Journal: Molecular Neurodegeneration

Article Title: Modulating the catalytic activity of AMPK has neuroprotective effects against α-synuclein toxicity

doi: 10.1186/s13024-017-0220-x

Figure Lengend Snippet: AMPKα reduces lysosome accumulation in primary cortical neurons overexpressing α-syn. a Primary cortical neurons transduced with α-syn-encoding vector (or non-coding vector as control) are co-transduced with AMPKα expressing vectors. Protein analysis by western blotting shows LC3-I, LC3-II and actin. Signal intensity is enhanced in the lower panel to show LC3-II immunoreactivity. b Relative quantification of LC3-I levels normalized to actin. AMPKα overexpression decreases LC3-I level in neurons expressing human α-syn. c Relative quantification of LC3-II levels normalized to actin. Note a significant decrease in LC3-II levels in neurons overexpressing AMPKα in the absence of human α-syn. d Representative image of a cortical neuron expressing the fusion protein LC3B-EGFP-mCherry. Note the presence of yellow (autophagosomes, yellow arrow) and red vesicles (autolysosomes, red arrow) in the cytosol. Scale bar: 10 μm. e Quantification of the average number of autophagosomes and autolysosomes per cell, normalized to cytosol area. Note the reduction in the total number of autophagic vesicles, in neurons overexpressing either AMPKα1 or α2. Overexpression of α-syn increases the number of autolysosomes in neuronal cells. AMPKα reduces the number of autolysosomes, even in presence of α-syn. Statistical analyses : (b, c) repeated measures two-way ANOVA with Fisher‘s LSD post hoc test; n = 3 per condition. (e) two-way factorial ANOVA with Newman-Keuls post hoc test; b, c: for each condition n = 22–30 neurons from 3 separately infected wells; * P < 0.05; ** P < 0.01; *** P < 0.001

Article Snippet: Neurons expressing the AAV-encoded LC3B-mCherry-EGFP probe were analyzed using Zeiss LSM 700 confocal microscope at 63× magnification.

Techniques: Transduction, Plasmid Preparation, Expressing, Western Blot, Over Expression, Infection