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β-adrenergic receptor (β-AR) stimulation represents a major mechanism of modulating cardiac output. In spite of its fundamental importance, its molecular basis on the level of cell signalling has not been characterised in detail yet. We employed mass spectrometry-based proteome and phosphoproteome analysis using SuperSILAC (spike-in stable isotope labelling by amino acids in cell culture) standardization to generate a comprehensive map of acute phosphoproteome changes in mice upon administration of isoprenaline (ISO), a synthetic β-AR agonist that targets both β1-AR and β2-AR subtypes. Our data describe 8597 quantitated phosphopeptides corresponding to 10,164 known and novel phospho-events from 2975 proteins. In total, 197 of these phospho-events showed significantly altered phosphorylation, indicating an intricate signalling network activated in response to β-AR stimulation. In addition, we unexpectedly detected significant cardiac expression and ISO-induced fragmentation of junctophilin-1, a junctophilin isoform hitherto only thought to be expressed in skeletal muscle. Data are available via ProteomeXchange with identifier PXD025569.

Aims: β-blockers are widely used in therapy for heart failure and hypertension. β-blockers are also known to evoke additional diversified pharmacological and physiological effects in patients. We aim to characterize the underlying molecular signalling and effects on cardiac inotropy induced by β-blockers in animal hearts., Methods and Results: Wild-type mice fed high-fat diet (HFD) were treated with carvedilol, metoprolol, or vehicle and echocardiogram analysis was performed. Heart tissues were used for biochemical and histological analyses. Cardiomyocytes were isolated from normal and HFD mice and rats for analysis of adrenergic signalling, calcium handling, contraction, and western blot. Biosensors were used to measure β-blocker-induced cyclic guanosine monophosphate (cGMP) signal and protein kinase A activity in myocytes. Acute stimulation of myocytes with carvedilol promotes β1 adrenergic receptor (β1AR)- and protein kinase G (PKG)-dependent inotropic cardiac contractility with minimal increases in calcium amplitude. Carvedilol acts as a biased ligand to promote β1AR coupling to a Gi-PI3K-Akt-nitric oxide synthase 3 (NOS3) cascade and induces robust β1AR-cGMP-PKG signal. Deletion of NOS3 selectively blocks carvedilol, but not isoproterenol-induced β1AR-dependent cGMP signal and inotropic contractility. Moreover, therapy with carvedilol restores inotropic contractility and sensitizes cardiac adrenergic reserves in diabetic mice with minimal impact in calcium signal, as well as reduced cell apoptosis and hypertrophy in diabetic hearts., Conclusion: These observations present a novel β1AR-NOS3 signalling pathway to promote cardiac inotropy in the heart, indicating that this signalling paradigm may be targeted in therapy of heart diseases with reduced ejection fraction., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2020. For permissions, please email: journals.permissions@oup.com.)

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Aims: β-adrenergic receptors (βARs) play pivotal roles in regulating cardiac excitation-contraction (E-C) coupling. Global signalling of β1ARs up-regulates both the influx of Ca2+ through sarcolemmal L-type Ca2+ channels (LCCs) and the release of Ca2+ from the sarcoplasmic reticulum (SR) through the ryanodine receptors (RyRs). However, we recently found that β2AR stimulation meditates 'offside compartmentalization', confining β1AR signalling into subsarcolemmal nanodomains without reaching SR proteins. In the present study, we aim to investigate the new question, whether and how compartmentalized β1AR signalling regulates cardiac E-C coupling., Methods and Results: By combining confocal Ca2+ imaging and patch-clamp techniques, we investigated the effects of compartmentalized βAR signalling on E-C coupling at both cellular and molecular levels. We found that simultaneous activation of β2 and β1ARs, in contrast to global signalling of β1ARs, modulated neither the amplitude and spatiotemporal properties of Ca2+ sparks nor the kinetics of the RyR response to LCC Ca2+ sparklets. Nevertheless, by up-regulating LCC current, compartmentalized β1AR signalling synchronized RyR Ca2+ release and increased the functional reserve (stability margin) of E-C coupling. In circumstances of briefer excitation durations or lower RyR responsivity, compartmentalized βAR signalling, by increasing the intensity of Ca2+ triggers, helped stabilize the performance of E-C coupling and enhanced the Ca2+ transient amplitude in failing heart cells., Conclusion: Given that compartmentalized βAR signalling can be induced by stress-associated levels of catecholamines, our results revealed an important, yet unappreciated, heart regulation mechanism that is autoadaptive to varied stress conditions., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2020. For permissions, please email: journals.permissions@oup.com.)

Females have increase in-hospital mortality and poorer outcomes following coronary artery bypass grafting (CABG). Biological differences in the reactivity of the graft conduits to circulating catecholamine may contribute to this sex difference. This study examined sex differences in the vasoconstrictor responses of internal mammary artery (IMA) and saphenous vein (SV) conduits to phenylephrine (PE) and endothelin-1 (ET-1). Functional IMA and SV were obtained from 78 male and 50 female patients undergoing CABG (67.7 ± 11 and 69 ± 10 years, respectively) and subjected to the following experimental conditions. (1) Concentration response curves for PE and ET-1 were generated in an intact IMA and SV and endothelium denuded IMA segments, (2) in the presence of the nitric oxide synthase inhibitor (L-NAME) or the cyclooxygenase inhibitor (indomethacin) in an endothelium-intact IMA and (3) the activity state (abundance and phosphorylation) of the α 1 -adrenergic receptor was investigated using Phos-tag™ western blot analysis. (1) Compared to male, female IMA and SV were hypersensitive to PE but not ET-1 (p < 0.05). The female IMA hypersensitivity response to PE was abolished following endothelial denudation, (2) persisted in the presence of L-NAME but was abolished in the presence of indomethacin and (3) there was no sex differences in the abundance and phosphorylation of the α 1 -adrenergic receptor in IMA. Female IMA and SV graft conduits are hypersensitive to α 1 -adrenergic stimuli. This endothelial cyclooxygenase pathway-mediated hypersensitivity may produce excessive IMA and SV graft constriction in females administered catecholamines and could contribute to their poorer CABG outcomes.

The β 1 -adrenergic regulation of cardiac myocyte contraction plays an important role in regulating heart function. Activation of this system leads to an increased heart rate and stronger myocyte contraction. However, chronic stimulation of the β 1 -adrenergic signaling system can lead to cardiac hypertrophy and heart failure. To understand the mechanisms of action of β 1 -adrenoceptors, a mathematical model of cardiac myocyte contraction that includes the β 1 -adrenergic system was developed and studied. The model was able to simulate major experimental protocols for measurements of steady-state force-calcium relationships, cross-bridge release rate and force development rate, force-velocity relationship, and force redevelopment rate. It also reproduced quite well frequency and isoproterenol dependencies for intracellular Ca 2+ concentration ([Ca 2+ ] i ) transients, total contraction force, and sarcomere shortening. The mathematical model suggested the mechanisms of increased contraction force and myocyte shortening on stimulation of β 1 -adrenergic receptors is due to phosphorylation of troponin I and myosin-binding protein C and increased [Ca 2+ ] i transient resulting from activation of the β 1 -adrenergic signaling system. The model was used to simulate work-loop contractions and estimate the power during the cardiac cycle as well as the effects of 4-aminopyridine and tedisamil on the myocyte contraction. The developed mathematical model can be used further for simulations of contraction of ventricular myocytes from genetically modified mice and myocytes from mice with chronic cardiac diseases. NEW & NOTEWORTHY A new mathematical model of mouse ventricular myocyte contraction that includes the β 1 -adrenergic system was developed. The model simulated major experimental protocols for myocyte contraction and predicted the effects of 4-aminopyridine and tedisamil on the myocyte contraction. The model also allowed for simulations of work-loop contractions and estimation of the power during the cardiac cycle.

Two common genetic polymorphisms in the beta-1 adrenergic receptor (ADRB1 Ser49Gly [rs1801252] and Arg389Gly [rs1801253]) significantly affect receptor function in vitro. The objective of this study was to determine whether ADRB1 Ser49Gly and Arg389Gly are associated with recovery of left ventricular ejection fraction (LVEF) in patients with heart failure. Patients with heart failure and baseline LVEF ≤ 40% were genotyped (n = 98), and retrospective chart review assessed the primary outcome of LVEF recovery to ≥ 40%. Un/adjusted logistic regression models revealed that Ser49Gly, but not Arg389Gly, was significantly associated with LVEF recovery in a dominant genetic model. The adjusted odds ratio for Ser49 was 8.2 (95% CI = 2.1-32.9; p = 0.003), and it was the strongest predictor of LVEF recovery among multiple clinical variables. In conclusion, patients with heart failure and reduced ejection fraction that are homozygous for ADRB1 Ser49 were significantly more likely to experience LVEF recovery than Gly49 carriers.

Repeated cycles of binge-like alcohol consumption and abstinence change the activity of several neurotransmitter systems. Some of these changes are consolidated following prolonged alcohol use and are thought to play an important role in the development of dependence. We have previously shown that systemic administration of the dual beta-adrenergic antagonist and 5-HT 1A/1B partial agonist pindolol selectively reduces long-term but not short-term binge-like consumption of ethanol and alters excitatory postsynaptic currents in basolateral amygdala (BLA) principal neurons. The aim of this study was to investigate the effects of pindolol microinfusions in the BLA on long-term ethanol intake using the drinking-in-the-dark paradigm in mice. We also microinfused RU24969 (5-HT 1A/1B receptor partial agonist) and CGP12177 (β 1/2 adrenergic antagonist) following long-term ethanol intake and determined the densities of 5-HT 1A/1B receptors and β 1/2 adrenergic in the BLA following short-term (4 weeks) and long-term ethanol (12 weeks) consumption. We show that intra-BLA infusion of pindolol (1000 pmol/0.5 μl), RU24969 (0.3 and 3 pmol/0.5 μl) and CGP12177 (500 pmol/0.5 μl) produce robust decreases in long-term ethanol consumption. Additionally, we identified reduced β 1/2 adrenergic receptor expression and no change in 5-HT 1A/1B receptor density in the BLA of long-term ethanol-consuming mice. Collectively, our data highlight the effects of pindolol on voluntary, binge-like ethanol consumption behavior following long-term intake., (© 2018 Society for the Study of Addiction.)

