Β-adrenoceptor Density Research Articles

Comparative characterization of β-adrenoceptors in the bladder, heart, and lungs of rats: Alterations in spontaneously hypertensive rats

The present study aimed to characterize and compare β-adrenoceptors in the rat bladder with those in the heart and lungs of SD rats (8–10 weeks old) using subtype-selective agonists and antagonists in a radioligand binding assay with (-)-[125I]cyanopindolol ([125I]CYP), and also to clarify alterations in β-adrenoceptors in the bladder of spontaneously hypertensive rats (SHR) at 14 weeks old, from those of Wistar-Kyoto rats (WKY) and Wistar rats at the same age. A radioligand binding assay with [125I]CYP was used to measure β-adrenoceptor binding activity in rat tissues. Metoprolol exhibited the highest affinity to specific binding sites of [125I]CYP in the rat heart, indicating the dominance of β1-adrenoceptors. β3-selective agonists (BRL37344 and CL316243) and antagonist (SR59230A) exhibited higher affinity to specific binding sites of [125I]CYP in the bladder than in the heart and lungs. Furthermore, the binding affinity of the β2-selective antagonist, ICI118551 was the highest in the bladder. The Bmax of specific [125]CYP binding in the bladder was significantly lower in WKY and SHR than in Wistar rats. The present study provides further evidence for the coexistence of β2-and β3-adrenoceptors in the rat bladder, and indicates that β-adrenoceptor density is lower in the bladders of WKY and SHR.

The Rho kinase (ROCK) inhibitor Y-27632 reduces the β2-adrenoceptor density but enhance cAMP formation in primary equine bronchial epithelial cells

The present study addresses the effect of the Rho-kinase (ROCK) inhibitor Y-27632 on the β2-adrenoceptor density and β-agonist-stimulated intracellular second messenger cAMP formation in primary equine bronchial epithelial cells (EBEC). Y-27632 significantly decreased the β2-adrenoceptor number (Bmax) without markedly affecting the receptor affinity (dissociation constant, KD) to the radioligand [125I]-iodocyanopindolol (ICYP). In contrast, Y-27632 augmented the β-agonist-stimulated intracellular cAMP production. Herein, Y-27632 markedly increased the maximal cAMP responses (Emax) (isoproterenol > epinephrine > norepinephrine) but did not shift the β-agonist concentration-effect curves to the left. The β2-selective antagonist ICI 118.551 and the β1/β2-antagonsit propranolol but not the β1-selctive antagonist CGP 20712A reversed the isoproterenol-induced cAMP formation equally in Y-27632-treated and control EBEC, suggesting the effect was merely related to the β2-subtype. These results show that Y-27632 differentially regulates the receptor density and function. Thus, these findings provide the first evidence that the functional interaction of the β2-adrenoceptor and Rho-kinase (ROCK) signaling pathways decreases the receptor expression but enhances receptor downstream cAMP formation. This differential regulation of the receptor density and function by Y-27632 should be further reconsidered with regard to the beneficial effect of the drug in asthma therapy.

Early postnatal environmental enrichment restores neurochemical and functional plasticities of the cerebral cortex and improves learning performance in hidden-prenatally-malnourished young-adult rats

Moderate reduction of dietary protein (from 25% to 8% casein) in pregnant rats, calorically compensated by carbohydrates, gives rise to ‘hidden prenatal malnutrition’ (HPM) in the offspring since it does not alter body and brain weights of pups at birth. However, this dietary treatment leads to decreased β-adrenoceptor signaling and brain derived neurotrophic factor (BDNF) levels in the pup' brain, altogether with defective cortical long-term potentiation (LTP) and lowered visuospatial memory performance. Since early postnatal environmental enrichment (EE) has been shown to exert plastic effects on the developing brain and neuroprotection both on cognition and on structural properties of the neocortex, in the present study we addressed the question of whether early postnatal EE during the lactation period could exert compensatory changes in the expression of ®-adrenergic receptors and BDNF in the neocortex of HPM rats, and if these effects are associated with an improvement or even a restore of both neocortical LTP in vivo and cognitive performance induced by HPM. The results obtained show that EE restored β-adrenoceptor density, BDNF expression and the ability to support LTP at prefrontal and occipital cortices of HPM rats. Besides, EE improved learning performance in visuospatial and operant conditioning tasks. The latter support the notion that adequate maternal protein nutrition during pregnancy is required for proper brain development and function. Further, the results highlight the role of environmental enrichment during early postnatal life in increasing later brain plasticity and exerting neuroprotection against brain deficits induced by prenatal malnutrition.

