Fear Responses In Male Rats Research Articles

Naloxone increases conditioned fear responses during social buffering in male rats.

Social buffering is the phenomenon in which the presence of an affiliative conspecific mitigates stress responses. We previously demonstrated that social buffering completely ameliorates conditioned fear responses in rats. However, the neuromodulators involved in social buffering are poorly understood. Given that opioids, dopamine, oxytocin and vasopressin play an important role in affiliative behaviour, here, we assessed the effects of the most well-known antagonists, naloxone (opioid receptor antagonist), haloperidol (dopamine D2 receptor antagonist), atosiban (oxytocin receptor antagonist) and SR49059 (vasopressin V1a receptor antagonist), on social buffering. In Experiment 1, fear-conditioned male subjects were intraperitoneally administered one of the four antagonists 25 min prior to exposure to a conditioned stimulus with an unfamiliar non-conditioned rat. Naloxone, but not the other three antagonists, increased freezing and decreased walking and investigation as compared with saline administration. In Experiment 2, identical naloxone administration did not affect locomotor activity, anxiety-like behaviour or freezing in an open-field test. In Experiment 3, after confirming that the same naloxone administration again increased conditioned fear responses, as done in Experiment 1, we measured Fos expression in 16 brain regions. Compared with saline, naloxone increased Fos expression in the paraventricular nucleus of the hypothalamus and decreased Fos expression in the nucleus accumbens shell, anterior cingulate cortex and insular cortex and tended to decrease Fos expression in the nucleus accumbens core. Based on these results, we suggest that naloxone blocks social buffering of conditioned fear responses in male rats.

The lateral intercalated cell mass of the amygdala is activated during social buffering of conditioned fear responses in male rats

The presence of an affiliative conspecific reduces stress responses to a wide variety of stimuli. This phenomenon is termed “social buffering”. We previously found that the presence of another Wistar rat (associate) suppressed activation of the lateral amygdala (LA) and ameliorated stress responses to an auditory conditioned stimulus (CS) in a fear-conditioned Wistar subject rat. Subsequent analyses suggested that activation of the posterior complex of the anterior olfactory nucleus (AOP) is responsible for the suppression of the LA. However, it remains unclear how the AOP suppresses the LA. To clarify this issue, a fear-conditioned Wistar subject was exposed to the CS either alone or with a Wistar associate. We also prepared a fear-conditioned Wistar subject that was tested with a Fischer344 (F344) associate as an additional control because F344 associates do not induce social buffering. We found that the presence of a Wistar associate induced a reduction of behavioral responses and Fos expression in the paraventricular nucleus of the hypothalamus (PVN) of the subject. Although Fos expression in the AOP was increased, the expression was not biased towards the GABAergic cells. In addition, Fos expression in the lateral intercalated cell mass of the amygdala (lITC) was increased. In contrast, the presence of a F344 associate did not affect Fos expression in subjects’ PVN or lITC, whereas behavioral responses were slightly reduced. These results suggest that the lITC was activated during social buffering. Based on these findings, we propose that the AOP indirectly suppresses the LA by activating the lITC.

A dyad shows mutual changes during social buffering of conditioned fear responses in male rats

The presence of an affiliative conspecific reduces stress responses to a wide variety of stimuli. This phenomenon is termed “social buffering”. We previously found that the presence of another naïve rat (associate) reduced conditioned fear responses to an auditory conditioned stimulus in a conditioned subject rat. Although we subsequently conducted a series of studies to examine behavioral, physiological, and neural changes during social buffering in the conditioned subject, the changes in the associate remained unclear. Therefore, in the present study, we investigated the behavioral and neural changes in the associate. Fear-conditioned and non-conditioned rats were re-exposed to the conditioned stimulus with an associate placed in the same enclosure (Experiment 1) or separated by a wire-mesh partition (Experiment 2). In Experiment 1, the associate exhibited increased anogenital contact and allo-grooming, which were accompanied by increased c-Fos expression in the paraventricular nucleus of the hypothalamus and central amygdala. These results suggest that the subject and associate mutually affected each other during social buffering. In contrast, in Experiment 2, we found only a difference in the time course of investigation between associates tested with the conditioned and non-conditioned subjects. These results suggest that the associate was unable to acquire a sufficient amount of signal from the conditioned subject behind the wire-mesh partition necessary to show clear changes in behavior and c-Fos expression. Taken together, the current findings suggest that a dyad shows mutual changes during social buffering of conditioned fear responses in male rats.

