Brain-wide analysis of the supraspinal connectome reveals anatomical correlates to functional recovery after spinal injury

Abstract

The supraspinal connectome is essential for normal behavior and homeostasis and consists of a wide range of sensory, motor, and autonomic projections from brain to spinal cord. Extensive work spanning a century has largely mapped the cell bodies of origin, yet their broad distribution and complex spatial relationships present significant challenges to the dissemination and application of this knowledge. Fields that study disruptions of supraspinal projections, for example spinal cord injury, have focused mostly on a handful of major populations that carry motor commands, with only limited consideration of dozens more that provide autonomic or crucial motor modulation. More comprehensive information is essential to understand the functional consequences of different injuries and to better evaluate the efficacy of treatments. Using viral retrograde labeling, 3D imaging, and registration to standard neuro-anatomical atlases we now provide a platform to profile the entire supraspinal connectome by rapidly visualizing and quantifying tens of thousands of supraspinal neurons, each assigned to more than 60 identified regions and nuclei throughout the brains of adult mice. We then use this tool to compare the lumbar versus cervically-projecting connectomes, to profile brain-wide the sensitivity of supraspinal populations to graded spinal injuries, and to correlate locomotor recovery with connectome measurements. To share these insights in an intuitive manner, we present an interactive web-based resource, which aims to spur progress by broadening understanding and analyses of essential but understudied supraspinal populations.

Adult spinal Dmrt3 neurons receive direct somatosensory inputs from ipsi‐ and contralateral primary afferents and from brainstem motor nuclei

“…Previous retrograde tracing experiments, albeit from cervical segments, have identified 27 brain centers projecting to the spinal cord (Nudo & Masterton, 1988 ) and 25 subregions were found from lumbar tracing experiments (Wang et al., 2021 ). In our whole‐brain analysis of Dmrt3‐Cre+ animals, we found traced cells in the motor/somatosensory cortical areas, in the red nuclei, PAG, pontine reticular nuclei, vestibulospinal nuclei, gigantocellular nuclei, and raphe nuclei (Figure 5g ).…”

Single nuclei analyses reveal transcriptional profiles and marker genes for diverse supraspinal populations

“…Starting from transcripts that were highly enriched in each cluster, we adopted a strategy of manual curation that compared ISH data from the Allen Brain Atlas to the known locations of supraspinal neurons. This approach was aided by our recently created 3D atlas of supraspinal populations (Wang et al, 2021), which registers retrogradely labeled cell nuclei to a digital neuroanatomical atlas based on the Allen Brain Atlas. In this way we could systemically examine locations in Allen Brain Atlas images that we knew to harbor supraspinal neurons, in search of putative marker genes from the scRNA-seq data.…”

“…To label supraspinal neurons, adult mice received lumbar injection of AAV2-retro expressing mScarlet fluorophore that was localized to the nucleus by fused histone protein 2B (AAV2-retro-H2B-mSc) (Figure 1A). We have shown previously that this procedure labels tens of thousands of supraspinal neurons distributed through forebrain, midbrain, and hindbrain (Wang et al, 2021(Wang et al, , 2018. Two weeks after injection animals were sacrificed, a sagittal series of brain slices were rapidly prepared and placed in ice-cold artificial cerebrospinal fluid and observed under fluorescent light.…”

“…The supraspinal connectome comprises a diverse and widely distributed set of neurons that project axons to spinal targets and which convey a wide range of motor, autonomic, and sensory modulatory commands. Work spanning more than a century has elucidated the anatomy of supraspinal projections in various model organisms, and in the mouse our recent work has provided a unified description of the supraspinal connectome in three-dimensional space (Wang et al, 2021). In contrast to the state of anatomical and physiological knowledge, much less is understood about the patterns of gene expression that characterize supraspinal neurons or which may distinguish different subtypes.…”