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Unravelling the cellular pathology leading to neurodegeneration in motor neuron disease
thesisposted on 2023-05-26, 17:38 authored by Anna KingAnna King
Despite advances in amyotrophic lateral sclerosis (ALS) research, important questions remain to be answered. Many degenerative processes have been attributed to the disease, but how these result in the selective degeneration of motor neurons and their links to observed pathological changes is not yet known. The subject of this thesis has been to investigate the relationship between the pathogenic factors in ALS, the neuroaxonal alterations to the motor neuron cytoskeleton and their selective degeneration. There is substantial evidence to implicate the cell selective vulnerability of motor neurons to excitotoxocity in the pathogenesis of ALS. However, the role of the different glutamate receptor subunits is still unknown. This thesis has used immunocytochemical techniques to investigate the cell type specific changes in glutamate receptor localisation and expression in relation to susceptibility to excitotoxicity in a cortical cell culture model. Vulnerability to N-methyl-D-aspartate (NMDA) receptor-mediated excitotoxicity occurred prior to their localisation in apposition to pre-synaptic terminals, indicating that excitotoxicity can be mediated by extrasynaptic receptor puncta. However, vulnerability to alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptor-mediated excitotoxicity was related to localisation ofreceptors at synapses and in spines. Additionally, a subclass of neurons was identified with a specific compliment of receptor subunits, indicating the presence of calcium permeable AMP A receptors and suggesting vulnerability to excitotoxicity. Mouse models of neurofilament disruption have demonstrated the importance of neurofilament pathology in ALS. However, the sequence of cytoskeletal changes in ALS is yet to be determined. This thesis has used an immunohistochemical approach to investigate the time course of cytoskeletal alterations in the G93A mouse model of ALS. Accumulations of neurofilament proteins occurred in the spinal cord prior to the development of symptoms, and this pathology was followed by neurofilament changes in the descending white matter tracts, indicative of upper motor neuron involvement. To investigate the causes of cytoskeletal disruption in ALS, this thesis has used a novel cell culture model of spinal motor and cortical neurons grown on an astrocyte feeder layer. Spinal motor neurons, but not cortical neurons, exposed to excitotoxins developed accumulations of neurofilament proteins in the distal portions of their axons, which showed a strong resemblance to the spinal neurofilament disruption seen in ALS. Additionally, a proximal axonopathy was induced in spinal motor neurons grown on mixed glial feeder layers, and ageing the feeder layer prior to plating the neurons exacerbated this pathology. These culture models are the first to replicate the neurofilament axonopathy of ALS and are likely to serve as useful tools for further investigations into the mechanisms of motor neuron degeneration and for the screening of potential therapeutic agents.
Rights statementCopyright 2008 the author - The University is continuing to endeavour to trace the copyright owner(s) and in the meantime this item has been reproduced here in good faith. We would be pleased to hear from the copyright owner(s). Chapter 3 appears to be the equivalent of the peer reviewed version of the following article: King, A., Chung, R., Vickers, J., Dickson, T., 2006. Localization of glutamate receptors in developing cortical neurons in culture and relationship to susceptibility to excitotoxicity, Journal of comparative neurology, 498(2), 277-294, which has been published in final form at http://dx.doi.org/10.1002/cne.21053. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions Chapter 5 appears to be the equivalent of the peer reviewed version of the following article: King, A., Dickson, T., Blizzard, C., Foster, S., Chung, R., West, A., Chuah, M., Vickers, J., 2007, Excitotoxicity mediated by non‚ÄövÑv™NMDA receptors causes distal axonopathy in long‚ÄövÑv™term cultured spinal motor neurons. European journal of neuroscience, 26(8) 2151-2159, which has been published in final form at https://doi.org/10.1111/j.1460-9568.2007.05845.x. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions