Beta-adrenoceptor Density Research Articles

Effects of Enalapril and L-158809 on Myocardial Beta-Adrenergic Receptor Density in Transplanted Rat Hearts

Background: We compare the effects of angiotensin-converting enzyme (ACE) inhibitor or angiotensin II type 1 (AT₁) receptor blocker on density of myocardial beta-adrenergic receptors (beta-ARs) in a heterotopic heart transplantation model. Methods: Hearts of F344 rats were heterotopically transplanted into Lewis rat recipients immunosuppressed with cyclosporine (10 mg/kg/day). Recipients were treated orally with the AT₁ receptor blocker L-158809 (3 mg/kg/day, n = 6), enalapril (3 mg/kg/day, n = 6), or vehicle only (n = 6) for 90 days. Density of myocordial beta-ARs was determined with an autoradiographic technique using [³H]CGP-12177. Results: Graft status, the sum of the functional score and the score for color, was preserved better in the L-158809-treated group (5.8 ± 0.9) and in the enalapril-treated group (5.6 ± 0.8) than in the vehicle-treated group (3.8 ± 0.9, p < 0.05). The grades of graft coronary artery disease in the L-158809-treated group and in the enalapril-treated group were significantly less than that seen in the vehicle-treated group. The density of myocardial beta-AR (fmol/mg of protein) was 3.5 ± 0.5 in the L-158809-treated group (p < 0.05 vs. vehicle-treated group) and 3.2 ± 0.5 (p < 0.05) in the enalapril-treated group but was 2.2 ± 0.4 in the vehicle-treated group. Conclusion: L-158809 is as effective as enalapril in restoring myocardial beta-AR density in immunosuppressed rat transplant model, and this efficacy, as well as the prevention of graft coronary arteriosclerosis, is probably associated with the preservation of graft status.

Bucindolol, a nonselective beta 1- and beta 2-adrenergic receptor antagonist, decreases beta-adrenergic receptor density in cultured embryonic chick cardiac myocyte membranes.

Bucindolol and carvedilol, nonselective beta1- and beta2-adrenergic receptor antagonists, have been widely used in clinical therapeutic trials of congestive heart failure. The aim of the current study was to investigate long-term effects of bucindolol or carvedilol on beta-adrenergic receptor protein and gene expression in cardiac myocytes. Embryonic chick cardiac myocytes were cultured and incubated with bucindolol (1 microM), carvedilol (1 microM), or norepinephrine (1 microM) for 24 h. 125I-iodocyanopindolol binding assays demonstrated that incubation with norepinephrine or bucindolol, but not carvedilol, significantly decreased beta-adrenergic receptor density in crude membranes prepared from the myocytes. Neither bucindolol nor carvedilol significantly stimulated adenylyl cyclase activity in membranes from drug-untreated cells. Unlike by norepinephrine, the receptor density reduction by bucindolol incubation was not accompanied by a change in beta1-adrenergic receptor messenger RNA abundance. A decrease in membrane beta-adrenergic receptor density without a change in cognate messenger RNA abundance was also observed in hamster DDT1 MF2 cell line incubated with bucindolol (1 microM, 24 h). We conclude that incubation with bucindolol, but not carvedilol, results in true reduction of beta-adrenergic receptor density in chick cardiac myocyte membranes by mechanisms that are distinct from those responsible for receptor density reduction by the agonist norepinephrine.

Inotropic response to beta-adrenergic receptor stimulation and anti-adrenergic effect of ACh in endothelial NO synthase-deficient mouse hearts.

