Streamlining drug discovery assays for cardiovascular disease using zebrafish

Figures & data

Table 1. Pharmacological impact of common human cardiovascular drugs on zebrafish.

drug	Targeted disease in humans	Pharmacological effect in humans	Major finding in zebrafish	Reference
Amiodarone	Heart rhythm disorder (tachycardia)	Inhibition of sodium ion currents	Reduced heart rate and prolonged QT interval in wild-type zebrafish	[52]
Propranolol	Heart rhythm disorder (tachycardia)	β-receptor antagonist	Reduced calcium currents and decreased heart rate in wild-type zebrafish	[26]
Isoproterenol	Heart rhythm disorder (bradycardia)	β-receptor agonist	Increased heart rate and increased ventricular systolic pump function in wild-type zebrafish	[26]
Dobutamine	Heart failure, bradycardia	β-receptor agonist	Increased heart rate in zebrafish mutant line hiphop with severe bradycardia	[51]
Spironolactone	Heart failure	Mineralocorticoid receptor antagonist	Increased systolic pump function in aristolochic acid induced heart failure model	[47]
Empagliflozin	Heart failure	Inhibition of sodium-glucose co-transport	Restoration of structural and functional defects in aristolochic acid induced heart failure model	[48]
Nifedipinie	Arterial hypertension	L-type calcium channel blocker	Reduction of calcium currents in both atrium and ventricle in embryonic zebrafish heart	[26,49]
Warfarin	Ischemic stroke	Vitamin K antagonist	Antagonims of vitamin K dependent gamma-carboxylation of hemostatic proteins leading to severe bleeding in wild-type zebrafish	[53,55]
Rivaroxaban	Ischemic stroke	Factor Xa antagonist	Inhibition of thrombin generation in wild-type zebrafish	[50]
Clopidogrel	Vascular thrombosis	Irreversible inhibition of platelet P2Y12-receptor	Reduced thrombus formation in an arachidonic acid induced thrombosis zebrafish model	[54]

Table 2. Selected zebrafish mutants as models for human CVD.

Zebrafish mutant	Mutated zebrafish gene	Phenotype	Human CVD gene	Human disease	Reference
pickwick	titin	Contractile dysfunction	TITIN	DCM, HCM, ARVC	[71,109–111]
silent heart	tnnt2	Contractile dysfunction	TNNT2	DCM, HCM	[72,73]
tell tale heart	myl7	Contractile dysfunction	MYL7	DCM, HCM	[74–76]
lazy susan	cmlc1	Contractile dysfunction	cMLC	HCM	[77,78]
heart strings	tbx5a	Heart development defect	TBX5	Holt-Oram syndrome	[79,80]
breakdance	kcnh6a/kcnh2	Arrhythmia, long QT syndrome	KCNH6	long QT syndrome	[81–83]
reggae	kcnh6a/kcnh2	Arrhythmia, short QT syndrome	KCNH6	short QT syndrome	[62,84]
hiphop	atp1a1a.1	Arrhythmia, bradycardia	ATP1A1	QT interval modulation	[51,118]
island beat	cacna1c	Atrial fibrillation	CACNA1C	Brugada syndrome, long/short QT syndrome	[85–87]
cardiofunk	acta1b	AVC defect	ACTA1	nemaline myopathy, DCM	[88–90]
jekyll	ugdh	AVC defect	UGDH	AVS defect	[91–93]
lost-a-fin	acvr1l	Heart looping defects	ACVRL1	AVS defect	[94,95]
apc	apc	AVC defect, EC hyperplasia	APC	AVS defect	[96,97]
weak atrium	myh6	atrial dysfunction	MYH6	DCM, HCM, atrial septal defect	[98,99]
weiches herz	tbx20	Contractile dysfunction, hypoplasia	TBX20	atrial septal defect, DCM	[70,100–103]
main squeeze/lost contact	ilk	Contractile dysfunction	ILK	DCM	[104,105]
T2EGEZ8	tpm4	Contractile dysfunction	TPM4	HCM	[106–108]

