Effects of under- and overcrowding on exploratory behavior in the elevated plus-maze

The present work investigated whether the number of rats housed in a cage affects exploration of an elevated plus-maze. Male Wistar-derived rats were kept 1, 2, 3, 4, 6, 8, 12, 16, or 24 to same size cages either for 1 or 14 days and tested in the elevated plus-maze. Rats kept 6 to a cage were arbitrarily considered controls because this is the housing condition adopted in many laboratories, ours included. In comparison to controls, 1-day housed rats kept 1, 2, 16, and 24 to a cage decreased the percentage of entries into the open arms. Similar decreases were also found in the time spent in the open arms, the only exception being the group with rats kept 16 to a cage which failed to show significant differences from the control group. Fourteen-day housed rats kept 1, 2, 16, or 24 to a cage decreased the percentage of entries and time spent in the open arms. We found plus-maze exploration to be similar in groups in which rats were kept from 4 to 12 to a cage. The present data indicate that anxiogenic effects resulting from under- and overcrowding should be taken into consideration in behavioral studies.     

Anxiolytic-like effect of the selective neuropeptide Y Y2 receptor antagonist BIIE0246 in the elevated plus-maze

The involvement of Neuropeptide Y (NPY) in the pathophysiology of mood disorders has been suggested by clinical and preclinical evidence. NPY Y1 and Y2 receptors have been proposed to mediate the NPY modulation of stress responses and anxiety related behaviors. To further investigate the role of Y2 receptors in anxiety we studied the effect of BIIE0246, a selective Y2 receptor antagonist, in the elevated plus-maze test. Rats treated with 1.0 nmol BIIE0246 showed an increase in the time spent on the open arm of the maze. In addition, to study the effects of the Y2 antagonism on NPY protein level, NPY-like immunoreactivity was measured in different brain regions following treatment with BIIE0246, but no statistically significant effects were observed. These results suggest that BIIE0246 has an anxiolytic-like profile in the elevated plus-maze.     

Individual differences in the elevated plus-maze and the forced swim test

The elevated plus-maze is an apparatus composed of enclosed and open (elevated) arms and time spent in the open arms by a rat can be increased/decreased by anxiolytic/anxiogenic agents. In the forced swim test, floating behavior is used as an index of behavioral despair and can be decreased by antidepressant agents. As the comorbidity between anxiety and depression is a remarkable issue in human behavioral disorders, a possible relationship between the behaviors seen in the cited tests is of great relevance. In the present study, fifty-four male rats (Rattus norvegicus) were submitted to a plus-maze session and to a 2-day forced swim protocol. According to their time in the open arms, they were divided into three groups: Low Open, Medium Open and High Open. Some plus-maze measures were found to be coherent with time in the open arms and are suggested to also be reliable anxiety indexes. In the forced swim test, the Low Open group showed decreases in floating duration from forced swim Session 1 to Session 2, an alteration opposite to that observed in the other groups (particularly, the Medium Open group). The Low Open group also showed increases in floating latency, again in sharp contrast with the alteration found in the other groups. Accordingly, positive and negative correlation were found between time in the open arms and floating duration and latency, respectively. Results are compared to previous studies and mediation of the effect by reactivity to aversive stimulation or alterations induced by open arm exposure is discussed.     

Characterization of behavioural responses in different test contexts after a single social defeat in male golden hamsters (Mesocricetus auratus)

The objective of the present study was to characterize behavioural responses of male hamsters in each of three test contexts after they had experienced either a single social defeat or a neutral encounter. In experiment 1, hamsters were observed in a familiar social context (i.e., their home cages), and defeated males displayed different amounts of time and submissive behaviours towards a known opponent than a novel intruder, whereas males in the neutral-encounter groups did not show such differences. In experiment 2, in an unfamiliar social context (i.e., a Y-maze), defeated males generated submissive behaviours and fear memory towards a known opponent that they re-encountered 5-min and 24-h after the defeat. The formation of long-term memory was interrupted by an injection of anisomycin (210 mg/kg). In experiment 3, in a non-social, anxiogenic context, hamsters that had previously had different social experiences did not demonstrate additional anxiety in an elevated plus-maze, with the exception of males that had previously experienced repeated social defeats. Our data suggested that hamsters’ behavioural changes following defeat are context-dependent and stimulus-specific. The experience of a single social defeat is sufficient to regenerate submissive behaviours and fear memory when reencountering a known opponent.     

