Diverse transcriptional alterations in V2a propriospinal neurons following spinal cord injury

Propriospinal neurons play crucial roles in recovery of function after spinal cord injury. The V2a class in particular contributes significantly to recovery of locomotor and respiratory function after injury in animal models. However, V2a are a diverse class with different properties and gene expression patterns, suggesting that V2a subtypes might have disparate responses to injury. Here, we used single nucleus RNA sequencing of enriched cervical V2a neurons from adult female mice to identify specific markers of 10 V2a subtypes. We found differences in medial-lateral location as well as rostral-caudal distribution of V2a belonging to specific clusters in the adult cord. Although adult V2a can be identified as belonging to N- and Z-group divisions similarly to neonatal V2a, there is a not a 1:1 correlation between most neonatal and adult clusters. We assessed changes in gene expression in each V2a subtype 5 days following a C2 hemisection injury. Remarkably, three subtypes of V2a neurons were found to be nearly absent ipsilateral and contralateral to injury, demonstrating that some V2a neurons are more vulnerable to changes in identity or cell loss than others. Further, the remaining V2a subtypes exhibited distinctive transcriptional alterations indicating a potential rewiring of cellular interactions within the spinal cord microenvironment. Gene expression changes common to most V2a subtypes included an upregulation of RNA splicing factors. This study reveals the diverse responses of propriospinal neurons to injury and provides a foundation for understanding changes in propriospinal neurons that may lead to adaptive (or maladaptive) changes in circuit function following injury. Significance statement This study is the first to describe markers of adult V2a propriospinal neuron subtypes and their relationship to developmental V2a neuron subtypes. In addition, we characterize the diverse responses of different V2a neurons to cervical spinal cord injury, including the loss of entire V2a subtypes. These analyses nominate new cellular targets for altering propriospinal neuron function, plasticity and regeneration following traumatic injury.