Figure 2. Organ maturation and functional cell remodelling from 36 to 52 hpf.

(a–c) Normalized calcium transients of all cells are shown at three different time-points: (a) 36, (b) 44, and (c) 52 hpf. The shortening of black arrows indicates the decrease in time between earliest and latest activation of cells along the whole heart. The gradual formation of atrial (1) and ventricular (2) populations with increasingly different calcium transients is highlighted by the dashed lines at 52 hpf. The color of each transient indicates its activation time (all three plots use the same color scale). (d–f) Changes in cardiac and cellular morphology at each developmental time point. Estimated cell shapes are visualized as scaled ellipsoids. The color of each ellipsoid indicates activation time (relative to the first-activated cell); color code as in (a–c). Two main populations of cells corresponding to highlighted clusters in (c) are indicated by numbers. (g–i) 3D pattern of biological conduction speeds across the heart. The formation of outer curvature clusters of cells over time is highlighted by arrows. (j–l) Development of conduction speeds across the myocardium, shown as 2D projections for the three time points. The color code indicates conduction speed. Isovelocity lines are shown in black (Δv = 50 cells/s). Corresponding outer curve cell populations are highlighted by arrows.

Figure 2—figure supplement 1. Structural organ maturation from 36 to 52 hpf.

(a) One and the same developing heart, imaged at three different time-points (36 (a), 44 (b), and 52 hpf (c), scale bar 50 μm. The fitted centerline visualizes the re-shaping of the myocardium during the ongoing looping/twisting process in early cardiac development. Local coordinate systems are shown along the midline (tangent – black, normal – grey, binormal – color), the twisting point is highlighted by an arrow. Total and chamber-specific cell numbers are indicated for each stage of development (A - atrial cells, V - ventricular cells).

Figure 2—figure supplement 2. Developmental changes of cellular characteristics along the heart.

(a) The distribution of activation times is shown for three time points: 36, 44 and 52 hpf. Dots indicate individual cell activation times. Smooth regression profiles are depicted as solid lines and show faster activation with progressive organogenesis. (b) The distribution of rise times during development. The probability density function of each developmental stage is shown by a smooth kernel density. The ordinate axis shows cell numbers and illustrates progressive emergence of cell sub-populations with different calcium transient properties. (c) The distribution of each cell’s conduction speed along the midline is shown as dots. Solid lines indicate smooth regression profiles for each stage. All hearts were normalized to a standard length, showing how speed of biological conduction in working myocardium rises by comparison with early developmental stages, whereas AVC conduction velocity is unchanged.

Figure 2—figure supplement 3. Cell shape changes during development.

Cell shapes in the atrium are represented by scaled ellipsoids for the same heart at 36 hpf (a), 44 hpf (b), and 52 hpf (c). Ventricular cells are shown with reduced opacity. (d, e, f) The same cells as in (a, b, c) with additional color code indicating cell size. Smaller cells are shown in blue, larger cells in red, highlighting the increase in cell size, in particular along the larger curvature of the atrium. Changes in cell size shown as 2D projections for 36 (g), 44 (h), and 52 hpf (i). The color code indicates normalized cell size as in (d, e, f). Isosize lines shown in black (ΔA = 0.1).