Background: Drugs targeting CB 1 and CB 2 receptors have been suggested to possess therapeutic benefit in cardiovascular disorders associated with elevated sympathetic tone. Limited data suggest cannabinoid ligands interact with postsynaptic β-adrenoceptors. The aim of this study was to examine the effects of CB 1 and CB 2 antagonists, AM251 and AM630, respectively, at functional cardiac β-adrenoceptors., Methods: Experiments were carried out in isolated spontaneously beating right atria and paced left atria where inotropic and chronotropic increases were induced by isoprenaline and selective agonists of β 1 and β 2 -adrenergic receptors., Results: We found four different effects of AM251 and AM630 on the cardiostimulatory action of isoprenaline: (1) both CB receptor antagonists 1 μM enhanced the isoprenaline-induced increase in atrial rate, and AM630 1 μM enhanced the inotropic effect of isoprenaline; (2) AM251 1 μM decreased the efficacy of the inotropic effect of isoprenaline; (3) AM251 0.1 and 3 μM and AM630 3 μM reduced the isoprenaline-induced increases in atrial rate; (4) AM630 0.1 and 3 μM enhanced the inotropic effect of isoprenaline, which was not changed by the same concentrations of AM251., Conclusions: Our results show that the CB 1 and CB 2 receptor antagonists AM251 and AM630 have bidirectional effects on the cardiostimulatory action of isoprenaline, most likely related to an interaction with β 1 -adrenoceptors. Provided that the results translate to human heart, caution should be taken when using CB 1 and CB 2 receptor antagonists, as an enhanced sympathetic tone accompanies many cardiovascular disorders., (Copyright © 2018 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier B.V. All rights reserved.)

On the basis of the evidence that the basolateral amygdala (BLA) modulates hippocampal memory processes via synaptic plasticity, here we report that adrenergic β receptor activation in the BLA rescues amyloid β 1-42 (Aβ 1-42 )-induced attenuation of long-term potentiation (LTP) at perforant pathway-dentate granule cell (DGC) synapses. When 500 μM isoproterenol (2 μl), an adrenergic β receptor agonist, was injected into the BLA 20 min before LTP induction, LTP was enhanced. Isoproterenol-mediated enhancement of LTP was blocked by co-injection with 100 μM ZnAF-2DA, an intracellular Zn 2+ chelator, suggesting that intracellular Zn 2+ is required for the intracellular signaling cascade after adrenergic β receptor activation in the BLA. Aβ 1-42 -induced attenuation of LTP, which was induced by Aβ 1-42 injection into the dentate gyrus 60 min before LTP induction, was rescued by isoproterenol injection into the BLA 20 min before LTP induction, but not by 500 μM phenylephrine (2 μl), an adrenergic α 1 receptor agonist, injection into the BLA, which did not enhance LTP unlike the case of isoproterenol injection. Interestingly, Aβ 1-42 -induced attenuation of LTP was also rescued by 100 μM isoproterenol injection into the BLA 20 min before LTP induction, which did not enhance LTP. The present study demonstrates that adrenergic β receptor activation in the BLA, which is linked with intracellular Zn 2+ signaling, rescues Aβ 1-42 -induced attenuation of dentate gyrus LTP. It is likely that adrenergic β receptor activation in the BLA is a strategy for rescuing Aβ 1-42 -induced cognitive decline that is associated with hippocampal synaptic plasticity., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Alpha1-adrenoceptors induce prostate smooth muscle contraction, and hold a prominent role for pathophysiology and therapy of lower urinary tract symptoms in benign prostatic hyperplasia. G protein-coupled receptors are regulated by posttranslational regulation, including phosphorylation by G protein-coupled receptor kinases 2 and 3 (GRK2/3). Although posttranslational adrenoceptor regulation has been recently suggested to occur in the prostate, this is still marginally understood. With the newly developed CMPD101, a small molecule inhibitor with assumed specificity for GRK2/3 is now available. Here, we studied effects of CMPD101 on smooth muscle contraction of human prostate tissue. Electric field stimulation caused frequency-dependent contractions, which were inhibited concentration-dependently by CMPD101 (5 µM, 50 µM). 50 µM of CMPD101 did not affect myosin light chain (MCL) phosphorylation or Rho kinase activity, and did not alter contractions induced by highmolar KCl. Noradrenaline, the α 1 -adrenoceptor agonist phenylephrine, endothelin-1, and the thromboxane A 2 analogue U46619 induced concentration-dependent contractions, which were inhibited by CMPD101 (50 µM). CMPD101 (50 µM) did not change phosphorylation of β 2 -adrenoceptors or β 2 -adrenergic relaxation of prostate strips. Molecular detection by Western blot and peroxidase staining suggested expression of GRK2 and GRK3 in human prostates. Double labeling in fluorescence staining confirmed that immunoreactivity for GRK2 and GRK3 was located to smooth muscle cells in the prostate stroma. In conclusion, CMPD101 inhibits adrenergic, neurogenic, and non-adrenergic smooth muscle contractions in the human prostate. Underlying mechanisms may be independent from GRK inhibition, and from inhibition of MLC kinase and Rho kinase. This may point to unknown properties of CMPD101., (Copyright © 2018 Elsevier B.V. All rights reserved.)

This study tested the hypothesis that glucagon-like peptide 1 (GLP-1) therapies improve cardiac contractile function at rest and in response to adrenergic stimulation in obese swine after myocardial infarction. Obese Ossabaw swine were subjected to gradually developing regional coronary occlusion using an ameroid occluder placed around the left anterior descending coronary artery. Animals received subcutaneous injections of saline or liraglutide (0.005-0.015 mg/kg/day) for 30 days after ameroid placement. Cardiac performance was assessed at rest and in response to sympathomimetic challenge (dobutamine 0.3-10 μg/kg/min) using a left ventricular pressure/volume catheter. Liraglutide increased diastolic relaxation (dP/dt; Tau 1 / 2 ; Tau 1 / e ) during dobutamine stimulation ( P < 0.01) despite having no influence on the magnitude of myocardial infarction. The slope of the end-systolic pressure volume relationship (i.e., contractility) increased with dobutamine after liraglutide ( P < 0.001) but not saline administration ( P = 0.63). Liraglutide enhanced the slope of the relationship between cardiac power and pressure volume area (i.e., cardiac efficiency) with dobutamine ( P = 0.017). Hearts from animals treated with liraglutide demonstrated decreased β1-adrenoreceptor expression. These data support that GLP-1 agonism augments cardiac efficiency via attenuation of maladaptive sympathetic signaling in the setting of obesity and myocardial infarction., (© 2017 by the American Diabetes Association.)

Aims: Postural tachycardia syndrome (POTS), a common and debilitating cardiovascular disorder, is characterized by an exaggerated heart rate increase during orthostasis and a wide spectrum of adrenergic-related symptoms. To determine the aetiology of POTS, we examined a possible pathophysiological role for autoantibodies against α1-adrenergic (α1AR) and β1/2-adrenergic receptors (β1/2AR)., Methods and Results: Immunoglobulin G (IgG) derived from 17 POTS patients, 7 with recurrent vasovagal syncope (VVS), and 11 normal controls was analysed for its ability to modulate activity and ligand responsiveness of α1AR and β1/2AR in transfected cells and to alter contractility of isolated rat cremaster arterioles in vitro. Immunoglobulin G activation of α1AR and β1/2AR was significantly higher in POTS compared with VVS and controls in cell-based assays. Eight, 11, and 12 of the 17 POTS patients possessed autoantibodies that activated α1AR, β1AR and β2AR, respectively. Pharmacological blockade suppressed IgG-induced activation of α1AR and β1/2AR. Eight of 17 POTS IgG decreased the α1AR responsiveness to phenylephrine and 13 of 17 POTS IgG increased the β1AR responsiveness to isoproterenol irrespective of their ability to directly activate their receptors. Postural tachycardia syndrome IgG contracted rat cremaster arterioles, which was reversed by α1AR blockade. The upright heart rate correlated with IgG-mediated β1AR and α1AR activity but not with β2AR activity., Conclusion: These data confirm a strong relationship between adrenergic autoantibodies and POTS. They support the concept that allosteric-mediated shifts in the α1AR and β1AR responsiveness are important in the pathophysiology of postural tachycardia., (© The Author 2016. Published by Oxford University Press on behalf of the European Society of Cardiology.)

