Fig 2: Western blotting for markers of early/late neuronal development, apoptosis, and axonal regeneration of the ipsilateral hippocampus in sham and HI rat pups treated with vehicle or sildenafil. Median with individual data points representation, with cropped representative western blots. Full-length blots are presented in Supplementary Material. (A) Sox2. (B) Nestin. (C) Doublecortin (DCX). (D) NeuN. (E) Calretinin. (F) Calbindin. (G) Cleaved PARP. (H) GAP-43. Significance: p value from Kruskal–Wallis test, with Dunn’s post-hoc comparison tests: *p < 0.05, **p < 0.01. Number of animals: n = 6, 8 and 7 sham vehicle, 6, 8 and 7 HI vehicle, and 7, 8 and 10 HI 50 mg/kg, respectively at P12, P17 and P30.

Fig 3: Immunolabeling of GAP-43 in the proximal urethra. The existence of neuronal sprouting was assessed by immunohistochemistry against Anti-Growth Associated Protein-43 (GAP-43), which stains sprouting axons. GAP43 was abundantly expressed in urethral sections from intact (A), 1-week (B) and 4-weeks post-SCI animals (C). The co-localization of GAP-43 with all markers of nerve fibres was observed (D–H), irrespective of the time point of disease progression and location within the urethral wall. We found no significant differences in GAP-43 content (I), irrespective of the time point of disease progression. Scale bars equal 50 µm. The quantification of labelling intensity by Image J in the extension of the proximal urethra (n = 4). One-way ANOVA followed by Tukey’s multiple comparison test.