Investigating Basal Autophagic Activity in Brain Regions Associated with Neurodegeneration using In Vivo and Ex Vivo Models

Abstract

Objective: Autophagic maintenance of protein turnover for neuronal homeostasis is of critical importance. Although autophagy dysfunction contributes to neurodegenerative pathology, it remains unclear why certain brain regions are initially targeted compared to others. In Alzheimer's disease, the hippocampus appears to be most severely and initially affected compared to regions such as the cerebellum, which seem to be spared initially and are only targeted during later stages of neurodegeneration. Here we hypothesize that brain-region specific variations in basal autophagic activity may underlie sensitivity to proteotoxicity and contribute towards pathology. We investigated the abundance of key autophagic markers in different regions of the mouse brain to determine whether variations in basal autophagic activity may underlie brain-region susceptibility to neurodegeneration. Methods: Autophagic lysosomal degradation was inhibited using chloroquine in vivo and bafilomycin ex vivo. We investigated the accumulation of LC3-II and p62 protein levels in different regions of the mouse brain following inhibition using western blot analysis, immunofluorescence and micro-computed tomography imaging techniques. Results: Results indicate clear and robust variation of autophagic marker abundance between different regions of the mouse brain, both in our in vivo and ex vivo models. Increased protein levels were particularly observed in the cerebellum compared to the hippocampus region, suggesting distinct and region specific changes in autophagic activity. Conclusion: Functional specificity and metabolic demands of different brain regions may translate into differential autophagic activities, which may vary from one region to the next. Here we report regional variations of key autophagic markers between different regions of the mouse brain when autophagosome degradation was inhibited. These findings indicate enhanced basal autophagic activity in the cerebellum compared to the hippocampus. We therefore conclude that enhanced basal autophagic activity may render certain brain regions better equipped to deal with imbalances in protein degradation and that lower levels of basal autophagic activity may underlie regional susceptibility towards pathological decline.