Aims: Cardiac β-adrenergic receptor (βAR) signalling is susceptible to heterologous desensitization by different neurohormonal stimuli in clinical conditions associated with heart failure. We aim to examine the underlying mechanism of cross talk between βARs and a set of G-protein coupled receptors (GPCRs) activated by hormones/agonists., Methods and Results: Rat ventricular cardiomyocytes were used to determine heterologous phosphorylation of βARs under a series of GPCR agonists. Activation of Gs-coupled dopamine receptor, adenosine receptor, relaxin receptor and prostaglandin E2 receptor, and Gq-coupled α1 adrenergic receptor and angiotensin II type 1 receptor promotes phosphorylation of β1AR and β2AR at putative protein kinase A (PKA) phosphorylation sites; but activation of Gi-coupled α2 adrenergic receptor and activation of protease-activated receptor does not. The GPCR agonists that promote β2AR phosphorylation effectively inhibit βAR agonist isoproterenol-induced PKA phosphorylation of phospholamban and contractile function in ventricular cardiomyocytes. Heterologous GPCR stimuli have minimal to small effect on isoproterenol-induced β2AR activation and G-protein coupling for cyclic adenosine monophosphate (cAMP) production. However, these GPCR stimuli significantly promote phosphorylation of phosphodiesterase 4D (PDE4D), and recruit PDE4D to the phosphorylated β2AR in a β-arrestin 2 dependent manner without promoting β2AR endocytosis. The increased binding between β2AR and PDE4D effectively hydrolyzes cAMP signal generated by subsequent stimulation with isoproterenol. Mutation of PKA phosphorylation sites in β2AR, inhibition of PDE4, or genetic ablation of PDE4D or β-arrestin 2 abolishes this heterologous inhibitory effect. Ablation of β-arrestin 2 or PDE4D gene also rescues β-adrenergic stimuli-induced myocyte contractile function., Conclusions: These data reveal essential roles of β-arrestin 2 and PDE4D in a common mechanism for heterologous desensitization of cardiac βARs under hormonal stimulation, which is associated with impaired cardiac function during the development of pathophysiological conditions., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2017. For permissions please email: journals.permissions@oup.com.)

Our previous studies have demonstrated that adrenaline dilates rat retinal arterioles by stimulating propranolol-sensitive β-adrenoceptors and β 3 -adrenoceptors, and selective stimulation of β 2 - or β 3 -adrenoceptors causes retinal vasodilator responses. In the present study, we compared the effects of β 1 - and β 2 -adrenoceptor stimulation on rat retinal arterioles in vivo. Rat ocular fundus images were captured using an original high-resolution digital fundus camera. Diameters of retinal arterioles contained in the images were measured. Systemic blood pressure and heart rate were recorded continuously. Denopamine, a β 1 -adrenoceptor agonist, increased the diameter of retinal arterioles and heart rate, and produced a small but statistically insignificant decrease in mean arterial pressure. CGP20712A, a β 1 -adrenoceptor antagonist, but not ICI118551, a β 2 -adrenoceptor antagonist, significantly prevented denopamine-induced retinal vasodilator and heart rate responses. Salbutamol, a β 2 -adrenoceptor agonist, increased the diameter of retinal arterioles and decreased mean arterial pressure without significantly changing heart rate. The effects of salbutamol were significantly prevented by ICI118551, but not by CGP20712A. These results suggest that stimulation of β 1 - and β 2 -adrenoceptors dilates retinal blood vessels and indicate that all three β-adrenoceptor subtypes (β 1 , β 2 , and β 3 ) may be involved in the retinal vasodilator response to adrenaline in rats.

Background: Class IIa histone deacetylase (HDAC) isoforms such as HDAC5 are critical signal-responsive repressors of maladaptive cardiomyocyte hypertrophy, through nuclear interactions with transcription factors including myocyte enhancer factor-2. β-Adrenoceptor (β-AR) stimulation, a signal of fundamental importance in regulating cardiac function, has been proposed to induce both phosphorylation-independent nuclear export and phosphorylation-dependent nuclear accumulation of cardiomyocyte HDAC5. The relative importance of phosphorylation at Ser259/Ser498 versus Ser279 in HDAC5 regulation is also controversial. We aimed to determine the impact of β-AR stimulation on the phosphorylation, localization, and function of cardiomyocyte HDAC5 and delineate underlying molecular mechanisms., Methods and Results: A novel 3-dimensional confocal microscopy method that objectively quantifies the whole-cell nuclear/cytoplasmic distribution of green fluorescent protein tagged HDAC5 revealed the β-AR agonist isoproterenol to induce β 1 -AR-mediated and protein kinase A-dependent HDAC5 nuclear accumulation in adult rat cardiomyocytes, which was accompanied by dephosphorylation at Ser259/279/498. Mutation of Ser259/Ser498 to Ala promoted HDAC5 nuclear accumulation and myocyte enhancer factor-2 inhibition, whereas Ser279 ablation had no such effect and did not block isoproterenol-induced nuclear accumulation. Inhibition of the Ser/Thr phosphatase PP2A blocked isoproterenol-induced HDAC5 dephosphorylation. Co-immunoprecipitation revealed a specific interaction of HDAC5 with the PP2A targeting subunit B55α, as well as catalytic and scaffolding subunits, which increased >3-fold with isoproterenol. Knockdown of B55α in neonatal cardiomyocytes attenuated isoproterenol-induced HDAC5 dephosphorylation., Conclusions: β-AR stimulation induces HDAC5 nuclear accumulation in cardiomyocytes by a mechanism that is protein kinase A-dependent but requires B55α-PP2A-mediated dephosphorylation of Ser259/Ser498 rather than protein kinase A-mediated phosphorylation of Ser279., (© 2017 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley Blackwell.)

Previous studies suggest beta-adrenergic receptor (β-AR) antagonists (β-blockers) decrease breast cancer progression, tumor metastasis, and patient mortality; however the mechanism for this is unknown. Immunohistochemical analysis of normal and malignant breast tissue revealed overexpression of β1-AR and β3-AR in breast cancer. A retrospective cross-sectional study of 404 breast cancer patients was performed to determine the effect of β-blocker usage on tumor proliferation. Our analysis revealed that non-selective β-blockers, but not selective β-blockers, reduced tumor proliferation by 66% (p < 0.0001) in early stage breast cancer compared to non-users. We tested the efficacy of propranolol on an early stage breast cancer patient, and quantified the tumor proliferative index before and after treatment, revealing a propranolol-mediated 23% reduction (p = 0.02) in Ki67 positive tumor cells over a three-week period. The anti-proliferative effects of β-blockers were measured in a panel of breast cancer lines, demonstrating that mammary epithelial cells were resistant to propranolol, and that most breast cancer cell lines displayed dose dependent viability decreases following treatment. Selective β-blockers alone or in combination were not as effective as propranolol at reducing breast cancer cell proliferation. Molecular analysis revealed that propranolol treatment of the SK-BR-3 breast cancer line, which showed high sensitivity to beta blockade, led to a reduction in Ki67 protein expression, decreased phosphorylation of the mitogenic signaling regulators p44/42 MAPK, p38 MAPK, JNK, and CREB, increased phosphorylation of the cell survival/apoptosis regulators AKT, p53, and GSK3β. In conclusion, use of non-selective β-blockers in patients with early stage breast cancer may lead to decreased tumor proliferation.

Background: Cardiac anaphylaxis is one of the features of anaphylactic hypotension. Patients treated with propranolol, a nonselective β-adrenoceptor (AR) antagonist, develop severe anaphylaxis, but the mechanism remains unknown. Under examination were the effects of β1- and β2-AR antagonist on anaphylaxis-induced coronary vasoconstriction and cardiac dysfunction in isolated blood-perfused rat hearts., Methods: Isolated hearts from ovalbumin-sensitized Wistar rats were subjected to coronary perfusion with blood at a constant pressure and measurements were made of coronary blood flow and left ventricu-lar (LV) pressure. Following pretreatment with selective β2-AR antagonist ICI118,551 or selective β1-AR antagonist atenolol, cardiac anaphylaxis was induced by intracoronary injections of ovalbumin antigen. LV contractility was evaluated by the maximum increasing rate of systolic LV pressure (dP/dtmax)., Results: In response to antigen administrations, ICI118,551 pretreated hearts showed a greater de-crease in coronary blood flow and consequently a greater increase in coronary vascular resistance than the atenolol pretreated hearts. Pretreatment with ICI118,551 caused a greater decrease in dP/dtmax than those with atenolol., Conclusions: Cardiac anaphylaxis-induced contractile dysfunction and coronary spasm are severe in b2-, rather than β1-AR antagonist, pretreated isolated blood-perfused rat hearts.

Handgrip-induced increases in blood flow through the contralateral artery that supplies the cortical representation of the arm have been hypothesized as a consequence of neurovascular coupling and a resultant metabolic attenuation of sympathetic cerebral vasoconstriction. In contrast, sympathetic restraint, in theory, inhibits changes in perfusion of the cerebral ipsilateral blood vessels. To confirm whether sympathetic nerve activity modulates cerebral blood flow distribution during static handgrip (SHG) exercise, beat-to-beat contra- and ipsilateral internal carotid artery blood flow (ICA; Doppler) and mean arterial pressure (MAP; Finometer) were simultaneously assessed in nine healthy men (27 ± 5 yr), both at rest and during a 2-min SHG bout (30% maximal voluntary contraction), under two experimental conditions: 1) control and 2) α1-adrenergic receptor blockade. End-tidal carbon dioxide (rebreathing system) was clamped throughout the study. SHG induced increases in MAP (+31.4 ± 10.7 mmHg, P < 0.05) and contralateral ICA blood flow (+80.9 ± 62.5 ml/min, P < 0.05), while no changes were observed in the ipsilateral vessel (-9.8 ± 39.3 ml/min, P > 0.05). The reduction in ipsilateral ICA vascular conductance (VC) was greater compared with contralateral ICA (contralateral: -0.8 ± 0.8 vs. ipsilateral: -2.6 ± 1.3 ml·min(-1)·mmHg(-1), P < 0.05). Prazosin was effective to induce α1-blockade since phenylephrine-induced increases in MAP were greatly reduced (P < 0.05). Under α1-adrenergic receptor blockade, SHG evoked smaller MAP responses (+19.4 ± 9.2, P < 0.05) but similar increases in ICAs blood flow (contralateral: +58.4 ± 21.5 vs. ipsilateral: +54.3 ± 46.2 ml/min, P > 0.05) and decreases in VC (contralateral: -0.4 ± 0.7 vs. ipsilateral: -0.4 ± 1.0 ml·min(-1)·mmHg(-1), P > 0.05). These findings indicate a role of sympathetic nerve activity in the regulation of cerebral blood flow distribution during SHG., (Copyright © 2016 the American Physiological Society.)