Differential regulation of the β-adrenoceptor density and cyclic AMP level with age and sex in turkey cardiac chambers

Decreased responses of the heart to β-adrenoceptor stimulation with aging have been shown to occur merely in selected heart chambers in relation to increased catecholamine levels. However, there are no systematic studies that investigate all cardiac chambers with regard to receptor density and cAMP (adenosine 3′, 5′-cyclic monophosphate) responses. We used meat-type turkey poults (British United Turkey (B.U.T.) Big 6) with increasing age because their heart seems to decrease in weight in relation to body weight and they are often used as an animal model for heart failure. The receptor density and distribution were quantified by radioligand binding analysis using (-)-[125I]-iodocyanopindolol and β-adrenoceptor subtype-specific antagonists (ICI 118.551 and CGP 20712 A) in membranes of four cardiac chambers (right and left atria and ventricles) of 6-week-, 12-week-, 16/21-week-, and 57-week-old B.U.T. BIG 6 turkeys. Receptor function was determined by measuring basal and stimulated cAMP production. In both sexes, the β-adrenoceptor density decreased significantly in all chambers with age without altered β-adrenoceptor subtype distribution. The receptor affinity (KD) to the radioligand was similar in hearts of all age groups. β-adrenoceptor-(isoproterenol and guanosine 5′-triphosphate), G-protein-(NaF) and catalytic unit of adenylate cyclase (forskolin, Mn2+) mediated cAMP responses were not chamber-dependent. Indeed, the cAMP level was significantly lower in 57-week-old hearts than in 6-week-, 12-week-, 16/21-week-old hearts. These data suggest that with increasing age and body weight, the β-adrenoceptor signal transduction pathway was highly blunted in all cardiac chambers, occurring by decreased receptor density and cAMP responses.

Preclinical characterization of a novel radiolabeled analog of practolol for the molecular imaging of myocardial β-adrenoceptor density

BackgroundThe great clinical potential of myocardial β-AR imaging has been shown by recent studies evaluating the β-AR-specific, non-selective agent [11C]-CGP12177 in the setting of idiopathic-dilated cardiomyopathy, and myocardial infarction. However, the short half-life of 11C hampers the potential of [11C]-CGP12177 for routine clinical use. AMI9 is an analog of the β-adrenoceptor ligand practolol that can readily be labeled using radioactive isotopes of iodine. The present study was aimed at characterizing the in vitro, ex vivo, and in vivo β-AR binding properties of [125I]-AMI9. Methods and ResultsNewborn rat cardiomyocytes were used for saturation and kinetic binding assays as well as for displacement and competition experiments. Isolated perfused rat hearts were used to evaluate the pharmacological activity of AMI9. The in vivo kinetics of [125I]-AMI9 were studied using biodistribution experiments in mice. [125I]-AMI9 displayed high specific affinity for β-AR with no β-AR subtype selectivity (KD, 5.6 ± 0.3 nM; Bmax, 231 ± 7 fmol·(mg protein)−1). AMI9 potently inhibited the inotropic effects of isoproterenol. The early in vivo cardiac and lung activities of [125I]-AMI9 compared favorably with those of the clinically validated tracer CGP12177. ConclusionIodine-labeled AMI9 is a promising agent for the molecular imaging of myocardial β-AR density.

Effects of long-term application of metoprolol and propranolol in a rat model of smoking.