Validation of a behavioral recording automated system in the elevated plus-maze test

The elevated plus-maze test has been widely used for screening of anxiolytic drugs and for exploring neurobiological bases of anxiety. In this study, we validated a new automated system that enables to record exploratory behavior in the elevated plus-maze test. This system, called cyberplus, consisted of ten pairs of photoelectric cells strategically located in several parts of the apparatus, and seemed to be sensitive to the position of the animal's forepaws, so it would yield scores in anxiety measurements and locomotor activity similar to those obtained by following the traditional procedure, that is, by analyzing videotapes by experienced observers. In order to assess this hypothesis, we exposed rats to the elevated plus-maze test and compared the scores obtained by cyberplus with the values recorded by two independent observers, conducting a correlational study with both kinds of recording procedures. The results obtained suggest the utility of cyberplus as a behavioral recording automated system in the elevated plus-maze test, making data collection and data analysis easier in exploring pharmacological and neurobiological bases of anxiety.     

Is full physical contact necessary for buffering effects of pair housing on social stress in rats?

Our previous study showed that pair housing with a familiar male prevented an increase in anxiety caused by social defeat in male rats. The present study attempted to identify the aspects of social interactions with a familiar male that are needed for the emergence of such a pair-housing effect. In Experiment 1, the subject was repeatedly exposed to the cage and bedding used by a familiar pairmate, after two instances of social defeat. Mere exposure to the soiled cage and bedding did not prevent an increase in anxiety in the elevated plus-maze test performed two weeks after social defeat. In Experiment 2, the subject was separated from a familiar pairmate with a wire mesh partition, which allowed visual, auditory, and limited physical contact, in addition to olfactory contact with the pairmate. The separation with a wire mesh partition abolished the buffering effect of pair housing on anxiety. These results indicate that visual, auditory, and olfactory contact with a familiar male was not sufficient in reducing the anxiogenic effect of social defeat in male rats. It was suggested that full physical contact is necessary for the emergence of the buffering effect of pair housing on social stress.     

Increases in plasma corticosterone and stretched-attend postures in rats naive and previously exposed to the elevated plus-maze are sensitive to the anxiolytic-like effects of midazolam

A single exposure to the elevated plus-maze test (EPM) reduces or abolishes the anxiolytic efficacy of benzodiazepines on a second trial. This phenomenon known as one-trial tolerance (OTT) is considered to be due to a shift in the emotional state of the animals across the test/retest sessions. Activation of the hypothalamic-pituitary-adrenocortical (HPA) axis has been considered to be an adaptive response to stressful or challenging situations such as height and openness of the EPM. This work looks at the phenomenon of OTT to the benzodiazepine compound midazolam through the conjoint examination of the novel ethological measures of the EPM and adrenocortical response of rats exposed to single and repeated sessions of the EPM. The results obtained confirmed that the approach/avoidance conflict on the first trial of the EPM is very sensitive to the anxiolytic effects of benzodiazepines. Moreover, stressful stimuli present upon initial exposure to the EPM render the standard measures of the EPM resistant to the anxiolytic effects of benzodiazepines on retest. However, the increases in plasma corticosterone and in risk assessment behavior observed in rats submitted to single or repeated sessions in the EPM were reversed by pretreatment with midazolam. The administration of metyrapone, a glucocorticoid synthesis blocker, decreased risk assessment but did not affect locomotion and anxiety-like behaviors. It is suggested that the detection of the dangerous environment through the stretched-attend postures in the second trial leads to the known low level of exploration and the consequent OTT upon retest. Moreover, in view of the similarity between the risk assessment and plasma corticosterone patterns in both naive and experienced rats, this hormone-behavior profile may be crucial for the expression of OTT to benzodiazepines in rodents exposed to the EPM.     