The β1-adrenergic signaling system is one of the most important protein signaling systems in cardiac cells. It regulates cardiac action potential duration, intracellular Ca(2+) concentration ([Ca(2+)]i) transients, and contraction force. In this paper, a comprehensive experimentally based mathematical model of the β1-adrenergic signaling system for mouse ventricular myocytes is explored to simulate the effects of moderate stimulations of β1-adrenergic receptors (β1-ARs) on the action potential, Ca(2+) and Na(+) dynamics, as well as the effects of inhibition of protein kinase A (PKA) and phosphodiesterase of type 4 (PDE4). Simulation results show that the action potential prolongations reach saturating values at relatively small concentrations of isoproterenol (∼0.01 μM), while the [Ca(2+)]i transient amplitude saturates at significantly larger concentrations (∼0.1-1.0 μM). The differences in the response of Ca(2+) and Na(+) fluxes to moderate stimulation of β1-ARs are also observed. Sensitivity analysis of the mathematical model is performed and the model limitations are discussed. The investigated model reproduces most of the experimentally observed effects of moderate stimulation of β1-ARs, PKA, and PDE4 inhibition on the L-type Ca(2+) current, [Ca(2+)]i transients, and the sarcoplasmic reticulum Ca(2+) load and makes testable predictions for the action potential duration and [Ca(2+)]i transients as functions of isoproterenol concentration., (Copyright © 2016 the American Physiological Society.)

Background/aims: We evaluated the association between coding region variants of adrenergic receptor genes and therapeutic effect in patients with congestive heart failure (CHF)., Methods: One hundred patients with stable CHF (left ventricular ejection fraction [LVEF] < 45%) were enrolled. Enrolled patients started 1.25 mg bisoprolol treatment once daily, then up-titrated to the maximally tolerable dose, at which they were treated for 1 year., Results: Genotypic analysis was carried out, but the results were blinded to the investigators throughout the study period. At position 389 of the β-1 adrenergic receptor gene (ADRB1), the observed minor Gly allele frequency (Gly389Arg + Gly389Gly) was 0.21, and no deviation from Hardy-Weinberg equilibrium was observed in the genotypic distribution of Arg389Gly (p = 0.75). Heart rate was reduced from 80.8 ± 14.3 to 70.0 ± 15.0 beats per minute (p < 0.0001). There was no significant difference in final heart rate across genotypes. However, the Arg389Arg genotype group required significantly more bisoprolol compared to the Gly389X (Gly389Arg + Gly389Gly) group (5.26 ± 2.62 mg vs. 3.96 ± 2.05 mg, p = 0.022). There were no significant differences in LVEF changes or remodeling between two groups. Also, changes in exercise capacity and brain natriuretic peptide level were not significant. However, interestingly, there was a two-fold higher rate of readmission (21.2% vs. 10.0%, p = 0.162) and one CHF-related death in the Arg389Arg group., Conclusions: The ADRB1 Gly389X genotype showed greater response to bisoprolol than the Arg389Arg genotype, suggesting the potential of individually tailoring β-blocker therapy according to genotype.

To determine the comprehensive G protein-coupled receptor (GPCR) expression profile in ghrelin-producing cells and to elucidate the role of GPCR-mediated signaling in the regulation of ghrelin secretion, we determined GPCR expression profiles by RNA sequencing in the ghrelin-producing cell line MGN3-1 and analyzed the effects of ligands for highly expressed receptors on intracellular signaling and ghrelin secretion. Expression of selected GPCRs was confirmed in fluorescence-activated cell-sorted fluorescently tagged ghrelin-producing cells from ghrelin-promoter CreERT2/Rosa-CAG-LSL-ZsGreen1 mice. Expression levels of GPCRs previously suggested to regulate ghrelin secretion including adrenergic-β1 receptor, GPR81, oxytocin receptor, GPR120, and somatostatin receptor 2 were high in MGN3-1 cells. Consistent with previous reports, isoproterenol and oxytocin stimulated the Gs and Gq pathways, respectively, whereas lactate, palmitate, and somatostatin stimulated the Gi pathway, confirming the reliability of current assays. Among other highly expressed GPCRs, prostaglandin E receptor 4 agonist prostaglandin E2 significantly stimulated the Gs pathway and ghrelin secretion. Muscarine, the canonical agonist of cholinergic receptor muscarinic 4, stimulated both the Gq and Gi pathways. Although muscarine treatment alone did not affect ghrelin secretion, it did suppress forskolin-induced ghrelin secretion, suggesting that the cholinergic pathway may play a role in counterbalancing the stimulation of ghrelin by Gs (eg, by adrenaline). In addition, GPR142 ligand tryptophan stimulated ghrelin secretion. In conclusion, we determined the comprehensive expression profile of GPCRs in ghrelin-producing cells and identified two novel ghrelin regulators, prostaglandin E2 and tryptophan. These results will lead to a greater understanding of the physiology of ghrelin and facilitate the development of ghrelin-modulating drugs.

Drug seeking is maintained by encounters with drug-associated cues, and disrupting retrieval of these drug-cue associations would reduce the risk of relapse. Retrieval of cocaine-associated memories is dependent on β-adrenergic receptor (β-AR) activation, and blockade of these receptors induces a persistent retrieval deficit. Whether retrieval of cocaine-associated memory is mediated by a specific β-AR subtype, however, remains unclear. Using a cocaine conditioned place preference (CPP) procedure, we examined whether retrieval of a cocaine CPP memory is mediated collectively by β1- and β2-ARs, or by one of these β-AR subtypes alone. We show that co-blockade of β1- and β2-ARs abolished CPP expression on that and subsequent drug-free CPP tests, resulting in a long-lasting retrieval deficit that prevented subsequent cocaine-induced reinstatement. To dissociate the necessity of either β1- or β2-ARs alone, we administered subtype-specific antagonists prior to retrieval. Administration of a β1-AR antagonist before the initial CPP trial dose-dependently reduced expression of a CPP on that and subsequent drug-free trials as compared to vehicle administration. In contrast, administration of a β2-AR antagonist had no effect on initial CPP expression, although the highest dose reduced subsequent CPP expression. Importantly, either β1- or β2-AR blockade prior to an initial retrieval trial prevented subsequent cocaine-induced reinstatement. Our findings indicate that the β1-AR subtype mediates retrieval of a cocaine CPP, and that acutely blocking either β1- or β2-ARs can prevent subsequent cocaine-induced reinstatement. Thus, β-AR antagonists, particularly β1-ARs antagonists, could serve as adjuncts for addiction therapies to prevent retrieval of drug-associated memories and provide protection against relapse., (Copyright © 2015 Elsevier B.V. All rights reserved.)

A high expression β1AR/cell membrane chromatography (β1AR-CMC) and offline UPLC/MS method has been developed for screening active ingredients from Coptis chinensis. In this study, the fractions retained by CMC column were separated and identified by UPLC/MS system. Using metoprolol as a positive control drug, coptisine from C. chinensis was identified as the active component which could inhibit β1AR. Compared with the control group: coptisine could attenuate the infarct size and release malondialdehyde (MDA) while increasing superoxide dismutase (SOD) activity, suggesting a role in reducing myocardial injury. In vitro, coptisine could decrease apoptosis, showing their protective effects upon cardiomyocytes. This β1AR-CMC-offline-UPLC/MS method can be applied for screening active components acting on β1AR from traditional Chinese medicines., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Although increase in heart rate is a crucial determinant for enhancement of cardiac output in the neonate, information on the chronotropic reactivity to catecholamines during postnatal development is scarce. The present study was aimed at investigating the role of β-adrenoceptor subtypes and catecholamine removal mechanisms in the adrenergic chronotropic response during the early post-natal period. Right atria isolated from immature (0-21 day old) and adult (4-6 month old) rats were used for determination of the responsiveness to agonists and quantitation of the transcripts of proteins involved in β-adrenergic signaling. The main results were: (a) the maximum response (Rmax) to norepinephrine increased with age, whereas sensitivity decreased; (b) age-dependent differences in sensitivity to norepinephrine were abolished by inhibition of the neuronal norepinephrine transporter; (c) Rmax to isoproterenol was similar in immature and adult atria, and depressed only in the former by β2-adrenoceptor blockade with ICI118,551; (d) neonatal atria showed greater β2-adrenoceptor mRNA levels, and more prominent positive chronotropic response to the β2- and β3-adrenoceptor agonists zinterol and YM178, respectively (nanomolar range); (e) in atria of immature rats, transcript levels of the extraneuronal monoamine transporter were lower, and its inhibition did not affect sensitivity to isoproterenol; and (f) reactivity to forskolin and 3-isobutyl-1-methylxanthine was not affected by age. The increased β2- and β3-adrenoceptor participation in the adrenergic chronotropic response, in addition to weaker catecholamine removal, may compensate for the immature cardiac innervation and the apparently reduced efficiency of β1-adrenoceptor signaling in the neonate, increasing the responsiveness to endogenous and exogenous β2-adrenoceptor agonists., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Background: Atrial fibrillation (AF) is the most common arrhythmias among old people. It causes serious long-term health problems affecting the quality of life. It has been suggested that the autonomic nervous system is involved in the onset and maintenance of AF in human. However, investigation of its pathogenesis and potential treatment has been hampered by the lack of suitable AF models in experimental animals., Objectives: Our aim was to establish a long-lasting AF model in mice. We also investigated the role of adrenergic receptor (AR) subtypes, which may be involved in the onset and duration of AF., Methods and Results: Trans-esophageal atrial burst pacing in mice could induce AF, as previously shown, but with only a short duration (29.0 ± 8.1 sec). We found that adrenergic activation by intraperitoneal norepinephrine (NE) injection strikingly increased the AF duration. It increased the duration to more than 10 minutes, i.e., by more than 20-fold (656.2 ± 104.8 sec; P<0.001). In this model, a prior injection of a specific β1-AR blocker metoprolol and an α1-AR blocker prazosin both significantly attenuated NE-induced elongation of AF. To further explore the mechanisms underlying these receptors' effects on AF, we assessed the SR Ca(2+) leak, a major trigger of AF, and consequent spontaneous SR Ca(2+) release (SCR) in atrial myocytes. Consistent with the results of our in-vivo experiments, both metoprolol and prazosin significantly inhibited the NE-induced SR Ca(2+) leak and SCR. These findings suggest that both β1-AR and α1-AR may play important roles in the development of AF., Conclusions: We have established a long-lasting AF model in mice induced by adrenergic activation, which will be valuable in future AF study using experimental animals, such as transgenic mice. We also revealed the important role of β1- and α1-AR-mediated signaling in the development of AF through in-vivo and in-vitro experiments.

