Hughes, MC; (2007) Neurone and astrocyte response to Aβ25-35: Role in glutathione in neuroprotection. Doctoral thesis , UCL (University College London).

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Abstract

Amyloid beta (AP) is strongly implicated in the pathogenesis of Alzheimer's disease and has been shown to cause oxidative stress and neurone death in vivo and in cell culture models. Astrocytes in cell culture conditions and in vivo appear to be more resistant to A(3 mediated toxicity, but do undergo morphological changes to adopt a stellate "activated" morphology. The experiments presented in this thesis have used the aggregating Ap fragment AJ325-35 to model A(3 toxicity to study why neurones are more vulnerable than astrocytes. Neurones and astrocytes were both shown to generate reactive oxygen species (ROS) in the presence of A025-35 although astrocytes contained higher levels of the antioxidant glutathione (GSH). It was shown that both astrocyte conditioned medium, and the GSH precursor y-glutamylcysteine raised neurone intracellular GSH levels and protected against AP25-35 mediated neurotoxicity to the same degree. In the brain, astrocytes provide neurones with the precursors needed for GSH synthesis. To test whether astrocyte support of neurone GSH synthesis was maintained in the presence of A(325-35, intracellular GSH was measured in both cell types after A(325- 35 treatment. It was shown that intracellular GSH was lowered in neurones but was maintained in astrocytes. The ability of astrocytes to maintain their GSH levels appeared to be dependent on an increase in the activity of glutathione reductase, the enzyme that recycles oxidised glutathione (GSSG) to its reduced form GSH. Furthermore, the amount of GSH released by astrocytes was increased after treatment with AP25-35. Conditioned medium from Ap25-35 treated astrocytes raised neurone intracellular GSH to the same degree, and gave similar neuroprotection as conditioned medium from control astrocytes. A co-culture protocol was developed in which neurones could be treated with A 325-35 and then transferred to co-culture with astrocytes. Astrocytes co-cultured with Ap25-35 treated neurones showed a decrease in intracellular GSH. This suggests that although Ap25-35 does not affect the ability of astrocytes to protect neurones by releasing GSH, signals from damaged neurones could limit the amount of antioxidant support neurone

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