Analysis of CST contacts onto cervical interneurons after SCI.

<p>(A,B) Confocal images of contacts (arrows, defined as boutons in apposition to neuronal cell bodies) between hindlimb CST collaterals (YFP, green) and the cell bodies of cervical interneurons (Neurotrace, red) at 4 weeks (A) and 12 weeks (B) following SCI. (C) Quantification of the number of contacts a given hindlimb CST collateral makes with the cell body of a single interneuron at different timepoints after SCI as well as in the lumbar spinal cord of unlesioned animals. (D) Quantification of the percentage of Neurotrace (NT)-stained interneurons contacted by collaterals emerging from the different CST components at multiple timepoints following the lesion. Asterisks indicate significant difference compared to main CST collaterals. Scale bar in A (also for B), 15 µm.</p

Schematic representation of hindlimb CST remodeling following SCI.

<p>Scheme illustrating the formation of cervical collaterals derived from the main CST (upper row) and the minor dorsolateral (2^nd row) and ventral (3^rd row) CST components at 10 days (blue), 3–4 weeks (green) and 12 weeks (red) after SCI. Bottom row illustrates the refinement over time of the contacts between CST collaterals and cervical interneurons.</p

Synaptic differentiation of newly formed CST boutons.

<p>(A–B) Confocal images of bassoon immmunostaining (green) in the cervical spinal cord of mice with a traced hindlimb CST (BDA, red) perfused 10 days (A) and 3 weeks following SCI (B). Yellow arrows indicate boutons that were immunoreactive for bassoon, white arrows indicate those that were not. (A′–A′″) Single plane confocal image of the boutons boxed in A showing the collateral (A′; BDA, white), bassoon immunostaining (A″; white) and the overlay (A′″; BDA, red; bassoon, green) at 10 days after SCI. (B′–B′″) Single plane confocal image of the bouton boxed in B showing the collateral (B′; BDA, white), bassoon immunostaining (B″, white) and the overlay (B′″; BDA, red; bassoon, green) at 3 weeks after SCI. (C) Quantification of the number of boutons on hindlimb CST collaterals that were immunopositive for bassoon at 10 days and 3 weeks following SCI in the cervical cord. The percentages were normalized to the expression pattern in the lumbar cord (L) of control animals (which was set to 100%). (D–E) Confocal images of synapsin I immunostaining in the cervical spinal cord of mice with a traced hindlimb CST (BDA, red) perfused 10 days following SCI (D) and at 3 weeks post-injury (E). Yellow arrows indicate boutons that were immunoreactive for synapsin I, white arrows indicate those that were not. (D′–D′″) Single plane confocal image of the bouton boxed in D showing the collateral (D′, BDA, white), the synapsin I staining (D″; white) and the overlay (D′″; BDA, red; synapsin I, green) at 10 days after SCI. (E′–E′″) Single plane confocal image of the bouton boxed in E showing the collateral (E′; BDA, white), the bassoon staining (E″; white) and the overlay (D′″; BDA, red; synapsin I, green) at 3 weeks after SCI. (F) Quantification of the number of boutons on hindlimb CST collaterals that were immunnopositive for synapsin I at 10 days and 3 weeks following SCI in the cervical cord. The percentages were normalized to the expression pattern in the lumbar cord (L) of control animals (which was set to 100%). (G) Quantification of the co-expression of bassoon and synapsin I in boutons of CST collaterals of animals perfused at 3 weeks after injury (expressed as percentages of all immunoreactive boutons). Scale bar in A (also for B, D, E), 10 µm and in A′ (also for A″–E′″), 3 µm.</p

Population analysis of hindlimb CST collateral formation at different timepoints after SCI.

