational changes in G protein are conserved in living cells as well as across different receptors.We set out to understand the movement of the G(s) heterotrimer after activation of β2AR in intact cells. Using bioluminescence resonance energy transfer, we made observations on movement within the G protein and between the receptor and the G protein. Using a library of novel G(s) biosensors with either luciferase or GFP inserted at various positions throughout the structure, we studied conformational changes in cells and compared the results to the crystal structures of the inactive and active conformations. We also studied conformational changes in G(s) protein induced by the D1 dopamine receptor and A2A adenosine receptor.Our analysis using these G(s) biosensors suggests that conformational changes within the G(s) heterotrimer are largely conserved among different G(s)‐coupling receptors. Using this set of G(s) biosensors, partiality of agonists can be studied in relation to structural changes and subsequent effector activation. Thus, these biosensors represent a novel pharmacological tool to study structure‐function relationships.

β2 Adrenergic Receptor, Protein Kinase A (PKA) and c-Jun N-terminal Kinase (JNK) Signaling Pathways Mediate Tau Pathology in Alzheimer Disease Models

Alzheimer disease (AD) is characterized by neurodegeneration marked by loss of synapses and spines associated with hyperphosphorylation of tau protein. Accumulating amyloid β peptide (Aβ) in brain is linked to neurofibrillary tangles composed of hyperphosphorylated tau in AD. Here, we identify β2-adrenergic receptor (β2AR) that mediates Aβ-induced tau pathology. In the prefrontal cortex (PFC) of 1-year-old transgenic mice with human familial mutant genes of presenilin 1 and amyloid precursor protein (PS1/APP), the phosphorylation of tau at Ser-214 Ser-262 and Thr-181, and the protein kinases including JNK, GSK3α/β, and Ca(2+)/calmodulin-dependent protein kinase II is increased significantly. Deletion of the β2AR gene in PS1/APP mice greatly decreases the phosphorylation of these proteins. Further analysis reveals that in primary PFC neurons, Aβ signals through a β2AR-PKA-JNK pathway, which is responsible for most of the phosphorylation of tau at Ser-214 and Ser-262 and a significant portion of phosphorylation at Thr-181. Aβ also induces a β2AR-dependent arrestin-ERK1/2 activity that does not participate in phosphorylation of tau. However, inhibition of the activity of MEK, an upstream enzyme of ERK1/2, partially blocks Aβ-induced tau phosphorylation at Thr-181. The density of dendritic spines and synapses is decreased in the deep layer of the PFC of 1-year-old PS1/APP mice, and the mice exhibit impairment of learning and memory in a novel object recognition paradigm. Deletion of the β2AR gene ameliorates pathological effects in these senile PS1/APP mice. The study indicates that β2AR may represent a potential therapeutic target for preventing the development of AD.

Environmental Novelty Activates β2-Adrenergic Signaling to Prevent the Impairment of Hippocampal LTP by Aβ Oligomers

A central question about human brain aging is whether cognitive enrichment slows the development of Alzheimer changes. Here, we show that prolonged exposure to an enriched environment (EE) facilitated signaling in the hippocampus of wild-type mice that promoted long-term potentiation. Akey feature of the EE effect was activation of β2-adrenergic receptors and downstream cAMP/PKA signaling. This EE pathway prevented LTP inhibition by soluble oligomers of amyloid β-protein (Aβ) isolated from AD cortex. Protection by EE occurred in both young and middle-aged wild-type mice. Exposure to novelty afforded greater protection than did aerobic exercise. Mice chronically fed a β-adrenergic agonist without EE were protected from hippocampal impairment by Aβ oligomers. Thus, EE enhances hippocampal synaptic plasticity by activating β-adrenoceptor signaling and mitigating synaptotoxicity of human Aβ oligomers. These mechanistic insights support using prolonged exposure to cognitive novelty and/or oral β-adrenergic agonists to lessen the effects of Aβ accumulation during aging.

