Interaction of background Ca²⁺ influx, sarcoplasmic reticulum threshold and heart failure in determining propensity for Ca²⁺ waves in sheep heart

Ventricular arrhythmias can cause death in heart failure (HF). A trigger is the occurrence of Ca²⁺ waves which activate a Na⁺ -Ca²⁺ exchange (NCX) current, leading to delayed after-depolarisations and triggered action potentials. Waves arise when sarcoplasmic reticulum (SR) Ca²⁺ content reaches a threshold and are commonly induced experimentally by raising external Ca²⁺ , although the mechanism by which this causes waves is unclear and was the focus of this study. Intracellular Ca²⁺ was measured in voltage-clamped ventricular myocytes from both control sheep and those subjected to rapid pacing to produce HF. Threshold SR Ca²⁺ content was determined by applying caffeine (10  mM) following a wave and integrating wave and caffeine-induced NCX currents. Raising external Ca²⁺ induced waves in a greater proportion of HF cells than control. The associated increase of SR Ca²⁺ content was smaller in HF due to a lower threshold. Raising external Ca²⁺ had no effect on total influx via the L-type Ca²⁺ current, I_Ca-L , and increased efflux on NCX. Analysis of sarcolemmal fluxes revealed substantial background Ca²⁺ entry which sustains Ca²⁺ efflux during waves in the steady state. Wave frequency and background Ca²⁺ entry were decreased by Gd³⁺ or the TRPC6 inhibitor BI 749327. These agents also blocked Mn²⁺ entry. Inhibiting connexin hemi-channels, TRPC1/4/5, L-type channels or NCX had no effect on background entry. In conclusion, raising external Ca²⁺ induces waves via a background Ca²⁺ influx through TRPC6 channels. The greater propensity to waves in HF results from increased background entry and decreased threshold SR content. KEY POINTS: Heart failure is a pro-arrhythmic state and arrhythmias are a major cause of death. At the cellular level, Ca²⁺ waves resulting in delayed after-depolarisations are a key trigger of arrhythmias. Ca²⁺ waves arise when the sarcoplasmic reticulum (SR) becomes overloaded with Ca²⁺ . We investigate the mechanism by which raising external Ca²⁺ causes waves, and how this is modified in heart failure. We demonstrate that a novel sarcolemmal background Ca²⁺ influx via the TRPC6 channel is responsible for SR Ca²⁺ overload and Ca²⁺ waves. The increased propensity for Ca²⁺ waves in heart failure results from an increase of background influx, and a lower threshold SR content. The results of the present study highlight a novel mechanism by which Ca²⁺ waves may arise in heart failure, providing a basis for future work and novel therapeutic targets.