Ethnopharmacological Relevance: Ge-Gen-Tang (GGT) is a traditional Chinese medicinal formula composed of Puerariae radix (Pueraria lobata Ohwi), Ephedrae Herba (Ephedra sinica Stapf), Cinnamomi Ramulus (Cinnamomum cassia Blume), Paeoniae Radix (Paeonia lactiflora Pallas), Glycyrrhizae Radix preparata (Glycyrrhiza uralensis Fischer), Zingiberis Rhizoma (Zingiber officinale Roscoe), and Zizyphi Fructus (Ziziphus jujuba Mill. var. inermis Rehder) and is widely used to ameoliorate the symptoms of gastrointestinal (GI) disorders related to diarrhea and intestinal mucosal immunity and for anti-cold, antipyretic and analgesic in Eastern Asia., Aim of the Study: Interstitial cells of Cajal (ICCs) are pacemaker cells in the GI tract that generate rhythmic oscillations in membrane potentials known as slow waves. We investigated the effects of GGT on pacemaker potentials in cultured ICCs from the mouse small intestine, and sought to identify the receptors and the action mechanisms involved., Materials and Methods: Enzymatic digestions were used to dissociate ICCs from mouse small intestine tissues. All experiments on ICCs were performed on within 12h after culture. A whole-cell patch-clamp configuration was used to record potentials (current clamp) from cultured ICCs. Intracellular Ca(2+) ([Ca(2+)]i) increase was studied in cultured ICCs using fura-2AM. All of the experiments were performed at 30-32°C., Results: Under the current clamping mode, GGT decreased the amplitude and frequency of pacemaker potentials; however, these effects were blocked by intracellular GDPβS, a G-protein inhibitor, and glibenclamide, a specific ATP-sensitive K(+) channels blocker. Prazosin (α1-adrenoceptor antagonist) and butoxamine (β2-adrenoceptor antagonist) did not block the GGT-induced effects, whereas atenolol (β1-adrenoceptor antagonist) blocked the GGT-induced effects. Also, yohimbine (α2-adrenoceptor antagonist) partially blocked the GGT-induced effects. Pretreatment with SQ-22536, an adenylate cyclase inhibitor, did not block the GGT-induced effects, whereas pretreatment with ODQ, a guanylate cyclase inhibitor, or L-NAME, an inhibitor of nitric oxide (NO) synthase, did. Additionally, [Ca(2+)]i analysis showed that GGT decreased [Ca(2+)]i., Conclusion: These results suggest that GGT inhibits pacemaker potentials in ICCs in a G protein-, cGMP- and NO-dependent manner through stimulation of α2 and β1-adrenoceptors., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Background: Rikkunshito (RKT) is a gastroprotective herbal medicine. In this study, we investigated the role of RKT in the relaxation of the gastric body (fundus and corpus) and antrum., Methods: We used Suncus murinus, a unique small model animal with similar gastrointestinal motility to humans and dogs. RKT was added at 0.1, 1.0, and 5.0 mg/mL to induce relaxation in vitro; the outcome measure was the intensity of relaxation. The number of spontaneous antral contractions in the absence or the presence of RKT was also counted., Key Results: Rikkunshito induced the relaxation of the gastric body and antrum and decreased the number of spontaneous antral contractions in a dose-dependent manner. The responses to RKT (1.0 mg/mL) were not affected by pretreatment with atropine, N-nitro-l-arginine methyl ester, ritanserin, or ondansetron. On the other hand, timolol almost completely reversed the relaxation induced by RKT (1.0 mg/mL) on the gastric body and antrum and the occurrence of the spontaneous antral contractions. Both butoxamine, a β(2) -adrenoreceptor antagonist, and L 748337, a β(3) -adrenoreceptor antagonist, but not CGP 20712, a β(1) -adrenoreceptor antagonist, significantly reversed the RKT-induced (1.0 mg/mL) gastric relaxation., Conclusions & Inferences: These results indicate that RKT stimulates and modulates gastric relaxation through β(2) - and β(3) -adrenergic, but not β(1) -adrenergic, pathways in S. murinus., (© 2015 John Wiley & Sons Ltd.)

New Findings: What is the central question of this study? Can bilateral renal denervation, an effective antihypertensive treatment in clinical and experimental studies, improve cardiac β-adrenoceptor responsiveness in a diabetic model with underlying hypertension? What is the main finding and its importance? Bilateral renal denervation did not affect β-adrenergic responsiveness in the diabetic hypertensive rat heart, but denervation reduced the hypertension-induced concentric hypertrophic remodelling. This suggests that the positive haemodynamic changes induced by renal denervation are most likely to reflect an attenuation of sympathetic effects on the systemic vasculature and/or the renal function rather than direct sympathetic modulation of the heart. Bilateral renal denervation (BRD) has been shown to normalise blood pressure in clinical and experimental studies of hypertension by reducing systemic sympathetic output. This study determined the effect of BRD on cardiac β-adrenoceptor (AR) responsiveness in a diabetic model with underlying hypertension using the transgenic (mRen-2)27 rats. Bilateral renal denervation or sham surgeries were conducted repeatedly at 3, 6 and 9 weeks in Ren-2 rats with or without streptozotocin (STZ)-induced diabetes (4 × n = 7); Sprague-Dawley rats (n = 6) served as control animals. Cardiac function was determined in isolated hearts at 18 weeks of age. Normalised left ventricular developed pressure and relaxation was recorded in response to incremental concentrations of the β-AR agonist isoprenaline (from 10-10 to 10-7 m) or the β3 -AR agonist BRL37344 (from 10(-13) to 10(-6 ) m). Expression levels of β1 -AR were determined by Western blot. Both inotropic and lusitropic β-AR responsiveness was reduced in the hypertensive diabetic hearts, but these responses were unaltered after BRD. Expression levels of β1 -AR were increased after BRD (Sham, 0.85 ± 0.11 versus 1.01 ± 0.05 a.u.; BRD, 1.45 ± 0.11 versus 1.46 ± 0.07 a.u.; Ren-2 versus Ren-2 STZ, P < 0.05 versus Sham). No effect of β3 -AR agonist stimulation with BRL37344 was observed. Interestingly, BRD increased left ventricular diastolic volume in both the Ren-2 and the Ren-2 STZ groups. Bilateral renal denervation did not restore the attenuated cardiac β-AR responsiveness in the diabetic hypertensive rats, but it reduced the extent of hypertension-induced concentric hypertrophic remodelling. Thus, the haemodynamic protection offered by renal denervation appears to reflect an attenuated sympathetic innervation of the systemic vasculature and/or kidney rather than a direct cardiac effect., (© 2015 The Authors. Experimental Physiology © 2015 The Physiological Society.)

Rationale: Cyclic nucleotides are second messengers that regulate cardiomyocyte function through compartmentalized signaling in discrete subcellular microdomains. However, the role of different microdomains and their changes in cardiac disease are not well understood., Objective: To directly visualize alterations in β-adrenergic receptor-associated cAMP and cGMP microdomain signaling in early cardiac disease., Methods and Results: Unexpectedly, measurements of cell shortening revealed augmented β-adrenergic receptor-stimulated cardiomyocyte contractility by atrial natriuretic peptide/cGMP signaling in early cardiac hypertrophy after transverse aortic constriction, which was in sharp contrast to well-documented β-adrenergic and natriuretic peptide signaling desensitization during chronic disease. Real-time cAMP analysis in β1- and β2-adrenergic receptor-associated membrane microdomains using a novel membrane-targeted Förster resonance energy transfer-based biosensor transgenically expressed in mice revealed that this unexpected atrial natriuretic peptide effect is brought about by spatial redistribution of cGMP-sensitive phosphodiesterases 2 and 3 between both receptor compartments. Functionally, this led to a significant shift in cGMP/cAMP cross-talk and, in particular, to cGMP-driven augmentation of contractility in vitro and in vivo., Conclusions: Redistribution of cGMP-regulated phosphodiesterases and functional reorganization of receptor-associated microdomains occurs in early cardiac hypertrophy, affects cGMP-mediated contractility, and might represent a previously not recognized therapeutically relevant compensatory mechanism to sustain normal heart function., (© 2015 American Heart Association, Inc.)

Background: Arrhythmia is the major cause of death in patients with heart failure, for which β-adrenergic receptor blockers are a mainstay therapy. But the role of β-adrenergic signaling in electrophysiology and arrhythmias has never been studied in human ventricles., Methods and Results: We used optical imaging of action potentials and [Ca(2+)]i transients to compare the β1- and β2-adrenergic responses in left ventricular wedge preparations of human donor and failing hearts. β1-Stimulation significantly increased conduction velocity, shortened action potential duration, and [Ca(2+)]i transients duration (CaD) in donor but not in failing hearts, because of desensitization of β1-adrenergic receptor in heart failure. In contrast, β2-stimulation increased conduction velocity in both donor and failing hearts but shortened action potential duration only in failing hearts. β2-Stimulation also affected transmural heterogeneity in action potential duration but not in [Ca(2+)]i transients duration. Both β1- and β2-stimulation augmented the vulnerability and frequency of ectopic activity and enhanced substrates for ventricular tachycardia in failing, but not in donor, hearts. Both β1- and β2-stimulation enhanced Purkinje fiber automaticity, whereas only β2-stimulation promoted Ca-mediated premature ventricular contractions in heart failure., Conclusions: During end-stage heart failure, β2-stimulation creates arrhythmogenic substrates via conduction velocity regulation and transmurally heterogeneous repolarization. β2-Stimulation is, therefore, more arrhythmogenic than β1-stimulation. In particular, β2-stimulation increases the transmural difference between [Ca(2+)]i transients duration and action potential duration, which facilitates the formation of delayed afterdepolarizations., (© 2015 American Heart Association, Inc.)

