Sex differences in fear regulation and reward-seeking behaviors in a fear-safety-reward discrimination task

Highlights

• Significant sex differences were seen in a fear-safety-reward discrimination task.

• Females did not inhibit conditioned freezing in the presence of a safety cue.

• Females showed higher darting levels than males.

• Females did not show extinction of cued freezing, whereas male rats did.

• Females had elevated reward seeking to a reward cue which declined after footshock.

Abstract

Reward availability and the potential for danger or safety potently regulate emotion. Despite women being more likely than men to develop emotion dysregulation disorders, there are comparatively few studies investigating fear, safety and reward regulation in females. Here, we show that female Long Evans rats did not suppress conditioned freezing in the presence of a safety cue, nor did they extinguish their freezing response, whereas males did both. Females were also more reward responsive during the reward cue until the first footshock exposure, at which point there were no sex differences in reward seeking to the reward cue. Darting analyses suggest females were able to regulate this behavior in response to the safety cue, suggesting they were able to discriminate between fear and safety cues but did not demonstrate this with conditioned suppression of freezing behavior. However, levels of darting in this study were too low to make any definitive conclusions. In summary, females showed a significantly different behavioral profile than males in a task that tested the ability to discriminate among fear, safety and reward cues. This paradigm offers a great opportunity to test for mechanisms that are generating these behavioral sex differences in learned safety and reward seeking.

Introduction

Clinical disorders arising from maladaptive emotion regulation present a large burden on society worldwide. Many of these disorders show comorbidity, for example, addiction with anxiety disorders [1]. Cues predicting aversive outcomes elicit avoidance and fear behaviors whereas cues predicting reward elicit approach and reward-seeking behaviors. Cues signifying safety have the power to modulate fear and reward-seeking behaviors by informing the organism whether or not the environment is safe [2]. Thus, safety, fear and reward behaviors, and the circuitries governing these behaviors, are intertwined. The majority of studies on reward and fear processing have been conducted in parallel, investigating the circuitries separately in primarily male subjects. If we hope to understand and treat comorbid disorders resulting from maladaptive emotion regulation, increased efforts in investigating how these circuitries integrate their functions to influence behavior is needed in both male and female subjects.

Our laboratory has designed and validated a behavioral task in which fear, safety and reward cues are learned within the same session allowing us to assess the animal's ability to discriminate among these cues [3], [4], [5], [6], [7]. In this task, rats are exposed to cues associated with safety, fear (fear cue paired with footshock), and reward (reward cue paired with sucrose). Male rats consistently learn to discriminate among safety, fear and reward cues to (1) suppress conditioned freezing in the presence of a safety cue (fear + safety cue), and (2) increase reward seeking when reward is available (reward cue) [3], [4], [5], [6], [7]. This paradigm also allows us to investigate how safety cues can regulate both fear and reward behaviors. Evidence suggests that reward learning mechanisms overlap at least partially with safety learning [2], [3], [8], [9], [10], [11], [12]. For example, learned safety can act as a behavioral antidepressant in mice [9], and animals will perform certain behaviors in order to turn on a safety signal [10], [11]. Within the amygdala we have shown a subpopulation of neurons responding with the same level of excitation or inhibition during both the reward and safety cues [3]. We have also shown a dissociation between reward and safety discrimination; inactivation of the prelimbic or infralimbic cortices of the ventromedial prefrontal cortex have differential effects on reward and safety discrimination, respectively [5]. Thus, in male rats, our prior work has already shown a critical involvement of the corticoamygdalar circuit in learning this fear-safety-reward cue discrimination.

Much of the research investigating emotion regulation mechanisms have exclusively used male subjects. In a study using male Vietnam veterans, Post-Traumatic Stress Disorder (PTSD) patients show impairments in suppressing their fear response in the presence of a safety cue [13]. Importantly though, women are more than twice as likely to develop PTSD than men, with women having a lifetime prevalence of 8.5% in contrast to 3.4% in men [14]. In fear studies that have included female rats, it has been shown that females exhibit lower levels of freezing behavior than male rats after repeated fear cue presentations [15], [16]. These findings have been thought to indicate a difficulty in fear conditioning in female rats. A more recent study has identified that approximately 40% of female rats tested exhibit an alternate fear behavior in the form of fast paced movements called ‘darting’; this was only seen in approximately 10% of male rats tested [17]. There is also evidence of sex differences in the seeking of natural rewards, where it has been reported that female rats consume more sucrose pellets than males and are willing to work harder for them [18]. Dopamine signaling during reward tasks has also been demonstrated to be different between sexes. For example, Conway et al. [19] showed females continued to perform intracranial self-stimulation for brain stimulation reward while under the influence of a kappa-opioid receptor agonist, which suppresses dopamine release, whereas males decreased this behavior. Their data suggest that female rats may have an increased capacity to produce and release dopamine compared to males, under these conditions. Our prior work has also shown, in males, that dopamine signaling in the basolateral amygdala contributes to effective discrimination among fear, safety and reward cues [6].

Taken together, we hypothesized there would be sex differences in the ability to express clear discrimination among fear, safety and reward cues. The inability of male PTSD patients to learn safety signaling has been labeled a biomarker of the disorder [20]. Due to sex-related differences in the human diagnosis of PTSD, with women diagnosed at rates twice that of men [21], any differences female rats have in the learning or retention of safety signals could steer future research on the neurobiological processes underlying these variations.