Accumulation of autophagosomes and motor neuron degeneration in the dnc-1(RNAi) worms.

(A, B) Representative kymographs of Lgg1::DsRed in the ventral nerve cord from the control(RNAi (A) and dnc-1(RNAi) (B) worms derived from time-lapse images. Vertical lines represent stationary/docked Lgg1 puncta, while the oblique lines (labeled with arrowheads) represent the tracks of moving Lgg1 puncta. The slope of this track is an indicator of velocity. (C) The number of Lgg1 puncta within a single kymograph image. (D, E) Quantitative analyses of the mobility of puncta. The number of puncta that moved more than 2 μm was counted (D). The ratio of moving puncta was calculated by dividing the number of moving puncta by the total number of puncta (E). (F) The mean velocities of Lgg1 puncta. A total of 20 time-lapse images were analyzed for each strain in C–F. (G) The number of Lgg1 puncta was increased in the dnc-1(RNAi) worms compared with the control(RNAi) worms (n = 15 for each group). (H, I) Accumulation of autophagosomes in the dnc-1(RNAi) worms was correlated with the severity of axonal defasciculation (H) and locomotor function (I) (n = 20 for each analysis). (J–L) Ultrastructural images of ventral motor neurons from the dnc-1(RNAi) worms. Aberrant membranous vesicles including degenerated mitochondria were observed in the cytoplasm (J) and axons (K) (arrows). Autophagosome-like, double membrane vesicles (asterisk in L) were also found in the axons of the dnc-1(RNAi) worms (L). Scale bar = 500 nm (A–C) or 10 μm (D). Statistical analyses were performed using Student's t test (*p<0.05 and **p<0.0001) and Pearson's correlation coefficient in H and I. The error bars are S.E.M.