Studies suggest that increased activity of Gi contributes to the reduced β-adrenoceptor-mediated inotropic response (βAR-IR) in failing cardiomyocytes and that β2AR-IR but not β1AR-IR is blunted by dual coupling to Gs and Gi. We aimed to clarify the role of Gi upon the β1AR-IR and β2AR-IR in Sham and failing myocardium by directly measuring contractile force and cAMP accumulation. Contractility was measured ex vivo in left ventricular strips and cAMP accumulation in cardiomyocytes from rats with post-infarction heart failure (HF) or sham operates (Sham). The β2AR-IR in Sham and HF was small and was amplified by simultaneously inhibiting phosphodiesterases 3 and 4 (PDE3&4). In HF, the inotropic response and cAMP accumulation evoked by β1AR- or β2AR-stimulation were reduced. Inactivation of Gi with pertussis toxin (PTX) did not restore the β1AR-IR or β2AR-IR in HF to Sham levels but did enhance the maximal β2AR-IR. PTX increased both β1AR- and β2AR-evoked cAMP accumulation more in Sham than that in HF, and HF levels approached those in untreated Sham. The potency of agonists at β1 and at β2ARs (only under PDE3&4 inhibition) was increased in HF and by PTX in both HF and Sham. Without PDE3&4 inhibition, PTX increased only the maximal β2AR-IR, not potency. We conclude that Gi regulates both β1AR- and β2AR-IR independent of receptor coupling with Gi. Gi together with PDE3&4 tonically restrict the β2AR-IR. Gi inhibition did not restore the βAR-IR in HF despite increasing cAMP levels, suggesting that the mechanism of impairment resides downstream to cAMP signalling.

Cardiac disorders are the main cause of mortality in autosomal-dominant polycystic kidney disease (ADPKD). However, how mutated polycystins predispose patients with ADPKD to cardiac pathologies before development of renal dysfunction is unknown. We investigate the effect of decreased levels of polycystin 2 (PC2), a calcium channel that interacts with the ryanodine receptor, on myocardial function. We hypothesize that heterozygous PC2 mice (Pkd2(+/-)) undergo cardiac remodeling as a result of changes in calcium handling, separate from renal complications. We found that Pkd2(+/-) cardiomyocytes have altered calcium handling, independent of desensitized calcium-contraction coupling. Paradoxically, in Pkd2(+/-) mice, protein kinase A (PKA) phosphorylation of phospholamban (PLB) was decreased, whereas PKA phosphorylation of troponin I was increased, explaining the decoupling between calcium signaling and contractility. In silico modeling supported this relationship. Echocardiography measurements showed that Pkd2(+/-) mice have increased left ventricular ejection fraction after stimulation with isoproterenol (ISO), a β-adrenergic receptor (βAR) agonist. Blockers of βAR-1 and βAR-2 inhibited the ISO response in Pkd2(+/-) mice, suggesting that the dephosphorylated state of PLB is primarily by βAR-2 signaling. Importantly, the Pkd2(+/-) mice were normotensive and had no evidence of renal cysts. Our results showed that decreased PC2 levels shifted the βAR pathway balance and changed expression of calcium handling proteins, which resulted in altered cardiac contractility. We propose that PC2 levels in the heart may directly contribute to cardiac remodeling in patients with ADPKD in the absence of renal dysfunction.

Aims: Cardiac β-adrenergic receptors (β-AR) are key regulators of cardiac haemodynamics and size. The scaffolding protein A-kinase anchoring protein 79/150 (AKAP5) is a key regulator of myocardial signalling by β-ARs. We examined the function of AKAP5 in regulating cardiac haemodynamics and size, and the role of β-ARs and Ca(2+)-regulated intracellular signalling pathways in this phenomenon., Methods and Results: We used echocardiographic, histological, genetic, and biochemical methods to examine the effect of ablation of AKAP5 on cardiac haemodynamics, size, and signalling in mice. AKAP5(-/-) mice exhibited enhanced signs of cardiac dilatation and dysfunction that progressed with age. Infusions of isoprenaline worsened cardiac haemodynamics in wild-type (WT) mice only, but increased the ratio of heart-to-body weight equally in WT and in AKAP5(-/-) mice. Mechanistically, loss of AKAP5 was associated with enhanced activity of cardiac calmodulin kinase II (CaMKII) and calcineurin (CaN) as indexed by nuclear factor of activated T-cell-luciferase activity. Loss of AKAP5 interfered with the recycling of cardiac β1-ARs, which was mediated in part by CaN binding to AKAP5. Carvedilol reversed cardiac hypertrophy and haemodynamic deficiencies in AKAP5(-/-) mice by normalizing the activities of cardiac CaN and CaMKII., Conclusions: These findings identify a novel cardioprotective role for AKAP5 that is mediated by regulating the activities of cardiac CaN and CaMKII and highlight a significant role for cardiac β-ARs in this phenomenon., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2014. For permissions please email: journals.permissions@oup.com.)

The expression and effects of β-adrenergic receptors (β-ARs) on the neurons of the bulbospinal rostral ventrolateral medulla (RVLM) have been limitedly examined to date. The objective of this study was to examine the expression of β1- and β2-ARs on the bulbospinal RVLM neurons electrophysiologically and histologically. To directly investigate whether RVLM neurons display sensitivity to metoprolol (a β1-AR antagonist), dobutamine (a β1-AR agonist), butoxamine (a β2-AR antagonist), and salbutamol (a β2-AR agonist), we examined changes in the membrane potentials of the bulbospinal RVLM neurons using the whole-cell patch-clamp technique during superfusion of these drugs. During metoprolol superfusion, 16 of the 20 RVLM neurons were hyperpolarized, and 5 of the 6 RVLM neurons were depolarized during dobutamine superfusion. During butoxamine superfusion, 11 of the 16 RVLM neurons were depolarized, and all of the 8 RVLM neurons were hyperpolarized during salbutamol superfusion. These results suggest the expression of β1- and β2-ARs on the RVLM neurons. To determine the presence of β1- and β2-ARs histologically, immunofluorescence examination was performed. Five metoprolol-hyperpolarized neurons were examined for β1-AR and tyrosine hydroxylase (TH) immunoreactivity. All of the neurons displayed β1-AR immunoreactivity, whereas three of the neurons displayed TH immunoreactivity. All of the five RVLM neurons that became depolarized during metoprolol superfusion and hyperpolarized during butoxamine superfusion displayed β1- and β2-AR immunoreactivity. Our findings suggest that β1-AR antagonists or β2-AR agonists may decrease blood pressure through decreasing the activity of the bulbospinal RVLM neurons.

Aims: Noradrenaline plays an important role in the modulation of atrial electrophysiology. However, the identity of the modulated channels, their mechanisms of modulation, and their role in the action potential remain unclear. This study aimed to investigate the noradrenergic modulation of an atrial steady-state outward current (IKss)., Methods and Results: Rat atrial myocyte whole-cell currents were recorded at 36°C. Noradrenaline potently inhibited IKss (IC50 = 0.90 nM, 42.1 ± 4.3% at 1 µM, n = 7) and potentiated the L-type Ca(2+) current (ICaL, EC50 = 136 nM, 205 ± 40% at 1 µM, n = 6). Noradrenaline-sensitive IKss was weakly voltage-dependent, time-independent, and potentiated by the arachidonic acid analogue, 5,8,11,14-eicosatetraynoic acid (EYTA; 10 µM), or by osmotically induced membrane stretch. Noise analysis revealed a unitary conductance of 8.4 ± 0.42 pS (n = 8). The biophysical/pharmacological properties of IKss indicate a TREK-like K(+) channel. The effect of noradrenaline on IKss was abolished by combined β1-/β2-adrenoceptor antagonism (1 µM propranolol or 10 µM β1-selective atenolol and 100 nM β2-selective ICI-118,551 in combination), but not by β1- or β2-antagonist alone. The action of noradrenaline could be mimicked by β2-agonists (zinterol and fenoterol) in the presence of β1-antagonist. The action of noradrenaline on IKss, but not on ICaL, was abolished by pertussis toxin (PTX) treatment. The action of noradrenaline on ICaL was mediated by β1-adrenoceptors via a PTX-insensitive pathway. Noradrenaline prolonged APD30 by 52 ± 19% (n = 5; P < 0.05), and this effect was abolished by combined β1-/β2-antagonism, but not by atenolol alone., Conclusion: Noradrenaline inhibits a rat atrial TREK-like K(+) channel current via a PTX-sensitive mechanism involving co-operativity of β1-/β2-adrenoceptors that contributes to atrial APD prolongation., (© The Author 2014. Published by Oxford University Press on behalf of the European Society of Cardiology.)