1. The functional consequences of a lack of endothelial nitric oxide synthase (eNOS) on left ventricular force development and the anti-adrenergic effect of acetylcholine (ACh) were investigated in isolated hearts and cardiomyocytes from wild type (WT) and eNOS knockout (eNOS-/-) mice. 2.eNOS expression in cardiac myocytes accounted for 20 % of total cardiac eNOS (Western blot analysis). These results were confirmed by RT-PCR analysis. 3. In the unstimulated perfused heart, the left ventricular pressure (LVP) and maximal rate of left ventricular force development (dP/dtmax) of eNOS-/- hearts were not significantly different from those of WT hearts (LVP: 97 +/- 11 mmHg WT vs. 111 +/- 11 mmHg eNOS-/-; dP/dtmax: 3700 +/- 712 mmHg s(-1) WT vs. 4493 +/- 320 mmHg s)-1) eNOS-/-). 4. The dobutamine (10-300 nM)-induced increase in LVP was enhanced in eNOS-/- hearts. In contrast, L-type Ca2+ currents (ICa,L) in isolated cardiomyocytes of WT and eNOS-/- hearts showed no differences after beta-adrenergic stimulation. Dibutyryl-cGMP (50 microM) reduced basal ICa,L in WT cells to 72 +/- 12 % while eNOS-/- ICa,L was insensitive to the drug. The pre-stimulated ICa,L (30 nM isoproterenol) was attenuated by dibutyryl-cGMP in WT and eNOS-/- cells to the same extent. 5. The Ca2+ (1.5-4.5 mM)-induced increase in inotropy was not different between the two experimental groups and beta-adrenergic receptor density was increased by 50% in eNOS-/- hearts. 6. The contractile effects of dobutamine could be inhibited almost completely by ACh or adenosine. The extent of the anti-adrenergic effect of both compounds was identical in WT and eNOS-/- hearts. Measurement of ICa,L in isolated cardiac myocytes yielded similar results. 7. These data demonstrate that in the adult mouse (1) lack of eNOS is associated with increased cardiac contractile force in response to beta-adrenergic stimulation and with elevated -adrenergic receptor density, (2) the unaltered response of ICa,L in eNOS-/- cardiac myocytes to beta-adrenergic stimulation suggests that endothelium-derived NO is important in mediating the whole-organ effects and (3) eNOS is unimportant for the anti-adrenergic effect of ACh and adenosine.

Diabetes reduces right atrial β-adrenergic signaling but not agonist stimulation of heart rate in swine

This study assessed the effects of streptozotocin diabetes in swine on the heart rate response to beta-adrenergic stimulation the adenylyl cyclase signal transduction pathway. Diabetic animals (n = 9) were hyperglycemic compared to the control group (n = 10) (12.6 +/- 1.0 vs. 3.53 +/- 0.29 mM). There were no significant differences between the diabetic and nondiabetic groups in the heart rate response to isoproterenol, however, there was a significant reduction (14%) in beta-adrenergic receptor density in the right atrium in the diabetic (61 +/- 3 fmol/mg protein) versus the nondiabetic group (71 +/- 3) (P < 0.05). The content of guanosine triphosphate binding regulatory proteins (Gs and Gi) in the right atrium was not affected by diabetes, nor was adenylyl cyclase activity under unstimulated conditions or with receptor-dependent stimulation with isoproterenol. On the other hand, adenylyl cyclase activity was 34% lower when directly stimulated with forskolin, and it was reduced by 23% when stimulated through Gs with Gpp(NH)p. In conclusion, beta-adrenergic stimulation of heart rate with isoproteronol and the receptor-dependent signal transduction pathway remained intact in the right atrium of diabetic swine despite reduced beta-adrenergic receptor density, G-protein content, and direct stimulation of adenylyl cyclase activity.

The effect of streptozotocin-induced diabetes on cardiac beta-adrenoceptor subtypes in the rat.

1. The present study investigates the effect of short-term experimental diabetes of 14-days duration on the beta-adrenoceptor subtypes of the rat heart. 2 .beta-adrenoceptor-mediated functional responses to submaximal doses of isoprenaline were enhanced in Langendorff-perfused hearts from diabetic rats, manifested as greater changes in tension, heart rate and rates of tension development (+dT/dt) and decline (-dT/dt). 3. Radioligand binding data demonstrated that total cardiac beta-adrenoceptor density and affinity for [3H]-dihydroalprenolol was unchanged by diabetes, although a decrease in beta1-adrenoceptor density and increase in beta2-adrenoceptor density was observed. 4. In conclusion, hearts from 14-day streptozotocin-induced diabetic rats demonstrate a number of alterations within the beta-adrenoceptor system. However, the enhanced beta-adrenoceptor-mediated responses to isoprenaline were not caused by an overall increase in density of beta-adrenoceptors, but were accompanied by changes in the ratio of the beta-adrenoceptor subtypes.

