Development of tools for embryonic ECG for heart dysfunction in zebrafish

In the past decade, the zebrafish, Danio rerio, has risen to much greater prominence as a vertebrate model system for drug discovery and toxicity testing. Zebrafish larvae represent an in vivo vertebrate model, with high throughput potential and proven efficacy as a model of human disease and drug responses. Specifically, zebrafish have been used for cardiotoxicity studies and cardiac arrhythmia modelling. This project aims to develop and optimise tools for electrocardiographic recording in zebrafish larvae for use in cardiotoxicity screening and human arrhythmia modelling. Steps were taken towards a high throughput ECG system for zebrafish larvae through establishing viability of non-contact capillary electrode recording and development of microfabricated electrode arrays capable of replacing glass capillary electrodes. In depth analysis of atrium and ventricle signals using wavelet analysis was performed and the frequency profiles for these chambers examined. The utility of the larval ECG recording system for characterisation of a cardiac mutant (cacna1c sa6050) was demonstrated through a set of drug treatments and demonstration of drug discovery through ECG analysis of chemical rescue of the mutant phenotype. Overall, the results presented highlight the zebrafish as an effective, robust and reliable model for cardiac arrhythmia and assessing drug responses in vivo

3D finite element electrical model of larval zebrafish ECG signals

Assessment of heart function in zebrafish larvae using electrocardiography (ECG) is a potentially useful tool in developing cardiac treatments and the assessment of drug therapies. In order to better understand how a measured ECG waveform is related to the structure of the heart, its position within the larva and the position of the electrodes, a 3D model of a 3 days post fertilisation (dpf) larval zebrafish was developed to simulate cardiac electrical activity and investigate the voltage distribution throughout the body. The geometry consisted of two main components; the zebrafish body was modelled as a homogeneous volume, while the heart was split into five distinct regions (sinoatrial region, atrial wall, atrioventricular band, ventricular wall and heart chambers). Similarly, the electrical model consisted of two parts with the body described by Laplace’s equation and the heart using a bidomain ionic model based upon the Fitzhugh-Nagumo equations. Each region of the heart was differentiated by action potential (AP) parameters and activation wave conduction velocities, which were fitted and scaled based on previously published experimental results. ECG measurements in vivo at different electrode recording positions were then compared to the model results. The model was able to simulate action potentials, wave propagation and all the major features (P wave, R wave, T wave) of the ECG, as well as polarity of the peaks observed at each position. This model was based upon our current understanding of the structure of the normal zebrafish larval heart. Further development would enable us to incorporate features associated with the diseased heart and hence assist in the interpretation of larval zebrafish ECGs in these conditions

A comparison between model relative peak heights and measured relative peak heights.

<p>A comparison between model relative peak heights and measured relative peak heights.</p

Comparison between model and recorded action potential properties.

<p>Comparison between model and recorded action potential properties.</p

Activation wave conduction velocities in an adult human and a 3 dpf zebrafish heart.

<p>Activation wave conduction velocities in an adult human and a 3 dpf zebrafish heart.</p

Maximum R wave voltage and R wave percentage change.

<p>The point at which the R wave is at its maximum at 0.84 s from the start of the simulation. A) zoomed out ventral view of the model with the point of maximum voltage intensity labelled, B) close-up ventral view with the maximum point labelled, C) close-up ventral view showing ten different points distributed around the ventricle as seen on the surface of the body, D) percentage of the maximum R wave voltage at each point.</p

A comparison between model ECG (model 2 from Fig 8) parameters and measured ECG parameters.

<p>A comparison between model ECG (model 2 from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0165655#pone.0165655.g008" target="_blank">Fig 8</a>) parameters and measured ECG parameters.</p

Temporal sequence of transmembrane potential (V_m).

<p>The propagation of an action potential through the heart is shown at different times. Nine time steps were chosen to show the progression of the action potentials through all stages of the cardiac cycle. A) the heart at rest B) atrial depolarisation C) ventricular depolarisation D) the end of atrial repolarisation and ventricular plateau stage E,F,G,H) ventricular repolarisation I) returning to the rest state. The wave originates at the sinoatrial region then it progresses across the atrium, through the atrioventricular band and into the ventricle. An animation is given in <b><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0165655#pone.0165655.s005" target="_blank">S2 File</a></b>.</p

Initial conditions.

<p>Initial conditions.</p