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Wnt/β-catenin activation rescues AC phenotypes in Dsp zebrafish mutants. A -ab mutant zebrafish larvae in Wnt/β-catenin Tg(7xTCF-Xla.Siam:EGFP)ia4 transgenic background, treated for 2 days with 5 µM XAV939 (XAV, Wnt inhibitor) or 40 µM SB216763 (SB, Wnt agonist), showed a downregulation or a full recovery of the pathway activity (GFP reporter) when compared to WT controls or untreated mutants. All embryos are at 3 dpf and displayed in lateral view, anterior to the left. Sample size: WT n = 45; -ab n = 55; -ab + XAV n = 17; -ab + SB n = 21. R.I. relative intensity. Error bars: SEM. **p < 0.01; ****p < 0.0001. Test: One-way ANOVA followed by Tukey’s test. B The -ab larvae treated with 40 µM SB showed a significant reduction of cardiac dysmorphisms, while 5 µM XAV induced a worsening of the condition when compared to WT controls or untreated mutants. All embryos are at 3 dpf and displayed in lateral view, anterior to the left. Sample size: n = 100. C The -ab larvae treated for 2 days with 40 µM SB216763 showed a significant rescue of the main morphological alterations. The treatment with 5 µM XAV939 induced a worsening of the condition. Sample size Cardiac region size: WT n = 41; -ab n = 41; -ab + XAV n = 37; -ab + SB n = 82. Sample size Body length WT n = 41; -ab n = 46; -ab + XAV n = 54; -ab + SB n = 82. Sample size Eye size: WT n = 41; -ab n = 46; -ab + XAV n = 36; -ab + SB n = 81. R.R. relative ratio. Error bars: SEM. ns not significant; **p < 0.01; ***p < 0.001; ****p < 0.0001. Test: One-way ANOVA followed by Tukey’s test. D At 5 dpf, treated -ab larvae showed a statistically significant recovery of the bradycardia phenotype. Sample size: WT n = 70; -ab n = 66; -ab + XAV n = 40; -ab + SB n = 77. Error bars: SEM. ****p < 0.0001. Test: One-way ANOVA followed by Tukey’s test. E One-month survival rate of WT and -ab larvae untreated or treated with 40 µM SB216763 for 4 days; a statistically significant decreased mortality in mutants is observed after treatment, compared to untreated mutants (p < 0.0001). Sample size: n = 100. Error bars: SEM. Test: Log-rank (Mantel-Cox) test. F: 5-day survival analysis, from 3 to 8 dpf, of untrained or trained WT and -ab, as well as -ab simultaneously SB-treated and trained (training induced in 1% methylcellulose). Trained -ab, treated with SB, showed a statistically significant decrease in mortality when compared with -ab subjected to training in the absence of SB (p < 0.01). Sample size: n = 50. Error bars: SEM. Test: Test: Log-rank (Mantel–Cox) test.
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