Popeye domain-containing (POPDC) proteins selectively bind cAMP and mediate cellular responses to sympathetic nervous system (SNS) stimulation. The first discovered human genetic variant (POPDC1^S201F) is associated with atrioventricular (AV) block, which is exacerbated by increased SNS activity. Zebrafish carrying the homologous mutation (popdc1^S191F) display a similar phenotype to humans. To investigate the impact of POPDC1 dysfunction on cardiac electrophysiology and intracellular calcium handling, homozygous popdc1^S191F and popdc1 knock-out (popdc1^KO) zebrafish larvae and adult isolated popdc1^S191F hearts were studied by functional fluorescent analysis. It was found that in popdc1^S191F and popdc1^KO larvae, heart rate (HR), AV delay, action potential (AP) and calcium transient (CaT) upstroke speed, and AP duration were less than in wild-type larvae, whereas CaT duration was greater. SNS stress by β-adrenergic receptor stimulation with isoproterenol increased HR, lengthened AV delay, slowed AP and CaT upstroke speed, and shortened AP and CaT duration, yet did not result in arrhythmias. In adult popdc1^S191F zebrafish hearts, there was a higher incidence of AV block, slower AP upstroke speed, and longer AP duration compared to wild-type hearts, with no differences in CaT. SNS stress increased AV delay and led to further AV block in popdc1^S191F hearts while decreasing AP and CaT duration. Overall, we have revealed that arrhythmogenic effects of POPDC1 dysfunction on cardiac electrophysiology and intracellular calcium handling in zebrafish are varied, but already present in early development, and that AV node dysfunction may underlie SNS-induced arrhythmogenesis associated with popdc1 mutation in adults.

In recent decades, the use of zebrafish to study cardiac electrophysiology has expanded significantly, based on striking similarities between zebrafish and human action potentials, as well as the underlying ion channels involved. Here, we developed a detailed mathematical model of the zebrafish ventricular cardiomyocyte action potential. The model is based on a previously developed human cardiomyocyte framework, with a simple calcium dynamics component that allows realistic modelling of calcium transients and excitation–contraction coupling in zebrafish. It was reparameterized using published patch clamp data and newly generated L‐type calcium current recordings from single cells to adjust the biophysical properties of the principal ionic currents. The principal ionic current conductances in the model were then calibrated and validated using new experimental data, including microelectrode measurements of membrane potential and optical measurements of intracellular calcium in isolated hearts during steady‐state and restitution pacing protocols. The model was used to explore components underlying the zebrafish action potential and calcium transient, highlighting that: (1) the T‐type calcium current contributes to the action potential upstroke; (2) the L‐type calcium current strongly affects the plateau and is a greater contributor to the intracellular calcium transient than sarcoplasmic reticulum calcium release; and (3) both rapid and slow delayed rectifier potassium currents make significant contributions to action potential repolarization. Overall, the novel zebrafish‐specific computational model presented here provides a valuable tool for studying cardiac electrophysiology in zebrafish and may be adapted in future work for use in large‐scale models to study whole heart electrical activity. image Key points: We have developed the first zebrafish‐specific computational ventricular action potential model, based on new and existing patch clamp data from single cells, with model calibration and validation performed using newly generated voltage and calcium measurements in the whole heart. The model reinforces experimental findings, highlighting key roles of T‐ and L‐type calcium currents in sustaining action potential depolarization and the intracellular calcium transient. Despite conflicting evidence regarding the existence of the slow delayed rectifier potassium current in zebrafish, the model suggested its important role in repolarization. While single‐cell and tissue model simulations produced similar results, depolarization‐related parameters (i.e. action potential upstroke speed and amplitude) varied, highlighting the importance of tissue‐based simulations for accurate comparison with tissue‐derived data. The model accurately predicted action potential prolongation with individual current block, aligning with experimental data. The effects of multi‐channel block were greater than in human, emphasizing the need for caution when translating zebrafish pharmacology.

In this study, we show that the intracardiac nervous system, a key integrative pathway for autonomic control of the heart, can modulate pacemaker rate in the absence of extracardiac innervation in the isolated adult zebrafish heart. Pharmacological blockade of muscarinic and β-adrenergic receptors reveals opposing inhibitory and excitatory influences on heart rate, and these effects are eliminated when intracardiac nicotinic synaptic transmission is disrupted with hexamethonium.