Differential effects of carvedilol and metoprolol on isoprenaline-induced changes in β-adrenoceptor density and systolic function in rat cardiac myocytes

beta-Blockers improve cardiac function and survival in heart failure patients. The underlying mechanisms are not completely elucidated. Differences between agents might be important for the development of more specific therapeutical approaches. This study investigated whether metoprolol or carvedilol alter beta-adrenergic signaling differently. beta-Adrenoceptor density and systolic function were determined in rat adult ventricular cardiac myocytes. 12 h isoprenaline-treatment (Iso, 1 micromol/l) reduced beta-adrenoceptor density by 33% (P<0.01). The effect was abolished by incubation with isoprenaline plus metoprolol (3 micromol/l), but was more pronounced after coincubation with carvedilol (0.003 micromol/l, P<0.05 Carv vs. Iso). Metoprolol alone had no effect on beta-adrenoceptor density, but carvedilol induced a decrease in receptor density even in absence of isoprenaline (P<0.05 Carv vs. ctr.). The isoprenaline (0.0003-10 micromol/l) induced concentration-dependent increase in myocyte shortening was blunted after 12 h preincubation with Iso (1 micromol/l, P<0.001). This reduction was abolished or partly prevented by coincubation with metoprolol or carvedilol, respectively. Carvedilol decreased the number of receptors which had to be occupied by isoprenaline in order to obtain 50% and 90% increase in myocyte cell shortening. Comparison of guanine nucleotide-dependent binding characteristics of isoprenaline, carvedilol and metoprolol revealed beta-receptor agonist like binding characteristics for carvedilol, but antagonist like binding characteristics for metoprolol. Metoprolol but not carvedilol prevents isoprenaline-induced downregulation of myocyte beta-adrenoceptors. The difference might be due to specific binding properties of the beta-blockers. Restoration of isoprenaline responsiveness by carvedilol might be due to improved coupling of beta-receptors to postreceptor effects.

Pulmonary beta adrenoceptor density in arrhythmogenic right ventricular cardiomyopathy and idiopathic tachycardia.

In recent in vivo studies using positron emission tomography (PET) our group demonstrated that the myocardial beta adrenoceptor (betaAR) density is reduced in arrhythmogenic right ventricular cardiomyopathy (ARVC) and idiopathic right ventricular outflow tract tachycardia (RVO-VT) associated with an increased presynaptic catecholamine washout. It was hypothesised that the reduction of myocardial betaAR density is secondary to an increase of local catecholamines in the myocardium resulting from the presynaptic dysfunction since circulating plasma catecholamines were demonstrated to be unchanged in these conditions. To further prove this hypothesis of an organ-limited adrenergic nervous dysfunction of the heart, this study aimed to investigate betaAR density in another thoracic organ, the lung. Pulmonary and myocardial betaAR density was measured in 7 ARVC patients, 8 RVO-VT patients and in a group of healthy controls (n = 13) using the non-selective beta-blocker [11C]-CGP 12177 and PET. Pulmonary betaAR density was similar in controls (12.4 +/- 1.7 pmol/g tissue), ARVC (11.6 +/- 1.7 pmol/g tissue, p = ns) and RVO-VT (12.8 +/- 2.0 pmol/g tissue, p = ns), whereas myocardial betaAR density was significantly reduced in ARVC (6.3 +/- 1.1 pmol/g tissue, p = 0.006) and RVO-VT (6.8 +/- 1.2 pmol/g tissue, p=0.02) as compared to controls (8.8+/-1.5 pmol/g tissue). The unchanged pulmonary betaAR density in the presence of a previously described significant reduction in myocardial betaAR density in the same patient principally supports our pathophysiological hypothesis that the myocardial betaAR density may be reduced in ARVC and RVO-VT because of an increase in local synaptic catecholamine levels due to an organ-limited presynaptic adrenergic dysfunction of the heart. Since in the present study only pulmonary betaAR density was measured, future functional studies excluding pulmonary betaAR desensitisation are required to finally prove the unchanged pulmonary sympathetic innervation in ARVC and RVO-VT.

Antidepressant activity and calcium signaling cascades.

