(A) Representative images from cardiac sympathetic nerves (TH+) co-immunostained with the marker of axon fasciculation PSA-NCAM. SB=15 μm. (A’) Quantification of the sympathetic nerve area (TH+) that co-localizes with PSA-NCAM relative to the total TH+ area. After ablation of Postn+ SC and ECF from P0 to P6 by injecting TAM in PostnMCM/+;R26DTA mice, reduced PSA-NCAM is observed in the sympathetic nerves. Yellow arrows point to fasciculation areas (TH+/PSA-NCAM+). N=6. (B) Representative images of sympathetic nerves in the epicardial area. After ablation of Postn+ cells from P0-P7, FEZ1+ areas can be observed inside the sympathetic nerves. After Postn+ cell ablation, parallel sympathetic nerve trajectories are observed (pink arrows). SB=15 μm. (C) Representative images of sympathetic nerves around coronary blood vessels in the LV. After ablation parallel sympathetic nerve trajectories can be observed, suggesting reduced fasciculation (pink arrows). White circles mark coronary blood vessels identified morphologically. (D) Representative pictures of sympathetic nerve cross sections. SB=15 μm (D’) Quantification of sympathetic nerve cross-section area shows a statistically significant reduction in nerve thickness after Postn+ cell ablation, which is suggestive of reduced nerve fasciculation. (E) Higher magnification of sympathetic nerve cross-section of similar size. The inset shows an example of the “closest bright point” automatic measurements performed to measure the distance between axons indicated by the TH+ signal in the immunofluorescence. (E’) Measurement of “closest bright point” performed to measure the distance between axons. After Postn+ cell ablation from P0 to P7, a trend towards bigger separation between axons was found. N=7. (F) Representative pictures of intracardiac neuronal ganglia. Some sympathetic neuronal bodies (DAPI surrounded by TH+ signal) with intranuclear expression of FEZ1 can be observed. SB=15 μm. (F’) No statistically significant differences in the number of FEZ1+ sympathetic neuron nuclei were observed. DAPI was used to stain nuclei. LA=Left Atria, LV=Left Ventricle, RV=Right Ventricle. *P < 0.05 by Mann-Whitney test.