Effects of carvedilol on RyR2-mediated spontaneous calcium oscillation

AIM：To investigate the effects of carvedilol and other β-blockers on ryanodine receptor 2(RyR2)-mediated spontaneous calcium oscillation.
METHODS：HEK293 cells, which steadily and inducibly expressed RyR2, were generated using the Flp-In T-REx Core Kit. Single rat ventricular myocyte was isolated by the method of collagenase. The extracellular Ca2+ concentration was increased step by step to induce calcium oscillation. Single cell calcium imaging was used to test the effects of carvedilol and other β-blockers on RyR2-mediated spontaneous cal-cium oscillation in cardiac cells and HEK293 cells. RESULTS：Carvedilol at concentration of 30 μmol/L obviously suppressed the spontaneous cal-cium oscillation in cardiac cells and HEK293 cells expressing RyR2. The inhibitory rate was (6530±230)% and (6908±530)%, respectively. Metoprolol and other β-blockers had no effects on spontaneous calcium oscillation in cardiac cells and HEK293 cells. CONCLUSION：Carvedilol is the only β-blocker that suppresses RyR2-mediated spontaneous calcium oscillation. This may be one of the mechanisms that carvedilol is better than other β-blockers in reducing the mortality of heart failure.     

Effects of carvedilol,cilazapril and their combination on left ventricular remodeling after acute myocardial infarction in rats

AIM:To compare the effects of carvedilol, cilazapril and their combination on left ventricular remodeling(LVRM) after acute myocardial infarction(AMI) in rats. METHODS: Twenty-four hours after AMI operation, 100 surviving rats were randomly assigned to: ①AMI control(n= 25), ②AMI+carvedilol(1 mg·kg^-1 ·d^-1, n= 25)(C1), ③AMI+cilazapril(1 mg·kg^-1 ·d^-1, n= 25)(Z1), and ④ AMI+combination(n= 25) groups. Sham-operated group(n= 17) were selected randomly. After 4 weeks of therapy with the drugs gastric gavage, hemodynamic and pathological studies were performed. RESULTS: There were no significant differences in MI size among the four AMI groups(all P> 0.05) Left ventricular(LV) end diastolic pressure(LVEDP), volume(LVV), weight(LVW) and septal thickness(STh) were all higher and left ventricular pressure maximal rate of rise and fall(±d p /d t) were lower(all P< 0.01) in AMI group than sham-operated group. The LVEDP, LVV, LVW and STh were all lower and ±dp /dt were higher in Z1, C1, and combination groups than those in AMI group(P< 0.05, P< 0.01), with LVEDP and STh were more lower in the combination group than in the two monotherapy group(P< 0.05, P< 0.01), but there were no significant differences in other variables among the three therapy groups. CONCLUSION: Carvedilol, cilazapril and their combination all can prevent from LVRM after AMI in rats, improve hemodynamics and LV function, with the combination superior.     

Effect of carvedilol on store overload-induced Ca2+ release in rat pacing cardiomyocytes

AIM：To explore the effect of carvedilol on store overload-induced Ca2+ release (SOICR) in pacing cardiomyocytes. METHODS：Single rat cardiomyocyte with rapid pacing was perfused with isoprenaline and caffeine to induce calcium overload. Spontaneous calcium releases through sarcoplamic reticulum calcium release channel (ryanodine receptor 2, RyR2) were investigated by the method of fluorescence imaging. The cardiomyocytes were divided into control (DMSO) group, carvedilol group, metoprolol group, phentolamine group and nifedipine group. RESULTS：In control group, the incidence of SOICR in cardiomyocytes was significantly increased under the condition of calcium overload by perfusing with isoprenaline and caffeine in addition to the enhancement of calcium transient. The incidence of SOICR in carvedilol group was significantly lower than that in control group at the pacing frequency of 1 Hz to 4 Hz (2.00%, 6.00%, 10.00% and 16.00% vs 43.59%, 74.36%, 87.18% and 89.71%, respectively, P<0.01). The inhibitory effect of carvedilol was not significantly different at variant pacing frequency (P>0.05). The incidences of SOICR in metoprolol group, phentolamine group and nifedipine group had no significant difference compared with control group (P>0.05). The amplitude of calcium transient and caffeine peaking value of pacing cardiomyocytes had no significant difference among different groups (P>0.05). CONCLUSION：Carvedilol effectively suppresses SOICR in pacing cardiomyocytes due to its direct inhibition on the spontaneous opening of the cardiac RyR2 channel rather than the α1, β1 receptor or L-type calcium channel blockade.     