Social buffering enhances extinction of conditioned fear responses in male rats

In social species, the phenomenon in which the presence of conspecific animals mitigates stress responses is called social buffering. We previously reported that social buffering in male rats ameliorated behavioral fear responses, as well as hypothalamic-pituitary-adrenal axis activation, elicited by an auditory conditioned stimulus (CS). However, after social buffering, it is not clear whether rats exhibit fear responses when they are re-exposed to the same CS in the absence of another rat. In the present study, we addressed this issue using an experimental model of extinction. High stress levels during extinction training impaired extinction, suggesting that extinction is enhanced when stress levels during extinction training are low. Therefore, we hypothesized that rats that had received social buffering during extinction training would not show fear responses to a CS, even in the absence of another rat, because social buffering had enhanced the extinction of conditioned fear responses. To test this, we subjected male fear-conditioned rats to extinction training either alone or with a non-conditioned male rat. The subjects were then individually re-exposed to the CS in a recall test. When the subjects individually underwent extinction training, no responses were suppressed in the recall test. Conversely, when the subjects received social buffering during extinction training, freezing and Fos expression in the paraventricular nucleus of the hypothalamus and lateral amygdala were suppressed. Additionally, the effects of social buffering were absent when the recall test was conducted in a different context from the extinction training. The present results suggest that social buffering enhances extinction of conditioned fear responses.

A familiar conspecific is more effective than an unfamiliar conspecific for social buffering of conditioned fear responses in male rats

In social animals, the presence of an affiliative conspecific alleviates acute stress responses, and this is called social buffering. We previously reported in male rats that the presence of a conspecific mitigates conditioned fear responses to auditory conditioned stimulus paired with foot shocks. Subsequent studies revealed that we could observe this social buffering when rats were tested in a box odorized by a conspecific. Because we previously used an unfamiliar conspecific, the effects of familiarity with a conspecific on the intensity of social buffering remain unclear. Here, we examine this question by preparing a familiar conspecific that had been housed with a subject for 3 weeks in the same cage. We exposed fear-conditioned subjects to a conditioned stimulus in either a clean control box or a box odorized beforehand by either an unfamiliar or a familiar conspecific. When the subjects were tested in the control box, they showed freezing and Fos expression in the paraventricular nucleus. These responses were suppressed when we placed rats in the box odorized by a conspecific. However, the suppression was greater when the box was odorized by a familiar conspecific rather than by an unfamiliar conspecific. Fos expression in the lateral amygdala was also suppressed in the same manner. These results suggest that a familiar conspecific is more effective for social buffering of conditioned fear responses.

Social buffering reduces male rats' behavioral and corticosterone responses to a conditioned stimulus

In social species, the presence of an affiliative same-sex conspecific ameliorates acute stress responses in threatening conditions. We previously found that the presence of an unfamiliar male rat separated by a wire mesh barrier blocks the behavioral responses and Fos expression in the paraventricular nucleus of the hypothalamus (PVN) in a male subject rat that had previously been exposed to an auditory conditioned stimulus (CS) paired with foot shocks. Based on the Fos expression in the PVN, we hypothesized that the presence of a conspecific ameliorated the hypothalamic–pituitary–adrenal (HPA) axis activation and induced social buffering of conditioned fear responses. The direct evidence for this hypothesis, however, is still lacking. To clarify this point, we exposed fear-conditioned and non-conditioned subjects to the CS either alone or with a conspecific separated by a wire mesh barrier. When the fear-conditioned subject alone was re-exposed to the CS, it exhibited increased freezing, decreased sniffing, and elevated corticosterone levels. In contrast, the presence of the conspecific suppressed these behavioral and HPA axis responses to a level similar to those observed in the non-conditioned subjects. These results suggest that the presence of a conspecific suppressed the behavioral responses and HPA axis activation to the CS. The present results provide direct evidence for the existence of social buffering of conditioned fear responses in male rats.

Olfactory signals mediate social buffering of conditioned fear responses in male rats

In social animals, the presence of an affiliative conspecific alleviates acute stress responses, and this is called social buffering. We previously reported that social buffering mitigates the fear responses of male rats to auditory conditioned stimuli that had been paired with foot shocks. Subsequent studies revealed that signals that are perceived by the main olfactory epithelium are important for social buffering. Because olfactory signals are the signal perceived by the main olfactory epithelium, we hypothesized that we could induce the social buffering of conditioned fear responses by presenting olfactory signals that were derived from a conspecific. In order to test this hypothesis, we exposed fear-conditioned subjects to a conditioned stimulus either in a clean test box or in a test box that was odorized by keeping a conspecific in it as an odor donor beforehand. When the subjects were tested in the clean test box, they showed behavioral fear responses and enhanced Fos expression in the paraventricular nucleus. In contrast, the presence of conspecific olfactory signals blocked these fear responses and Fos expression. These results suggested that olfactory signals suppress conditioned fear responses. Fos expression in the posteromedial region of the olfactory peduncle and amygdala suggested that this suppression involves the same neural mechanisms as those of social buffering. Taken together, we concluded that olfactory signals mediate the social buffering of conditioned fear responses.

