Fig. 4.

Involvement of the MEK/ERK cascade in fisetin-induced facilitation of LTP in rat hippocampal slices. (A) Effect of fisetin (1 μM, n= 6) on NMDA receptor-mediated fEPSP in Mg²⁺-free medium. The hippocampal slices were exposed to fisetin (black bar) and the NMDA receptor antagonist 2-amino-5-phosphonovalerate (APV, white bar). The NMDA receptor-mediated fEPSP area is expressed as the percentage of the value immediately before addition of fisetin. (A Insets) Representative records 5 min before (Inset 1) and 25 min after (Inset 2) exposure to fisetin (Inset 2) and 10 min after exposure to APV (Inset 3). Fisetin has no effect on NMDA receptor-mediated synaptic responses. (B and C) MEK inhibitors PD98059 and U0126 block fisetin-dependent facilitation of LTP. The hippocampal slices were untreated (n= 12) or exposed to 1 μM fisetin alone (n= 7) or to a weak tetanic stimulation applied at time 0 after a 10-min pretreatment with PD98059 (50 μM, n= 6) or U0126 (20 μM, n= 5). (B) Time course of changes in the fEPSP slope. To compare the data among the groups, the averages of the fEPSP slopes 30–60 min after tetanic stimulation were calculated as an index of LTP magnitude and are shown in C. (C) None: 104.5 ± 4.0%; fisetin: 143.9 ± 11.4%, ∗∗, P < 0.01 vs. none; fisetin + PD98059: 103.9 ± 5.5%, , P < 0.05 vs. fisetin; fisetin + U0126: 104.7 ± 8.3%, , P < 0.05 vs. fisetin (ANOVA followed by Tukey–Kramer test). All data are mean ± SEM. (D and E) Effect of actinomycin D on fisetin-induced facilitation of LTP. The hippocampal slices were untreated (n= 10) or exposed to 1 μM fisetin alone (n= 7) or after a 10-min pretreatment with 40 μM actinomycin D (ACTD, n= 6) and weak tetanic stimulation was applied at time 0. The presentation of the data are as in B and C. ACTD does not block fisetin-dependent facilitation of LTP. [None: 104.6 ± 4.3%; fisetin: 141.9 ± 8.6%, ∗∗, P < 0.01 vs. none; fisetin + ACTD 139.7 ± 13.6%, not significant vs. fisetin (ANOVA followed by Tukey–Kramer test).] All data are the mean ± SEM.