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Fig. 5 | Acta Neuropathologica Communications

Fig. 5

From: Slow motor neurons resist pathological TDP-43 and mediate motor recovery in the rNLS8 model of amyotrophic lateral sclerosis

Fig. 5

Cross-reinnervation surgery manipulates slow MNs to innervate TA muscle in place of FF MNs. a Schematic representation of unilateral cross reinnervation surgery. b The experimental approach for using cross-reinnervation surgery to investigate the significance of MN identity in axonal regeneration following hTDP-43ΔNLS disease. rNLS8 mice were subjected to unilateral cross-innervation or self-innervation surgery (the latter as a procedural control) and allowed to recover for 6, 10 and 24 wks. Recovery was confirmed using bilateral TA muscle CMAP and TreadScan analysis. At 6, 10 and 24 wks. post-operation, the TA muscle was bilaterally injected with CTB-488 3 days before sacrificed mice to validate the reinnervation surgery. At 10 wks. post-operation, Dox was withdrawn to activate hTDP-43ΔNLS expression for a period of 8 wks. c Representative fluorescence images of sections from the lumbar spinal cord highlighting the ipsilateral (cross-reinnervation surgery; “S”) and contralateral (non-surgical; “N.S.”) sides of rNLS8 mice 10 wks. after surgery. Visible are FF MNs positive for Mmp9 immunostaining (red) and re-innervated TA MNs retrogradely labelled with CTB-488 (green). Mmp9-negative/CTB-488-positive MNs were either slow type or fatigue-resistant TA MNs (white arrows), whereas Mmp9-positive/CTB-488-positive MNs were fast-fatigable TA MNs (yellow arrows). Scale bar: 50 µm. d Representative fluorescence images of sections from the lumbar spinal cord highlighting the ipsilateral (cross-reinnervation surgery; “S”) and contralateral (non-surgical; “N.S.”) sides of rNLS8 mice 10 wks. after surgery. Visible are MNs positive for SK3 immunostaining (red) and re-innervated TA MNs retrogradely labelled with CTB-488 (green). White arrows mark SK3-negative, CTB-488-positive presumptive FF TA MNs. The yellow arrow marks as SK3-positive, CTB-488-positive slow type or fatigue-resistant TA MN. Scale bar: 50 µm. e Quantification of reinnervated FF type TA MNs (Mmp9-positive/CTB-488-positive) as a proportion of total re-innervated TA MNs (all CTB-488-positive) across the study time course outlined in (b). Two-to-four mice were assayed per time point, with 70–80 MNs scored per side. Two-way ANOVA, ***p = 0.0003. f Quantification of reinnervated slow type TA MNs (SK3-positive/CTB-488-positive MNs) as a proportion of total re-innervated TA MNs (all CTB-488-positive) across the study time course outlined in (b). Three-to-five mice were assayed per timepoint, with 70–80 MNs scored per side. Two-way ANOVA, ****p < 0.0001

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