Molecular mechanisms underlying the memory-enhancing effects of estradiol

Abstract

This article is part of a Special Issue "Estradiol and cognition". Since the publication of the 1998 special issue of Hormones and Behavior on estrogens and cognition, substantial progress has been made towards understanding the molecular mechanisms through which 17β-estradiol (E2) regulates hippocampal plasticity and memory. Recent research has demonstrated that rapid effects of E2 on hippocampal cell signaling, epigenetic processes, and local protein synthesis are necessary for E2 to facilitate the consolidation of object recognition and spatial memories in ovariectomized female rodents. These effects appear to be mediated by non-classical actions of the intracellular estrogen receptors ERα and ERβ, and possibly by membrane-bound ERs such as the G-protein-coupled estrogen receptor (GPER). New findings also suggest a key role of hippocampally-synthesized E2 in regulating hippocampal memory formation. The present review discusses these findings in detail and suggests avenues for future study.

Keywords: Cell signaling; DNA methylation; ERK; Epigenetic; Estrogen; Estrogen receptor; GPER; Hippocampus; Histone acetylation; mTOR.

Figure 1

Classical (genomic) and non-classical (non-genomic) mechanisms of E₂ action. In the classical mechanism (left), E₂ binds to ERα and ERβ in the cytoplasm, and then the E₂-ER complex translocates into the nucleus and binds to an estrogen response element (ERE) on the DNA. Together with histone acetyltransferases (HAT) and other co-regulators (Co), the ERs facilitate gene transcription. Non-classical mechanisms (right) involve action at or near the plasma membrane. ERα and ERβ in the dorsal hippocampus interact with metabotropic glutamate receptor 1a (mGluR1a) to rapidly activate extracellular signal-regulated kinase (ERK) cell signaling, which triggers epigenetic alterations such as histone acetylation, local protein synthesis via the mammalian target of rapamycin (mTOR) cell-signaling pathway, and gene expression via the transcription factor cAMP response element binding protein (CREB). NMDA receptor activation in the dorsal hippocampus is also necessary for E₂ to activate ERK. G-protein-coupled estrogen receptor (GPER) rapidly activates c-Jun N-terminal kinase (JNK) cell signaling in the dorsal hippocampus, although E₂ does not appear to mediate this effect. Effects of GPER activation on epigenetic processes, gene expression, and protein translation are not yet known.

Figure 2

Schematic illustration of the non-classical mechanisms required for E₂ and ERs to enhance hippocampal memory consolidation. Phosphorylation of the p42 isoform of ERK is necessary for E₂ to enhance object recognition memory consolidation. This phosphorylation is triggered by numerous upstream events including interactions between mGluR1a and the canonical ERs (ERα and ERβ), and activation of NMDA receptors, protein kinase A (PKA), and phosphatidylinositol-3-kinase (PI3K). E₂-induced phosphorylation of ERK, PI3K, and Akt elicits mTOR signaling, promoting local protein synthesis. E₂-activated ERK also transduces into the nucleus to phosphorylate the transcription factor CREB. Activation of ERK is also necessary for E₂ to increase histone H3 acetylation; E₂ increases H3 acetylation at the pII and pIV promoters of the Bdnf gene. DNA methylation is also necessary for E₂ to enhance memory consolidation, although the specific genes methylated are unknown. Finally, GPER enhances memory consolidation by activating JNK, which facilitates gene expression via transcription factors such as ATF2.

Figure 3

Dorsal hippocampal ERK activation is necessary for E₂ to enhance object recognition memory consolidation. (A) Illustration of the object recognition protocol used in our laboratory. During training, mice accumulate 30 seconds of time exploring two identical objects. Immediately after training, mice receive an infusion of E₂ or other drugs into the dorsal hippocampus or dorsal third ventricle. Forty-eight hours later, mice accumulate 30 seconds with a novel object and an object identical to that explored during testing (familiar). Mice that remember the familiar object spend significantly more time than chance (15 sec) with the novel object. Adapted with permission from (Fortress and Frick, 2014; Frick, 2013). (B) ERK activation is necessary for E₂ to enhance object recognition memory consolidation in ovariectomized 8–12 week-old C57BL/6 mice. Forty-eight hours after training, mice infused with 10 μg E₂, but not vehicle, into the dorsal third ventricle spend significantly more time than chance (dashed line at 15 sec) with the novel object (*p < 0.05), demonstrating enhanced object recognition. This effect is blocked by dorsal hippocampal infusion of the ERK activation inhibitor U0126 (0.5 μg/hemisphere), which has no detrimental effect on memory on its own at this dose (Fernandez et al., 2008). Error bars represent the mean ± standard error of the mean (SEM). Reprinted with permission from (Zhao et al., 2010).

