FIGURE 1. Estrogen Induces Phosphorylation of CaMKII, ERK, and GluR1 in Primary Neuronal Culture.

E2 (10 nM) temporally regulated the phosphorylation of CaMKII, ERK, and GluR1 in primary cortical neurons. A) Phosphorylation was seen within 5 min and was maximum at 20 min for CaMKII (592 ± 183%), while B) ERK phosphorylation was seen within 5 min but was maximum at 30 min (1827 ± 73%). E2 also induced phosphorylation of CaMKII and ERK in the cell body and dendritic extensions of primary cortical neurons. Cortical neurons untreated or treated with E2 (10 nM) for either 20 min or 30 min were labeled with C) phospho-CaMKII or D) phospho-ERK, respectively, and visualized by fluorescence microscopy. E) E2 (10 nM) temporally regulated the phosphorylation of GluR1 in primary cortical neurons. GluR1 was phosphorylated at 30 min but was maximal at 60 min (403 ± 88%). F) Primary cortical neurons were treated with E2 (10 nM) in the presence or absence of CaMKII inhibitor (KN-93, 5 µM), MEK inhibitor (UO126, 10 µM), and PI3K inhibitor (LY29402, 10 µM) for 30 and 60 min. Induction of phosphorylation of GluR1 at serine 831 was inhibited by KN-93 (169 ± 19%) but not U0126 (442 ± 40%) and LY29402 (401 ± 40%) compared to 60 min vehicle-treated (DMSO) control. Data are mean ± SD. *, p<0.05, **, p<0.01, ***, p<0.001, versus time 0 or 30 min or groups connected by bars as determined by one-way ANOVA followed by Newman-Keuls Multiple Comparison Test or two-way ANOVA followed by Bonferroni Test, n=3 (3 independent experiments).