Abbreviations: DCM, Dilated Cardiomyopathy; HCM, Hypertrophic Cardiomyopathy; ARVC, Arrhythmogenic Right Ventricular Cardiomyopathy; AVC, Atrioventricular Canal; AVS, Atrioventricular Valve and Septum; EC, Endocardial Cushions

Figure 1. Essential electrocardiographic parameters including p-wave, QRS complex, and T-wave, between (a) humans and (b) zebrafish are similar. Also, de- and repolarizing ion currents during action potential of human and zebrafish cardiomyocyte share common characteristics. (c) QT interval measurement by electrocardiogram (ECG) in embryonic wild-type zebrafish with normal QT interval in comparison to (D) prolonged QT interval in a zebrafish mutant with prolonged myocardial repolarization. (adapted from [14] and [51] with permission of Bentham Science Publishers and Elsevier, respectively).

Lin MH, Chou HC, Chen YF, et al. Development of a rapid and economic in vivo electrocardiogram platform for cardiovascular drug assay and electrophysiology research in adult zebrafish. Sci Rep. 2018;8: 15986-018-33577-7.

 Web of Science ®Google Scholar

van Opbergen CJM, Koopman CD, Kok BJM, et al. Optogenetic sensors in the zebrafish heart: a novel in vivo electrophysiological tool to study cardiac arrhythmogenesis. Theranostics. 2018;8:4750–4764.

 PubMed Web of Science ®Google Scholar

Pott A, Bock S, Berger IM, et al. Mutation of the Na(+)/K(+)-ATPase Atp1a1a.1 causes QT interval prolongation and bradycardia in zebrafish. J Mol Cell Cardiol. 2018;120:42–52.

 PubMed Web of Science ®Google Scholar

Huang CC, Monte A, Cook JM, et al. Zebrafish heart failure models for the evaluation of chemical probes and drugs. Assay Drug Dev Technol. 2013;11:561–572.

 PubMed Web of Science ®Google Scholar

Shi X, Verma S, Yun J, et al. Effect of empagliflozin on cardiac biomarkers in a zebrafish model of heart failure: clues to the EMPA-REG OUTCOME trial? Mol Cell Biochem. 2017;433:97–102.

 PubMed Web of Science ®Google Scholar

Hou JH, Kralj JM, Douglass AD, et al. Simultaneous mapping of membrane voltage and calcium in zebrafish heart in vivo reveals chamber-specific developmental transitions in ionic currents. Front Physiol. 2014;5:344.

 PubMed Web of Science ®Google Scholar

Jagadeeswaran P, Sheehan JP. Analysis of blood coagulation in the zebrafish. Blood Cells Mol Dis. 1999;25:239–249.

 PubMed Web of Science ®Google Scholar

Hanumanthaiah R, Thankavel B, Day K, et al. Developmental expression of vitamin K-dependent gamma-carboxylase activity in zebrafish embryos: effect of warfarin. Blood Cells Mol Dis. 2001;27:992–999.

 PubMed Web of Science ®Google Scholar

Schurgers E, Moorlag M, Hemker C, et al. Thrombin generation in zebrafish blood. PLoS One. 2016;11:e0149135.

 PubMed Web of Science ®Google Scholar

Zhu XY, Liu HC, Guo SY, et al. A zebrafish thrombosis model for assessing antithrombotic drugs. Zebrafish. 2016;13:335–344.

 PubMed Web of Science ®Google Scholar

Huttner IG, Wang LW, Santiago CF, et al. A-band titin truncation in zebrafish causes dilated cardiomyopathy and hemodynamic stress intolerance. Circ Genom Precis Med. 2018;11:e002135.

 PubMed Web of Science ®Google Scholar

Xu X, Meiler SE, Zhong TP, et al. Cardiomyopathy in zebrafish due to mutation in an alternatively spliced exon of titin. Nat Genet. 2002;30:205–209.

 PubMed Web of Science ®Google Scholar

Herman DS, Lam L, Taylor MR, et al. Truncations of titin causing dilated cardiomyopathy. N Engl J Med. 2012;366:619–628.

