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RNAseq of motor neurons with differential vulnerability to ALS isolated from the SOD1G93A mouse model of ALS

ABSTRACT: ALS is characterised by thedegeneration of somaticmotor neurons (MNs), that control voluntary muscle, while visceral motor neurons remain preserved. Some somatic motor neurons are also relatively resilient to degeneration, including oculomotor and trochlear motor neurons that regulate eye movement.In this study we aim to investigate the longitudinal transcriptional dynamics of resistant and vulnerable MN populations in response to the SOD1G93A ALS-causative mutation. We have conducted bulk RNA sequencing of MNs isolated from the SOD1G93A mouse at presymptomatic (p56) and symptomatic (p112) stages using laser capture microdissection followed by Smart-seq2 RNA sequencing. Oculomotor and trochlear (CN3/4) MN, and visceral MNs from the vagus nucleus (CN10) represent resilient neuron groups that were analysed. MNs from the lumbar spinal cord (SC) and hypoglossal (CN12) nucleus were utilised as two vulnerable populations.

ORGANISM(S): Mus musculus

PROVIDER: GSE244538 | GEO | 2025/07/07

REPOSITORIES: GEO

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Project description:ALS is a uniformly fatal neurodegenerative disease in which motor neurons in the spinal cord and brain stem are selectively lost. Individual motor - groups of motor neurons innervating single muscles - show widely varying degrees of disease resistance: in the final stages of ALS, nearly all voluntary movement is lost but eye movement and eliminative and sexual functions remain relatively unimpaired. These functions are controlled by motor neurons of the oculomotor (III), trochlear (IV) and abducens (VI) nuclei in the midbrain and brainstem, and by Onuf's nucleus in the lumbosacral spinal cord, respectively. Correspondingly, in ALS autopsies the oculomotor and Onuf's nuclei are almost completely preserved. We used microarray profiling of isolated wildtype mouse motor neurons to identify genes whose expression was characteristic of both oculomotor and Onuf's nuclei but not of vulnerable lumbar spinal neurons, or vice versa. Three wild-type C57BL/6J P7 male animals were perfused with 30% sucrose, lumbosacral spinal cord and midbrain regions were rapidly recovered, embedded in OCT compound, and frozen in liquid nitrogen. 12 um-thick cryosections were mounted on RNAse-free, PEN-foil covered glass slides (Zeiss), fixed for 2 min in 100% EtOH, rinsed in 50% EtOH, stained with 1% cresyl violet for 2 min, rinsed with 50% EtOH, dehydrated in graded solutions of ethanol and air dried prior to LCM using PALM Microbeam system (Zeiss). From each animal, ~200 DL, L5, and oculomotor motor neurons were collected directly in lysis buffer. RNA was purified using Absolutely RNA, NanoPrep kit (Stratagene, La Jolla, CA). RNA integrity was assessed on the Bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA). At least 1.5 ng of purified RNA was the starting material used in the WT-Ovation Pico RNA Amplification System (Nugen, San Carlos,CA) with the FL-Ovation cDNA Biotin Module V2 (Nugen) to generate labeled probe. 10 ug of biotinylated cRNA from three independent samples for each motor neuron group isolated by LCM was hybridized to on Affymetrix (Santa Clara, CA) Mouse Genome 430 2.0 Arrays. Gene ontology and pathway analysis was performed using DAVID Bioinformatics Resources 6.7 (National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD).

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RNAseq of motor neurons with differential vulnerability to ALS isolated from the SOD1G93A mouse model of ALS

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