Figure 3. Intramuscular Neurotrophin-3 treatment improved functional recovery and reduced spasms after bilateral pyramidotomy.
Please note that for clarity in describing our model of spasticity, Figure 1 contained information from Figure 3 relating to the uninjured naïve and bPYX GFP groups. (A) Neurotrophin-3 treatment reduced signs of spasticity (RM two-way ANOVA, group F = 19.8, p<0.001; bPYX NT3 versus bPYX GFP at Week 6, 8 and 10, p-values<0.05). (B) Neurotrophin-3 caused rats to make fewer errors on the horizontal ladder with their treated forelimb as a percentage of the total steps taken (RM two-way ANOVA, group F = 123.4, p<0.001; bPYX NT3 versus bPYX GFP p<0.0001; bPYX NT3 versus bPYX GFP at Week 6, 8 and 10, p-values<0.05). (C) Unilateral Grip Strength Test. Neurotrophin-3 treatment slightly improved grip strength of the treated forepaw at 10 weeks (RM two-way ANOVA, group F = 145.0, p<0.001; bPYX NT3 vs bPYX GFP p = 0.11; bPYX NT3 vs bPYX GFP at Week 10, p-value = 0.026). (D) Cutaneous mechanical hypersensitivity was not affected by NT3 treatment assessed using the automated von Frey test (RM two-way ANOVA, group F = 5.2 p=0.019; bPYX NT3 versus bPYX GFP p = 0.29). (E) Dexterity was assessed using the staircase test. The two treatment groups were impaired relative to uninjured naïve rats, and no differences were detected between two treatment groups post-injury (RM two-way ANOVA, group F = 100.3, p<0.0001; bPYX NT3 versus bPYX GFP p = 0.69) confirming that the corticospinal tract is essential for recovery of fine motor function (Weidner et al., 2001). (F) Schematic showing the H-reflex paradigm. The ulnar nerve was stimulated distally and EMGs were recorded from a homonymous hand muscle (abductor digiti quinti). (G) Graph shows changes over time in hyper-reflexia, measured as the area under the curve of the frequency-dependent depression (Figure 1—figure supplement 3F). Injury groups had exaggerated reflexes from 2 weeks post-injury, but NT3 treated rats had normal H-wave depression from 6 weeks onwards. (RM two-way ANOVA, group F = 5.9 p<0.001; bPYX NT3 versus bPYX GFP p = 0.024; bPYX NT3 versus bPYX GFP at Week 6 and 10, p-values<0.05). (H) At week 10, the radial, median and ulnar nerves were exposed for stimulation and recording. The radial nerve (blue) innervates extensor muscles (blue) whereas the median and ulnar nerves (pink and magenta) innervate synergist flexor muscles that were injected with AAV (pink and magenta). (I) Stimulation of afferents in the median nerve evoked responses in the (synergist, flexor) ulnar nerve (J) The polysynaptic compound action potentials were quantified by measuring the absolute integral (area under the rectified curve) from 3.5 ms to 12 ms. Neurotrophin-3 treatment restored the exaggerated reflexes to normal (one-way ANOVA F-value = 4.8, p=0.02, bPYX NT3 versus bPYX GFP, p-values = 0.01) (K) Stimulation of afferents in the radial nerve evoked polysynaptic responses in the (antagonistic) ulnar nerve. (L) The polysynaptic compound action potentials were quantified by measuring the absolute integral (area under the rectified curve) from 3.5 ms to 12 ms. Neurotrophin-3 treatment did not restore the exaggerated reflexes to normal (one-way ANOVA F-value = 4.2, p=0.03, bPYX NT3 versus bPYX GFP, p-values = 0.63) (A–L) n = 10 or 11 per group. Data are represented as mean ± SEM.
Figure 3—figure supplement 1. Changes in polysynaptic reflexes after bilateral pyramidotomy and with Neurotrophin-3.
