FIGURE 5.

BDNF increases LC3 puncta number through both mTOR-dependent and -independent pathways. A, immunofluorescence images of neurons after 24-h survival treatment with and without Baf A1. Green, MAP2; red, LC3; blue, DAPI. The black and white images were obtained by automated LC3 puncta identification and analysis. rap, rapamycin. B, quantification of LC3 puncta number after 24-h treatment. C, quantification of individual LC3 puncta size. D, LC3 puncta flux calculated by comparison of total puncta load with or without Baf A1. Baf A1 (100 nm) was used for the last 3 h of treatment. Data are from six slides, with 10 neurons analyzed per group per slide. E, neurons transfected with p70S6K mutants were stained for LC3 and analyzed by Volocity; red, LC3; green, HA; blue, DAPI. F, quantification of LC3 puncta number from four experiments for a total of 70 WT neurons, 38 ΔCT neurons, and 30 F5A neurons in withdrawal medium, and 67 WT neurons, 42 ΔCT neurons, and 33 F5A neurons in BDNF-containing medium. For all graphs,* is compared with control; # is compared with BDNF; *, or #, p < 0.05; **, p < 0.01; scale bars, 20 μm.