Medial prefrontal cortical estradiol rapidly alters memory system bias in female rats: ultrastructural analysis reveals membrane-associated estrogen receptors as potential mediators

Abstract

High plasma levels of estradiol (E2) are associated with use of a place memory system over a response memory system. We examined whether infusing estradiol into the medial prefrontal cortex (mPFC) or anterior cingulate cortex (AC) could affect memory system bias in female rats. We also examined the ultrastructural distribution of estrogen receptor (ER)-α, ERβ, and G protein-coupled estrogen receptor 1 (GPER1) in the mPFC of female rats as a mechanism for the behavioral effects of E2 in the mPFC. Each rat was infused bilaterally with either E2 (0.13 μg) or vehicle into the mPFC or AC. The majority of E2 mPFC rats used place memory. In contrast, the majority of mPFC vehicle rats and AC E2 or vehicle rats used response memory. These data show that mPFC E2 rapidly biases females to use place memory. Electron microscopic analysis demonstrated that ERα, ERβ, and GPER1 are localized in the mPFC, almost exclusively at extranuclear sites. This is the first time that GPER1 has been localized to the mPFC of rats and the first time that ERα and ERβ have been described at extranuclear sites in the rat mPFC. The majority of receptors were observed on axons and axon terminals, suggesting that estrogens alter presynaptic transmission in the mPFC. This provides a mechanism via which ERs could rapidly alter transmission in the mPFC to alter PFC-dependent behaviors, such as memory system bias. The discrete nature of immunolabeling for these membrane-associated ERs may explain the discrepancy in previous light microscopy studies.

Figure 1.

A, Maze orientation during training trials. B, Maze orientation during probe trial used to determine what memory system is being used to navigate the maze.

Figure 2.

Light microscopic examination of ER localization in the mPFC. A, Light nuclear IR for ERα was observed. B, Dense ERα-IR in the ventromedial hypothalamus. C, Light ERβ-IR was observed in the mPFC. D, Dense nuclear ERβ-IR in the hypothalamus. E, Dense extranuclear GPER1-IR is detected in the neuropil. F, A coronal schematic depicting the area of the mPFC (gray trapezoid) analyzed by EM.

Figure 3.

Percentage of rats that used a place or response strategy after the microinfusions of E2 or vehicle in the medial prefrontal cortex (A) and the anterior cingulate cortex (B). The number of rats per group is shown on the bar, and cannula placements are shown in the image beside the graph.

Figure 4.

Electron micrographs showing examples of profiles containing ERα-IR in the mPFC. These photomicrographs show IR for ERα in (A) an axon (AX) and in a terminal (TER), in which IR is observed at small synaptic vesicles and on the membrane of a mitochondrion (mit) (B), ERα-IR associated with the membrane of a glial cell (GL) (C), ERα-IR filling an axon (AX), and IR for ERα in a dendrite, observed at the plasma membrane and associated with microtubules (D). Bar, 500 nm.

Figure 5.

Electron micrographs showing examples of profiles containing IR for ERβ in the mPFC. These photomicrographs show ERβ in (A) an axon (AX) and in a dendritic spine (SP) that forms an asymmetrical synapse with an unlabeled terminal (uTER) (B), ERβ-IR filling an axon profile (AX) (C), ERβ-IR associated with vesicles and the plasma membrane of an axon terminal (TER), and (D) GPER1 in a glial cell that is in apposition to an unlabeled dendritic spine (uSP). Bar, 500 nm.

Figure 6.

Electron micrographs showing examples of profiles containing IR for GPER1 in the mPFC. These photomicrographs show GPER1-IR associated with (A) small synaptic vesicles in a terminal (TER) that is adjacent to an unlabeled terminal (uTER) and spine (uSP) that form an asymmetric synapse, (B) GPER1-IR associated with the plasma membrane of a dendritic spine that is forming a synapse with an unlabeled terminal (uTER) (C), GPER1 in a soma (SOM) in which it is localized to the endoplasmic reticulum and the membrane of mitochondria (mit) and IR for GPER1 in two axons (AX) (D), and GPER1 in an axon (AX) and in a dendrite (DEN), where it is associated with microtubules and the cell membrane. Bar, 500 nm.