Figure 8.

AAV-labeled Ia afferent axons after SCI in rats. A, Adult rat CNS with injured spinal cord at T8 (red arrow) showing the site of virus injection (yellow arrow) and the regions from where the images in H−K were taken (brackets with corresponding letters). B−D, 3D reconstruction of confocal images from cleared DRG at C3, T9, and L5 levels labelled with AAV8-UbC-GFP. Confocal images of labeled neurons and their axon projections show similar quality between whole cleared DRG (E) and DRG tissue sections (F). Tissue sections from the cervical (H), thoracic (I), and lumbar (J) spinal cord as well from L5 nerve (G) show that this method labels both the central and peripheral branches of the DRG (see also Movie 7). 3D reconstruction of LSFM images from the cleared injury site (K, dorsal view) shows the caudal (left, box 1) and rostral (right, box 2) portions of the lesioned ascending sensory axons disrupted by the injury site as outlined by the cavity (magenta, see also Movie 8). Arrows in K indicate dorsal roots that are still attached to the spinal cord. Arrows in inset 2 indicate afferent collaterals that are present in greater numbers in the rostral regions compared to caudal regions (inset 1). This is quantified in L, which displays the number of axon bundles emanating from the dorsal columns in a similar region of interest drawn rostral and caudal to the injury site in 3D reconstructions. Error bars represent SEM. p = 0.009 using unpaired two-tailed Student’s t test. All images are from rat spinal cord 8-9 weeks after AAV injection and SCI. Scale bars: B−D, 200 μm; E, 40 μm; F, 50 μm; G, 10 μm; H−J, 300 μm; K, 500 μm; Insets 1, 2, 100 μm. Gr, gracile; Cu, cuneate; VI and IX indicate the Rexed laminae.