Ryanodine receptor 2 inhibition improves cardiac repolarization reserve and contractile function and prevents sudden arrhythmic death in failing hearts

Reviewer #1 (Public Review):

The current study tests the hypothesis that inhibition of ryanodine receptor 2 (RyR2) in failing arrhythmogenic hearts reduces sarcoplasmic Ca leak, ventricular arrhythmias and improves contractile function. A guinea pig model of nonischemic heart failure (HF) was used and randomized to receive dantrolene (DS) or placebo in early or chronic HF. The authors show that DS treatment prevented ventricular arrhythmias and sudden cardiac death by decreasing dispersion of repolarization. The authors conclude that inhibition of RyR2 hyperactivity with DS mitigates the vicious cycle of sarcoplasmic Ca leak-induced increases in diastolic Ca and reactive oxygen species-mediated RyR2 oxidation. Moreover, the consequent increase in sarcoplasmic Ca2+ load improves contractile function.

In general, the study is well designed and the findings are likely to be of interest to the field. The characterization of the phenotypes is comprehensive, however, the study appears relatively weak in terms of the proposed mechanisms. Only in vivo functional analyses were presented with no in vitro analyses. The rationale for only using the male animals remains unclear. Data presented in Supplemental Figure 1 lacks the HF with DS group. As presented, the manuscript appears relatively descriptive in nature.

Reviewer #2 (Public Review):

Joshi et al. investigated the use of dantrolene, an RyR stabilizing drug, in improving contractile function and slowing pathological progression of pressure-overload heart failure. In a guinea pig model, they found that dantrolene treatment reduced cytosolic Ca2+ levels, improved contractility, reduced the incidence of arrhythmias, reduced fibrosis, and slowed the progression of heart failure. Importantly, delaying treatment until 3 weeks after aortic banding (when heart failure was already established) also resulted in improvements in function and decreased arrhythmogenesis. While some of the mechanistic details remain to be worked out, the data suggest that improving intracellular Ca2+ handling can break the vicious cycle of sympathetic activation, ROS production, and further deterioration of cardiac function.

The functional ECG and echo data are convincing, and very clearly demonstrate the positive effects of dantrolene in heart failure. This is important because dantrolene is already FDA-approved to treat malignant hyperthermia and muscle spasms, so repurposing this drug as a heart failure therapeutic might have a straightforward path to clinical implementation. This also highlights the non-specific nature of dantrolene to interact with RyR1, and therefore, potential side effects. However, this does not detract from the main proof-of-concept demonstrated here.

The guinea pig model employed here is also a strength, as the guinea pig has intracellular Ca2+ handling and ionic currents that are much more similar to human (vs. a murine model, for example).

One weakness is the exclusion of female animals from the study. The authors report more heterogeneity in the progression of HF in the female guinea pig model, however it will be very important to determine effects of dantrolene in the female heart, as there are considerable known sex differences in intracellular Ca2+ handling and contractility. Therefore, it is possible that dantrolene could have sex-dependent effects.

The title and parts of the discussion of the manuscript focus on 'repolarization reserve'. This term is often used in the realm of safety pharmacology, and 'reserve' refers to the fact that blocking a single K+ channel (for example) may not impact action potential duration because there may be enough other K+ currents to ensure proper repolarization. The repolarization reserve refers to this overall balance of depolarizing and repolarizing currents and potential redundancies to ensure proper repolarization. Although the present study clearly demonstrates QT shortening with dantrolene (thus, there must be a change in the balance of depolarizing and repolarizing currents), the study does not definitively demonstrate changes in any membrane currents. While this may seem like a minor point of terminology, it may mislead readers as to the main focus of the study, which is not at all on ionic currents, but on functional outcomes.