Beta-blockers, especially selective β1 -adrenoceptor antagonists, are often used to treat cardiovascular disease, even when complicated by chronic obstructive pulmonary disease. The association of beta-blocker selectivity and treatment effects is disputed, and the curative effects and side-effects of various antagonists may differ. Herein we investigated the effects of 1months treatment with the selective β1 -adrenoceptor antagonist metoprolol and the non-selective β-adrenoceptor antagonist propranolol on pulmonary function and pathology in a 4-month rat model of passive cigarette smoke exposure and explored potential mechanisms of action. Lung function and general pathological changes were evaluated after 4months exposure to cigarette smoke, with metoprolol and propranolol treatment (50 and 25mg/kg per day, respectively; intragastrically) during the last month. Cytokine and mucin levels in bronchoalveolar lavage fluid (BALF) were determined by ELISA, whereas β1 - and β2 -adrenoceptor expression in the lungs was evaluated by immunohistochemistry and western blot analysis. Chronic treatment with metoprolol and propranolol did not exacerbate peak expiratory flow or intra-airway pressure in rats exposed to cigarette smoke. Propranolol significantly attenuated inflammatory cell infiltration, cytokine levels (tumour necrosis factor-α and interleukin-8) in BALF or mucus secretion, whereas metoprolol reduced only smooth muscle proliferation. Moreover, propranolol treatment was associated (albeit not significantly) with restoring β2 -adrenoceptor expression in airway epithelia. Propranolol had a more beneficial effect on cigarette smoking-induced lung damage than metoprolol in a smoking rat model that may be associated with restoration of endogenous β2 -adrenoceptor density in the airway epithelial cells.

Increased α2- and β1-adrenoceptor densities in postmortem brain of subjects with depression: Differential effect of antidepressant treatment

BackgroundBrain α2- and β-adrenoceptor alterations have been suggested in suicide and major depressive disorder. MethodsThe 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 [3H]UK14304 and [3H]CGP12177. β1- and β2-adrenoceptor subtype densities were dissected by means of β1-adrenoceptor selective antagonist CGP20712A. ResultsBoth, 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. LimitationsAntidepressant-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. ConclusionsThese 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.

Activation of Cardiomyocytes Depending on Their Proximity to Human Bone Marrow Stem Cells

Our study aimed to elucidate whether bone marrow stem cell (BMC) treatment might result in a cellular response in cardiomyocytes IN VITRO. Subconfluent neonatal rat cardiomyocyte cultures were cocultured for three days with Vybrant CM-DiI labeled BMC from human sternal bone marrow and underwent immunohistological staining for the proto-oncogene c-Myc and the cell cycle proteins CDK2, CDK4 and ATF-3. β-adrenoceptor density was analyzed using [125I]-iodocyanopindolol (ICYP) histoautoradiography. Quantitative analysis of immunohistochemical images revealed significantly increased expression and upregulation of c-Myc, and its downstream targets ATF-3, CDK2 and CDK4 in neighboring cardiomyocytes to BMC, depending on their distance to the BMC compared to cardiomyocytes far from the BMC. Histoautoradiography revealed a significantly higher β-adrenoceptor density in cardiomyocytes in the immediate vicinity to the BMC. With increasing distance to the BMC, β-adrenoceptor density in cardiomyocytes declined. Thus, a small number of BMC can affect a larger number of cardiomyocytes by activating an intracellular signaling cascade and enhancing β-adrenoceptor density.

Modulation of intracellular calcium transient in response to β-adrenoceptor stimulation in the hearts of 4-wk-old rats during simulated weightlessness