The neural pathway underlying social buffering of conditioned fear responses in male rats

In social animals, the presence of an affiliative conspecific alleviates acute stress responses, and this is termed social buffering. However, the neural mechanisms underlying social buffering have not been elucidated. We have reported that the main olfactory system mediates social buffering of conditioned fear responses in male rats, and this is accompanied by suppression of the lateral and central amygdala. Therefore, olfactory signals are probably transmitted from the main olfactory system to the amygdala. Because the lateral and central amygdala do not receive projections from the main olfactory bulb, the site that links the main olfactory bulb and amygdala was presumed to be located within the main olfactory system. To find the linkage site, we generated lesions within the main olfactory system, and found that a bilateral lesion in the posteromedial region of the olfactory peduncle (pmOP) blocked social buffering. Next, we determined that the pmOP receives direct projections from the main olfactory bulb. Finally, we demonstrated that the connection between the pmOP and ipsilateral amygdala is important for social buffering, and that the pmOP projects directly to the ipsilateral amygdala, including the lateral and central amygdala. On the basis of these results, we suggest that the pmOP links the main olfactory blub to the amygdala and enables social buffering of conditioned fear responses. These results provide the first comprehensive picture of the neural pathway underlying the social buffering phenomenon.

Anterior olfactory nucleus plays an important role in the social buffering of conditioned fear responses in male rats

Anterior olfactory nucleus plays an important role in the social buffering of conditioned fear responses in male rats

Main olfactory system mediates social buffering of conditioned fear responses in male rats

We previously reported that the presence of a conspecific animal blocked freezing of a male rat in response to an auditory conditioned stimulus that had been paired with foot shocks, as well as associated Fos expression in the paraventricular nucleus. Here we investigated how this 'social buffering' is mediated by examining the contributions of both physical contact and the main olfactory system. Fear-conditioned rats exposed to the conditioned stimulus alone responded by freezing and increased Fos expression in the paraventricular nucleus. However, the presence of another rat, but not a guinea pig, dramatically mitigated these responses, even if the dyad was separated by a wire mesh or a pair of wire meshes 5 cm apart. In contrast, social buffering was absent when a transparent acrylic board was inserted between the double wire mesh. Lesioning of the main olfactory epithelium by injection of ZnSO(4) intranasally also abolished social buffering. Thus, we conclude that the main olfactory system is essential for the social buffering but does not require physical contact between the dyad.

Main olfactory system is essential for the social buffering of conditioned fear responses in male rats

Main olfactory system is essential for the social buffering of conditioned fear responses in male rats

Two types of social buffering differentially mitigate conditioned fear responses

In a phenomenon known as 'social buffering' in various species, signals from a conspecific animal can mitigate stress responses. This buffering can be achieved either by 'pair-housing' after a stressful event or by 'pair-exposure' to an acute stressor with a conspecific animal. In this study, we compared the impacts of these two types of social buffering on auditory conditioned fear responses in male rats. When subjects were exposed to an auditory conditioned stimulus (CS) that had been paired with foot shocks on the previous day, they clearly exhibited behavioral (freezing), autonomic (aggravated stress-induced hyperthermia) and neural (Fos expression) responses. Pair-housing for 24 h with an unfamiliar rat following fear conditioning resulted in a suppressed autonomic, but not behavioral, response, with Fos expression in the lateral nucleus of the amygdala and ventrolateral periaqueductal gray. On the other hand, pair-exposure to the CS with an unfamiliar rat eliminated the behavioral, but not the autonomic, response, with Fos expression in the basal nucleus of the amygdala and infralimbic region of the prefrontal cortex. Furthermore, subjects that had been pair-housed and then pair-exposed showed no behavioral, autonomic or neural responses, suggesting that the combination of the two procedures can completely block the fear conditioning sequence. These results demonstrate that two types of social buffering differentially relieve conditioned fear responses, by influencing different neural pathways in the amygdala.

Two types of social buffering differentially attenuates conditioned fear responses in male rats

Two types of social buffering differentially attenuates conditioned fear responses in male rats

Face pictures reduce behavioural, autonomic, endocrine and neural indices of stress and fear in sheep.

Faces are highly emotive stimuli and we find smiling or familiar faces both attractive and comforting, even as young babies. Do other species with sophisticated face recognition skills, such as sheep, also respond to the emotional significance of familiar faces? We report that when sheep experience social isolation, the sight of familiar sheep face pictures compared with those of goats or inverted triangles significantly reduces behavioural (activity and protest vocalizations), autonomic (heart rate) and endocrine (cortisol and adrenaline) indices of stress. They also increase mRNA expression of activity-dependent genes (c-fos and zif/268) in brain regions specialized for processing faces (temporal and medial frontal cortices and basolateral amygdala) and for emotional control (orbitofrontal and cingulate cortex), and reduce their expression in regions associated with stress responses (hypothalamic paraventricular nucleus) and fear (central and lateral amygdala). Effects on face recognition, emotional control and fear centres are restricted to the right brain hemisphere. Results provide evidence that face pictures may be useful for relieving stress caused by unavoidable social isolation in sheep, and possibly other animal species, including humans. The finding that sheep, like humans, appear to have a right brain hemisphere involvement in the control of negative emotional experiences also suggests that functional lateralization of brain emotion systems may be a general feature in mammals.