Single cell and ensemble activity of lumbar intermediate and ventral horn interneurons in the spinal air-stepping cat

We explored the relationship between population interneuronal network activation and motor output in the adult, in-vivo, air stepping, spinal cat. By simultaneously measuring the activity of large numbers of spinal interneurons, we explored ensembles of coherently firing interneurons and their relation to motor output. Additionally, the networks were analyzed in relation to their spatial distribution along the lumbar enlargement for evidence of localized groups driving particular phases of the locomotor step cycle. We simultaneously recorded hindlimb EMG activity during stepping and extracellular signals from 128 channels across two polytrodes inserted within lamina V-VII of two separate lumbar segments. Results indicated that spinal interneurons participate in one of two ensembles that are highly correlated with the flexor or the extensor muscle bursts during stepping. Interestingly, less than half of the isolated single units were significantly unimodally tuned during the step cycle while >97% of the single units of the ensembles were significantly correlated with muscle activity. These results show the importance of population scale analysis in neural studies of behavior as there is a much greater correlation between muscle activity and ensemble firing than between muscle activity and individual neurons. Finally, we show that there is no correlation between interneurons' rostrocaudal locations within the lumbar enlargement and their preferred phase of firing or ensemble participation. These findings indicate that spinal interneurons of lamina V-VII encoding for different phases of the locomotor cycle are spread throughout the lumbar enlargement in the adult spinal cord.

RNA Profiling of Mouse Ependymal Cells after Spinal Cord Injury Identifies the Oncostatin Pathway as a Potential Key Regulator of Spinal Cord Stem Cell Fate

Ependymal cells reside in the adult spinal cord and display stem cell properties in vitro. They proliferate after spinal cord injury and produce neurons in lower vertebrates but predominantly astrocytes in mammals. The mechanisms underlying this glial-biased differentiation remain ill-defined. We addressed this issue by generating a molecular resource through RNA profiling of ependymal cells before and after injury. We found that these cells activate STAT3 and ERK/MAPK signaling post injury and downregulate cilia-associated genes and FOXJ1, a central transcription factor in ciliogenesis. Conversely, they upregulate 510 genes, seven of them more than 20-fold, namely Crym, Ecm1, Ifi202b, Nupr1, Rbp1, Thbs2 and Osmr—the receptor for oncostatin, a microglia-specific cytokine which too is strongly upregulated after injury. We studied the regulation and role of Osmr using neurospheres derived from the adult spinal cord. We found that oncostatin induced strong Osmr and p-STAT3 expression in these cells which is associated with reduction of proliferation and promotion of astrocytic versus oligodendrocytic differentiation. Microglial cells are apposed to ependymal cells in vivo and co-culture experiments showed that these cells upregulate Osmr in neurosphere cultures. Collectively, these results support the notion that microglial cells and Osmr/Oncostatin pathway may regulate the astrocytic fate of ependymal cells in spinal cord injury.

Adult Spinal Cord Injury without Major Bone Injury: Effects of Surgical Decompression and Predictors of Neurological Outcomes in American Spinal Injury Association Impairment Scale A, B, or C

The cervical spine can be injured even in the absence of radiographic abnormality, and the best surgical treatment for adult spinal cord injury without bone injury is debated. The aim of this study was to retrospectively investigate the effect of surgical decompression for severe adult spinal cord injury without major bone injury and to establish predictors of good neurological outcome. We analyzed 11 patients who underwent surgical decompression in severe adult spinal cord injury without major bone injury patients classified as American Spinal Injury Association Impairment Scale (AIS) grade A, B, or C. Neurological assessments were performed using AIS at preoperative and postoperative 1-year follow-up. Radiological evaluations were performed using cervical magnetic resonance imaging (MRI) at preoperative. Seven cases were classified as AIS grade A; two cases as AIS grade B; and two cases as AIS grade C. Five of 11 (45.5%) patients showed improved neurological grade 1-year postoperatively. Intramedullary lesion length (IMLL) (p = 0.047) and compression rate (p = 0.045) had the most powerful effect on AIS grade conversion. This study shows that the fate of the injured spinal cord is determined at the time of the injury, but adequate decompression may have limited contribution to the recovery of neurological function. Compression rate and IMLL on MRI can be used as a predictor of neurological recovery.

PATH-23. ADULT SPINAL CORD ASTROBLASTOMA WITH EWSR1-BEND2 FUSION

The most recurrent fusion of CNS high-grade neuroepithelial tumor with MN1alteration(HGNET-MN1) is MN1- BEN Domain Containing 2(BEND2) fusion. Recently, there was a report of a 3-month-old boy with spinal astroblastoma, classified as CNS HGNET-MN1 by DKFZ methylation classification but positive for EWSR1-BEND2 fusion(Yamasaki, 2019). Here, we report a 36-year old man with a spinal cord astroblastoma with EWSR1 alternation. The patient presented with back pain, gait disorder and dysesthesia in lower extremities and trunk was referred to our hospital. MRI showed intramedullary tumor in Th3-5 level, displaying low-intensity on T1 weighted image, high-intensity on T2 weighted image, and homogeneous gadolinium enhancement. Partial removal was performed with the laminectomy. The tumor extended to extramedullary and its boundary was unclear. Histological examinations showed the epithelium-like tumor cells with eosinophilic cytoplasm with high cellularity palisade, intracellar fibrosis, and mitosis. Immunohistochemical staining showed positive for Olig2, GFAP, EMA, SSTR2, S-100, but negative for p53, PgRAE1/AE3. The tumor was diagnosed as astroblastoma, and was classified as HGNET-MN1 by the DKFZ methylation classifier. However, the MN1 alternation was not detected by fluorescence in situ hybridization, instead EWSR1 and BEND2 alternations which suggested EWSR1-BEND2 fusion were detected. After radiation therapy of 54Gy/30fr with bevacizumab and temozolomide, the residual tumor reduced the size and his symptoms improved. This case provides evidence that EWSR1-BEND2 fusion is recurrent in HGNET-MN1 and, as previously reported, suggests the importance of BEND2 in this entity. These two cases suggested that it may be the BEND2 alteration that biologically defines the HGNET-MN1 subclass rather than MN1.

Single-cell transcriptomic analysis of the adult mouse spinal cord

The spinal cord is a fascinating structure responsible for coordinating all movement in vertebrates. Spinal motor neurons control the activity of virtually every organ and muscle throughout the body by transmitting signals that originate in the spinal cord. These neurons are remarkably heterogeneous in their activity and innervation targets. However, because motor neurons represent only a small fraction of cells within the spinal cord and are difficult to isolate, the full complement of motor neuron subtypes remains unknown. Here we comprehensively describe the molecular heterogeneity of motor neurons within the adult spinal cord. We profiled 43,890 single-nucleus transcriptomes using fluorescence-activated nuclei sorting to enrich for spinal motor neuron nuclei. These data reveal a transcriptional map of the adult mammalian spinal cord and the first unbiased characterization of all transcriptionally distinct autonomic and somatic spinal motor neuron subpopulations. We identify 16 sympathetic motor neuron subtypes that segregate spatially along the spinal cord. Many of these subtypes selectively express specific hormones and receptors, suggesting neuromodulatory signaling within the autonomic nervous system. We describe skeletal motor neuron heterogeneity in the adult spinal cord, revealing numerous novel markers that distinguish alpha and gamma motor neurons—cell populations that are specifically affected in neurodegenerative disease. We also provide evidence for a novel transcriptional subpopulation of skeletal motor neurons. Collectively, these data provide a single-cell transcriptional atlas for investigating motor neuron diversity as well as the cellular and molecular basis of motor neuron function in health and disease.