Modulation of intracellular calcium ([Ca(2+)](i)) transient in response to beta-adrenoceptor stimulation in the hearts of hindlimb unweighted (HLU) rats during simulated weightlessness has not been reported. In the present study, we adopted the rat tail suspension for 4 wk to simulate weightlessness. Effects of simulated microgravity on beta-adrenoceptor responsiveness were then studied. Mean arterial blood pressure, left ventricular pressure (LVP), systolic function [maximum positive change in pressure over time (+dP/dt(max))], and diastolic function [maximum negative change in pressure over time (-dP/dt(max))] were monitored during the in vivo experiment. beta-Adrenoceptor density was quantitated by radioactive ligand binding. Single rat ventricular myocyte was obtained by enzymatic dissociation method. +/-dP/dt(max), myocyte contraction, intracellular [Ca(2+)](i) transient, and L-type calcium current in response to beta-adrenoceptor stimulation with isoproterenol were measured. Compared with the control group, no significant changes were found in heart weight, body weight, and mean arterial blood pressure, whereas LVP and +/-dP/dt(max) were significantly reduced. LVP and +/-dP/dt(max) were significantly attenuated in the HLU group in response to isoproterenol administration. In the in vitro study, the beta-adrenoceptor density was unchanged. Effects of isoproterenol on electrically induced single-cell contraction and [Ca(2+)](i) transient in myocytes of ventricles in HLU rats were significantly attenuated. The enhanced L-type Ca(2+) current elicited by isoproterenol in cardiomyocytes was significantly decreased in the HLU group. The above results indicate that impaired function of L-type Ca(2+) current and decreased [Ca(2+)](i) transient cause the depressed responsiveness of the beta-adrenoceptor stimulation, which may be partially responsible for the depression of cardiac function.

Influence of β 2-adrenoceptor gene polymorphisms on β 2-adrenoceptor expression in human lung

Background The aim of the present study was to establish whether polymorphisms, especially those within the promoter region, of the β 2-adrenoceptor gene ( ADRB2) influence β 2-adrenoceptor expression in human lung. Methods The density of β-adrenoceptors in human lung tissue ( n = 88) was determined by saturation binding using the radioligand, iodinated cyanopindolol. Discrimination of β 1- and β 2-adrenoceptors was determined using the highly selective β 1-adrenoceptor antagonist, CGP20712A. Genotype was determined at 5 positions of ADRB2 previously reported as polymorphic. Potential influences of single nucleotide polymorphisms (SNPs) within the promoter region (−367, −47) and coding block (46, 79, 491) of ADRB2 on β 2-adrenoceptor expression were investigated. Results The density of β 2-adrenoceptors was variable among the 88 lung preparations studied ranging from 17 to 177 fmol/mg protein (mean ± S.E.M., 72 ± 4 fmol/mg protein). There was no influence of genotype on β 2-adrenoceptor expression for any of the polymorphisms studied except at position 491. The polymorphism at position 491C > T, leading to a change from thr to ile at amino acid 164, is uncommon. Preparations genotyped as heterozygous (126 ± 15 fmol/mg protein; n = 5) expressed significantly ( P = 0.0005) higher levels of β 2-adrenoceptor than those that were homozygous (69 ± 4 fmol/mg protein; n = 83). Conclusion With the exception of position 491, these data indicate that polymorphisms of ADRB2 do not influence β 2-adrenoceptor expression in human lung.

Effects of dexamethasone on the expression of β1-, β2- and β3-adrenoceptor mRNAs in skeletal and left ventricle muscles in rats

Glucocorticoids are known to increase the density and mRNA levels of beta-adrenoceptors (beta-AR) via the glucocorticoid receptor (GR) in many tissues. However, the effects of these changes in the skeletal and cardiac muscles remain relatively unknown. We have investigated the effects of dexamethasone on the expression of the beta(1)-, beta(2)-, and beta(3)-AR mRNAs and GR mRNA in fast-twitch fiber-rich extensor digitorum longus (EDL), slow-twitch fiber-rich soleus (SOL), and left ventricle (LV) muscles by real-time quantitative RT-PCR. Male rats were divided into a dexamethasone group and control group. The weight, RNA concentration, and total RNA content of EDL muscle were 0.76-, 0.85-, and 0.65-fold lower, respectively, in the dexamethasone group than in the control group. The weight, RNA concentration, and total RNA content of SOL muscle were 0.92-, 0.87-, and 0.81-fold lower, respectively, in the dexamethasone group than in the control group; these differences were significant. However, the weight/body weight and total RNA content/body weight of LV muscle were 1.38- and 1.39-fold higher, respectively, in the dexamethasone group than in the control group, respectively; these differences were also significant. Dexamethasone significantly decreased GR mRNA expression in EDL muscle without changing the expression of the beta(1)-, beta(2)-, and beta(3)-AR mRNAs. However, dexamethasone significantly decreased the expressions of beta(2)-AR and GR mRNAs in SOL muscle and significantly increased beta(1)-AR mRNA expression in LV muscle-without changing GR mRNA expression. These results suggest that the effects of dexamethasone on the expression of beta(1)- and beta(2)-AR mRNAs and muscle mass depend on the muscle contractile and/or constructive types.