One-trial tolerance to the effects of chlordiazepoxide in the elevated plus-maze is not due to acquisition of a phobic avoidance of open arms during initial exposure

A single exposure to the elevated plus-maze (EPM) test of anxiety reduces or abolishes the anxiolytic-like efficacy of benzodiazepines. This phenomenon called one-trial tolerance has been suggested to represent the acquisition of a phobic-like response to the open arms during trial 1. The present study was designed to examine the effects of chlordiazepoxide (5 mg/kg, ip) on the behaviour of rats in a conventional EPM apparatus after previous exposure to a four-open-arm EPM, a four-enclosed arm EPM or a conventional EPM, as well as in naive rats. Chlordiazepoxide had clear-cut anxiolytic-like effects (increased percentage of time spent on the open arms) in a traditional EPM in naive rats and in animals previously exposed to a four-open-arm EPM. However, it was ineffective in rats previously exposed to a traditional or a four-closed-arm EPM. Thus, the phenomenon of one-trial tolerance does not depend upon initial open-arm experience.     

Modulation of elevated plus maze behavior after chronic exposure to the anabolic steroid 17alpha-methyltestosterone in adult mice

Exposure to supraphysiological doses of androgens may disrupt affective components of behavior. In this study, behavior of adult C57Bl/6 male mice was studied after exposure to the anabolic androgenic steroid (AAS) 17alpha-methyltestosterone (17alpha-meT; 7.5 mg/kg) via a subcutaneous osmotic pump for 17 days. Controls received vehicle implants (0.9% NaCl + 30% cyclodextrine). On day 15, experimental animals were challenged with an ethanol (EtOH) injection (i.p.; 1 g/kg) while controls received saline injections. Five minutes after the injection, animals were tested in an automated elevated plus maze (EPM) or in automated activity chambers. In addition, injection-free animals were tested for ethanol consumption on day 16 after an overnight water deprivation period. Whereas chronic exposure to 17alpha-meT did not modulate open arm behavior, EtOH-exposed animals made more entries into the open arms than controls (P < 0.05). A significant reduction of risk assessment behaviors (rearing, flat approach behavior, and stretch attended posture) over the EPM was noted for EtOH-exposed animals whereas a reduction in stretch attended postures was observed among 17alpha-meT-exposed animals. Locomotor activity, and light-dark transitions in activity chambers remained unaltered. Exposure to AAS did not modulate EtOH consumption. Our data suggest that exposure to a supraphysiological dose of 17alpha-meT has minimal effects on exploratory-based anxiety.     

CDK5-mediated phosphorylation of Sirt2 contributes to depressive-like behavior induced by social defeat stress

Major depressive disorder (MDD) is a common, severe and recurrent psychiatric disorder worldwide; however, the underlying neuropathological mechanisms remain elusive. Histone deacetylases (HDACs) appear to play an essential role in depression. As the class III HDACs, Sirt1 and Sirt2 have attracted the most interest in the nervous system. Indeed, chronic stress decreased Sirt1 activity and down-regulated Sirt1 gene expression in MDD. Nevertheless, there is a paucity of literature on the role of Sirt2. To study the role of Sirt2 we established a MDD mouse model in wild type and Sirt2 knockout C57BL/6 mice using social defeat stress (SDS). We found that a lack of Sirt2 blocked the development of SDS-induced depressive-like behavior. Moreover, SDS led to Sirt2 phosphorylation in the amygdala without changing total Sirt2 levels, and blocking the phosphorylation of Sirt2 by CDK5 at serine residues 368 and 372 prevented SDS-induced depressive-like behavior and Sirt2 nuclear import. We also discovered that SDS-induced Sirt2 phosphorylation was involved in VTA-amygdala modulation using TetTag-pharmacogenetic method. These results suggest that CDK5 mediates phosphorylation of Sirt2 in the amygdala and contributes to the depressive-like behavior induced by SDS. This study highlights that inhibiting CDK5-dependent phosphorylation of Sirt2 at serine residues 368 and 372 by myristoylated membrane-permeabilising peptide (Sirt2-p), rather than using non-specific sirtuin inhibitors, may be a novel strategy for treating depression.     

