Arp2/3 is necessary for spine head formation. (A) Mouse hippocampal neurons were transfected with GFP or GFP + p34 siRNA at DIV 10. At DIV 11, the cells were fixed and stained with anti-p34 antibodies (see Fig. S4). Reduced p34 levels resulted in a loss of spine heads. Bars, 5 µm. (B) Dendritic protrusion density and length of p34 siRNA-transfected neurons was analyzed with NeuronIQ software (Cheng et al., 2007). Dendritic protrusion density was decreased and dendritic protrusion length increased in p34 siRNA-transfected cells compared with wild-type cells. See Tables I and II for numerical data. (C) Dendritic protrusion morphology analysis revealed a clear reduction of thin, mushroom, and stubby spines. See Tables I and II for numerical data. (D) Mouse hippocampal neurons were transfected with GFP (wt) or inactive Rif + Scar1-WA (inact Rif + WA) constructs at DIV 10. At DIV 11, cells were fixed and stained with anti-myc antibodies. Inhibition of Arp2/3 and Rif induced spine head loss. Bars, 5 µm. (E) Dendritic protrusion density was reduced, whereas mean dendritic protrusion length was not affected in cells expressing inactive Rif and Scar1-WA. See Table IV for numerical data. (F) Morphology analysis revealed a significant reduction in thin, mushroom, and stubby spines in cells expressing inactive Rif and Scar1-WA. See Tables I and II for numerical data. Graphs represent mean ± SEM.