Changes in spine morphology during peripheral neuropathic pain. (A) Development of tactile allodynia (~6 d) and its prolonged maintenance (6 d~) in the injured paw of PSL mice (n = 6). Sham, sham-operated mice (n = 6). (B) Left, schematic of in vivo imaging. Middle, low-magnification z-projection image of the S1 cortex layer 5 pyramidal cell (scale bar, 50 μm) in a sham control mouse. Right, lateral view of the same cell (scale bar, 100 μm). (C) Left, high-resolution time-lapse image of the same dendritic segment shown in B (red box). Scale bar, 3 μm. Right, lengths of the seven spines numbered in the left panel. (D) Representative in vivo time-lapse image of the dendrite taken at PSL+3 d. Note that highly motile thin spines (yellow asterisks) could be seen with high proportion. In addition, a subset of spines exhibited elimination (blue asterisk) and generation (red asterisks) during the 2-h imaging period. (E) Time-lapse image of the dendrite taken at PSL+9 d. (F) Spine motility significantly increased at PSL+3 d (n = 212 spines, 5 mice) and then returned to control (n = 355 spines, 7 mice) level at PSL+9 d (n = 172 spines, 4 mice). ***P < 0.001, one-way ANOVA followed by a Dunnet's multiple comparison test. (G) Proportion of the mushroom (left panel) and thin (right panel) spines significantly decreased and increased, respectively, at PSL+3 d. Such change in spine types was restored at PSL+9 d. *P < 0.05 vs ShamCont or PSL+9 d, one-way ANOVA followed by a Dunnet's multiple comparison test. Middle, representative image showing different types of spines: mushroom (M), stubby (S) and thin (T) spines. Stubby spines were excluded in analysis because they might be in the middle of elimination or formation. Scale bar, 1 μm.