Spinal cord regeneration at single-cell resolution restores walking after paralysis

Although axon regeneration can now be induced experimentally across anatomically complete spinal cord injury (SCI), restoring meaningful function after such injuries has been elusive. This failure contrasts with the spontaneous, naturally occuring repair that restores walking after severe but incomplete SCI. Here, we applied projection-specific and comparative single-nucleus RNA sequencing to uncover the transcriptional phenotype and connectome of neuronal subpopulations involved in natural spinal cord repair. We identified a molecularly defined population of excitatory projection neurons in the thoracic spinal cord that extend axons to the lumbar spinal cord where walking execution centers reside. We show that regrowing axons from these specific neurons across anatomically complete SCI and guiding them to reconnect with their appropriate target region in the lumbar spinal cord restores walking in mice. These results demonstrate that mechanism-based repair strategies that recapitulate the natural topology of molecularly defined neuronal subpopulations can restore neurological functions. Expanding this principle to different classes of neurons across the central nervous system may unlock the framework to achieve complete repair of the injured spinal cord. We used single-nucleus RNA sequencing to profile the low thoracic spinal cord of mice, we devised three experiments to identify  and target the specific neuron populations involved in natural recovery following spinal cord injury. Experiment 1: projection specific sequencing of neurons in the low thoracic spinal cord with projections to walking execution centres in the lumbar spinal cord. Experiment 2: comparative snRNAseq experiment of neurons in the low thoracic spinal cord from uninjured mice and mice that underwent natural spinal cord repair. Experiment 3: projection specific sequencing of regenerating neurons.