Moderate MT destabilization selectively blocks the formation of minor neurites. (A–D) Rat hippocampal neurons (3 DIV) cultured in the presence of various low concentrations of nocodazole or 0.02% DMSO (treatment after 1 DIV) stained for Tau-1 (B and D). (A and B) Control neurons have formed one axon (A, arrow) with the typical Tau-1 gradient toward its distal part (B, arrow) and several Tau-1–negative minor neurites (arrowheads). (C and D) The number of minor neurites (arrowhead) is reduced under growth conditions that slightly destabilize MTs, however, neurons are still able to form an axon (arrow). Bars, 20 μm. (E) Nocodazole reduces the number of minor neurites formed in a concentration-dependent manner (neurites per cell: 4.0 ± 0.1, 3.6 ± 0.1, 2.6 ± 0.1, and 2.2 ± 0.2 for 0.02% DMSO and 15, 45, and 75 nM nocodazole, respectively; P < 0.001 by ANOVA; n > 750 neurons from three independent experiments per condition). (F) Neurite extension is not blocked by low concentrations of nocodazole. Total neurite length increased from day 1 to 3 under all conditions (P < 0.001 by t test), though to a lesser extent when treated with 45 or 75 nM nocodazole (increase from 57.5 ± 7.8 to 255.8 ± 9.7, 209.3 ± 8.6, or 130.3 ± 3.6 μm for 0.02% DMSO and 45 or 75 nM nocodazole, respectively; n > 275 neurons per condition from three independent experiments). Data is presented as mean ± SEM.