α1A‐Adrenergic regulation of inhibition in the olfactory bulb

By regulating inhibition at dendrodendritic synapses between mitral and granule cells (GCs), noradrenergic neurons extending from the brainstem provide an input essential for odour processing in the olfactory bulb (OB). In the accessory OB (AOB), we have recently shown that noradrenaline (NA) increases GABA inhibitory input on to mitral cells (MCs) by exciting GCs. Here, we show that GCs in the main OB (MOB) exhibit a similar response to NA, indicating a common mechanism for noradrenergic regulation of GCMC inhibition throughout the OB. In GCs of the MOB, NA (10 μM) produced a robust excitatory effect that included a slow afterdepolarization that followed a train of action potentials evoked by a current stimulus. The depolarization and slow afterdepolarization in GCs were blocked by the α1A-adrenergic receptor (AR) selective antagonist WB 4101 (30 nm) and mimicked by the α(1A)-AR selective agonist A 61603 (1 μM). In recordings from MCs, A 61603 (30 nm-1 μM) produced a sizeable increase in the frequency of spontaneous and miniature IPSCs, an effect completely abolished by the GABAA receptor antagonist gabazine (5 μM). Likewise, activation of β-ARs increased the frequency of spontaneous IPSCs; however, this effect was smaller and confined to the first postnatal weeks. NA enhanced inhibition in MCs across a broad concentration range (0.1-30 μM) and its effects were completely abolished by a mixture of α1- and β-AR antagonists (1 μM prazosin and 10 μM propranolol). Furthermore, the general α2-AR agonist clonidine (10 μM) failed to affect sIPSC frequency. Thus, the NA-mediated increase in GCMC inhibition in the OB results mostly from activation of the α1A-AR subtype.

In This Issue

HomeCirculation ResearchVol. 112, No. 3In This Issue Free AccessResearch ArticlePDF/EPUBAboutView PDFView EPUBSections ToolsAdd to favoritesDownload citationsTrack citationsPermissions ShareShare onFacebookTwitterLinked InMendeleyReddit Jump toFree AccessResearch ArticlePDF/EPUBIn This Issue Originally published1 Feb 2013https://doi.org/10.1161/RES.0b013e318287730bCirculation Research. 2013;112:407β-Adrenergic Signaling and CPC Function (p 476)Download figureDownload PowerPointβ-Adrenergic stimulation of cardiac progenitor cells is a double-edged sword, say Khan et al.During heart failure, β-adrenergic stimulation of the heart is increased to aid in contraction, providing temporary relief to weakened heart muscles. In the long-term however, this stimulation causes cardiac myocytes to die. To prevent this damage β-adrenergic blockers are often used to treat heart failure patients. When cardiac myocytes die, they are replaced, albeit slowly, by cardiac progenitor cells. However, it is unclear how exactly these progenitors are affected by β-blockers, or indeed, by β-adrenergic stimulation. Now, Khan et al report that β-adrenergic stimulation induces mouse cardiac progenitor cells to proliferate. But, just like adult cardiac myocytes, the progenitor cells underwent apoptosis as soon as they were induced to differentiate. The investigators found that progenitor cells expressed only one type of β-adrenergic receptor, β1-AR, but upon differentiation, they started to express a second, β2-AR—also found in mature cardiac myocytes. And because these 2 receptors convey separate effects, the authors suggest that a bipartite approach, involving β1-AR activation and β2-AR blockade, could be more effective in treating heart failure.Pro- and Anti-Apoptotic Signaling of cAMP (p 498)Download figureDownload PowerPointZhang et al suggest replacing β-blockers with a PKA inhibitor to treat heart failure.Much like Khan and colleagues, Zhang et al were interested in the effects of β-adrenergic stimulation and β-blockers on the heart. They knew that long-term β-adrenergic stimulationduring heart failure causes heart cells to die. They also knew that β-blockers prevent this cell death. However, the molecular details of these processes were unclear. β-Adrenergic–induced cell death is thought to be mediated by the stimulation of protein kinase A (PKA), but the effects of β-adrenergic stimulation could also be PKA-independent. To differentiate between PKA and non-PKA effects, the team created transgenic mice expressing a PKA inhibitor. They found that β-adrenergic stimulation in these mice prevented cardiac myocyte death and that the level of protection offered by PKA inhibition was similar to that provided by β-blockers. There was one important distinction, however. The hearts of the transgenic mice when stimulated also exhibited marked ERK activation, which offered further protection to cardiac myocytes. Following myocardial infarction, these PKA-inhibited mice also showed greater improvement in cardiac function than wild-type mice treated with β-blockers. The authors suggest that for treating heart failure patients, PKA-inhibitors may be more effective than β-blockers.Remodeling of Diabetic Bone Marrow (p 510)Download figureDownload PowerPointVascular problems beget more vascular problems in the bone marrow of patients with diabetes, report Spinetti et al.Cardiovascular complications, such as atherosclerosis and capillary damage are common in diabetes and can have far reaching clinical consequences. In mouse models such capillary damage can affect even the bone marrow. Worse still, in the bone marrow, the failing microvasculature, which feeds the stem cell niche, could reduce the production of progenitors, including pro-angiogenic cells. While it has been shown that circulating progenitor cell levels are suppressed in people with diabetes it is unclear whether, like mice, human bone marrow microvasculature is also affected. Spinetti et al show that bone specimens from diabetes patients do indeed exhibit decreased capillary densities and also contain fewer progenitor cells. The team found that while impaired blood supply did contribute to the reduction in progenitor cell populations, high glucose itself also caused these cells to upregulate a pro-apoptosis pathway. This involved inactivation of microRNA miR-155, and the upregulation of its target - the transcription factor, FOXO3a. Taken together, the data suggest that specific treatments designed to preserve the integrity of capillaries of the bone marrow and the function of progenitor cells could prove important for managing diabetes. Previous Back to top Next FiguresReferencesRelatedDetails February 1, 2013Vol 112, Issue 3 Advertisement Article InformationMetrics © 2013 American Heart Association, Inc.https://doi.org/10.1161/RES.0b013e318287730b Originally publishedFebruary 1, 2013 PDF download Advertisement