 PubMed Web of Science ®Google Scholar

Huang W, Zhang R, Xu X. Myofibrillogenesis in the developing zebrafish heart: a functional study of tnnt2. Dev Biol. 2009;331:237–249.

 PubMed Web of Science ®Google Scholar

Watkins H, McKenna WJ, Thierfelder L, et al. Mutations in the genes for cardiac troponin T and alpha-tropomyosin in hypertrophic cardiomyopathy. N Engl J Med. 1995;332:1058–1064.

 PubMed Web of Science ®Google Scholar

Chen Z, Huang W, Dahme T, et al. Depletion of zebrafish essential and regulatory myosin light chains reduces cardiac function through distinct mechanisms. Cardiovasc Res. 2008;79:97–108.

 PubMed Web of Science ®Google Scholar

Hernandez OM, Jones M, Guzman G, et al. Myosin essential light chain in health and disease. Am J Physiol Heart Circ Physiol. 2007;292:H1643–54.

 PubMed Web of Science ®Google Scholar

Rottbauer W, Wessels G, Dahme T, et al. Cardiac myosin light chain-2: a novel essential component of thick-myofilament assembly and contractility of the heart. Circ Res. 2006;99:323–331.

 PubMed Web of Science ®Google Scholar

Poetter K, Jiang H, Hassanzadeh S, et al. Mutations in either the essential or regulatory light chains of myosin are associated with a rare myopathy in human heart and skeletal muscle. Nat Genet. 1996;13:63–69.

 PubMed Web of Science ®Google Scholar

Ahn DG, Kourakis MJ, Rohde LA, et al. T-box gene tbx5 is essential for formation of the pectoral limb bud. Nature. 2002;417:754–758.

 PubMed Web of Science ®Google Scholar

Basson CT, Bachinsky DR, Lin RC, et al. Mutations in human TBX5 [corrected] cause limb and cardiac malformation in Holt-Oram syndrome. Nat Genet. 1997;15:30–35.

 PubMed Web of Science ®Google Scholar

Balijepalli SY, Anderson CL, Lin EC, et al. Rescue of mutated cardiac ion channels in inherited arrhythmia syndromes. J Cardiovasc Pharmacol. 2010;56:113–122.

 PubMed Web of Science ®Google Scholar

Meder B, Scholz EP, Hassel D, et al. Reconstitution of defective protein trafficking rescues Long-QT syndrome in zebrafish. Biochem Biophys Res Commun. 2011;408:218–224.

 PubMed Web of Science ®Google Scholar

Hassel D, Scholz EP, Trano N, et al. Deficient zebrafish ether-a-go-go-related gene channel gating causes short-QT syndrome in zebrafish reggae mutants. Circulation. 2008;117:866–875.

 PubMed Web of Science ®Google Scholar

Schimpf R, Wolpert C, Gaita F, et al. Short QT syndrome. Cardiovasc Res. 2005;67:357–366.

 PubMed Web of Science ®Google Scholar

Pfeufer A, Sanna S, Arking DE, et al. Common variants at ten loci modulate the QT interval duration in the QTSCD Study. Nat Genet. 2009;41:407–414.

 PubMed Web of Science ®Google Scholar

Splawski I, Timothy KW, Sharpe LM, et al. Ca(V)1.2 calcium channel dysfunction causes a multisystem disorder including arrhythmia and autism. Cell. 2004;119:19–31.

 PubMed Web of Science ®Google Scholar

Li W, Zheng NZ, Yuan Q, et al. NFAT5-mediated CACNA1C expression is critical for cardiac electrophysiological development and maturation. J Mol Med (Berl). 2016;94:993–1002.

 PubMed Web of Science ®Google Scholar

Sztal TE, Zhao M, Williams C, et al. Zebrafish models for nemaline myopathy reveal a spectrum of nemaline bodies contributing to reduced muscle function. Acta Neuropathol. 2015;130:389–406.

 PubMed Web of Science ®Google Scholar

D’Amico A, Graziano C, Pacileo G, et al. Fatal hypertrophic cardiomyopathy and nemaline myopathy associated with ACTA1 K336E mutation. Neuromuscul Disord. 2006;16:548–552.

