Characterizing the effects of tonic 17β-estradiol administration on spatial learning and memory in the follicle-deplete middle-aged female rat

Abstract

17β-estradiol (E2)-containing hormone therapy is a safe, effective way to alleviate unwanted menopause symptoms. Preclinical research has focused upon the role of E2 in learning and memory using a surgically menopausal rodent model whereby the ovaries are removed. Given that most women retain their reproductive tract and undergo a natural menopause transition, it is necessary to understand how exogenous E2 impacts a structurally intact, but follicle-deplete, system. In the current study, 8 month old female rats were administered the ovatoxin 4-vinylcyclohexene diepoxide (VCD), which accelerates ovarian follicular depletion, to model the human menopause transition. After follicular depletion, at 11 months old, rats were administered Vehicle or tonic E2 treatment for 12 days prior to behavioral evaluation on spatial working and reference memory tasks. Results demonstrated that E2 had both enhancing and impairing effects on taxed working memory depending upon the learning or retention phases of the water radial-arm maze, with no impact on reference memory. Relationships between memory scores and circulating estrogen levels were specific to follicle-depleted rats without E2 treatment. Collectively, findings demonstrate the complexity of E2 administration in a follicle-depleted background, with cognitive effects specific to working memory; furthermore, E2 administration altered circulating hormonal milieu and relationships between hormone profiles and memory. In sum, menopausal etiology impacts the parameters of E2 effects on cognition, complementing prior work with other estrogen compounds. Deciphering estrogenic actions in a system wherein the reproductive tract remains intact with follicle-depleted ovaries, thus modeling the majority or menopausal women, is critical for translational perspectives.

Keywords: Estrogen; Hormone therapy; Learning; Memory; Menopause; VCD.

Fig. 1.

(A) An illustration of the WRAM apparatus. Learning curves for working memory correct errors (1B), working memory incorrect errors (1C), and reference memory errors (1D) across the 12 days of the WRAM task. Day 1 performance was considered training and was excluded from analyses. VCD-Vehicle: n = 11; VCD-E2: n = 11.

Fig. 2.

Working memory errors during the Learning Phase of WRAM. VCD-Vehicle: n = 11; VCD-E2: n = 11. (A) Across Trials 2–4, a Treatment × Trial interaction occurred for WMC errors (p < 0.05). (B) VCD-E2 rats made significantly fewer WMC errors compared to VCD-Vehicle rats when memory load was maximally burdened during learning (p < 0.05). Individual data points are overlaid on the mean ± SEM. (C) Across all trials, a Treatment × Trial interaction occurred for WMI errors (p < 0.01). (D) VCD-E2 rats made significantly fewer WMI errors compared to VCD-Vehicle rats when memory load was maximally burdened during learning (p < 0.05). Individual data points are overlaid on the mean ± SEM.

Fig. 3.

Working memory errors during the Asymptotic Phase of WRAM. VCD-Vehicle: n = 11; VCD-E2: n = 11. (A) Across Trials 2–4, a marginal Treatment × Trial interaction occurred for WMC errors. (B) VCD-E2 rats made significantly more WMC errors compared to VCD-Vehicle rats when memory load was maximally burdened during the latter block of testing (p < 0.05). Individual data points are overlaid on the mean ± SEM. (C) Across all trials, a marginal Treatment × Trial interaction occurred for WMI errors. (D) There was no statistically significant difference between VCD-E2 rats and VCD-Vehicle rats for WMI errors when memory load was maximally burdened during the Asymptotic Phase. Individual data points are overlaid on the mean ± SEM.

Fig. 4.

Eight-hour delayed memory retention WRAM test. VCD-Vehicle: n = 11; VCD-E2: n = 11. Individual data points are overlaid on the mean ± SEM (some data point icons are overlapping in the figure). (A) The VCD-Vehicle group exhibited a delay-induced impairment in WMC errors following an 8 hour delay interval compared to the previous day’s performance. (B) The VCD-E2 group working memory performance was not significantly affected by the 8 hour delay interval.

Fig. 5.

Morris water maze performance. VCD-Vehicle: n = 11; VCD-E2: n = 11. (A) A schematic of the MM apparatus. (B) Average swim distance to platform decreased across days and did not interact with Treatment, indicating that reference memory was intact and not differentially impacted by exogenous hormone treatment following follicular depletion. (C-D) Analysis of the probe trial indicated that both groups swam a greater percent of total distance in the maze quadrant that previously contained the hidden escape platform compared to swim distance in the opposite quadrant. Individual data points are overlaid on the mean ± SEM.

Fig. 6.

(A) A schematic of the apparatus used for the VP task. (B) Visible Platform Performance: VCD-Vehicle and VCD-E2 groups significantly decreased latency to the visible platform across trials of the control task. VCD-Vehicle: n = 11; VCD-E2: n = 11.

Fig. 7.

Serum ovarian hormone levels. Individual data points are overlaid on the mean ± SEM. (A) E2 levels were elevated in rats that received exogenous E2 treatment following follicular depletion (p < 0.05). VCD-Vehicle: n = 6; VCD-E2: n = 11. (B) E1 levels were elevated in rats that received exogenous E2 treatment following follicular depletion (p < 0.001). VCD-Vehicle: n = 10; VCD-E2: n = 11. (C) Androstenedione levels were decreased in rats that received exogenous E2 treatment following follicular depletion (p < 0.05). VCD-Vehicle: n = 8; VCD-E2: n = 9.

Fig. 8.

Markers of follicular depletion and hormone status. VCD-Vehicle: n = 11; VCD-E2: n = 11 for all follicle evaluations. An ovary-intact, age-matched, Vehicle control for VCD treatment Reference Group from an independent data set (n = 10) has been included to demonstrate follicular depletion. Individual data points are overlaid on the mean ± SEM (some data point icons are overlapping in the graphs) (A) Primordial follicles were reduced in both VCD-treated groups compared to the Reference Group. VCD-treated groups did not differ in primordial follicle counts. (B) Primary follicles were reduced in both VCD-treated groups compared to the Reference Group. Primary follicles were also decreased in VCD-E2 rats compared to VCD-Vehicle rats (p < 0.05). (C) Secondary follicles were reduced in both VCD-treated groups compared to the Reference Group. VCD-treated groups did not differ in secondary follicle counts. (D) Antral follicles were reduced in both VCD-treated groups compared to the Reference Group. VCD-treated groups did not differ in antral follicle counts. (E) Corpora lutea were reduced in both VCD-treated groups compared to the Reference Group. VCD-treated groups did not differ in corpora lutea counts.

Fig. 9.

Body weight and uterine weight. Individual data points are overlaid on the mean ± SEM. (A) Overall body weight did not differ between groups at the end of the experiment, regardless of exogenous hormone treatment. (B) Follicle-deplete rats treated with exogenous E2 had significantly heavier uterine weights compared to the VCD-Vehicle group that did not receive exogenous hormone treatment, indicating a stimulatory effect of E2 on uterine tissue.

Fig. 10.

Correlations among hormone markers and cognitive scores (FDR correction set at 0.10). (A–B) Better working memory performance (fewer errors) at the end of WRAM testing was associated with lower E2 levels in the VCD-Vehicle group, but not the VCD-E2 group, regardless of whether the outlying value was included. VCD-Vehicle: n = 6 (some data point icons are overlapping in the figure); VCD-E2: n = 11. (C) Better working memory performance (fewer errors) at the end of WRAM testing was associated with higher E1 levels in the VCD-Vehicle group, but not the VCD-E2 group. VCD-Vehicle: n = 10; VCD-E2: n = 11.