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dc.contributor.advisorMahad, Donen
dc.contributor.advisorLyons, Daviden
dc.contributor.authorLicht-Mayer, Simonen
dc.date.accessioned2018-10-17T10:22:28Z
dc.date.available2018-10-17T10:22:28Z
dc.date.issued2018-11-30
dc.identifier.urihttp://hdl.handle.net/1842/33134
dc.description.abstractAxonal degeneration is the major cause of disability in progressive multiple sclerosis (MS). It has been shown that in MS and relevant disease models, demyelinated axons harbor an increased number of mitochondria, which is reflected in bigger stationary sites of mitochondria, increased mitochondrial activity and increased transport speed of mitochondria. This axonal response of mitochondria to demyelination (ARMD) is protective, as there is an increase in energy demand due to the redistribution of sodium channels along the axon following demyelination. However, it remains to be determined how this ARMD is mounted and how mitochondrial dynamics are involved. By using in vivo and in vitro systems we are determined to elucidate the transport and fusion dynamics of the ARMD and where these additional mitochondria come from. Using a cerebellar slice culture system with lysolecithin induced demyelination, we show that the increase in mitochondrial occupancy of the axon already occurs at 24 hours after demyelination and plateaus around 3 to 4 days after demyelination. At 24 hours, there was a steep increase in the mitochondrial numbers inside the axon, which is then followed by an increase in mitochondrial size over the following days. All parameters decrease again over the following days, but remain elevated compared to baseline even 12 days after demyelination. To determine the source of these additional mitochondria and to assess the fusion dynamics within the axon, we used a lentivirus expressing a mitochondrial targeted photoconvertible dye (mEOS2) to label mitochondria in Purkinje cells. The mitochondria that are labelled green in the Purkinje cell axons are then photoconverted to red by illuminating the initial part of the axon with a 405-nm laser and imaged over the following 20 minutes to determine the transport and fusion dynamics. This showed an increased number of mitochondria moving from the cell body into the axon, as well as an increase in retrograde transport of mitochondria in the demyelinated compared to the myelinated axons. Furthermore the size of newly transported mitochondria and their speed was increased in the anterograde direction. Furthermore, the fusion rate of newly transported mitochondria with stationary converted mitochondria was increased in the demyelinated axons compared to myelinated control. These changes can also be observed in unmyelinated axons, as well as axons of cerebellar slices of the dysmyelinating shiverer mutant with or without lysolecithin treatment. The manipulation of mitochondrial dynamics after demyelination with the fission inhibitor mdivi-1 and the ATPase inhibitor oligomycin both showed an increasing or decreasing effect on the mitochondrial parameters after demyelination respectively. The effect on the axonal health after demyelination was detrimental with both of these treatments. Increasing mitochondrial biogenesis with pioglitazone increased axonal mitochondrial parameters, as well as ameliorated axonal damage after demyelination with lysolecithin. As the neuronal cell bodies in MS harbour mitochondrial DNA deletions, which affects their physiology, including energy production efficiency, another aim of this thesis was to model this deficiency in vitro. As it was not possible to model these mitochondrial defects in vitro within the experiments of this thesis, the characterization of a mitochondrial mutant in vivo model was done as a contribution to a greater set of experiments performed by other members of the Mahad lab.en
dc.language.isoen
dc.publisherThe University of Edinburghen
dc.subjectaxonsen
dc.subjectmultiple sclerosisen
dc.subjectdemyelinationen
dc.subjectmitochondrial deficiencyen
dc.subjectaxonal lossen
dc.titleMitochondrial dynamics in demyelinated axons in a cerebellar slice culture systemen
dc.typeThesis or Dissertationen
dc.type.qualificationlevelDoctoralen
dc.type.qualificationnamePhD Doctor of Philosophyen
dc.rights.embargodate2019-11-29
dcterms.accessRightsRestricted Accessen


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