Although antidepressant treatments produce clear effects on monoaminergic neuronal function, the link between these effects and therapeutic response to treatment is controversial. Previous studies have demonstrated that antagonists of the NMDA receptor-gated calcium ionophore result in antidepressant-like responses in rodents and humans. Likewise, antidepressant treatments produce regionally selective adaptation of the NMDA receptor suggestive of diminished capacity to gate calcium into receptive neurons. Similarly, voltage-dependent calcium channel antagonists have been reported to produce antidepressant-like effects in rodents. A major target of increases in subcellular calcium concentration is nitric oxide synthase (NOS) which liberates NO in response to stimulation. Recently, we have demonstrated that nitric oxide synthase antagonists produced antidepressant-like response in both in vivo preclinical screening procedures and in post-mortem in vitro studies of beta-adrenoceptor density. We propose: 1) that interruption of the Ca(2+)-calmodulin-NOS-guanylyl cyclase subcellular signaling pathway at any point will produce antidepressant-like effects; 2) that the acute actions of antidepressants in preclinical screening procedures are a consequence of their ability to disrupt Ca(2+)-calmodulin-NOS-guanylyl cyclase signaling; 3) that chronic but, not acute treatment with antidepressants results in adaptation of the Ca(2+)-calmodulin-NOS-guanylyl cyclase signaling pathway; 4) that this adaptation is necessary for the achievement of the therapeutic actions of antidepressants and; 5) that major depression is accompanied by an alteration (hyperactivity?) of subcellular Ca(2+) signaling. Copyright 2001 John Wiley & Sons, Ltd.

Impairment of cardiac beta-adrenoceptor cellular signaling by decreased expression of G(s alpha) in septic rabbits.

Abnormalities in the beta-adrenergic control of cardiac function play a role in the pathogenesis of several disease states. Because circulatory failure in patients with septic shock is known to be less responsive to catecholamines, we investigated whether the beta-adrenoceptor-linked signal transduction mechanisms are altered in the heart of a septic animal model Rabbits were rendered endotoxemic by an intravenous injection of 100 microg/kg Escherichia coli lipopolysaccharide. Three and 6 h later, the myocardial tissues were used for the experiments. The positive inotropic response to isoproterenol was significantly impaired in papillary muscles isolated from septic rabbits compared with those from controls. The impaired inotropic responsiveness to isoproterenol was not prevented by the nitric oxide synthase inhibitor N(G)-nitro-L-arginine, indicating no involvement of nitric oxide overproduction. Adenylate cyclase activity stimulated with isoproterenol and 5'-guanylyl imidodiphosphate was markedly reduced in septic myocardium. The contractile and adenylate cyclase responses to colforsin daropate, a direct adenylate cyclase activator, were unaffected by sepsis. Radioligand binding experiments with (-)[125I]iodocyanopindolol revealed no significant alteration in myocardial beta-adrenoceptor density or affinity in sepsis. Determination of cardiac G(s alpha) level by Western blotting showed a reduction of approximately 50% in sepsis. The relative content of G(s alpha) messenger RNA in septic myocardium also was reduced from the control level by about 50%, as determined by Northern blot analysis. Little change was found in protein and messenger RNA levels of G(s alpha) in septic myocardium. Impairment of myocardial functional responsiveness to beta-adrenoceptor stimulation appears in the early stage of sepsis. The impaired response to beta-adrenoceptor stimulation in the heart in this pathologic state may result in part from a decreased level of G(s alpha) protein which occurs at the level of gene expression.

Effects of beta-blockers on neurohormonal activation in patients with congestive heart failure.

The effect of beta-adrenoceptor antagonists (beta-blockers) on neurohormonal activation in patients with congestive heart failure has been the subject of study in numerous small clinical trials. Short term therapy with beta-blockers is associated with a variable acute neurohormonal response which may be determined by the pharmacology of the agent under study and the baseline characteristics of the patient population. Long term therapy with beta-blockers devoid of intrinsic sympathomimetic activity (partial agonist activity) is associated with evidence of decreased plasma markers of activation of the sympathetic nervous system, the renin-angiotensin system, and endothelin-1. Beta1-selective and nonselective beta-blockers appear to be associated with evidence of decreased neurohormonal activation, with differential effects on beta-adrenoceptor density. Agents with partial agonist activity appear to differ from pure antagonists, with some studies reporting evidence of increased neurohormonal activation. The mechanisms by which beta-blockers reduce neurohormonal activation and the clinical relevance of changes in adrenergic function to their use in the treatment of heart failure require further investigation.

