Observations of increased CSF glutamate in ALS, together with findings that motor neurons (MNs) are selectively vulnerable to glutamate receptor mediated (""""""""excitotoxic"""""""") injury support an excitotoxic contribution to MN loss in the disease. Past studies have highlighted factors that may underlie this vulnerability;in comparison to most other neurons, excitotoxic activation of MNs induces greater mitochondrial Ca2+ overload and exceptionally strong reactive oxygen species (ROS) generation. However, while astroglial glutamate transporters account for most glutamate uptake in the CMS,and their damage appears to underlie extracellular glutamate elevations in ALS, the reason for their dysfunction has been unclear. Providing a possible clue, recent studies suggest that this ROS generated within MNs in response to excitotoxic activation may in itself cause disruption of glutamate uptake in surrounding astrocytes. These observations suggest a mechanism that causatively links excitotoxic MN damage with oxidative disruption of glutamate transport, and provide the basis the for a feed forward model of ALS, in which a range of inciting factors could lead into a common disease pathway. The broad aim of this proposal is to use culture and slice models to extend these studies and further examine the hypothesis that ROS generation within MNs contributes to the loss of regional glutamate transport in vivo. Excitotoxic ROS generation within MNs and its ability to penetrate surrounding tissue and disrupt glutamate transport will be examined in dissociated and slice culture models from wild type mice as well as mice harboring superoxide dismutase (SOD) mutations associated with familial forms of ALS (which provide the best animal models of the disease). SOD mutant mice will be used to examine the degree to which oxidative changes and disruption of transport occurs in regions immediately surrounding large ventral horn MNs, as would be predicted if the MNs are the source of the ROS. Finally, the ability of selected pharmacological interventions to decrease these pathological processes will be tested in slice and animal models. It is hoped that these studies will further clarify sequences of events culminating in selective MN loss in ALS, and thereby facilitate development of new treatment strategies.
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