Axonal F-actin dynamics are Formin dependent. (A) Kymographs from an axon transfected with GFP:Utr-CH (to label F-actin) and imaged before (left) and after (right) treatment with the formin-inhibitor SMIFH2. Note dramatic attenuation of the F-actin trails with little effect on F-actin hotspots, suggesting that the nucleation of F-actin at hotspots is spontaneous, and not formin dependent. (B) Similar experiments as in A, except that neurons were treated with the Arp2/3 inhibitor CK-666. Note that addition of CK-666 essentially has no effects on axonal F-actin dynamics. (C–F) Quantification of F-actin dynamics from all experiments. Note that treatment with SMIFH2 attenuates various kinetic behaviors of the F-actin trails, whereas CK-666 has no effects. Also note that SMIFH2 treatment leads to an increase in the hotspot duration, perhaps a compensatory response to attenuated actin trails (also see Results). For SMIFH2 treatment, n = 9 axons and for CK-666 treatment, n = 6 axons were imaged. At least three independent repeats were performed for each condition. All values represent means ± SEM. *, P < 0.05; **, P < 0.01; ***, P < 0.001; paired t test. For detailed statistics, see Table S1. Arrows between images represent passage of time in before/after experiments.