<p>(A–C) Reconstruction of hindlimb CST collaterals (black) from 5 consecutive sections in the cervical spinal cord of control mice (A) and of mice perfused 10 days (B) and 24 weeks (C) following SCI. (D) Quantification of the total numbers of collaterals emerging from all CST components in the cervical gray matter of control mice and of mice at different timepoints following SCI. (E–G) Confocal images of main CST (BDA, yellow) and the adjacent gray matter (Neurotrace, blue; border shown by dashed white line) in control mice (E) and in mice perfused 10 days (F, arrow indicates CST collateral emerging from main CST) and 24 weeks (G, arrow indicates CST collateral emerging from main CST) following SCI. (H) Quantification of the number of collaterals emerging from the main CST component at different timepoints following SCI. (I–K) Confocal images of the minor dorso-lateral CST (BDA, yellow) and the adjacent gray matter (Neurotrace, blue; border shown by dashed white line) in control mice (I) and in mice perfused 10 days (J) and 24 weeks (K, arrow indicates CST collateral emerging from dorso-lateral CST) following SCI. (L) Quantification of the number of collaterals emerging from the minor dorso-lateral CST component at different timepoints following SCI. (M–O) Confocal images of the minor ventral CST (BDA, yellow) and the adjacent gray matter (Neurotrace, blue; border shown by dashed white line) in control mice (M, arrow indicates ventral CST fiber) and in mice perfused 10 days (N) and 24 weeks (O, arrow indicates collateral emerging from ventral CST) following SCI. (P) Quantification of the number of collaterals emerging from the minor ventral CST component at different timepoints following SCI. Asterisks indicate significance compared to the unlesioned controls. Pound signs indicate significance compared to the 10-day timepoint. Scale bar in A (also for B,C), 500 µm; in M (also for E–O), 100 µm.</p

Illustration of a dorsal hemisection of the thoracic spinal cord.

<p>(A) Confocal image of a cross-section of the thoracic (T8) spinal cord of a mouse perfused 12 weeks after dorsal hemisection (counterstained with Neurotrace). Dashed line indicates lesion border. (B) Schematic representation of the location of the different CST components (highlighted in different shades of green) in relation to this lesion (outlined by dashed line from A). Scale bar in A, 200 µm.</p

Strategies for labeling individual CST collaterals.

<p>(A, B) Confocal images of the cortex of a <i>Thy1-Brainbow</i> (A; YFP, yellow; CFP, blue) and <i>Thy1-Stp-YFP</i> mouse (B; YFP, green; Neurotrace, red) after local injection of rAAV-Cre. Boxed areas are magnified 2 times in insets. (C,D) Confocal images of CST collaterals in the cervical spinal cord of a <i>Thy1-Brainbow</i> (C; YFP, yellow; CFP, blue) and <i>Thy1-Stp-YFP</i> (D; YFP, green; Neurotrace, red) mouse after injection of rAAV-Cre in the cortex. Boxed areas are magnified 2 times in insets. Arrows in inset in C indicate different collaterals expressing either CFP (blue), YFP (yellow) or a combination of both (white). Dashed white line indicates the outline of the spinal gray matter. (E–G) Confocal images of individual collaterals (white) emerging from the main CST (F), the dorso-lateral CST (E) and the ventral CST (G) following SCI. Arrows indicate individual collaterals. Dashed white lines indicate the outline of the spinal gray matter. Scale bar in B (also for A),100 µm; Scale bar in D (also for C),100 µm; Scale bar in G (also for E,F), 50 µm.</p

Brain mappings of beta-ERD.

<p><b>A–D:</b> Beta-ERD mapping results for control subjects were shown in four sub-stages. <b>E–H:</b> Beta-ERD mapping results for stroke patients were illustrated which were corresponding to four sub-stages respectively. All other three sub-stages had lower ERD than Baseline, and deeper blue color represented higher phase-unlocked activation (i.e., more negative compared with Baseline).</p

Schematic diagram of experimental procedure and topological cortical regions.

<p><b>A:</b> Experimental procedure with red wafer indicated the correct response button was illustrated. ISI denoted interstimulus interval (i.e., 800 ms with black crosshair “+” in this study). <b>B:</b> Grouped thirteen cortical regions with their abbreviations, (i.e., PF (prefrontal area, Fp1 and Fp2), LF (left frontal area, F3 and F7), MF (middle frontal area, Fz), RF (right frontal area, F4 and F8), LT (left temporal area, T7), LC (left central area, FC1, FC5, CP1, CP5 and C3), MC (middle central area, Cz), RC (right central area, FC2, FC6, CP2, CP6 and C4), RT (right temporal area, T8), LP (left parietal area, P3 and P7), MP (middle parietal area, Pz), RP (right parietal area, P4 and P8), O (occipital area, O1, Oz and O2)) were shown.</p

Demography of stroke patients.

<p>M =  Male.</p><p>F =  Female.</p><p>NIHSS  =  National Institute of Health Stroke Scale.</p

Brain mappings of beta-ERD for control group at different angles in response sub-stage.

<p>Higher ERD in larger brain areas particularly in right hemisphere was found with larger angles in control subjects (i.e., more brain areas with deeper blue color represented higher phase-unlocked cortical activation).</p