Inhibiting mitochondrial Na+/Ca2+ exchange prevents sudden death in a Guinea pig model of heart failure

Abstract

Rationale: In cardiomyocytes from failing hearts, insufficient mitochondrial Ca(2+) accumulation secondary to cytoplasmic Na(+) overload decreases NAD(P)H/NAD(P)(+) redox potential and increases oxidative stress when workload increases. These effects are abolished by enhancing mitochondrial Ca(2+) with acute treatment with CGP-37157 (CGP), an inhibitor of the mitochondrial Na(+)/Ca(2+) exchanger.

Objective: Our aim was to determine whether chronic CGP treatment mitigates contractile dysfunction and arrhythmias in an animal model of heart failure (HF) and sudden cardiac death (SCD).

Methods and results: Here, we describe a novel guinea pig HF/SCD model using aortic constriction combined with daily β-adrenergic receptor stimulation (ACi) and show that chronic CGP treatment (ACi plus CGP) attenuates cardiac hypertrophic remodeling, pulmonary edema, and interstitial fibrosis and prevents cardiac dysfunction and SCD. In the ACi group 4 weeks after pressure overload, fractional shortening and the rate of left ventricular pressure development decreased by 36% and 32%, respectively, compared with sham-operated controls; in contrast, cardiac function was completely preserved in the ACi plus CGP group. CGP treatment also significantly reduced the incidence of premature ventricular beats and prevented fatal episodes of ventricular fibrillation, but did not prevent QT prolongation. Without CGP treatment, mortality was 61% in the ACi group <4 weeks of aortic constriction, whereas the death rate in the ACi plus CGP group was not different from sham-operated animals.

Conclusions: The findings demonstrate the critical role played by altered mitochondrial Ca(2+) dynamics in the development of HF and HF-associated SCD; moreover, they reveal a novel strategy for treating SCD and cardiac decompensation in HF.

Keywords: calcium; energy metabolism; heart failure; mitochondria; oxidative stress; reactiveoxygen species; sudden cardiac death.

Figure 1. The effects of CGP-37157 on [Ca²⁺]_m dynamics, NAD(P)H oxidation, and ROS production in myocytes isolated from ACi hearts

A) Representative recording of MityCam fluorescence (1-F/F0) in cardiomyocytes isolated from SHAMi (black) and ACi with (light gray) or without (gray) the presence of 1μmol/L CGP-37157 (CGP) in the perfusate. B) Mean peak MityCam fluorescence (1-F/F0) during 1Hz stimulation in SHAMi (n=30), ACi (n=29), and ACi + CGP (n=13). *, p<0.001 as compared to SHAMi; **, p<0.001 as compared to ACi. In the presence of 100nmol/L isoproterenol, NAD(P)H autofluorescence (A) and CM-DCF oxidation (B) were recorded in myocytes isolated from SHAMi hearts (n=3) or ACi hearts, with (n=3) or without (n=3) 1μmol/L CGP-37157 (CGP) in the perfusate. Box indicates duration of 4Hz field-stimulation.

Figure 2. The effects of CGP-37157 treatment on hypertrophic remodeling and cardiac function

A) Heart weight, HW (left panel), and Lung weight, LW (right panel), were measured at 4 weeks after surgery and normalized to tibia length (TL). Isoproterenol challenge had no effects on either HW/TL or LW/TL in sham-operated animals (SHAMi vs. SHAM). Pressure overload with daily isoproterenol challenge (ACi) induced cardiac hypertrophy and lung congestion, and CGP-37157 treatment attenuated both cardiac hypertrophy and lung congestion (ACi+CGP). B) Interstitial fibrosis was analyzed with Masson’s Trichrome staining. Left panel: representative images from each experimental group; right panel: measurements of interstitial fibrous tissue showing that pressure overload, together with isoproterenol challenge, induced significant fibrosis in ACi group, which was attenuated by CGP-37157 treatment (ACi+CGP). C) Cardiac function is preserved by CGP-37157. Upper panel: representative pictures of echocardiographic study; lower panel: measurements of fractional shortening (FS) (left) and diastolic LV internal dimension (LVIDd) (right) showing reduced FS and LV dilation in ACi group but not in CGP-37157 treated group. D) Left panel: representative LV pressure-volume loops. Heart from ACi group shows increased end-diastolic volume and depressed end-systolic pressure-volume relation. Right panel: summary data of end-systolic LV pressure (upper panel) and dP/dt_ip (dP/dt normalized to pressure) (lower panel). Isoproterenol treatment in the absence of aortic constriction (SHAMi) had no effect on cardiac function.

Figure 3. The antiarrhythmic effect of CGP-37157

A and B) Left panels: Representative telemetric ECG recordings at week 1 (A) and week 4 (B) after aortic constriction showing baseline records before isoproterenol injection, records after isoproterenol injection (post-ISO), and during recovery 4 to 5 hours after isoproterenol injection. Black arrows indicate PVBs; white arrows indicate VF. Right panels: PVB incidence at baseline and after isoproterenol injection in week 1 and week 4. *, p<0.05 compared to SHAMi group, †, p<0.05 ACi+CGP compared to ACi group. C) Left panels: representative ECG recordings at baseline for each group at weeks 1 and 4; right panels: summary measurements of QTc at week 1 and week 4 showing that long QT was induced by pressure overload. Neither CGP-37157 treatment nor isoproterenol challenge had any effect on QTc.

Figure 4. Survival curves over 4 weeks after the time of surgery for animals in SHAM, SHAMi, ACi, and ACi+CGP groups

Survival of animals with pressure overload plus isoproterenol challenge was significantly increased by treatment with CGP-37157.

Figure 5. Effects of acute CGP-37157 treatment on isolated perfused failing hearts

A) Representative traces of LV pressure and ECG simultaneously recorded before (pre-ISO), during (ISO), and after (post-ISO) isoproterenol (10μM) exposure in hearts of shami (n=3), ACi (n=3), and ACi with CGP-37157 (1μM) treatment (ACi+CGP) (n=3). B) Representative Poincaré plots of SHAMi, ACi, and ACi+CGP hearts during pre-ISO, ISO, and post-ISO states, respectively. Ectopic activity is evident as clustered of points only in the ACi heart post-ISO (middle right panel).