Figure 4

Dorsal hippocampal mTOR activation is necessary for E₂ to enhance object recognition memory consolidation. The phosphorylation of S6K (A), 4E-BP1 (B), and p42 ERK (C) were significantly increased in young ovariectomized mice 5 minutes after bilateral dorsal hippocampal infusion of 5 μg/hemisphere E₂ (*p < 0.05 relative to vehicle). This effect was blocked by the ERK inhibitor U0126 (0.5 μg/hemisphere), the PI3K inhibitor LY298002 (0.005 μg/hemisphere), or the mTOR inhibitor rapamycin (0.25 μg/hemisphere) (A–C). (D) All three inhibitors also prevented E₂ from enhancing object recognition memory consolidation, as indicated by the fact that only ice infused with E₂ + vehicle spent more time than chance with the novel object (*p < 0.05). Error bars in all panels represent the mean ± standard error of the mean (SEM). Phosphorylated proteins were normalized to total protein or β-actin. Insets are representative Western blots of phosphorylated and total protein. Adapted with permission from (Fortress et al., 2013).

Figure 5

E₂ facilitates dorsal hippocampal histone acetylation and BDNF protein expression in middle-aged ovariectomized mice. (A) Bilateral dorsal hippocampal infusion of 5 μg/hemisphere E₂ increased H3 acetylation 30 and 60 min later (***p < 0.001, **p < 0.01 relative to vehicle). Acetylated H3 protein was normalized to total H3 protein. Insets are representative Western blots of acetylated and total histone protein. (B,C) Hippocampal infusion of E₂ decreased protein levels of HDAC2 (B) four hours after infusion, and of HDAC3 (C) four and six hours after infusion (**p < 0.01, *p < 0.05 relative to vehicle). (D) Conversely, BDNF protein levels were increased in the dorsal hippocampus four and six hours after bilateral dorsal hippocampal infusion of E₂ (*p < 0.05 relative to vehicle). HDAC and BDNF proteins in panels B–D were normalized to β-actin protein. Insets are representative Western blots of protein and β-actin. (E) Chromatin immunoprecipitation analysis showed that hippocampal E₂ infusion increased acetylation of Bdnf promoters pII and pIV in both young and middle-aged ovariectomized mice (*p < 0.05 relative to age-matched controls). Among vehicle-infused controls, acetylation of pI and pIV was significantly lower in middle-aged females than in young females (#p < 0.05 relative to young vehicle-infused controls), indicating an age-related reduction in acetylation of these promoters. Data were normalized to LINE1 for each sample and then normalized to young vehicle-infused mice for each promoter region and represented as fold of control. For all panels, each bar represents the mean ± SEM. Adapted with permission from (Fortress et al., 2014).

Figure 6

Dorsal hippocampal mGluR1a activation is necessary for ERα and ERβ to enhance object recognition and object placement memory consolidation, and to increase dorsal hippocampal ERK phosphorylation. (A,B) Immediate post-training infusion of PPT (0.2 pg) or DPN (20 pg) into the dorsal third ventricle significantly increased the time spent with the novel object (A) and moved object (B) relative to chance (dashed line at 15 s, **p < 0.01); these effects were blocked by dorsal hippocampal infusion of the mGluR1a antagonist LY367385 (10 pg/hemisphere). Bars represent the mean ± SEM time spent with each object. (C) Five min after infusion, PPT and DPN significantly increased phospho-p42 ERK levels (*p < 0.05; **p < 0.01 relative to vehicle); this effect was completely abolished by dorsal hippocampal infusion of LY367385. Bars represent mean ± SEM % change from vehicle. Reprinted with permission from (Boulware et al., 2013).