Single-cell transcriptomic analysis of the adult mouse spinal cord reveals molecular diversity of autonomic and skeletal motor neurons

Abstract

The spinal cord is a fascinating structure responsible for coordinating movement in vertebrates. Spinal motor neurons control muscle activity by transmitting signals from the spinal cord to diverse peripheral targets. We profiled 43,890 single-nucleus transcriptomes from the adult mouse spinal cord using fluorescence-activated nuclei sorting to enrich for motor neuron nuclei. We identified 16 sympathetic motor neuron clusters, which are distinguishable by spatial localization and expression of neuromodulatory signaling genes. We found surprising skeletal motor neuron heterogeneity in the adult spinal cord, including transcriptional differences that correlate with electrophysiologically and spatially distinct motor pools. We also provide evidence for a novel transcriptional subpopulation of skeletal motor neuron (γ*). Collectively, these data provide a single-cell transcriptional atlas ( http://spinalcordatlas.org ) for investigating the organizing molecular logic of adult motor neuron diversity, as well as the cellular and molecular basis of motor neuron function in health and disease.

Evidence for glutamine synthetase function in mouse spinal cord oligodendrocytes

“…Furthermore, we found no expression of GS in NeuN + neurons or in IBA1 + microglia at the protein level ( Figure 2D, E ). These findings were consistent with Glul enrichment analysis in a published single nuclei (sn) RNA-seq data set from adult mouse spinal cord (Blum et al, 2021) ( Figure 2F ). We found that Glul was expressed in two main cell populations: mature OL, identified with Mobp and astrocytes (AS), identified with Aqp4 expression.…”

“…Normalized and scaled data setss (featurebarcode matrices) from published mouse spinal cord single-cell RNA sequencing (scRNA- seq) (Blum et al, 2021) and human leukocortical single-nucleus RNA-sequencing (snRNA- seq) (Schirmer et al, 2019) and tissues were obtained (https://cells.ucsc.edu/ and http://spinalcordatlas.org/) and used without any processing. In addition, another mouse CNS scRNA-seq data sets (Marques et al, 2016) was reconstructed using the available count matrix and corresponding metadata (GSE75330).…”

Integrative genetic analysis of the amyotrophic lateral sclerosis spinal cord implicates glial activation and suggests new risk genes

“…New approaches will be needed to disentangle effects of tissue composition from cell-intrinsic changes in state, such as single nucleus RNA-seq (Lake et al 2016) or spatial transcriptomics (Maniatis et al 2019). The relative paucity of motor neurons in the spinal cord is an obstacle to understanding cell-intrinsic effects in motor neurons, so viable strategies include enriching spinal cord samples for motor neurons (Blum et al 2021) or for inducing pluripotent stem cells from ALS patients into motor neurons (Ho et al 2021).…”

Single-cell transcriptomic analysis unveils spinal motor neuron subtype diversity underpinning the water-to-land transition in vertebrates

“…These molecular codes may serve not only as markers, but also provide insights into how precise axon targeting and neural circuit patterning is achieved. This principle seems to be applicable to other neuronal types 81 and adult MNs 26 . Unexpectedly, either by means of trypsin- or papain-based disscociation approaches, we acquired equivalent numbers of MMC (2354 cells) and LMC (2248 cells) MNs, which is inconsistent with the population ratio of brachial LMC to MMC MNs (4:1) in E13.5 mouse embryos 82 .…”

“…Recent studies have utilized single-cell analysis to demonstrate the diversity of cholinergic neurons in the adult spinal cord 26, 27 , yet the embryonic origin of spinal MN diversity has not been determined. We tackled this topic by using Mnx1-GFP transgenic mice and established a robust optimized method to enrich for and characterize spinal MNs.…”