Extended Data Fig. 7: Characterization of neurons in the LH and motor cortex with projections to vGi^Vglut2 neurons.

a, Schematic of the experimental scheme to understand projection inputs on vGi^Vglut2 neurons with spared projections to the lumbar spinal cord. Vglut2^Cre mice underwent a contusion SCI and were allowed to spontaneously recover for one month (n = 3 mice). To understand the inputs onto vGi^Vglut2 neurons with spared projections to the lumbar spinal cord, rAAV2-hSyn-GFP was injected into the lumbar spinal cord bilaterally. AAV-DIO-tdTomato was concomitantly injected into the right LH of these mice, which was allowed to express for 4 weeks. Photomicrographs were acquired with immunohistochemical staining for Homer and vGlut2, in addition to the viral labelling. b, Photomicrographs from each of the three tested mice, demonstrating synaptic-like appositions of LH^Vglut2 projections to vGi^Vglut2 with spared axonal projections to the lumbar spinal cord. Sections have been labelled with immunohistochemistry for the presynaptic marker vGlut2 and the postsynaptic marker Homer. c, Quantification of the number of vGi^Vglut2 neurons with at least one synaptic-like apposition from LH^Vglut2 neurons. d, Schematic of the experimental scheme to label neurons in the motor cortex and LH with monosynaptic projections to vGi^Vglut2 neurons. Vglut2^Cre mice were injected with AAV8-DIO-TC66T-eGFP-oG in the vGi, followed by injections of EnvA G-deleted Rabies-mCherry two weeks later. Photomicrographs demonstrate neurons in the motor cortex and LH with monosynaptic projections to vGi^Vglut2 neurons. Right, quantification of the number of labelled neurons in each region. e, Schematic of the experimental scheme demonstrating injection of AAV5-Con/Fon-ChR2-YFP into the right LH with insertion of an optic fiber and of retro-AAV2-DIO-FLPo into the vGi of Vglut2^Cre mice. Mice underwent a contusion injury at T9, and behavioral recordings were performed at 6 weeks post-SCI. Middle, Representative image demonstrating AAV5-Con/Fon-ChR2-YFP expression in the right LH and position of the optic fiber. Right, Contusion lesion epicenter reconstructions with percent spared tissue. Mean sparing = 25.5%. 3 V: third ventricle. f, As in Extended Data Fig. 6b, for mice following optogenetic activation of LH^Vglut2 neurons with direct projects to the vGi (n ≥ 10 gait cycles per mouse, n = 7 mice per group). Statistics are provided in Supplementary Data 1.