Male Wistar rats are more susceptible to lasting social anxiety than Wild-type Groningen rats following social defeat stress during adolescence

Adolescence is an important period for the development of adult social competences. Social stress during adolescence may contribute not only to an inadequate social development but also to adult vulnerability to social anxiety. There seems to be a clear individual differentiation, however, in the vulnerability to the long-term negative consequences of social stress. The current study further explores this individual vulnerability and is aimed at the influence of social stress during adolescence on adult social anxiety and its context specificity. Rats from different strains (Wistar and Wild-type Groningen rats) were exposed to the resident-intruder paradigm five times during 10 min each in the period between postnatal day 45 and 58. Three and 7 weeks later, the animals were re-exposed to the context in the presence of either a dominant male or an anestrous female behind a wire mesh screen. Wistar rats that were socially defeated spent less time exploring the social stimulus in comparison with socially defeated Wild-type rats and their non-defeated controls. We conclude that the stressed Wistar rat shows signs of generalized social anxiety indicating that the Wistar rat can be considered as a vulnerable phenotype to effects of adolescent social stress.     

Interaction between isolation rearing and social development on exploratory behavior in male rats

The effect of isolation on exploratory behavior has been shown to differ depending on the developmental stages of male rats. However, there has been little systematic comparison of the frequencies and the patterns of exploratory behavior across the developmental stages. The present study assessed the frequencies of exploration using the emergence test and exploratory patterns in the open-field test in three developmental stages of male rats: juvenile, post-puberty, and adult. A lower propensity for exploration was observed in rats isolated during the juvenile stage, as assessed by increased latency and decreased duration of exploratory behaviors compared to pair-reared rats, and this tendency was maintained in adulthood. Altered patterns of exploratory behavior were demonstrated both in rats isolated in adulthood, who showed an increased active pattern, and those pair-reared following puberty, who shifted to a more passive pattern. However, rats isolated during the juvenile stage did not change their exploratory patterns following puberty. These results suggest that the changes in the exploratory pattern, which can be observed in adulthood, are associated with the emergence of adult-like dominance relationships. Juvenile-isolated rats did not show these changes following puberty, suggesting the importance of social interaction as juveniles for the ontogenetic emergence of behavioral flexibility implicated in the regulation of exploratory patterns.     

Individual differences in the effect of social defeat on anhedonia and histone acetylation in the rat hippocampus

Major depression is a growing problem worldwide with variation in symptoms and response to treatment. Individual differences in response to stress may contribute to such observed individual variation in behavior and pathology. Therefore, we investigated depressive-like behavior following exposure to repeated social defeat in a rat model of individual differences in response to novelty. Rats are known to exhibit either high locomotor activity and sustained exploration (high responders, HR) or low activity with minimal exploration (low responders, LR) in a novel environment. We measured anhedonia using the sucrose preference test in HR and LR rats following exposure to social defeat stress or in basal, non-defeated conditions. We then compared histone acetylation in the hippocampus in HR and LR defeat and non-defeated rats and measured mRNA levels of histone deacetylases (HDAC) 3, 4, 5, and Creb binding protein (CBP). We found that basally, HR rats consumed more sucrose solution than LR rats, but reduced consumption after exposure to defeat. LR rats' preference was unaffected by social defeat. We found that HR rats had higher levels of histone acetylation on H3K14 and H2B than LR rats in non-stress conditions. Following defeat, this acetylation pattern changed differentially, with HR rats decreasing acetylation of H3K14 and H2B and LR's increasing acetylation of H3K14. Acetylation on histone H4 decreased following defeat with no individual variation. Basal differences in CBP expression levels may underlie the observed acetylation pattern; however we found no significant effects of defeat in levels of HDACs 3, 4, 5 in the hippocampus.     

Behavioral specificity of non-genomic glucocorticoid effects in rats: effects on risk assessment in the elevated plus-maze and the open-field

The rapid effects of glucocorticoids on various behaviors suggest that these hormones play a role in rapidly coping with challenging situations. The variety of behaviors affected in different situations raise, however, questions regarding the specificity and roles of glucocorticoids in controlling behavior. To clarify this issue, we assessed the rapid behavioral effects of glucocorticoids in the elevated plus-maze (EPM) and the open-field (OF) tests in male rats. Both tests measure three different kinds of behavioral responses: locomotion, anxiety-like behaviors (central area and open arm exploration in the OF and EPM tests, respectively), and risk assessment (investigating aversive areas in a stretched attend posture). The acute inhibition of glucocorticoid synthesis by metyrapone decreased risk assessment but did not affect locomotion and anxiety-like behaviors. Corticosterone administration increased risk assessment, without affecting locomotion and anxiety-like behaviors. Moreover, plasma corticosterone levels measured immediately after testing strongly correlated with the intensity of risk assessment. The effects of corticosterone were rapid, as occurred even when the hormone was injected 2 min before behavioral testing. In addition, the effect was resistant to protein synthesis inhibition. These data demonstrate that glucocorticoids are able to increase specifically risk assessment behaviors by non-genomic mechanisms in two different, novelty-related, non-social challenging situations. Thus, glucocorticoids appear to rapidly induce specific behavioral adjustments to meet immediate requirements set by the challenge. These data support earlier assumptions on the role of glucocorticoids in coping, and it can be hypothesized that the rapid activation of the HPA-axis may play a role in forming coping responses.     