Effects of carvedilol on metabolic mechanism of endogenous NOS inhibitor in human umbilical vein endothelial cells treated with oxidized free radical

AIM: To observe the effects of oxidized free radical (OFR) on dimethylarginine dimethylaminohydrolase (DDAH) activity and concentration in human umbilical vein endothelial cells (HUVECs), and to investigate the metabolic mechanism of endogenous NOS inhibitor and the role of carvedilol. METHODS: HUVECs of 3-6th passage, cultured with modified Jaffes method, were divided into three groups: (1) cells cultured with equivalent DMEM medium as control; (2) OFR intervention groups, 0.01 mmol/L, or 0.1 mmol/L OFR was added respectively; (3) drug intervention groups: 0.1 mmol/L OFR plus 10 μmol/L carvedilol. ADMA, nitric oxide (NO), endothim (ET), L-citrulline concentrations and the activity of NOS in conditioned medium were measured after 24 h exposure. ADMA concentration in the conditioned medium was determined by high-performance liquid chromatography. Western blotting was performed to evaluate DDAH expression. RESULTS: Compared with control group, ADMA and ET concentrations were increased, while the level of NO and the activity of NOS decreased and relevant to the concentrations of OFR. We assayed DDAH activity by determining L-citrulline formation from ADMA. The concentration of L-citrulline was decreased, while the DDAH expression had no obvious change. With the role of carvedilol, ADMA, ET concentrations were decreased, while the level of NO, L-citrulline and the activity of NOS increased. CONCLUSION: Endothelial dysfunction induced by OFR is associated with the increase in ADMA concentration and reduction of DDAH activity, but not DDAH expression. Carvedilol promotes the degradation of ADMA through increasing activity of DDAH and improving endothelium function.     

Pharmacodynamics of carvedilol in conscious, healthy dogs

The purpose of the study reported here was to determine the magnitude and duration of beta-blocking efficacy, determine an effective dose and dosing interval, and document safety and tolerability of carvedilol given orally in clinically normal dogs. Pharmacodynamic data were evaluated in conscious, unrestrained, healthy hound dogs at baseline and after long-term oral administration of carvedilol (1.5 mg/kg of body weight PO q12h for >5 days). At baseline, heart rate (HR) and blood pressure (BP) data were collected continuously for 24 hours, and complete echocardiography was performed. This protocol was repeated after long-term oral carvedilol administration. Additionally, isoproterenol was administered to evaluate the magnitude and duration of the nonselective beta-blocking efficacy of carvedilol. An isoproterenol challenge was performed 0.75, 1.5, 2.25, 4, 6, 12, and 24 hours after carvedilol administration, with echocardiography being performed once at 2 hours. Plasma samples were obtained prior to each challenge time point for determination of plasma carvedilol concentration. Time series regression analysis indicated no difference between baseline and carvedilol-induced HR or BP trend lines in 6 of 8 dogs. In 2 of 8 dogs, HR, after long-term carvedilol administration, was reduced. Carvedilol attenuated isoproterenol-induced changes in HR by 54-76% through 12 hours and by 30% at 24 hours. The BP changes were attenuated by 80-100% through 12 hours. These data suggest that carvedilol (1.5 mg/kg PO q12h) in healthy, conscious dogs confers nonselective beta blockade for 12 hours, with minimal effects on resting HR, BP, and echocardiographic variables. Additionally, the magnitude of beta blockade correlated strongly to peak plasma carvedilol concentration, suggesting that therapeutic drug monitoring may be clinically useful.