The estrogen receptor (ER) subtypes, ERα and ERβ, modulate numerous signaling cascades in the brain to result in a variety of cell fates including neuronal differentiation. We report here that 17β-estradiol (E2) rapidly stimulates the autophosphorylation of α-Ca 2+ /calmodulin-dependent kinase II (αCaMKII) in immortalized NLT GnRH neurons, primary hippocampal neurons, and Cos7 cells cotransfected with ERα and αCaMKII. The E2-induced αCaMKII autophosphorylation is ERα-and Ca 2+ /calmodulin (CaM)-dependent. Interestingly, the hormone-dependent association of ERα with αCaMKII attenuates the positive effect of E2 on αCaMKII autophosphorylation, suggesting that ERα plays a complex role in modulating αCaMKII activity and may function to fine-tune αCaMKII-triggered signaling events. However, it appears as though the activating signal of E2 dominates the negative effect of ER since there is a clear, positive downstream response to E2-activated αCaMKII; pharmacological inhibitors and RNAi technology show that targets of ERα-mediated αCaMKII signaling include extracellular signal-regulated kinase 1/2 (ERK1/2), cAMP response elementbinding protein (CREB), and microtubule associated protein 2 (MAP2). These findings suggest a novel model for the modulation of αCaMKII signaling by ERα, which provides a molecular link as to how E2 might influence brain function.