Anxiolytic-like effects of extracts from Albizzia julibrissin bark in the elevated plus-maze in rats

The purpose of the this study was to characterize the putative anxiolytic-like effects of the aqueous extract of Albizzia julibrissin stem bark using the elevated plus maze (EPM) in rats. The water extract of Albizzia julibrissin was orally administered at 10, 50, 100 or 200 mg/kg to adult male SD rats, 1 h before behavioral evaluation in an EPM, respectively. Control rats were treated with an equal volume of saline, and positive control rats buspirone (1 mg/kg). Single or repeated treatment (for 7 days) of the water extract of Albizzia julibrissin (at 100 or 200 mg/kg) significantly increased time-spent and arm entries into the open arms of the EPM, and decreased time-spent and arm entries in the closed arms of the EPM versus saline controls (P < 0.05). However, no changes in the locomotor activity and myorelaxant effect were seen in any group versus the saline control. In addition, the anxiolytic-like effects of Albizzia julibrissin extract were abolished by pindolol (10 mg/kg, i.p), a 5-HT(1A/1B) receptor antagonist. These results suggest that Albizzia julibrissin might proved to be an effective anxiolytic agent, and that it acts via the serotonergic nervous system.     

Transgenerational effects of social stress on social behavior,corticosterone, oxytocin,and prolactin in rats

Social stressors such as depressed maternal care and family conflict are robust challenges which can have long-term physiological and behavioral effects on offspring and future generations. The current study investigates the transgenerational effects of an ethologically relevant chronic social stress on the behavior and endocrinology of juvenile and adult rats. Exposure to chronic social stress during lactation impairs maternal care in F0 lactating dams and the maternal care of the F1 offspring of those stressed F0 dams. The overall hypothesis was that the male and female F2 offspring of stressed F1 dams would display decreased social behavior as both juveniles and adults and that these behavioral effects would be accompanied by changes in plasma corticosterone, prolactin, and oxytocin. Both the female and male F2 offspring of dams exposed to chronic social stress displayed decreased social behavior as juveniles and adults, and these behavioral effects were accompanied by decreases in basal concentrations of corticosterone in both sexes, as well as elevated juvenile oxytocin and decreased adult prolactin in the female offspring. The data support the conclusion that social stress has transgenerational effects on the social behavior of the female and male offspring which are mediated by changes in the hypothalamic–pituitary–adrenal axis and hypothalamic–pituitary–gonadal axis. Social stress models are valuable resources in the study of the transgenerational effects of stress on the behavioral endocrinology of disorders such as depression, anxiety, autism, and other disorders involving disrupted social behavior.     

Chronic social defeat stress-induced enhancement of T-type calcium channels increases burst-firing neurons in the ventral subiculum

The ventral subiculum (vSub), a representative output structure of the hippocampus, serves as a main limbic region in mediating the brain's response to stress. There are three subtypes of subicular pyramidal neurons based on their firing patterns: regular-spiking (RS), weak-bursting (WB) and strong-bursting (SB) neurons, located differently along proximal–distal axis. Here, we found that chronic social defeat stress (CSDS) in mice increased the population of SB neurons but decreased RS neurons in the proximal vSub. Specific blockers of T-type calcium channels inhibited the burst firings with a concomitant reduction of afterdepolarization, suggesting that T-type calcium channels underlie the burst-spiking activity. Consistently, CSDS increased both T-type calcium currents and expression of Cav3.1 proteins, a subtype of T-type calcium channels, in the proximal vSub. Therefore, we conclude that CSDS-induced enhancement of Cav3.1 expression increased bursting neuronal population in the vSub, which may contribute to stress-related behaviors.     

