Figure 7.

Genistein blocked E2-induced pNR2B and pERK1/2 upregulation, and partially blocked E2-potentiated glutamate-evoked pNR2B and pERK1/2 upregulation in the hippocampus. OVX rats were assigned to 4 groups for western blot analyses: (1) no injections, (2) E2 replacement, (3) genistein treatment, and (4) genistein pretreatment combined with E2 replacement. (A,B) Representative western blots of pNR2B and pERK1/2 after E2 and genistein treatment. Quantification of protein levels was normalized against loading control β-actin and presented as the relative density compared with the control group (OVX). *P < 0.05 vs. the rest groups; n = 5, per group. To examine the effect of genistein on E2 and glutamate-evoked upregulation of pNR2B and pERK1/2, OVX rats were assigned to 3 groups respectively for protein quantitative detection and immunohistofluorescence staining: (1) no injections, (2) E2 replacement combined with glutamate injections, and (3) genistein pretreatment combined with E2 and glutamate injections. (C, D) Representative western blots of pNR2B and pERK1/2 after E2, glutamate and genistein treatment. Quantification of protein levels was normalized against loading control β-actin and presented as the relative density compared with the control group. *P < 0.05, **P < 0.01 vs. control group (OVX); ^#P < 0.05 vs. E2 + Glu group; n = 5, per group. (E,G) Representative fluorescence photomicrographs showing the pNR2B and pERK1/2 positive neurons in the CA3 region of hippocampus after E2, glutamate and genistein treatment; Scale bar = 50 μm. (F,H) The percentages of pNR2B and pERK1/2 positive neurons in the CA3 region of hippocampus after E2, glutamate and genistein treatment; *P < 0.05 comparison among the three groups; n = 4, per group. OVX = ovariectomy; E2 = 17β-estradiol; Glu = glutamate; Gen = genistein; pNR2B = phosphorylated NR2B subunit of the N-methyl-D-aspartate receptor; pERK1/2, phosphorylated (p44/42) mitogen-activated protein kinase; IR, immunoreactive.