Relation of tissue Doppler derived myocardial velocities to myocardial structure and beta-adrenergic receptor density in humans

OBJECTIVESWe sought to evaluate the relation of segmental tissue Doppler (TD) velocities to both the regional amount of interstitial fibrosis and the myocyte beta-adrenergic receptor density in humans.BACKGROUNDThe systolic myocardial velocity (Sm) and early diastolic myocardial velocity (Em) acquired by TD are promising new indexes of left ventricular function. However, their structural and functional correlates in humans are still unknown.METHODSTen patients with coronary artery disease underwent echocardiographic examination including TD imaging, along with transmural endomyocardial biopsy at the time of coronary bypass surgery (two biopsies per patient for a total of 20 specimens). The specimens were analyzed for percent interstitial fibrosis and beta-adrenergic receptor density.RESULTSNormal segments (n = 8) had a higher beta-adrenoceptor density (2,280 ± 738 vs. 1,373 ± 460, p = 0.03) and a lower amount of interstitial fibrosis (13 ± 3.3% vs. 28 ± 11.5%, p = 0.002) than dysfunctional segments (n = 12). Myocardial systolic velocity and Em were also significantly higher (9.5 ± 2.7 vs. 5.9 ± 1.8 cm/s, p = 0.025 and 11.3 ± 2.8 vs. 6.4 ± 2.1 cm/s, p = 0.002, respectively) in normal segments. A significant relationship was present between Em and the beta-adrenergic receptor density (r = 0.78, p < 0.001) and percent interstitial fibrosis (r = −0.7, p = 0.0026), which together accounted for 81% of the variance observed in Em. Likewise, a significant relationship was present between Sm and the beta-adrenergic receptor density (r = 0.68, p < 0.001) and the percent interstitial fibrosis (r = −0.66, p = 0.004) and together accounted for 62% of the variance observed in Sm.CONCLUSIONSSystolic myocardial velocity and Em are strongly dependent on both the number of myocytes and the myocardial beta-adrenergic receptor density.

Beta-adrenergic desensitization after burn excision not affected by the use of epinephrine to limit blood loss.

Burn patients have impaired myocardial function and decreased beta-adrenergic responsiveness. Further beta-adrenergic dysfunction from systemic absorption of topically administered epinephrine that is given to limit blood loss during burn excision could affect perioperative management. The authors evaluated the effect of topical epinephrine administration to patients during burn excision on the lymphocytic beta-adrenergic response. Fifty-five patients (age, 2-18 yr) with 20-90% body surface area burns received a standardized anesthetic for a burn excision procedure. Lymphocyte samples were taken at baseline and 1 and 3 h after the initial use of epinephrine (n = 43) or thrombin (controls, n = 12). Plasma epinephrine levels were measured by high-performance liquid chromatography. Lymphocyte beta-adrenergic responsiveness was assessed by measuring production of cyclic adenosine monophosphate (cAMP) after stimulation with isoproterenol, prostaglandin E1 (PGE1), and forskolin. beta-adrenergic receptor binding assays using iodopindolol and CGP12177 yielded beta-adrenergic receptor density. Epinephrine levels were elevated at 1 h (P < 0.01) and 3 h (P < 0.01) after epinephrine use but not in control patients. Production of cAMP in lymphocytes 1 h after epinephrine was greater in patients receiving epinephrine than in control patients on stimulation with isoproterenol (P < 0.05) and PGE1 (P < 0.05). Three hours after epinephrine administration, production of cAMP decreased when compared with baseline in both control patients and those receiving epinephrine after stimulation with isoproterenol (P < 0. 05), PGE1(P < 0.05), and forskolin (P < 0.05). Lymphocytic beta-adrenergic receptor content was not changed. Topical epinephrine to limit blood loss during burn excision resulted in significant systemic absorption and increased plasma epinephrine levels. Acute sensitization of the lymphocytic beta-adrenergic cascade was induced by the administration of epinephrine reflected by increased cAMP production after stimulation with isoproterenol and PGE1. The lymphocytic beta-adrenergic cascade exhibited homologous and heterologous desensitization 3 h after the use of epinephrine or thrombin, indicating that epinephrine administration was not a causative factor.

