Blebbistatin Effectively Uncouples the Excitation-Contraction Process in Zebrafish Embryonic Heart

Abstract

Background/Aims: The zebrafish is an emerging model system for the study of cardiac electrophysiology and human arrhythmias. High resolution imaging techniques are powerful tools for the study of zebrafish cardiac electrophysiology, but these methods require the complete absence of cardiac contraction. Many pharmacological agents that uncouple cardiac contraction also markedly alter the cardiac action potential (AP). In this study, we compared the effects two uncoupling agents, 2,3-Butanedione monoxime (BDM) and blebbistatin, on contractility and AP parameters in embryonic zebrafish heart. Methods: Zebrafish hearts were explanted (48 hpf) and superfused with either BDM (15 mM) or blebbistatin (1, 5 or 10 µM), while recording atrial or ventricular APs with the disrupted patch technique. Calcium transients were recorded with a high-speed confocal scanning microscope in hearts loaded intracellularly with 10 µM fluo-4 and superfused with 10 µM blebbistatin. Results: Despite abolishing cardiac contractility, BDM altered ventricular AP morphology and inhibited spontaneous APs. In contrast, blebbistatin (10 µM) abolished contractility without significantly altering AP morphology or generation of spontaneous APs. Blebbistatin allowed for high fidelity measurements of atrial and ventricular calcium transients. Conclusion: Blebbistatin is a potent and effective excitation-contraction uncoupling agent in embryonic zebrafish heart.