The sympathetic nervous system (SNS) plays an important role in mediating bone remodeling. However, the exact role that beta-1 adrenergic receptors (beta1AR) have in this process has not been elucidated. We have previously demonstrated the ability of dobutamine (DOB), primarily a beta1AR agonist, to inhibit reductions in cancellous bone formation and mitigate disuse-induced loss of bone mass. The purpose of this study was to characterize the independent and combined effects of DOB and hindlimb unloading (HU) on cancellous bone microarchitecture, tissue-level bone cell activity, and osteocyte apoptosis. Male Sprague-Dawley rats, aged 6-mos, were assigned to either normal cage activity (CC) or HU (n = 18/group) for 28 days. Animals were administered either daily DOB (4 mg/kg BW/d) or an equal volume of saline (VEH) (n = 9/gp). Unloading resulted in significantly lower distal femur cancellous BV/TV (-33%), Tb.Th (-11%), and Tb.N (-25%) compared to ambulatory controls (CC-VEH). DOB treatment during HU attenuated these changes in cancellous bone microarchitecture, resulting in greater BV/TV (+29%), Tb.Th (+7%), and Tb.N (+21%) vs. HU-VEH. Distal femur cancellous vBMD (+11%) and total BMC (+8%) were significantly greater in DOB- vs. VEH-treated unloaded rats. Administration of DOB during HU resulted in significantly greater osteoid surface (+158%) and osteoblast surface (+110%) vs. HU-VEH group. Furthermore, Oc.S/BS was significantly greater in HU-DOB (+55%) vs. CC-DOB group. DOB treatment during unloading fully restored bone formation, resulting in significantly greater bone formation rate (+200%) than in HU-VEH rats. HU resulted in an increased percentage of apoptotic cancellous osteocytes (+85%), reduced osteocyte number (-16%), lower percentage of occupied osteocytic lacunae (-30%) as compared to CC-VEH, these parameters were all normalized with DOB treatment. Altogether, these data indicate that beta1AR agonist treatment during disuse mitigates negative changes in cancellous bone microarchitecture and inhibits increases in osteocyte apoptosis.

Endogenous catecholamines such as adrenaline (A) and noradrenaline (NA) are released from the adrenal gland and sympathetic nervous system during exposure to stress. The adrenergic system plays a central role in stress signaling, and excessive stress was found to be associated with increased production of reactive oxygen species (ROS). Overproduction of ROS induces oxidative damage in tissues and causes the development of diseases such as cancer. In this study, we investigated the effects of quercetin-3-O-glucuronide (Q3G), a circulating metabolite of quercetin, which is a type of natural flavonoid, on the catecholamine-induced β2-adrenergic receptor (β2-AR)-mediated response in MDA-MB-231 human breast cancer cells expressing β2-AR. Treatment with A or NA at concentrations above 1μM generated significant levels of ROS, and NA treatment induced the gene expression of heme oxygenase-1 (HMOX1), and matrix metalloproteinase-2 (MMP-2) and -9 (MMP9). Inhibitors of p38 MAP kinase (SB203580), cAMP-dependent protein kinase (PKA) (H-89), activator protein-1 (AP-1) transcription factor (SR11302), and NF-κB and AP-1 (Tanshinone IIA) decreased MMP2 and MMP9 gene expression. NA also enhanced cAMP induction, RAS activation and phosphorylation of ERK1/2. These results suggested that the cAMP-PKA, MAPK, and ROS-NF-κB pathways are involved in β2-AR signaling. Treatment with 0.1μM Q3G suppressed ROS generation, cAMP and RAS activation, phosphorylation of ERK1/2 and the expression of HMOX1, MMP2, and MMP9 genes. Furthermore, Q3G (0.1μM) suppressed invasion of MDA-MB-231 breast cancer cells and MMP-9 induction, and inhibited the binding of [(3)H]-NA to β2-AR. These results suggest that Q3G may function to suppress invasion of breast cancer cells by controlling β2-adrenergic signaling, and may be a dietary chemopreventive factor for stress-related breast cancer., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Increased plasma catecholamine levels are associated with a high risk of perioperative cardiac events in aged individuals undergoing non-cardiac surgical interventions. Given the different effects of β1-adrenoreceptor (β1AR) and β2-adrenoreceptor (β2AR) stimulation by catecholamine in cardiomyocytes, this study evaluated whether simultaneous inhibition of β1AR and activation of β2AR is better than separate application in reducing the risk of perioperative cardiac events in aged rats undergoing non-cardiac surgery. Male aged Sprague-Dawley (SD) rats were divided into five groups. Normal group received no treatment. Surgery group received an abdominal surgery with hypoxia. β1- group, β2+ group, β2+ group and β1+β2+ group received surgery and hypoxia with metoprolol (100 mg/kg·d), fenoterol (250 μg/kg·d) or both, respectively. The drugs were given three days before surgery with treatment continued through post-surgical day 7. The results showed that simultaneous activation of β2AR with a β2AR agonist and inhibition of β1AR with a selective β1AR blocker normalized myocardial oxygen consumption, decreased myocardial damage, augmented cardiomyocyte survival, improved cardiac function, reduced the incidence of arrhythmia, thus decreasing the occurrence of cardiac events in perioperative aged rats undergoing non-cardiac surgery. The results demonstrated that combined use of β2AR agonist and β1AR blocker achieved better general effects than use of either one alone. Our results provide a new insight into preventing perioperative cardiac events for elderly patients undergoing surgical stress.

Gastrointestinal dysmotility is one of the nonmotor symptoms of Parkinson's disease (PD). Gastroparesis and upregulated β-adrenoceptors (β-ARs) have been reported in rats with bilateral microinjection of 6-hydroxydopamine (6-OHDA) in the substantia nigra, but the underlying mechanism is unclear. The aim of the current study is to investigate the role of β-ARs in gastroparesis in 6-OHDA rats. Gastric motility was studied through strain gauge measurement. Immunofluorescence, real-time reverse transcription-polymerase chain reaction and Western blotting were performed to examine the expression of β-ARs. Norepinephrine (NE) inhibited gastric motility in a dose-dependent fashion in both control and 6-OHDA rats, but much stronger adrenergic reactivity was observed in the 6-OHDA rats. The inhibition of gastric motility by NE in both control and 6-OHDA rats was not affected by tetrodotoxin, a neural sodium channel blocker. Blocking β1-AR or β2-AR did not affect the inhibition of strip contraction by NE in control rats, but β1-AR blockage obviously enhanced the half maximal inhibitory concentration value of NE in 6-OHDA rats. Selective inhibition of β3-AR blocked the effect of NE significantly in both control and 6-OHDA rats. The protein expression of β1-AR, but not β2-AR and β3-AR in gastric muscularis externa was increased significantly in 6-OHDA rats. In conclusion, β3-AR involves the regulation of gastric motility in control rats, whereas the upregulation of β1-AR is responsible for enhanced NE reactivity in 6-OHDA rats and therefore is involved in the formation of gastroparesis. The effect of both β1-AR and β3-AR on gastric motility is independent of the enteric nervous system., (Copyright © 2014 Mosby, Inc. All rights reserved.)

Purpose: Relaxin, a new drug for heart failure therapy, exerts its cardiac actions through relaxin family peptide receptor 1 (RXFP1). Factors regulating RXFP1 expression remain unknown. We have investigated effects of activation of adrenoceptors (AR), an important modulator in the development and prognosis of heart failure, on expression of RXFP1 in rat cardiomyocytes and mouse left ventricles (LV)., Methods: Expression of RXFP1 at mRNA (real-time PCR) and protein levels (immunoblotting) was measured in cardiomyocytes treated with α- and β-AR agonists or antagonists. RXFP1 expression was also determined in the LV of transgenic mouse strains with cardiac-restricted overexpression of α1A-, α1B- or β2-AR. Specific inhibitors were used to explore signal pathways involved in α1-AR mediated regulation of RXFP1 in cardiomyocytes., Results: In cultured cardiomyocytes, α1-AR stimulation resulted in 2-3 fold increase in RXFP1 mRNA (P < 0.001), which was blocked by specific inhibitors for protein kinase C (PKC) or mitogen-activated protein kinases/extracellular signal-regulated kinases (MAPK/ERK). Activation of β1-, but not β2-AR, significantly inhibited RXFP1 expression (P < 0.001). Relative to respective wild-type controls, RXFP1 mRNA levels in the LV of mice overexpressing α1A- or α1B-AR were increased by 3- or 10-fold, respectively, but unchanged in β2-AR transgenic hearts. Upregulation by α1-AR stimulation RXFP1 expression was confirmed at protein levels both in vitro and in vivo., Conclusions: Expression of RXFP1 was up-regulated by α1-AR but suppressed by β-AR, mainly β1-AR subtype, in cardiomyocytes. Future studies are warranted to characterize the functional significance of such regulation, especially in the setting of heart failure.

Rationale: β-Adrenoceptor activation contributes to sudden death risk in heart failure. Chronic β-adrenergic stimulation, as occurs in patients with heart failure, causes potentially arrhythmogenic reductions in slow delayed-rectifier K(+) current (IKs)., Objective: To assess the molecular mechanisms of IKs downregulation caused by chronic β-adrenergic activation, particularly the role of exchange protein directly activated by cAMP (Epac)., Methods and Results: Isolated guinea pig left ventricular cardiomyocytes were incubated in primary culture and exposed to isoproterenol (1 μmol/L) or vehicle for 30 hours. Sustained isoproterenol exposure decreased IKs density (whole cell patch clamp) by 58% (P<0.0001), with corresponding decreases in potassium voltage-gated channel subfamily E member 1 (KCNE1) mRNA and membrane protein expression (by 45% and 51%, respectively). Potassium voltage-gated channel, KQT-like subfamily, member 1 (KCNQ1) mRNA expression was unchanged. The β1-adrenoceptor antagonist 1-[2-((3-Carbamoyl-4-hydroxy)phenoxy)ethylamino]-3-[4-(1-methyl-4-trifluoromethyl-2-imidazolyl)phenoxy]-2-propanol dihydrochloride (CGP-20712A) prevented isoproterenol-induced IKs downregulation, whereas the β2-antagonist ICI-118551 had no effect. The selective Epac activator 8-pCPT-2'-O-Me-cAMP decreased IKs density to an extent similar to isoproterenol exposure, and adenoviral-mediated knockdown of Epac1 prevented isoproterenol-induced IKs/KCNE1 downregulation. In contrast, protein kinase A inhibition with a cell-permeable highly selective peptide blocker did not affect IKs downregulation. 1,2-Bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetate-AM acetoxymethyl ester (BAPTA-AM), cyclosporine, and inhibitor of nuclear factor of activated T cell (NFAT)-calcineurin association-6 (INCA6) prevented IKs reduction by isoproterenol and INCA6 suppressed isoproterenol-induced KCNE1 downregulation, consistent with signal-transduction via the Ca(2+)/calcineurin/NFAT pathway. Isoproterenol induced nuclear NFATc3/c4 translocation (immunofluorescence), which was suppressed by Epac1 knockdown. Chronic in vivo administration of isoproterenol to guinea pigs reduced IKs density and KCNE1 mRNA and protein expression while inducing cardiac dysfunction and action potential prolongation. Selective in vivo activation of Epac via sp-8-pCPT-2'-O-Me-cAMP infusion decreased IKs density and KCNE1 mRNA/protein expression., Conclusions: Prolonged β1-adrenoceptor stimulation suppresses IKs by downregulating KCNE1 mRNA and protein via Epac-mediated Ca(2+)/calcineurin/NFAT signaling. These results provide new insights into the molecular basis of K(+) channel remodeling under sustained adrenergic stimulation.

