Experiential contributions to social dominance in a rat model of fragile-X syndrome.

Social withdrawal is one phenotypic feature of the monogenic neurodevelopmental disorder fragile-X. Using a 'knockout' rat model of fragile-X, we examined whether deletion of the Fmr1 gene that causes this condition would affect the ability to form and express a social hierarchy as measured in a tube test. Male fragile-X 'knockout' rats living together could successfully form a social dominance hierarchy, but were significantly subordinate to wild-type animals in mixed group cages. Over 10 days of repeated testing, the fragile-X mutant rats gradually showed greater variance and instability of rank during their tube-test encounters. This affected the outcome of future encounters with stranger animals from other cages, with the initial phenotype of wild-type dominance lost to a more complex picture that reflected, regardless of genotype, the prior experience of winning or losing. Our findings offer a novel insight into the complex dynamics of social interactions between laboratory living groups of fragile-X and wild-type rats. Even though this is a monogenic condition, experience has an impact upon future interactions with other animals. Gene/environment interactions should therefore be considered in the development of therapeutics.

Repeated social stress leads to contrasting patterns of structural plasticity in the amygdala and hippocampus.

Previous studies have demonstrated that repeated immobilization and restraint stress cause contrasting patterns of dendritic reorganization as well as alterations in spine density in amygdalar and hippocampal neurons. Whether social and ethologically relevant stressors can induce similar patterns of morphological plasticity remains largely unexplored. Hence, we assessed the effects of repeated social defeat stress on neuronal morphology in basolateral amygdala (BLA), hippocampal CA1 and infralimbic medial prefrontal cortex (mPFC). Male Wistar rats experienced social defeat stress on 5 consecutive days during confrontation in the resident-intruder paradigm with larger and aggressive Wild-type Groningen rats. This resulted in clear social avoidance behavior one day after the last confrontation. To assess the morphological consequences of repeated social defeat, 2 weeks after the last defeat, animals were sacrificed and brains were stained using a Golgi-Cox procedure. Morphometric analyses revealed that, compared to controls, defeated Wistar rats showed apical dendritic decrease in spine density on CA1 but not BLA. Sholl analysis demonstrated a significant dendritic atrophy of CA1 basal dendrites in defeated animals. In contrast, basal dendrites of BLA pyramidal neurons exhibited enhanced dendritic arborization in defeated animals. Social stress failed to induce lasting structural changes in mPFC neurons. Our findings demonstrate for the first time that social defeat stress elicits divergent patterns of structural plasticity in the hippocampus versus amygdala, similar to what has previously been reported with repeated physical stressors. Therefore, brain region specific variations may be a universal feature of stress-induced plasticity that is shared by both physical and social stressors.

A role for the extended amygdala in the fear-enhancing effects of acute selective serotonin reuptake inhibitor treatment.

Selective serotonin reuptake inhibitors (SSRIs) are reported to exacerbate symptoms of anxiety when treatment is initiated. These clinical findings have been extended to animal models wherein SSRIs also potentiate anxiety and fear learning, which depend on the amygdala. Yet, little is known about the role of specific amygdalar circuits in these acute effects of SSRIs. Here, we first confirmed that a single injection of fluoxetine 1 h before auditory fear conditioning potentiated fear memory in rats. To probe the neural substrates underlying this enhancement, we analyzed the expression patterns of the immediate early gene, Arc (activity-regulated cytoskeleton-associated protein). Consistent with previous reports, fear conditioning induced Arc protein expression in the lateral and basal amygdala. However, this was not enhanced further by pre-treatment with fluoxetine. Instead, fluoxetine significantly enhanced expression of Arc in the central amygdala (CeA) and the bed nucleus of the stria terminalis (BNST). Next, we tested whether direct targeted infusions of fluoxetine into the CeA, or BNST, leads to the same fear-potentiating effect. Strikingly, direct infusion of fluoxetine into the BNST, but not the CeA, was sufficient to enhance fear memory. Moreover, this behavioral effect was also accompanied by robust Arc expression in the CeA, similar to the systemic injection. Our results identify a novel role for the BNST in the acute fear-enhancing effects of SSRIs. These findings highlight the need to look beyond the traditional focus on input nuclei of the amygdala and add to accumulating evidence implicating these microcircuits in gating fear and anxiety.

Disruption of fatty acid amide hydrolase activity prevents the effects of chronic stress on anxiety and amygdalar microstructure.

