(A and B) Section images of uninjured (A) and regenerated (30 dpa; B) ventricular apices visualized for endothelial cells (green) and nerves (red). The approximate regenerated area is indicated by a dashed line. Grayscale images indicating nerves, positive for acetylated alpha-tubulin, are shown in (A′) and (B′). Scale bar represents 100 μm.

(C) Whole mount images of hearts from wild-type (WT; left) and cmlc2:sema3aa (right) animals, immunostained for acetylated alpha-tubulin to indicate cardiac nerves, which are reduced by sema3aa overexpression. Scale bar represents 100 μm.

(D and E) Section images of 7 dpa ventricular apices of wild-type (D) or cmlc2:sema3aa (E) animals, stained for Mef2+PCNA+ cells. Wounds are indicated by dashed lines. Scale bar represents 100 μm.

(F) Quantification of cardiomyocyte proliferation at 7 dpa from hypo-(cmlc2:sema3aa). Wild-type clutchmates (n = 16) were used as controls for cmlc2:sema3aa animals (n = 13). Data are represented as mean ± SEM. ∗p < 0.05, Mann-Whitney rank sum.

(G) Quantification of surface innervation as measured by acetylated alpha-tubulin staining, in cmlc2:sema3aa (n = 6) and wild-type clutchmate controls (n = 5). Data are represented as mean ± SD. ∗p < 0.05, Mann-Whitney rank sum.

(H–J) Section images of 30 dpa ventricular apices of wild-type (D) or cmlc2:sema3aa (E) animals stained with acid-fuschin orange. Scale bar represents 100 μm.

(K) Quantification of regeneration between cmlc2:sema3aa and wild-type siblings were compared at 30 dpa. Hearts (H–J) were scored for regeneration, with 1 indicating complete regeneration (H), 2 indicating partial regeneration (I), and 3 indicating a block in regeneration (J). Data represent percent of total heart per score. ∗p < 0.05, Fisher’s exact.