Blockade of estrogen by hormonal contraceptives impairs fear extinction in female rats and women

Abstract

Background: Fear extinction is a laboratory model of fear inhibition and is the basis of exposure therapy for anxiety disorders. Emerging evidence from naturally cycling female rodents and women indicates that estrogens are necessary to the consolidation of fear extinction. Hormonal contraceptives (HCs) inhibit estrogen production; yet, their effects on fear extinction are unknown.

Methods: We used a cross-species translational approach to investigate the impact of HCs and estradiol supplementation on fear extinction in healthy women (n=76) and female rats (n = 140).

Results: Women using HCs exhibited significantly poorer extinction recall compared with naturally cycling women. The extinction impairment was also apparent in HC-treated female rats and was associated with reduced serum estradiol levels. The impairment could be rescued in HC-treated rats either by terminating HC treatment after fear learning or by systemic injection of estrogen-receptor agonists before fear extinction, all of which restored serum estradiol levels. Finally, a single administration of estradiol to naturally cycling women significantly enhanced their ability to recall extinction memories.

Conclusions: Together, these findings suggest that HCs may impact women's ability to inhibit fear but that this impairment is not permanent and could potentially be alleviated with estrogen treatment.

Fig. 1

(A) Mean (±SEM) differential SCRs during conditioning (average across trials) and the end of extinction training (average of last two trials) on Day 1 in Experiment 1. * = p<0.05; cond. vs. end of ext. (B) Mean (±SEM) percent resovery of SCRs at extinction recall in Experiment 1. Ns = 16 per group for H-EST and L-EST, N = 13 for HC users. * = p<0.05; H-EST vs. L-EST, and HC.

Fig. 2

(A) Mean (±SEM) freezing responses across experimental phases in Experiment 2. Ns = 11 (Veh-Pro), 7 (Veh-Met), 12 (LEV-100), 12 (LEV-500). Day 2 * = p<0.05; LEV-500 vs. Veh-Pro. Day 3 * = p<0.05; Veh-Pro vs. Veh-Met, LEV-100, and LEV-500; and LEV-100 vs. Veh-Met and LEV-500. (B) Mean (±SEM) freezing responses across experimental phases in Experiment 3. Ns = 8–9 per group. Day 3 * = p<0.05, HC/Veh vs. HC/HC. (C) Mean (±SEM) freezing responses across experimental phases in Experiment 4. Ns = 12–14/ group. Day 3 * = p<0.05; HC/Veh vs. Veh/Veh, HC/ERα, and HC/ERβ.

Fig. 3

(A) Representative vaginal cytology of a naturally cycling rat that received HC treatment for 5 days, and was then switched to vehicle treatment for another 5 days, during Experiment 3 (arrows denote cell types referred to in the following descriptions). A naturally cycling rat progresses through different stages of the estrous cycle on a daily basis (row 1). Nucleated epithelial cells characterize proestrus; cornified cells characterize estrus, leukocytes characterize metestrus, and scant leukocytes along with some non-nucleated and nucleated cells characterize diestrus. HC treatment (row 2) causes persistent metestrus-like cytology (predominantly leukocytes, emerging on HC Days 2–5). Switching the rat from HC to vehicle treatment (row 3) results in a gradual return to normal cycling. Note the reduction in leukocytes by Day 3, and the emergence of cytology consistent with diestrus and then proestrus on Days 4 and 5, respectively. (B) Serum estradiol from naturally cycling rats (N = 5/group) sacrificed during proestrus (Pro) and metestrus (Met), and from HC-treated rats sacrificed after 5 days of HC treatment (HC), 5 days of HC treatment and 1 hr after s.c. injection of an ERα (HC/ERα) or ERβ (HC/ERβ) agonist, and 5 days of HC treatment followed by a switch to vehicle treatment for another 5 days (HC/switch). Estradiol levels are presented as a percentage of those obtained from rats during proestrus. * = p<0.05; HC and Met vs. Pro, HC/ERα, HC/ERβ, and HC/switch.

Fig. 4

(A) Experimental protocol for Experiment 5. (B) Mean (±SEM) serum estradiol levels during the different phases of the experiment. T2 * = p< 0.05; Placebo vs. Estradiol. (C) Mean (±SEM) differential SCRs during conditioning, and differential SCRs during early and late extinction. * = p<0.05; Early vs. Late Ext. (D) Mean (±SEM) percent recovery of SCRs at extinction recall in Experiment 5 (Ns = 15–16/group). * = p<0.05; Placebo vs. Estradiol (E) A selection of estradiol (N = 13) and placebo-treated (N = 10) women were matched for conditioning strength. Estradiol-treated women exhibited significantly less percent recovery of fear at extinction recall than placebo-treated women when matching for conditioning strength. * = p< 0.05; Estradiol vs. Placebo.