Extended Data Fig. 2: AAV targeting, axon tracing and axon quantification. | Nature

Extended Data Fig. 2: AAV targeting, axon tracing and axon quantification.

From: Required growth facilitators propel axon regeneration across complete spinal cord injury

Extended Data Fig. 2

a, AAV targeting of green fluorescent protein (GFP) to propriospinal neurons. Multi-fluorescent, survey (left) and detail (right, boxed area) confocal images of horizontal section through mouse grey (gm) and white (wm) matter. Essentially all NeuN+ propriospinal neurons targeted with AAV express GFP. b, Multi-fluorescent, orthogonal 3D confocal images show that AAV-targeted propriospinal neurons express GDNFR. c, Multi-fluorescent, survey images show tract-tracing of propriospinal axons using biotinylated dextran amine (BDA) in tiled confocal scans of horizontal section from uninjured mouse. Hatched area indicates densely labelled location of BDA injections. d, e, Multiple channel fluorescent images compare BDA-labelled propriospinal axons and immunohistochemically stained serotonin (5HT) axons in mice after SCI + AAV-OIC + 1D + FGF + EGF + GDNF. d, Survey and orthogonal 3D confocal detail from an area proximal to the SCI lesion shows a complete lack of overlap of BDA-labelling and 5HT immunohistochemistry, indicating that BDA-tracing did not label 5HT axons of passage. e, Survey images of the same field examined with different filters show BDA-labelled propriospinal axons (bottom image) regrowing robustly past the astrocyte scar proximal border and through the non-neural lesion core; by contrast, 5HT axons (top and bottom image) did not regrow into or through the lesion core. f, Open field hindlimb locomotor score at various times after SCI assessed using a 6-point scale in which 5 is normal walking and 0 is no movement of any kind. Data are mean ± s.e.m., n = 6 mice per group.

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