Effect of carvedilol on β-adrenoceptor density on peripheral lymphocytes from patients with chronic heart failure

Objective To explore the effect of carvedilol on a-adrenoceptor density on peripheral lymphocytes from patients with chronic heart failure(CHF).Methods 112 CHF patients[New York Heart Association(NYHA) class Ⅱ ~Ⅳ) were randomly received either low dose carvedilol(12.5mg/d,n=56) or high dose carvedilol(50mg/d,n=56) for three months. Echocardiographic examinations were performed before and after drug administration. The maximum density of a-adrenoceptor( β max) on peripheral lymphocytes was measured by radioligand binding assay using 3H-dihydroalprenolol.Results The total study population consisted of 105 patients. The left ventricular end-diastolic dimension(LVEDD) and left ventricular end-systolic dimension(LVESD) were significantly decreased, the left ventricular ejection fraction(LVEF) was significantly increased after drug treatment. Compared with the low dose carvedilol group, the high dose carvedilol groun showed that LVEDD and LVESD were significantly decreased(P< 0.05), whereas LVEF significantly increased(P<0.01). The level of β max was much lower in those patients with more severe symptoms(NYHA class Ⅱ ~ Ⅴ). The level of β max was not significantly changed before and after drug treatment in the low dose carvedilol group, but was significantly decreased after drug treatment in the high dose carvedilol group(P < 0.05).Conclusion High dose carvedilol is more effective than low dose in attenuating left ventricular remodeling and in improving systolic function after heart failure. The beneficial effect of earvedilol is not depending on the change of β -adrenoceptor density. Key words: Heart failure ; β3-adrenoceptor ; Lymphocyte ; Carvedilol

High beta-adrenoceptor density on peripheral blood mononuclear cells in progressive multiple sclerosis: a manifestation of autonomic dysfunction?

In multiple sclerosis (MS) up-regulation of beta-adrenoceptors on peripheral blood mononuclear cells (PBMCs) has been attributed to either autonomic dysfunction, inflammation or a combination of the two. We have compared secondary progressive MS patients with normal subjects (NS) and two models of autonomic dysfunction; pure autonomic failure (PAF) and multiple system atrophy (MSA, Shy-Drager syndrome). There was up-regulation of beta-adrenoceptors on PBMCs in MS and PAF patients but not in MSA patients. Only in PAF patients beta-adrenoceptor up-regulation was correlated with low plasma levels of noradrenaline (NA) and adrenaline (Ad). In addition to studies in the basal state, measurements also were made after the centrally acting sympatholytic agent clonidine. These were combined with haemodynamic and neurohormonal measurements. After clonidine, there was a fall in blood pressure in NS and MSA patients but not in MS and PAF patients; a rise in growth hormone (GH) in NS and PAF patients but not in MS and MSA patients; and an up-regulation in PBMCs beta-adrenoceptors in NS but not in MS, MSA and PAF patients. Up-regulation of beta-adrenoceptors on PBMCs in MS could be attributed to autonomic dysfunction but the disparity between MS and PAF patients when considering their plasma levels of NA and Ad argue against. Although the neurohormonal responses to clonidine and the physiological assessment of autonomic function in progressive MS patients, demonstrate central autonomic dysfunction resembling that of the MSA patients, the normal basal beta-adrenoceptor densities in the latter, suggests that the up-regulation of these receptors is independent of the central autonomic dysfunction in MS.