Cardiac beta-adrenoceptors in chronic uremia: studies in humans and rats

OBJECTIVESThe purpose of this study was to elucidate whether cardiac beta-adrenergic effects may be blunted in patients on maintenance hemodialysis (HD) and may help to explain autonomic dysfunction.BACKGROUNDPatients on HD often suffer from autonomic dysfunction.METHODSWe investigated the cardiovascular response of five HD patients (age: 46.1 ± 7.9 years) and six healthy volunteers (age: 48.2 ± 7.5 years) to isoprenaline, pirenzepine and phenylephrine. For analysis of underlying mechanisms of beta-adrenoceptor hyporesponsiveness, six-week-old male Wistar rats were rendered uremic by 5/6-nephrectomy (n = 9; SNX) and were killed for removal of the heart after six to seven weeks. Sham-operated rats (n = 15) served as controls.RESULTSIn the patient study, isoprenaline (3.5, 7, 17, 35 ng/kg/min, i.v.) led to an increase in heart rate, and shortening of the heart rate corrected duration of the electromechanical systole (QS2c), both of which were significantly reduced in HD patients. Baroreflex sensitivity was significantly reduced in HD patients. The response to low parasympathomimetic doses of pirenzepine was unchanged. In the rat study, left ventricular strips were placed in an organ bath, electrically driven and exposed to isoprenaline (10−11 to 10−6 mol/liter). While pD2 values were unchanged, maximum effect at the highest concentration was significantly reduced in SNX rats. The response to carbachol was not altered, nor was the M2-cholinoceptor density. There was no difference in beta-adrenoceptor density, or in immunodetectable amount of Gs and Gi protein. Activation of adenylyl cyclase evoked by isoprenaline was significantly reduced in left ventricular membranes of SNX rats, whereas effects of 10 μmol/liter GTP, 10 mmol/liter NaF, 10 μmol/liter forskolin and 10 mmol/liter Mn2+ were not altered.CONCLUSIONSCardiac beta-adrenergic responses are blunted in chronic uremia due to reduced isoprenaline-dependent activation of adenylyl cyclase. This might be caused by an “uncoupling” of the receptor or by an inhibition of the receptor by uremic toxins.

Perioperative beta-blockers. Part one: fundamentals

To review the pharmacologic and pathophysiologic information necessary to prescribe beta-blockers (BB) in perioperative medicine. Manual retrieval and electronic research of the literature using MEDLINE (key-words: anesthesia and beta- blocker; surgery and beta-blocker). Cardioselective BB inhibit preferentially beta-1 receptors, inducing a decrease in heart rate and cardiac inotropism leading to reduction of oxygen myocardial consumption. Non-cardioselective BB inhibit also beta-2 receptors, increasing bronchial and peripheral vascular resistances and uterine contractions. However, some BB are also vasodilators (carvedilol, celiprolol, labetalol). Contraindications to BB result logically from their pharmacological effects. Treatment with BB increases membrane beta-receptor density; this explains sympathetic overactivity observed during weaning of treatment. Since the discovery of propranolol in 1964, the use of BB has been controversial in anesthesia. Formerly, the adverse effects of partial sympatholysis during anesthesia and surgery were feared. However, since 1973, experimental and clinical data have suggested a protective hemodynamic effect. Continued administration of BB up to the time of anesthesia has been encouraged except in patients with signs of intolerance such as hypotension or excessive bradycardia.

Prostaglandin deficiency promotes sensitization of adenylyl cyclase.

Inhibition of prostaglandin synthesis by the drug indomethacin suppresses the synthesis of the cyclic AMP antagonist, prostaglandylinositol cyclic phosphate (cyclic PIP), and leads to a metabolic state comparable to type II diabetes. It was of interest whether prostaglandin-deficiency likewise causes sensitization of adenylyl cyclase, as this has been reported for the diabetic state. In liver plasma membranes of indomethacin-treated male rats, basal and forskolin-stimulated cyclic AMP synthesis remained unchanged when compared to untreated control rats. In control rats, stimulation of cyclic AMP synthesis by fluoride (2.2-fold) or glucagon (3.5-fold) was much lower than stimulation by forskolin (6.6-fold). In contrast, in indomethacin-treated rats, stimulation of cAMP synthesis by fluoride (4.6-fold) or glucagon (5.2-fold) nearly matched the stimulation by forskolin (6.4-fold). The level of alpha1-adrenergic receptors was slightly reduced, from 450 to 320 fmol/mg protein, by the indomethacin treatment. Independent of the treatment by indomethacin, stimulation of cyclic AMP synthesis by adrenaline failed, in agreement with the low density of adrenergic beta-receptors. In conclusion, PGE deficiency sensitizes adenylyl cyclase in rat liver for G protein-coupled receptors (glucagon) and also for fluoride.

