Knocking down tau with lentivirus-shRNA prevents deficits in axonal transport of mitochondria in wild-type (Tau+/+) primary hippocampal neurons caused by exposure to Aβ oligomers. (A) Neurons were transduced with lentivirus expressing anti-tau shRNA and EGFP (Lenti-shTau-GFP; left, monochrome; right, green) on DIV 0 and transfected with a mitochondrial marker (mito-RFP; middle, monochrome; right, red) at DIV 6. Arrowheads indicate mitochondria in axon. Bar, 20 µm. (B) Total tau levels in neurons transduced with Lenti-shTau-GFP (shTau) or a similar construct expressing a scrambled shRNA (shScr) were determined by Western blot analysis with the Tau-5 antibody. Top, representative blot; bottom, quantitation of blot signals. Mean Tau/actin ratios in scrambled shRNA–expressing neurons were arbitrarily defined as 1.0. n = 6–7 wells from four to five experiments per condition at DIV 14. *, P < 0.05 (unpaired t test). (C) The percentage of mitochondria moving anterogradely or retrogradely relative to all mitochondria in the axons of neurons transduced as in A and B before (baseline) and 10–60 min after adding vehicle (Veh) or Aβ1–42 oligomers (final concentration equivalent to 2-µM monomer) to the medium. n = 25–40 axons recorded during three to four independent sessions at 12–14 DIV. ***, P < 0.001 (paired t test, Bonferroni). Data are means ± SEM.