 PubMed Web of Science ®Google Scholar

Hyde AS, Farmer EL, Easley KE, et al. UDP-glucose dehydrogenase polymorphisms from patients with congenital heart valve defects disrupt enzyme stability and quaternary assembly. J Biol Chem. 2012;287:32708–32716.

 PubMed Web of Science ®Google Scholar

Warren KS, Fishman MC. “Physiological genomics”: mutant screens in zebrafish. Am J Physiol. 1998;275:H1–7.

 PubMedGoogle Scholar

Shore EM, Xu M, Feldman GJ, et al. A recurrent mutation in the BMP type I receptor ACVR1 causes inherited and sporadic fibrodysplasia ossificans progressiva. Nat Genet. 2006;38:525–527.

 PubMed Web of Science ®Google Scholar

LaBonty M, Pray N, Yelick PC. A zebrafish model of human fibrodysplasia ossificans progressiva. Zebrafish. 2017;14:293–304.

 PubMed Web of Science ®Google Scholar

Masuelli L, Bei R, Sacchetti P, et al. Beta-catenin accumulates in intercalated disks of hypertrophic cardiomyopathic hearts. Cardiovasc Res. 2003;60:376–387.

 PubMed Web of Science ®Google Scholar

Hurlstone AF, Haramis AP, Wienholds E, et al. The Wnt/beta-catenin pathway regulates cardiac valve formation. Nature. 2003;425:633–637.

 PubMed Web of Science ®Google Scholar

YH S, Zhang Y, Ding Y, et al. Cardiac transcriptome and dilated cardiomyopathy genes in zebrafish. Circ Cardiovasc Genet. 2015;8:261–269.

 PubMedGoogle Scholar

Tomita-Mitchell A, Stamm KD, Mahnke DK, et al. Impact of MYH6 variants in hypoplastic left heart syndrome. Physiol Genomics. 2016;48:912–921.

 PubMed Web of Science ®Google Scholar

Just S, Raphel L, Berger IM, et al. Tbx20 is an essential regulator of embryonic heart growth in zebrafish. PLoS One. 2016;11:e0167306.

 PubMed Web of Science ®Google Scholar

Mittal A, Sharma R, Prasad R, et al. Role of cardiac TBX20 in dilated cardiomyopathy. Mol Cell Biochem. 2016;414:129–136.

 PubMed Web of Science ®Google Scholar

Chakraborty S, Sengupta A, Yutzey KE. Tbx20 promotes cardiomyocyte proliferation and persistence of fetal characteristics in adult mouse hearts. J Mol Cell Cardiol. 2013;62:203–213.

 PubMed Web of Science ®Google Scholar

Bendig G, Grimmler M, Huttner IG, et al. Integrin-linked kinase, a novel component of the cardiac mechanical stretch sensor, controls contractility in the zebrafish heart. Genes Dev. 2006;20:2361–2372.

 PubMed Web of Science ®Google Scholar

Knoll R, Postel R, Wang J, et al. Laminin-alpha4 and integrin-linked kinase mutations cause human cardiomyopathy via simultaneous defects in cardiomyocytes and endothelial cells. Circulation. 2007;116:515–525.

 PubMed Web of Science ®Google Scholar

Alcalai R, Seidman JG, Seidman CE. Genetic basis of hypertrophic cardiomyopathy: from bench to the clinics. J Cardiovasc Electrophysiol. 2008;19:104–110.

 PubMed Web of Science ®Google Scholar

Zhao L, Zhao X, Tian T, et al. Heart-specific isoform of tropomyosin4 is essential for heartbeat in zebrafish embryos. Cardiovasc Res. 2008;80:200–208.

 PubMed Web of Science ®Google Scholar

Pott A, Rottbauer W, Just S. Functional genomics in zebrafish as a tool to identify novel antiarrhythmic targets. Curr Med Chem. 2014;21:1320–1329.

 PubMed Web of Science ®Google Scholar