Rapid electrical stimulation of contraction reduces the density of beta-adrenergic receptors and responsiveness of cultured neonatal rat cardiomyocytes. Possible involvement of microtubule disassembly secondary to mechanical stress.

Although tachycardia is commonly present in patients with congestive heart failure, its role in the development of congestive heart failure remains unclear. We studied the effect of rapid electrical stimulation of contraction on beta-adrenergic receptor (beta-AR) signal pathway in cultured cardiomyocytes of neonatal rats. Contraction of cardiomyocytes was induced by electrical stimulation at 50 V with twice the threshold pulse width. beta-ARs were identified by [(3)H]CGP-12177 and [(3)H]dihydroalprenolol. Electrical stimulation reduced cell-surface but not total beta-AR density; the effect was dependent on pacing frequency (a reduction of 11%, 28%, and 18% in cells paced at 2.5, 3. 0, and 3.3 Hz, respectively). This reduction was apparent at 3 hours, in contrast to reduced beta-AR density after exposure to isoproterenol (ISP) for 1 hour. The fraction and inhibition constant of beta-AR binding agonist with high affinity were not affected by rapid electrical stimulation. In cardiomyocytes paced at 3.0 Hz for 24 hours, the response to ISP decreased compared with unpaced cells, 142% versus 204% of baseline with 1 micromol/L ISP, whereas the responses to forskolin or acetylcholine were not different. Treatment of cardiomyocytes with 2,3-butanedione monoxime (10 mmol/L) or taxol (10 micromol/L) inhibited the rapid pacing-induced reduction in beta-AR density. Our results suggest that contractile activity is involved in regulation of cardiac function by modulating the beta-AR system independently of hemodynamic and neurohormonal factors. This may help to elucidate the role of mechanical stress in the development of heart failure.

Properties of the ventricular adrenergic signal transduction system during ontogeny of spontaneous hypertension in rats.

The purpose of this study was to characterize adrenergic receptors and associated G proteins in ventricles of spontaneously hypertensive rats (SHRs) at different stages of development. The beta- and alpha1-adrenoceptor densities and subtype distribution, and beta-adrenoceptor-G protein coupling were studied by radioligand binding, and levels of G(Salpha), G(ialpha), and G(q/11alpha) protein species were determined by Western blotting in SHRs and Wistar-Kyoto (WKY) control rats aged 3.5 weeks, 3 months, and 8 months. In 3.5-week-old SHRs, both the beta-adrenoceptor density and the percentage of agonist high-affinity binding sites were higher than in age-matched WKY rats. The beta1/beta2-subtype distribution, the alpha1-adrenoceptor density, and the alpha1B/alpha1A-subtype distribution were similar in rats of both strains at all ages. Although essentially no differences in G(salpha) levels between SHRs and WKY rats were detected, higher G(ialpha) and lower Gq/1alpha concentrations were found in 3.5-week-old SHRs. In 3-month-old SHRs, increased levels of Gq/11alpha proteins were observed. In 8-month-old SHRs, none of the parameters was different from those of controls. We conclude that the differences in properties of the adrenergic signal transduction system between SHRs and WKY rats are exclusively observable before and at the onset of the overt hypertension. Moreover, the hypertensive genotype apparently affects G proteins more readily than adrenoceptors.

Relationship Between Myocardial Beta-Adrenergic Sensitivity and Heart Rate Variability