β-adrenoceptor affinity as a biological predictor of treatment response to paroxetine in patients with acute panic disorder

Background Few studies have reported on the functional differences of the β-adrenoceptor between treatment responders and non-responders in panic disorder (PD). The aim of this study was to compare the nature of the β-adrenoceptor function and clinical variables between treatment responders and non-responders to paroxetine treatment in acute PD patients. Method Paroxetine was administered to all of the panic patients for 12 weeks. The lymphocyte β-adrenoceptor density (Bmax), affinity (1/Kd), and sensitivity (cAMP ratio) were measured in 22 untreated outpatients with acute PD and 22 age, sex and BMI matched control subjects. Psychological assessments were conducted using the HAM-A, and HAM-D, STAI-S and STAI-T, Anxiety sensitivity index (ASI), and Acute panic inventory (API). Results A significantly higher Kd was observed in the panic patients before treatment as compared with the control subjects, but there was no significant difference in Kd between the panic patients and control subjects after the treatment. Among the 22 patients, the 11 treatment responders (50%) showed a significantly higher Kd and lower mean scores of HAM-D, STAI-S, STAI-T, and ASI at baseline, compared with the non-responders. Logistic regression revealed that the pretreatment Kd and HAM-D were significantly reliable predictors for treatment response ( p < 0.05). Conclusion The β-adrenoceptor affinity (1/Kd) was decreased and adaptively normalized after treatment with paroxetine in the acute panic patients. In addition, a low pretreatment β-adrenoceptor affinity (1/Kd) was found to predict the treatment response and can be suggested as a biological predictor of treatment response in acute PD.

Enhanced calcium cycling and contractile function in transgenic hearts expressing constitutively active Gαo* protein

In contrast to the other heterotrimeric GTP-binding proteins (G proteins) Gs and Gi, the functional role of G o is still poorly defined. To investigate the role of G alpha o in the heart, we generated transgenic mice with cardiac-specific expression of a constitutively active form of G alpha o1* (G alpha o*), the predominant G alpha o isoform in the heart. G alpha o expression was increased 3- to 15-fold in mice from 5 independent lines, all of which had a normal life span and no gross cardiac morphological abnormalities. We demonstrate enhanced contractile function in G alpha o* transgenic mice in vivo, along with increased L-type Ca2+ channel current density, calcium transients, and cell shortening in ventricular G alpha o*-expressing myocytes compared with wild-type controls. These changes were evident at baseline and maintained after isoproterenol stimulation. Expression levels of all major Ca2+ handling proteins were largely unchanged, except for a modest reduction in Na+/Ca2+ exchanger in transgenic ventricles. In contrast, phosphorylation of the ryanodine receptor and phospholamban at known PKA sites was increased 1.6- and 1.9-fold, respectively, in G alpha o* ventricles. Density and affinity of beta-adrenoceptors, cAMP levels, and PKA activity were comparable in G alpha o* and wild-type myocytes, but protein phosphatase 1 activity was reduced upon G alpha o* expression, particularly in the vicinity of the ryanodine receptor. We conclude that G alpha o* exerts a positive effect on Ca2+ cycling and contractile function. Alterations in protein phosphatase 1 activity rather than PKA-mediated phosphorylation might be involved in hyperphosphorylation of key Ca2+ handling proteins in hearts with constitutive G alpha o activation.

Different changes of plasma membrane β-adrenoceptors in rat heart after chronic administration of propranolol, atenolol and bevantolol

