Inhibition of activity-driven PAK phosphorylation does not block LTP or activity-induced spine F-actin. (A) Hippocampal slices were treated with 0.1 mM of the Rac inhibitor NSC23766 or vehicle (veh) for 1 h and processed for Rac pull-down assay. Representative blot and group mean (±SEM) values show that NSC23766 reduced active Rac1 levels by >30% (*, P < 0.01 vs. vehicle). (B) Slices were treated for 30 min with the Rac inhibitor before TBS and harvested 7 min after TBS for immunofluorescence. NSC23766 did not affect baseline levels of pPAK⁺ PSDs when applied alone but did block the effects of TBS on this measure (**, P < 0.01 vs. control). The right plot shows mean (±SEM) values of the same measures expressed as the difference between TBS + treatment and treatment alone groups (***, P < 0.001 vs. vehicle). (C) Images show in situ F-actin labeling in slices receiving TBS alone or TBS + NSC23766. The plot shows group mean (±SEM) numbers of labeled spines/100 µm³ (*, P < 0.01 vs. ACSF; #, P = 0.01 vs. NSC23766 alone). con, control. (D) The Rac inhibitor was applied to slices for 30 min before TBS (arrow) and remained throughout recording; no effects on LTP were observed in comparison to vehicle control slices. (E) LTP induced by TBS (arrow) in the presence of the Rac inhibitor did not show reversal after 6-min local adenosine treatment (gray shading) beginning 10 min after TBS. (D and E) Dashed lines indicate baseline levels. Bar, 5 µm.