The phosphorylation of c-jun N-terminal kinase (JNK) and the subsequent production of C-C chemokine CCL2 (monocyte chemoattractant protein; MCP-1) in spinal astrocytes contribute to the initiation of neurological disorders including chronic pain. Astrocytes express neurotransmitter receptors which could be targeted to ameliorate neurological disorders. In the current study, the involvement of the β-adrenergic system in the regulation of JNK activity and CCL2 production after stimulation with tumor necrosis factor (TNF)-α, one of many initiators of neuroinflammation, was elucidated. Treatment of cultured spinal astrocytes with isoproterenol (a β-adrenergic receptor agonist; 1 µM) reduced both TNF-α-induced JNK1 phosphorylation, as observed by Western blotting, and the subsequent increase of both CCL2 mRNA expression and CCL2 production, which were measured by real time-PCR and ELISA, respectively. The effects of isoproterenol were completely blocked by pretreatment with either propranolol (a β-adrenoceptor antagonist) or H89 (a protein kinase A [PKA] inhibitor). The current study revealed that the regulation of glycogen synthase kinase-3β (GSK-3β) activity is a crucial factor in the inhibitory action of isoproterenol. The TNF-α-induced JNK1 phosphorylation was significantly blocked by treatment with GSK-3β inhibitors (either LiCl or TWS119), and stimulation of β-adrenergic receptors induced the inhibition of GSK-3β through the phosphorylation of Ser(9) . Moreover, treatment with isoproterenol markedly suppressed the TNF-α-induced increase of CCL2 mRNA expression and CCL2 production through a β-adrenergic receptor-PKA pathway mediated by GSK-3β regulation. Thus, activation of β1/2 adrenergic receptors expressed in spinal astrocytes could be a novel method of moderating neurological disorders with endogenous catecholamines or selective agonists., (© 2013 Wiley Periodicals, Inc.)

Background: Clinical studies have now confirmed the link between short-term exposure to elevated levels of air pollution and increased cardiovascular mortality, but the mechanisms are complex and not completely elucidated. The present study was designed to investigate the hypothesis that activation of pulmonary sensory receptors and the sympathetic nervous system underlies the influence of pulmonary exposure to diesel exhaust particulate on blood pressure, and on the myocardial response to ischemia and reperfusion., Methods & Results: 6 h after intratracheal instillation of diesel exhaust particulate (0.5 mg), myocardial ischemia and reperfusion was performed in anesthetised rats. Blood pressure, duration of ventricular arrhythmia, arrhythmia-associated death, tissue edema and reperfusion injury were all increased by diesel exhaust particulate exposure. Reperfusion injury was also increased in buffer perfused hearts isolated from rats instilled in vivo, excluding an effect dependent on continuous neurohumoral activation or systemic inflammatory mediators. Myocardial oxidant radical production, tissue apoptosis and necrosis were increased prior to ischemia, in the absence of recruited inflammatory cells. Intratracheal application of an antagonist of the vanilloid receptor TRPV1 (AMG 9810, 30 mg/kg) prevented enhancement of systolic blood pressure and arrhythmia in vivo, as well as basal and reperfusion-induced myocardial injury ex vivo. Systemic β1 adrenoreceptor antagonism with metoprolol (10 mg/kg) also blocked enhancement of myocardial oxidative stress and reperfusion injury., Conclusions: Pulmonary diesel exhaust particulate increases blood pressure and has a profound adverse effect on the myocardium, resulting in tissue damage, but also increases vulnerability to ischemia-associated arrhythmia and reperfusion injury. These effects are mediated through activation of pulmonary TRPV1, the sympathetic nervous system and locally generated oxidative stress.

Inflammation reduces pharmacological response to β1-blockers by down-regulating the target receptor protein. This may contribute to the sub-optimal response to pharmacotherapy with β-blockers. Nebivolol is a third generation β-adrenoceptor (AR) blocker with high selectivity for blocking β1 and β3-agonistic properties. We studied whether response to nebivolol is also reduced by inflammation. Male Sprague-Dawley rats (Inflamed; Mycobacterium butyricum induced) and Control (healthy) were orally administered single doses of 2 mg/kg nebivolol (n=5) or 25 mg/kg propranolol (positive control, n=7-8); ECG recorded for PR and RR interval measurements; serial blood samples were collected for pharmacokinetic assessment. Subsequently, the myocardial β1, β2 and β3-AR levels were measured in homogenized hearts. For propranolol, inflammation resulted in increased concentration but reduced response and down-regulation of β1- AR. The action and disposition of nebivolol were, however, unaffected by inflammation despite the reduced β1-AR levels. The levels of β2 and β3-AR were unaffected by inflammation. The consistency of response to nebivolol despite inflammation may be due to the predominance of contribution of β2 and β3-AR. The lack of an inhibitory effect of inflammation on the clearance of nebivolol is suggestive of mechanisms other than an efficient hepatic metabolism for its low bioavailability. If extrapolated to human, nebivolol may be a more effective cardiovascular drug when inflammatory conditions are present.

Background: Brain α2- and β-adrenoceptor alterations have been suggested in suicide and major depressive disorder., Methods: The densities of α2-, β1- and β2-adrenoceptors in postmortem prefrontal cortex of 26 subjects with depression were compared with those of age-, gender- and postmortem delay-matched controls. The effect of antidepressant treatment on α2- and β-adrenoceptor densities was also evaluated. α2- and β-adrenoceptor densities were measured by saturation experiments with respective radioligands [(3)H]UK14304 and [(3)H]CGP12177. β1- and β2-adrenoceptor subtype densities were dissected by means of β1-adrenoceptor selective antagonist CGP20712A., Results: Both, α2- and β1-adrenoceptors densities were higher in antidepressant-free depressed subjects (n=14) than those in matched controls (Δ~24%, p=0.013 and Δ~20%, p=0.044, respectively). In antidepressant-treated subjects (n=12), α2-adrenoceptor density remained increased over that in controls (Δ~20%), suggesting a resistance of α2-adrenoceptors to the down-regulatory effect of antidepressants. By contrast, β1-adrenoceptor density in antidepressant-treated depressed subjects was not different from controls, suggesting a possible down-regulation by antidepressants. The down-regulation of β1-adrenoceptor density in antidepressant-treated depressed subjects differs from the unaltered β1-adrenoceptor density observed in citalopram-treated rats and in a group of non-depressed subjects also treated with antidepressants (n=6). β2-adrenoceptor density was not altered in depressed subjects independently of treatment., Limitations: Antidepressant-treated subjects had been treated with a heterogeneous variety of antidepressant drugs. The results should be understood in the context of suicide victims with depression., Conclusions: These results show the up-regulation of brain α2- and β1-adrenoceptors in depression and suggest that the regulation induced by chronic antidepressant treatment would be altered in these subjects., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Background and Purpose: The β1-adrenoceptor has at least two binding sites, high and low affinity sites (β1H and β1L, respectively), which mediate cardiostimulation. While β1H-adrenoceptor can be blocked by all clinically used β-blockers, β1L-adrenoceptor is relatively resistant to blockade. Thus, chronic β1L-adrenoceptor activation may mediate persistent cardiostimulation, despite the concurrent blockade of β1H-adrenoceptors. Hence, it is important to determine the potential significance of β1L-adrenoceptors in vivo, particularly in pathological situations., Experimental Approach: C57Bl/6 male mice were used. Chronic (4 or 8 weeks) β1L-adrenoceptor activation was achieved by treatment, via osmotic mini pumps, with (-)-CGP12177 (10 mg·kg(-1)·day(-1)). Cardiac function was assessed by echocardiography and micromanometry., Key Results: (-)-CGP12177 treatment of healthy mice increased heart rate and left ventricular (LV) contractility. (-)-CGP12177 treatment of mice subjected to transverse aorta constriction (TAC), during weeks 4-8 or 4-12 after TAC, led to a positive inotropic effect and exacerbated fibrogenic signalling while cardiac hypertrophy tended to be more severe. (-)-CGP12177 treatment of mice with TAC also exacerbated the myocardial expression of hypertrophic, fibrogenic and inflammatory genes compared to untreated TAC mice. Washout of (-)-CGP12177 revealed a more pronounced cardiac dysfunction after 12 weeks of TAC., Conclusions and Implications: β1L-adrenoceptor activation provides functional support to the heart, in both normal and pathological (pressure overload) situations. Sustained β1L-adrenoceptor activation in the diseased heart exacerbates LV remodelling and therefore may promote disease progression from compensatory hypertrophy to heart failure., (© 2013 The British Pharmacological Society.)