Recently, tissue segment binding method with a hydrophilic radioligand [ 3H]-CGP12177 was developed to detect plasma membrane β-adrenoceptors in rat heart (Horinouchi et al., 2006). In the present study, propranolol (40 mg kg − 1 day − 1 ), atenolol (40 mg kg − 1 day − 1 ) and bevantolol (200 mg kg − 1 day − 1 ) were administered to rats for 6 weeks, and the changes of plasma membrane β-adrenoceptors and their mRNA expression in rat ventricle were examined. Chronic administration of propranolol increased the β 1-adrenoceptors but decreased the β 2-adrenoceptors without changing total amount of plasma membrane β-adrenoceptors. Atenolol increased both plasma membrane β 1- and β 2-adrenoceptors, whereas bevantolol had no effect on the β-adrenoceptor density and their subtype proportions. In contrast, the density of β-adrenoceptors detected in conventional homogenate binding study was extremely low (approximately 60% of plasma membrane β-adrenoceptors detected with the tissue segment binding method) and the effects of chronic administration of β-adrenoceptor antagonists were not necessarily in accord with those at the plasma membrane β-adrenoceptors. The mRNA levels of β 1- and β 2-adrenoceptors were not altered by propranolol treatment, while β 1-adrenoceptor mRNA significantly decreased after administration of atenolol or bevantolol without changing the level of β 2-adrenoceptor mRNA. The present binding study with intact tissue segments clearly shows that the plasma membrane β 1- and β 2-adrenoceptors of rat heart, in contrast to the homogenate binding sites and the mRNA levels, are differently affected by chronic treatment with three β-adrenoceptor antagonists; up- and down-regulations of β 1- and β 2-adrenoceptors, respectively, by propranolol, and up-regulation of both the subtypes by atenolol, but no significant change in both the subtypes by bevantolol.

Effects of autologous bone marrow stem cell transplantation on beta-adrenoceptor density and electrical activation pattern in a rabbit model of non-ischemic heart failure

BackgroundSince only little is known on stem cell therapy in non-ischemic heart failure we wanted to know whether a long-term improvement of cardiac function in non-ischemic heart failure can be achieved by stem cell transplantation.MethodsWhite male New Zealand rabbits were treated with doxorubicine (3 mg/kg/week; 6 weeks) to induce dilative non-ischemic cardiomyopathy. Thereafter, we obtained autologous bone marrow stem cells (BMSC) and injected 1.5–2.0 Mio cells in 1 ml medium by infiltrating the myocardium via a left anterolateral thoracotomy in comparison to sham-operated rabbits. 4 weeks later intracardiac contractility was determined in-vivo using a Millar catheter. Thereafter, the heart was excised and processed for radioligand binding assays to detect β1- and β2-adrenoceptor density. In addition, catecholamine plasma levels were determined via HPLC. In a subgroup we investigated cardiac electrophysiology by use of 256 channel mapping.ResultsIn doxorubicine-treated animals β-adrenoceptor density was significantly down-regulated in left ventricle and septum, but not in right ventricle, thereby indicating a typical left ventricular heart failure. Sham-operated rabbits exhibited the same down-regulation. In contrast, BMSC transplantation led to significantly less β-adrenoceptor down-regulation in septum and left ventricle. Cardiac contractility was significantly decreased in heart failure and sham-operated rabbits, but was significantly higher in BMSC-transplanted hearts. Norepinephrine and epinephrine plasma levels were enhanced in heart failure and sham-operated animals, while these were not different from normal in BMSC-transplanted animals. Electrophysiological mapping revealed unaltered electrophysiology and did not show signs of arrhythmogeneity.ConclusionBMSC transplantation improves sympathoadrenal dysregualtion in non-ischemic heart failure.

Pharmacological evaluation of plasma membrane β-adrenoceptors in rat hearts using the tissue segment binding method

This study evaluates β-adrenoceptors in rat atria and ventricle using the tissue segment binding method and compares the results with those obtained using conventional homogenate binding assays. In studies with tissue segment binding, the hydrophilic radioligand [ 3H]-CGP12177 selectively bound to plasma membrane β-adrenoceptors, and the B max levels were significantly higher than those obtained with homogenate binding. However, both binding approaches revealed similar proportions of β 1- and β 2-adrenoceptors. The regional distribution of plasma membrane β 1- and β 2-adrenoceptors in rat hearts were also determined using tissue segment binding. Abundance of β-adrenoceptors and proportion of β 1-adrenoceptors were higher in atria than in ventricle, but there was no significant difference between right and left atria or within ventricle (right and left ventricle free walls, apex, and interventricular septum). To establish the ability of the tissue segment binding method to study β-adrenoceptor regulation such as the internalization of receptors, the effect of prolonged exposure of rat ventricle to (−)-isoprenaline was also investigated by using tissue segments and homogenate binding. Incubation with (−)-isoprenaline for 1 h in vitro caused a concentration-dependent decrease in the density of β-adrenoceptors, predominantly β 2-adrenoceptors, when assessed with tissue segment binding method. In contrast, the subtype-specific change after treatment with (−)-isoprenaline was not detected using homogenate binding. In summary, the tissue segment binding method with [ 3H]-CGP12177 enables a more precise quantitation of plasma membrane β 1- and β 2-adrenoceptors in rat hearts and is suitable for studying their regulation.

