Characterization of isolated ventricular myocytes from adult zebrafish (Danio rerio)

Abstract

The zebrafish is widely used for human related disease studies. Surprisingly, there is no information about the electrical activity of single myocytes freshly isolated from adult zebrafish ventricle. In this study, we present an enzymatic method to isolate ventricular myocytes from zebrafish heart that yield a large number of calcium tolerant cells. Ventricular myocytes from zebrafish were imaged using light and confocal microscopy. Myocytes were mostly rod shaped and responded by vigorous contraction to field electrical stimulation. Whole cell configuration of the patch clamp technique was used to record electrophysiological characteristics of myocytes. Action potentials present a long duration and a plateau phase and action potential duration decreases when increasing stimulation frequency (as observed in larger mammals). Together these results indicate that zebrafish is a species ideally suited for investigation of ion channels related mutation screening of cardiac alteration important in human.

Generating an In Vitro 3D Cell Culture Model from Zebrafish Larvae for Heart Research

“…Grunow et al (2011) and Mehnert et al (2013) showed previously that aggregates generated from larval trout have beat rates and action potential characteristics that closely match those of humans and that two commonly used hERG blockers, which are known to be pro-arrhythmic in humans, prolong action potentials in fish aggregates (Mehnert et al, 2013). Whilst we have not confirmed these findings in ZFHAs, our previous work (Brette et al, 2008) suggests ZF have a similar complement of cardiac ion channels to those of adult mammalian myocytes (Nemtsas et al, 2010) and hiPSC-CM (van den Heuvel et al, 2014). Thus, electrical activity between fish and humans is more similar than that between human and mouse, and, as such, ZFHA may be suitable for high/medium throughput pharmacological screening for human safety pharmacology (Brette et al, 2008;Nemtsas et al, 2010;Mehnert et al, 2013).…”

“…Whilst we have not confirmed these findings in ZFHAs, our previous work (Brette et al, 2008) suggests ZF have a similar complement of cardiac ion channels to those of adult mammalian myocytes (Nemtsas et al, 2010) and hiPSC-CM (van den Heuvel et al, 2014). Thus, electrical activity between fish and humans is more similar than that between human and mouse, and, as such, ZFHA may be suitable for high/medium throughput pharmacological screening for human safety pharmacology (Brette et al, 2008;Nemtsas et al, 2010;Mehnert et al, 2013).…”

“…The intrinsic contraction frequency of the ZFHAs is also akin to that of the adult human heart rate (60-90 beats min −1 at rest). The similarity in heart rate between ZF (at 20°C) and humans (at 37°C) is driven by an action potential with a very similar waveform; both have a prominent plateau phase, which reflects similarities in their underlying ion channel profiles (Brette et al, 2008;Nemtsas et al, 2010;Verkerk and Remme, 2012). Rodents, in contrast, have ∼10 times faster heart rates, and their action potential lacks a significant plateau phase, which is reflected in their ion channel expression (Li et al, 1999;Nerbonne et al, 2001).…”

“…This method builds on previous work by Grunow et al (2011) and Mehnert et al (2013) for salmonids but also offers new investigative potential by virtue of the role of ZF as a model organism for cardiac research. Work by us (Brette et al, 2008) and others (Nemtsas et al, 2010;Verkerk and Remme, 2012;Bovo et al, 2013) clearly shows that the ZF heart is a relevant model for human cardiac ion channel disorders, particularly those associated with cardiac repolarisation. ZF are also an established model species for environmental and (eco) toxicology testing (Strähle et al, 2011;Busch et al, 2011), with recent work focusing on the effect of pollutants on the ZF cardiovascular system (Parker et al, 2014).…”

Development of an In vitro Cultivated, Spontaneously and Long-term Contracting 3D Heart Model as a Robust Test System

“…Furthermore, the use of these SCC as a human drug test system has been described as their electrophysiological properties and their reaction to drugs are similar to human cardiomyocytes [8,9]. Fish cardiomyocytes could be an alternative because of their analogy to human heart electrophysiology [10][11][12].…”