Differential lysosomal motility inhibition phenotypes induced by NudE/L and LIS1 inhibition in neurons. Lysos/LEs in rat cortical neurons showed mostly retrograde transport in uninjected and IgG controls, whereas severe inhibition was observed with 74.1 Ab, NudE/L antibody, and LIS1 antibody–injected cells. (A) Kymographs showing cells 1–6 min after injection. Arrest of several large particles is observed by 6 min in LIS1 antibody–injected cells. (B) Time-lapse images of the same LIS1 antibody–injected cell showing large lysosome arresting at the axonal kink (yellow arrow). Boxed areas show LysoTracker red versus dextran fluorescence versus phase-contrast images of the region shown. (C and D) Long-duration videos of the same LIS1 antibody–injected axon and a NudE/L antibody–injected axon. Note the near-complete arrest of motility in the LIS1 antibody–injected case by 20 min as opposed to continued bidirectional motility in the NudE/L antibody–injected case. Note the severe arrest (yellow arrow) as well as the directional reversal (green arrow) in the first 20 min after injection using anti-NudE/L antibody but the extensive bidirectional movement persistent beyond 20 min (orange arrows in C). Lysos/LEs in LIS1 antibody–injected cells showed progressive linear arrest with distance from the cell body (dotted line) during the first 21 min after injection, which is consistent with the diffusion of the antibody along the axon. Although larger particles remained immobile, smaller particles showed rapid transport (orange arrows in D) and could bypass the larger particles (dotted circles). (E) In control cells, lysos/LEs moved mostly in retrograde direction. Kymographs were generated from as near the cell body as imaging could be performed to up to 140 µm away.