Colorectal cancer metastases affect the biochemical characteristics of the human liver β-adrenoceptor-G-protein-adenylate cyclase system

The sympathetic-catecholamine system is involved in the regulation of hepatic metabolic pathways mainly through cAMP-linked β 2-adrenoceptors (β 2-ARs) in humans and to a lesser extent through cAMP-independent mechanisms, but no information is available about the possible biochemical changes of β 2-ARs and their signalling pathways in human colorectal cancer (CRC) and colorectal cancer hepatic metastases (CRCHM). Changes in density and distribution of β-ARs as well as in post-receptor signalling components were studied in membranes of human liver with CRCHM, and for comparison, in membranes of nonadjacent, non-metastatic human liver (NA-NM) obtained from 13 patients, using binding and competition binding studies. Studies were also carried out using normal and cancerous human colon tissues. In CRCHM, the density of β-ARs ( B max) was significantly reduced, compared to NA-NM liver tissues (40.09 ± 2.83 vs. 23.09 ± 3.24 fmol/mg protein; P < 0.001). A similar decrease in the β-AR density was observed in the colon with primary colorectal cancer compared to healthy colon (37.6 ± 2.2 vs. 23.8 ± 3.5 fmol/mg protein), whereas the affinity of ICYP binding to the receptor remained unaffected. Desensitized β-ARs were uncoupled from stimulatory G-protein (G S), as total density of β-adrenoceptors in the high affinity state was significantly reduced. Concomitantly, CRCHM elicited decrease in the catalytic adenylate cyclase (AC) activity (cAMP formation) in response to isoproterenol plus GTP or forskolin or NaF. In NA-NM and CRCHM liver, the inhibition–concentration curves of ICI 118.551 showed the presence of a homogeneous population of the β 2-AR subtypes. Neither the binding patterns nor the inhibition constant ( K i) of ICI 118.551 were altered in CRCHM. In CRCHM, the hepatic β-AR-G-protein(s)-AC signalling system was markedly impaired, thus, these changes may well influence β-AR-mediated functions in both organs.

Positron Emission Tomography Studies of Human Airways Using an Inhaled β-Adrenoceptor Antagonist, S-11 C-CGP 12388

Positron emission tomography (PET) scanning may provide information on changes in the density and affinity of airway beta-adrenoceptors in lung diseases. However, the injection of a radiolabeled beta-blocker results in a pulmonary PET signal that reflects the binding of the ligand in the alveoli and not in the airways. Better discrimination between alveolar and airway beta-adrenoceptors may be possible with an inhaled radioligand. A nebulizer was used to administer the antagonist S-11C-CGP12388 in aerosol form. Eight volunteers inhaled the tracer twice, at baseline and after pretreatment with a beta-adrenergic drug. In both PET scan studies, a dynamic scan of the lungs was followed by a whole-body scan to assess the inhaled dose. Pulmonary uptake was quantified using a region-of-interest-based analysis. University hospital. Healthy volunteers. Pretreatment consisted either of inhaled salbutamol (400 microg, 20 min before the scan), or orally administered pindolol (3 x 5 mg during a period of 16 h before PET scanning). Drug pretreatment did not affect pulmonary deposition of the radioligand. The agonist salbutamol accelerated the monoexponential washout of 11C not only in the peripheral lung (mainly alveoli), but also in the central lung (mainly airways) and in the main bronchi. An even larger increase of the washout rate was induced by the antagonist pindolol. The similar effects of pindolol and salbutamol on tracer kinetics suggest that accelerated washout is due to the blockade of beta-adrenoceptors. Thus, the interaction of drugs with airway beta-adrenoceptors can be visualized using PET scanning and an inhaled radioligand.