Function and Mechanisms of Autophagy-Lysosomal Pathway in Traumatic Brain Injury

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 DESCRIPTION (provided by applicant): A major barrier to development of novel treatments against traumatic brain injury (TBI) is incomplete understanding of the mechanisms of injury and recovery. The overall aim of our research is to determine the molecular mechanisms and contribution of defects in the autophagy-lysosomal pathway to secondary injury after TBI, in order to allow future development of rational therapeutic approaches based on its manipulation. The autophagy-lysosomal pathway is an essential component of intracellular degradation and quality control, and plays a protective function against a variety of conditions including neurodegeneration. However, when lysosomal function is compromised autophagy can also contribute to cell death. Accumulation of autophagosomes has been noted after TBI, but its function and mechanisms remain unknown. Additionally, lysosomal function and the efficiency of lysosome-dependent autophagic degradation (flux), has not been assessed after TBI. Therefore, the purpose of this study is to test the hypothesis that early after TBI dysfunction of the autophagy-lysosomal pathway contributes to neuronal cell death and its restoration can promote long-term recovery. Our project will use autophagy-reporter (GFP-LC3) and autophagy-deficient (Beclin1+/-) transgenic mouse models to determine the function of autophagy-lysosomal pathway after TBI and employ both in vivo and in vitro model systems to determine the mechanisms by which disruption of this pathway affects outcomes after TBI. AIM 1 will determine the mechanisms of lysosomal and autophagy dysfunction after TBI. Complimentary in vivo and in vitro approaches, including pharmacological and genetic modulations, will be combined with novel techniques such as ex vivo autophagy flux analysis to test the hypothesis that autophagy flux is impaired early after TBI, reflecting cytoplasmic phospholipase A2 (cPLA2) mediated lysosomal membrane permeabilization (LMP). AIM 2 will determine the functional consequences of enhancing lysosomal function and autophagy flux after TBI. Chemical modulators of autophagy flux and lysosomal biogenesis, Rapamycin and Torin1, will be used in wild type and autophagy deficient mice to test the hypothesis that restoring function of the autophagy-lysosomal pathway after TBI will result in improved functional outcomes. AIM 3 will determine the influence of autophagy-lysosomal pathway on neuronal cell stress and survival after TBI. Pharmacological and genetic manipulation of autophagy, lysosomal function and ER stress in vivo and in vitro will be used to test the hypothesis that impaired lysosomal activity and autophagic clearance exacerbate ER stress resulting in neuronal apoptosis after TBI. Our study will for the first time determine the functio and the mechanisms of autophagy-lysosomal pathway after TBI. Additionally, we will demonstrate that increasing lysosomal function and autophagy flux can improve histological and behavioral outcomes after TBI, thus opening potential novel treatment avenues.