β1-Adrenergic receptor blockade extends the life span of Drosophila and long-lived mice

Chronic treatment with β-adrenergic receptor (βAR) agonists increases mortality and morbidity while βAR antagonists (β-blockers) decrease all-cause mortality for those at risk of cardiac disease. Levels of sympathetic nervous system βAR agonists and βAR activity increase with age, and this increase may hasten the development of age-related mortality. Here, we show that β-blockers extend the life span of healthy metazoans. The β-blockers metoprolol and nebivolol, administered in food daily beginning at 12 months of age, significantly increase the mean and median life span of isocalorically fed, male C3B6F1 mice, by 10 and 6.4%, respectively (P < 0.05). Neither drug affected the weight or food intake of the mice, indicating that induced CR is not responsible for these effects, and that energy absorption and utilization are not altered by the drugs. Both β-blockers were investigated to control for their idiosyncratic, off-target effects. Metoprolol and nebivolol extended Drosophila life span, without affecting food intake or locomotion. Thus, βAR antagonists are capable of directly extending the life span of two widely divergent metazoans, suggesting that these effects are phylogenetically highly conserved. Thus, long-term use of β-blockers, which are generally well-tolerated, may enhance the longevity of healthy humans.

Beta3-adrenergic receptors modulate vascular endothelial growth factor release in response to hypoxia through the nitric oxide pathway in mouse retinal explants

Beta-adrenergic receptors (β-ARs) play a role in angiogenic processes that characterize neovascularization-associated retinal diseases, but the role of β3-ARs has not been disclosed yet. We used ex vivo retinal explants to investigate the role of β3-ARs in regulating vascular endothelial growth factor (VEGF) release associated with hypoxia. Whether nitric oxide (NO) mediates β3-AR regulation of VEGF release was also investigated. β3-AR activation was obtained using BRL 37344, whereas SR59230A, L-748,337, or specific siRNAs were used to block β3-ARs. Pharmacological approaches were used to interfere with the NO pathway. Western blot was used to determine β-AR levels. Enzyme-linked immunosorbent assay was used to measure VEGF release. NO production was assessed by a colorimetric assay. We found that hypoxia upregulates β3-ARs. In addition, we observed that β3-AR activation with BRL 37344 increases VEGF release in response to hypoxia. Either β3-AR blocker or β3-AR silencing downregulates drastically hypoxic levels of VEGF. With experiments using NO synthase (NOS) blockade with L-NAME, NOS activation with fluvastatin or NO supplementation with SNAP, we demonstrated that β3-ARs and VEGF are functionally coupled via the NO pathway. In summary, the data presented here support the assumption that β3-ARs are involved in the regulation of angiogenic responses to hypoxia through the NO signalling, a key pathway in hypoxic/ischemic diseases. Although extrapolation of these data to the human situation is difficult, these findings may help to explore the possible role of β3-ARs in vascularization-associated disorders.