Hyperactivation of the amygdala following chronic stress is believed to be one of the primary mechanisms underlying the increased propensity for anxiety-like behaviors and pathological states; however, the mechanisms by which chronic stress modulates amygdalar function are not well characterized. The aim of the current study was to determine the extent to which the endocannabinoid (eCB) system, which is known to regulate emotional behavior and neuroplasticity, contributes to changes in amygdalar structure and function following chronic stress. To examine the hypothesis, we have exposed C57/Bl6 mice to chronic restraint stress, which results in an increase in fatty acid amide hydrolase (FAAH) activity and a reduction in the concentration of the eCB N-arachidonylethanolamine (AEA) within the amygdala. Chronic restraint stress also increased dendritic arborization, complexity and spine density of pyramidal neurons in the basolateral nucleus of the amygdala (BLA) and increased anxiety-like behavior in wild-type mice. All of the stress-induced changes in amygdalar structure and function were absent in mice deficient in FAAH. Further, the anti-anxiety effect of FAAH deletion was recapitulated in rats treated orally with a novel pharmacological inhibitor of FAAH, JNJ5003 (50 mg per kg per day), during exposure to chronic stress. These studies suggest that FAAH is required for chronic stress to induce hyperactivity and structural remodeling of the amygdala. Collectively, these studies indicate that FAAH-mediated decreases in AEA occur following chronic stress and that this loss of AEA signaling is functionally relevant to the effects of chronic stress. These data support the hypothesis that inhibition of FAAH has therapeutic potential in the treatment of anxiety disorders, possibly by maintaining normal amygdalar function in the face of chronic stress.

The neurobiological properties of tianeptine (Stablon): from monoamine hypothesis to glutamatergic modulation.

Tianeptine is a clinically used antidepressant that has drawn much attention, because this compound challenges traditional monoaminergic hypotheses of depression. It is now acknowledged that the antidepressant actions of tianeptine, together with its remarkable clinical tolerance, can be attributed to its particular neurobiological properties. The involvement of glutamate in the mechanism of action of the antidepressant tianeptine is consistent with a well-developed preclinical literature demonstrating the key function of glutamate in the mechanism of altered neuroplasticity that underlies the symptoms of depression. This article reviews the latest evidence on tianeptine's mechanism of action with a focus on the glutamatergic system, which could provide a key pathway for its antidepressant action. Converging lines of evidences demonstrate actions of tianeptine on the glutamatergic system, and therefore offer new insights into how tianeptine may be useful in the treatment of depressive disorders.

Lithium treatment prevents stress-induced dendritic remodeling in the rodent amygdala.

Amygdala function is altered in patients with bipolar disorder (BD), but may be normalized by treatment with mood stabilizers. Lithium remains the most effective mood stabilizing therapy for BD, but the relevance of its neuroprotective effects in pre-clinical studies to clinical outcomes is unknown, and the targeting of amygdalar neurons by therapeutic interventions for BD has not yet been examined. Chronic stress in rodents increases activation of the amygdala and induces dendritic hypertrophy, thus providing a quantifiable marker of neuronal structural pathology that may be reversed by lithium treatment. Rats underwent restraint stress for 21 days, with or without concurrent administration of lithium in their diet. The overall length and complexity of neuronal dendritic arbors of principal pyramidal neurons in the basolateral amygdala were quantified using Golgi-Cox impregnation and three-dimensional neuron tracing. Lithium treatment prevented stress-induced increases in dendritic branching of amygdalar pyramidal neurons by reducing total dendritic length (18.0%; P=0.006) and the number of dendritic branch points (21.0%; P=0.02). Despite its protective effect when administered during stress, lithium did not alter amygdalar dendritic morphology when administered to non-stressed control rats. Our results demonstrate that lithium attenuates structural remodeling in the amygdala during stress, but has contrasting effects on neuronal morphology under pathological versus healthy conditions. This may reflect an ability of lithium to stabilize excitatory neurotransmission in the amygdala of individuals with BD, reducing the need for compensatory adjustments of dendritic architecture.

Stress-induced changes in sleep and associated neuronal activity in rat hippocampus and amygdala.

Stress increases vulnerability to anxiety and depression. We have investigated the effect of acute immobilization stress in amygdalohippocampal circuits by measuring the electroencephalogram (EEG) in male Wistar rats during rapid eye movement (REM) sleep. Electrodes were implanted stereotaxically in the hippocampus (CA1 and CA3 subregions of the hippocampus) and the amygdala (lateral nucleus). Prior to the stress, two baseline recordings were taken. Twenty-four hours later rats were exposed once to acute immobilization stress (AIS) session for 2 h. After the release and on subsequent days, electrophysiological changes that occurred due to stress during REM sleep were analyzed by comparing them with baseline measurements. Our results suggest that acute immobilization stress induced significant increase in REM sleep in the first 24 h after the exposure. In addition to changes in the sleep patterns, we have observed increased theta oscillations in CA1 area of the hippocampus with decreased coherence at theta range (4-8 Hz) between hippocampus and amygdala. These results suggest that single exposure to aversive experience such as immobilization stress can lead to dynamic changes in neuronal activities with altered sleep morphology. The results obtained in the present study are comparable to those seen in human patients suffering from panic, and anxiety due to posttraumatic stress disorder (PTSD).

