The present study used a multi-modal, translational approach to show that estradiol facilitates extinction memory retention in female humans and rodents by modulating the functional reactivity of the vmPFC and amygdala—regions involved in fear expression and extinction in both species. In rodents, we show: a) estradiol administration facilitates extinction memory consolidation via ER-β activation in a time-dependent manner, and b) estradiol administration increases and decreases c-fos expression in the IL/vmPFC and amygdala, respectively, during extinction recall. In parallel to human studies, we show that natural variation in estradiol levels in women are associated with both the degree to which women retain extinction memory and the functional activation of the vmPFC and amygdala during extinction recall. Together, these data suggest that estradiol might improve the ability to retain extinguished fear memories by modulating neuronal activity and synaptic plasticity in the IL/vmPFC and amygdala.
The rodent experiments begin to illustrate the mechanisms by which estradiol can facilitate fear extinction recall. Consistent with the finding that ER-β agonists enhances extinction recall, a recent study showed that estradiol facilitates extinction of contextual fear conditioning via activation of ER-β in the hippocampus(36
), and additional studies have shown that activation of ER-β induces anxiolytic behavior(39
). The time-dependent effect of estradiol on extinction consolidation and the change in c-fos expression in both the vmPFC and amygdala suggest that estradiol might enhance extinction-induced synaptic plasticity in these brain regions. Given that estradiol has been shown to increase NMDA receptor transmission as well as long-term potentiation(41
) and extinction consolidation involves NMDA receptors(42
), we hypothesize that the changes in c-fos expression observed here might be due to the influence of estradiol on NMDA receptors. Future studies are needed to elucidate the interplay between estradiol and NMDA receptors (as well as other molecular indices of extinction learning) during fear extinction.
The current series of studies translate rodent data into humans by using neuroimaging techniques to demonstrate that estradiol affects vmPFC and amygdala functional activation when recalling a safety memory. Previous functional imaging studies have reported increased amygdala activation during fear extinction(10
) and increased vmPFC-amygdala connectivity during extinction recall(10
). Interestingly, the present imaging data seem to contradict the observed reduction in c-fos expression in the amygdala during extinction recall in female rats. However it is challenging to interpret BOLD signal changes in the amygdala during fear extinction recall due to the well-documented role of the amygdala in both fear learning and fear extinction(2
). Previous studies have shown that enhanced GABAergic neural activation is critical for fear extinction(8
). In addition, Herry et al.(7
) recently demonstrated that there are two neural populations within the amygdala: one signals fear learning, and the other signals fear extinction. The tools used in rodents versus those in humans might not be directly comparable since both inhibitory and excitatory processes could both lead to an increase the BOLD signal. Nonetheless, these data show that estradiol does indeed influence the function of the amygdala across species, although the direction of the effect needs further elaboration to compare the results between species.
Given that results from our previous work suggest that progesterone may not facilitate fear extinction in healthy women(29
), we chose to focus primarily on the effects of estradiol in the present study. We did, however, find that a majority of women with high estradiol levels also had elevated progesterone levels, making it difficult to rule out the potential influence of progesterone on fear extinction. Previous studies(28
) have shown that estrogen and progesterone independently facilitate extinction recall in female rats. In fact, when these hormones are injected together, the effect is more pronounced, which is consistent with other data in the field(45
). Thus, further examination of progesterone independently as well as in interaction with estradiol on fear extinction in women is required to.
Though substantial effort in designing parallel rat and human experiments was made, there remain some differences between the two designs that could be viewed as potential limitations of the present study. One difference is that while women were conditioned and extinguished within the same phase of the menstrual cycle while female rats underwent fear conditioning and extinction training at two different phases of the estrus cycle (proestrus and metestrus, respectively). Though this limitation could have been mitigated by the use of ovariectomized female rats, we used naturally cycling female rats to allow for maximum translation between species. We understand that this may introduce effects of other hormones in relation to estradiol that may produce potential confounds. We have previously shown, however, that conducting conditioning or extinction recall at different phases of the estrus cycle had no effect on either fear conditioning or extinction recall(28
). Nonetheless, future studies of ovariectomized rats are needed to identify the specific mechanisms of estrogen’s influence on the fear extinction circuit. For example, it will be important to detail how estrogen influences c-fos expression as well as other molecular indices of neural plasticity within different nuclei of the amygdala and the infralimbic and prelimbic subdivisions of the vmPFC in rats. Additional studies using cannula-guided estrogen administration into vmPFC subregions will be critical.
The translational evidence presented in this study begins to establish a role of estradiol in modulating vmPFC and amygdala activation when recalling a safety memory and highlights the importance of this understudied area in neuroscience(25
). Our data suggest that estradiol may serve a protective function against elevated fear and anxiety and that transient periods of low estradiol levels may be associated with impaired retention of safety memory. The functional integrity of the fear extinction network in women using oral contraceptives and in menopausal women with and without hormone replacement therapy should be examined, especially given that the use of oral and intrauterine contraceptives is known to reduce endogenous cycling estradiol levels(46
). Moreover, women appear to be vulnerable to developing mood and anxiety disorders during postpartum(47
) and menopausal periods(48
) when endogenous estradiol levels are low. Additional studies investigating the effects of naturally cycling and exogenous hormone manipulations on the function of the fear extinction circuitry could ultimately introduce ways to adapt, improve or produce therapies specifically tailored to women.