Background: In congestive heart failure, despite activation of the sympathetic nervous system, heart rate variability parameters reflecting sympathetic modulation on sinus node are decreased. Our goal was to assess the role of beta-adrenergic sensitivity in the modulation of heart rate variability (HRV) in patients with valvular heart diseases. Methods and results: Ten patients with aortic stenosis, 10 patients with heart failure, and 12 controls were included. Baroreflex sensitivity was calculated by the sequency method. Noradrenaline plasma levels were assayed by HPLC. HRV was studied using 24-hour Holter monitoring. Cardiac beta-adrenergic receptivity was assessed by the chronotropic response to dobutamine. Right auricular samples were obtained for determination of beta-adrenergic receptor density by binding study with [125I]-iodocynaopindolol, and beta 1 and beta 2 densities, measured by competition between 125ICP binding by isoprenaline. In multivariate analysis, the dose of dobutamine that increases basal heart rate for 25 beats/min (ED25) is correlated with a parameter of global HRV: SDNN (r = 0.6, P < 0.001) and with indexes reflecting rather sympathetic modulation of HRV: SDANN (r = 0.62, P < 0.001) or SD (r = 0.47, P < 0.0001). All these relations were independent from mean NN, spontaneous baroreflex sensitivity, and noradrenaline plasma levels. No significant correlation existed between dobutamine ED25 and HRV indexes reflecting parasympathic tone. No relationship existed between HRV and beta-adrenergic receptor-binding characteristics. Conclusion: Cardiac beta-adrenergic sensitivity explored by dobutamine ED25 is an important determinant of HRV independent from mean NN, spontaneous baroreflex sensitivity, and noradrenaline plasma levels.

Dietary Selenium and Vitamin E Intakes Alter β-Adrenergic Response of L-Type Ca-Current and β-Adrenoceptor-Adenylate Cyclase Coupling in Rat Heart

Previously we have shown that both insufficient (combined with vitamin E deficiency) and excess intake of selenium (Se) impairs isoproterenol (ISO)-induced contractions of rat papillary muscle. In the present study, we used patch-clamp and biochemical techniques to investigate mechanisms of this effect in rats fed a Se- and vitamin E-deficient, a Se-excess or a normal diet. Whole-cell configuration of patch-clamp technique was used to investigate L-type Ca(2+) currents (I(Ca,L)) and their regulation by beta-adrenergic receptor stimulation in enzymatically isolated single rat ventricular myocytes. Alteration of Se and vitamin E intake did not affect peak I(Ca,L), but the threshold potential of activation was significantly different among groups. Maximal I(Ca,L) responses to ISO were depressed in both experimental groups, but the EC(50) values were not affected. In the Se-deficient group, basal, ISO- or forskolin-induced adenylate cyclase (AC) activity, measured in cardiac membrane preparations, was reduced when compared to the control, whereas 5' guanylyimidodphosphate (GppNHp) stimulated activity was unaffected. Decreased beta-adrenoceptor density and reduced GppNHp-induced affinity shift in ISO binding were also observed in the deficient group. No such differences were present in the excess group. These results suggest that combined Se and vitamin E deficiency interferes with beta-adrenoceptor-AC coupling, whereas excess intake of Se does not affect it. Thus, in the deficient group, the impairment of I(Ca) responses to ISO may be a result of a defect in beta-adrenoceptor-AC pathway. Impairment of I(Ca) response in the excess group, however, appears to have a different underlying mechanism.

Regional pre- and postsynaptic sympathetic system in the failing human heart--regulation of beta ARK-1.

Regional presynaptic sympathetic innervation varies considerably in the cardiomyopathic human heart, as shown in previous studies in vivo and in vitro. The goal of the present study was to correlate markers of presynaptic sympathetic innervation with local measurement of the postsynaptic beta-adrenergic system in failing human hearts. In nine left ventricular regions of hearts explanted from patients suffering from dilated cardiomyopathy, we measured the density of uptake(1) carriers ([3H]mazindol binding) as a marker of presynaptic function as well as beta-receptor density ([3H]CGP 12177 binding) and beta ARK-1 levels as the pivotal compounds of postsynaptic adrenergic signal transduction. Additionally, a subgroup of the patients was examined in vivo by HED-PET prior to heart transplantation. The density of uptake(1) was related to local hydroxyephedrine (HED) retention (as determined by pre-operative PET, r=0.65), whereas it was inversely correlated to regional beta ARK-1 levels (r=-0.61, P=0.04). In contrast, beta-adrenergic receptor density was not significantly correlated either to uptake(1) density or to local HED retention (r=0.15 and r=0.21). Regional beta ARK-1 levels rather than beta-adrenergic receptor density were correlated with presynaptic alterations in cardiomyopathic human left ventricles. It can be assumed that in the cardiomyopathic human heart, regional beta-adrenergic desensitization might be determined by differences in local beta ARK levels rather than by changes in beta-receptor density.