Assessment of Drug-Induced Cardiotoxicity in Zebrafish - Zebrafish: Methods for Assessing Drug Safety and Toxicity - page 48

4.4.10 Statistical Evaluation

To determine if a drug had a significant effect on heart rate, contractility,
or rhythmicity, ANOVA followed by Dunnett's test (based on raw data) was used
to identify concentrations that significantly altered heart rate; P < 0.05 was con-

sidered significant.

4.5 RESULTS

4.5.1 Heart Rate Assessment

Hatched 2dpf zebrafish were incubated with drugs for 4 h at 28 C. After incubation,
zebrafish (N

10) were visualized on a Zeiss dissecting scope, and ventricular
contractions for 30 s were counted manually. The number of contractions was
multiplied by 2 to calculate heart rate, reported in bpm.
To confirm that vehicle (0.1% DMSO) did not cause adverse effects, untreated
normal and 0.1% DMSO-treated zebrafish were used as controls. Terfenadine was
used both as a positive control drug and as a blinded test compound. Untreated
zebrafish heart rate is highly consistent with

¼

5% variation. 0.1% DMSO did not
cause significant change in heart rate. Terfenadine caused significant (P

<

0.0001)
reduction in heart rate as expected. Drug effects on zebrafish heart rate are summa-
rized in Fig. 4.1a and c.

<

4.5.2 Rhythmicity

The same zebrafish (n 10) that were used for heart rate assessment were used to

determine atrioventricular rhythmicity. Results showed that (1) untreated and 0.1%
DMSO-treated controls exhibited synchronized atrioventricular rhythmicity (AV
ratio

¼

1), and (2) positive control drug, terfenadine caused expected change in AV
ratio (AV ratio

¼

2), indicating blockage. Summary of drug effects on zebrafish heart
rhythmicity is shown in Fig. 4.1b and d.

¼

4.5.3 Summary of Observed Zebrafish Cardiotoxicity
on Function and Morphology

The most common drug-induced morphological defect was swelling of the heart
chambers (Fig. 4.2). If the ventricle stopped beating, blood frequently pooled and
clotted in the chamber. Circulation was frequently reduced or absent entirely. Note
that since sufficient oxygenation is acquired by passive diffusion, this effect does not
immediately kill zebrafish. Rates of pericardial edema were noted as well as the
presence and location of thrombi. Most thrombi occur at the yolk just posterior to the
entrance to the atrium.