Fig. 3: Actinification induces highly stable actin filaments that spontaneously disassemble upon stress removal.

a Prior latrunculin A (LatA) prevents actinification (LatA + NMDA), but does not reverse actinification when added for up to 2 h after NMDA (NMDA + LatA). n = 2 independent experiments; Kruskal–Wallis one-way ANOVA (p = 0.0159). b Phalloidin intensity within proximal dendrites of neurons exposed to NMDA for 30 min with LatA for the indicated times. n = 2 independent experiments; Kruskal–Wallis one-way ANOVA (p = 0.3471). c Selected phalloidin-stained dendrites illustrating that NMDA-induced actinification persists after 2 h incubation with LatA (dendrite borders outlined in blue). Scale bar, 4 µm. d Taxol (10 nM, 30 min) neither prevents NMDA-induced actinification, nor induces actinification in absence of NMDA. n = 3 independent experiments; two-way ANOVA [(−)NMDA vs (+) NMDA, F (1, 8) = 75.82; p = 0.0001; (−)taxol vs (+) taxol, F (1, 8) = 0.1954, n.s. p = 0.6702]. e Nocodazole neither prevents NMDA-induced actinification nor induces it in the presence of an NMDAR antagonist cocktail (MK-801, APV, CGP). n = 3 independent experiments; two-way ANOVA [(−) NMDA vs (+) NMDA, F (1, 8) = 28.35; p = 0.0007; (−) nocodazole vs (+) nocodazole, F (1, 8) = 0.2064, n.s. p = 0.6616]. Mann–Whitney test, two-tailed [nocodazole(+) vs nocodazole + NMDAR antagonists (ANT), n.s. p = 0.1]. f Reversibility of actinification after the addition of the NMDA antagonist MK-801 (25 µM) following 5 min NMDA. Total number of neurons per same field (blue circles) quantified to establish that the decrease in actinified neurons was not due to cell death. n = 2 independent experiments. g Time-lapse images collected at the indicated times (hours: minutes) from an actinified dendrite expressing mApple-F-tractin to monitor live the F-actin reorganization following NMDA stress removal (at t = 0) via MK-801 addition. Note the clearing of the accumulated shaft F-actin, and the re-emergence of F-actin in spine-like protrusions. Scale bar, 8 µm. All graphs: data presented as mean ± SEM, and source data are provided as source data file.