Figure 4

Suppression of heart contraction with para-aminoblebbistatin raises Ca²⁺ levels, Ca²⁺ transient amplitude, and induces bradycardia in 3 dpf larvae. (A) Ratiometric images of hearts and atrial and ventricular Ca²⁺ traces from representative 3 dpf Tg(myl7:Twitch-4) untreated larvae and larvae preincubated with para-aminoblebbistatin (PAB, 75 µM) for 2 h. Image of the untreated larva shows the mechanical systole of the ventricle. (B) Atrial and ventricular systolic Ca²⁺ (Twitch-4 emission ratio), diastolic Ca²⁺, and Ca²⁺ transient amplitude in untreated larvae (n = 15), and in larvae preincubated with PAB (n = 15). (C) Atrial CaT frequency (min⁻¹) in these larvae. (D) Time-averaged Ca²⁺ levels (L/L_max) measured by bioluminescence of 3 dpf Tg(myl7:GFP-aequorin) untreated larvae (n = 5), and in larvae preincubated with 75 µM PAB for 2 h (n = 10). Diacetyl h-coelenterazine was used as the aequorin substrate. Statistical analysis was performed using an unpaired Student’s t-test for parametric data and the Mann–Whitney test for non-parametric data. Data are shown as the mean ± SD (* for p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 for parametric statistics, and ^xxxx p < 0.0001 for non-parametric statistics).