Investigating the Effects of a Cardiotoxic Drug on Calcium Homeostasis in the Heart

Abstract

Contractile dysfunction and arrhythmogenic activity can arise if there are perturbations to calcium (Ca) homeostasis in the heart, such as in response to cardiotoxic substances. This investigation focuses on changes in cellular Ca homeostasis in response to the acute application of the cardiotoxic drug Clozapine (an atypical anti-psychotic).Rat ventricular myocytes were isolated and stimulated under voltage clamp control. Various parameters of Ca handling were measured both in control conditions and in the presence of 10 μM clozapine. Ca-sensitive indicators fluo3-AM or fura-2 were used to quantify cytosolic Ca. Sarcoplasmic reticulum (SR) Ca content was measured by applying 10 mM caffeine. Activity of Ca extrusion mechanisms (SERCA - sarco-endoplasmic reticulum ATP-ase, NCX - sodium-calcium exchanger) were calculated from Ca transient decay rates. The current-voltage (I-V) relationship of the L-type Ca channel was also measured.Acute application of clozapine significantly reduced mean Ca transient amplitude by 48% and L-type Ca current (ICaL) density by 50%. SR content was decreased by 18 % and SERCA activity (rate constant) was reduced by 34 %. NCX rate constant was reduced by 17%. The effect of clozapine on SERCA was also present in phospholamban knockout myocytes suggesting that the effect on SERCA is not dependent on phospholamban.It remains unclear if or how the observed effects on Ca handling contribute to the cardiotoxicity of clozapine observed in the clinical setting (arrhythmogenesis and myocarditis). We speculate that effects on ICaL may predispose to refractory heterogeneity and reduced SERCA activity may increase the chance for Ca wave propagation. Current work focuses on effects of clozapine on calcium spark activity and SR threshold for Ca waves.