Fig. 4.

E2-mediated increased actin polymerization is blocked by calpain inhibition in cultured neurons and acute hippocampal slices. (A) Cultured cortical neurons (DIV 14) were pretreated with or without the calpain inhibitor, calpeptin (10 μM) for 20 min before adding E2 (10 nM) for 5 min. At the end of incubation, cultures were subjected to the phalloidin fluorescent enhancement assay described under Materials and Methods. (B) Acute hippocampal slices (200-μm thick) were prepared from young male rats (2–4-month-old); area CA1 was dissected out and pretreated with or without a cell permeable calpain inhibitor, calpeptin (10 μM) for 20 min before adding vehicle of E2 (10 nM) for 5 min. At the end of incubation, actin polymerization was analyzed as described under Material and Methods with the rhodamine-phalloidin fluorescent enhancement assay. Results of the Alexa Fluor594-phalloidin fluorescence were normalized (subtraction of control fluorescence signal) and expressed as percentage of values found in untreated control slices and represent means ± SEM of 10 experiments. *, P < 0.05 (ANOVA followed by Bonferroni test).