The Role of bI-Spectrin in Dendritic Spines Figure 4. The ABD of bI-spectrin prevents acute actin depolymerization. A, Images of GFP fluorescence in spines from CA1 pyramidal neurons expressing actin-GFP alone (upper row), or co-expressing actin-GFP together with the ABD of bI-spectrin (lower row) before and after the application of 5 mm latrunculin-B. Note that fluorescence decreased within 5 minutes of latrunculin treatment in cells expressing GFP, but not in cells co-expressing the spectrin ABD. Scale bar = 2 mm. B, Normalized fluorescence emission of dendritic spines was imaged every 30 seconds before and after the addition of latrunculin-B. Fluorescence decreased rapidly (,3 min) in spines expressing actin-GFP alone and in spines in which the ABD of a- actinin was co-expressed, but not in spines co-expressing the ABD of bI-spectrin and actin-GFP (n = 5 neurons/construct; p,0.001, repeated measures ANOVA). doi:10.1371/journal.pone.0016197.g004 smaller than in cells expressing the bI-spectrin ABD alone (Fig. 8B,C). Spine head diameters in cells co-expressing the bI- spectrin ABD and dominant negative Rac3 or Rac3-DN alone were not significantly different than in cells expressing only GFP, suggesting that Rac3 signaling is required for the effects of the bI- spectrin ABD on spines. If so, then this effect should be mimicked PLoS ONE | www.plosone.org 8 January 2011 | Volume 6 | Issue 1 | e16197