Temperament moderates the influence of periadolescent social experience on behavior and adrenocortical activity in adult male rats

Adolescence is a period of significant behavioral and physiological maturation, particularly related to stress responses. Animal studies that have tested the influence of adolescent social experiences on stress-related behavioral and physiological development have led to complex results. We used a rodent model of neophobia to test the hypothesis that the influence of adolescent social experience on adult behavior and adrenocortical function is modulated by pre-adolescent temperament. Exploratory activity was assessed in 53 male Sprague–Dawley rats to classify temperament and then they were housed in one of the three conditions during postnatal days (PND) 28–46: (1) with familiar kin, (2) with novel social partners, or (3) individually with no social partners. Effects on adult adrenocortical function were evaluated from fecal samples collected while rats were individually-housed and exposed to a 1-hour novel social challenge during PND 110–114. Adolescent-housing with novel or no social partners led to reduced adult glucocorticoid production compared to adolescent-housing with familiar littermates. Additionally, highly-exploratory pre-weanling rats that were housed with novel social partners during adolescence exhibited increased exploratory behavior and a more rapid return to basal glucocorticoid production in adulthood compared to those housed with familiar or no social partners during adolescence and compared to low-exploratory rats exposed to novel social partners. In sum, relatively short-term adolescent social experiences can cause transient changes in temperament and potentially longer-term changes in recovery of glucocorticoid production in response to adult social challenges. Furthermore, early temperament may modulate the influence of adolescent experiences on adult behavioral and adrenocortical function.     

Differential long-term effects of social stress during adolescence on anxiety in Wistar and wild-type rats

Severe and chronic stress may interfere with adolescent neuronal plasticity that turns the juvenile brain into an adult brain increasing the vulnerability to develop anxiety disorders. It is well-known from adult stress research that there is a large individual differentiation in stress vulnerability. The current study is aimed at the individual resilience and vulnerability to adolescent social stress. Two strains of rats that differ in social behavioral skills were subjected to social stress during adolescence. In three experiments we studied short and long term effects of adolescent social stress using a water conflict test in different contexts. Wistar rats which had been socially defeated on postnatal days 45 and 46 showed, following water deprivation, a strong decrease in the total amount of water consumed and time spent drinking when tested 2 days and 3 weeks later in the context where they received the defeat experience. Also a strong increase in drinking latency was noticed in the context of the previous defeat. No differences in these parameters were found between defeated and non-defeated wild-type rats. The results of the water conflict test in an environment where no association with the previous defeat experience was present showed that the adolescent social stress did not induce a generalized anxiety.In conclusion, the water conflict test is a useful tool to measure the influence of social defeat on the motivation to obtain resources under conditions with different stimulus properties. In addition, our data suggest the importance of the strain used in adolescent stress experiments. The fact that Wistar rats showed a strong association with the context at adulthood whereas no effect was observed in the wild-type rats shows that victim characteristics are important determining factors for the long term effects of adolescent social stress.     

Gonadal hormones modulate the display of conditioned defeat in male Syrian hamsters

It has been widely reported that gonadal hormones influence the display of aggression in Syrian hamsters; conversely, much less is known about whether gonadal hormones modulate submissive/defensive behaviors in these animals. Following social defeat, male hamsters no longer display normal territorial aggression but instead display submissive/defensive behavior in the presence of a smaller opponent, a phenomenon we have termed conditioned defeat (CD). The purpose of the present study was to examine the effect of gonadal hormones on the display of CD in male hamsters. In Experiment 1, males were castrated or sham-operated. The castrated males were significantly more submissive following social defeat relative to their intact counterparts. The increased submissive behavior in the castrated males during CD testing was particularly surprising, given the fact that they were attacked significantly less during CD training. In Experiment 2a, males were castrated and given hormone replacement. Castrated males treated with testosterone or dihydrotestosterone displayed significantly less submissive behavior following social defeat than did those treated with cholesterol or estradiol. Finally, in Experiment 2b, there was no effect of hormone replacement on aggressive behavior in non-defeated hamsters suggesting that the decrease in submissive behavior in males treated with dihydrotestosterone or testosterone is specific to being previously defeated. Taken together the data indicate that the presence of androgens reduces the display of submission in defeated male hamsters. More importantly, these findings suggest that androgens may have a protective effect against the development of depression-like or anxiety-like behaviors following exposure to an ethologically relevant stressor.