Dynamics of axonal filopodia are reduced in Actα and Actγ knockdown motoneurons. (A) Time-lapse sequences representing axonal filopodia dynamics in motoneurons transduced with actin knockdown lentiviruses coexpressing GFP (Videos 1–4). Arrowheads mark dynamic filopodia. (B) Maximum projection of 120 frames representing alterations in filopodia length over time. (C) Rate of filopodia initiation is decreased by 60% after Actα or Actγ knockdown. (D) Lifetime of Actα-depleted filopodia is decreased compared with control (*, P < 0.010) and Actβ-depleted filopodia (***, P < 0.0008). Shown are mean ± SEM. (E) Graph shows mean rate of filopodia dynamic movement. Rate of filopodia movement is decreased in Actα- and Actγ-depleted neurons. (F) Graph showing mean total changes in filopodia length. Filopodia growth dynamics is reduced after Actα and Actγ knockdown (E and F: ***, P < 0.0001, shActα: n = 4, shActβ: n = 5, shActγ: n = 3, shCtrl: n = 6 for 50 filopodia per group). (G) Motoneurons were stained with phalloidin, and filopodia were counted per micrometer of axon length. Depletion of Actα reduces number of axonal filopodia (*, P < 0.043). Statistical analysis was performed using one-way ANOVA with Dunn’s post-test. Bars, 5 μm.