Hypobaric hypoxia-induced dendritic atrophy of hippocampal neurons is associated with cognitive impairment in adult rats.

Simulated hypobaric hypoxia (HBH), resembling high altitude hypoxia severely affects the CNS and results in several physiological changes. The hippocampus is closely associated with learning and memory and an insult to this region affects cognition. Previous studies suggest that rapid or prolonged exposures to HBH are associated with psychomotor and cognitive impairments. The defense personnel, mountain climbers and rescue teams are exposed to such harsh environment and thus it demands a systematic study emphasizing the subtle effects of such extreme environments on cognitive function. Accordingly, this study evaluated the effect of hypobaric hypoxia on structural changes in the principal neurons of the hippocampus and learning in eight-arm radial maze. Adult male Wistar rats, subjected to simulated hypobaric hypoxia equivalent to an altitude of 6000 m for a period of 2 or 7 days, in a hypoxic chamber served as hypoxic group (HY). Rats housed in a similar chamber for the same period of time, without hypoxic exposure served as sham control (SC), while normal control (NC) group of rats were housed in standard laboratory conditions. The dendritic morphology of neurons in cornu ammonis region 1 (CA1) and cornu ammonis region 3 (CA3) was studied in Golgi-impregnated hippocampal sections. Exposure for 2 days to hypobaric hypoxia had minimal deleterious effects on the CA1 pyramidal neurons, while exposure for 7 days resulted in a significant decrease in the number of branching points, intersections and dendritic length. Unlike the CA1 pyramidal neurons, the CA3 neurons exhibited dendritic atrophy following both 2 and 7 days of hypoxic exposure. Further, hippocampal-dependent spatial learning was affected marginally following 2 day exposure, while 7 day exposure severely affected learning of the partially baited radial arm maze task. Our study suggests that dendritic atrophy in the hippocampus on exposure to HBH could be one of the bases for the cognitive deficits exhibited under such conditions.

Stress-induced spine loss in the medial amygdala is mediated by tissue-plasminogen activator.

The amygdala, which exerts a regulatory influence on the stress response, is itself affected by stress. It has been reported that the serine protease tissue-plasminogen activator (tPA), a key mediator of spine plasticity, is required for stress-induced facilitation of anxiety-like behavior. Importantly, tPA is also involved in stress-induced activation of molecular signals that have the potential to contribute to neuronal remodeling in the medial amygdala (MeA). However, little is known about the precise nature of, and specific role played by tPA in, stress-induced structural plasticity in the MeA. Hence, we compared the impact of chronic restraint stress on spine density of medium spiny stellate neurons in MeA in wild-type mice with mice in which the tPA gene is disrupted (tPA-/-). In wild-type mice, chronic stress caused significant reduction in MeA spine density, which was in contrast to enhanced spine density in the neighboring basolateral amygdala (BLA). Strikingly, tPA-/- mice exhibited significant attenuation of stress-induced spine retraction in the MeA, but BLA spinogenesis was not affected. Therefore, tPA-dependence of stress-induced modulation in spine density was restricted to the MeA. Further, MeA neurons in tPA-/- mice, even when challenged with repeated stress, were able to maintain levels of spine density that were comparable to that of wild-type mice without stress. Our findings provide novel evidence for a permissive role for tPA in amygdalar spine plasticity elicited by behavioral stress.

Prolonged behavioral stress enhances synaptic connectivity in the basolateral amygdala.

Recently identified cellular and molecular correlates of stress-induced plasticity suggest a putative link between neuronal remodeling in the amygdala and the development of anxiety-like behavior. Rodent models of immobilization stress, applied for 10 consecutive days, have been reported to enhance anxiety, and also cause dendritic elongation and spine formation in the basolateral amygdala (BLA). Paradoxically, longer exposure to stress, which is also anxiogenic, fails to affect key molecular markers of neuronal remodeling in the BLA. This has raised the possibility of homeostatic mechanisms being triggered by more prolonged stress that could potentially dampen the morphological effects of stress in the BLA. Therefore, we examined the cellular and behavioral impact of increasing the duration of stress in rats. We find that prolonged immobilization stress (PIS), spanning 21 days, caused significant enhancement in dendritic arborization of spiny BLA neurons. Spine density was also enhanced along these elongated dendrites in response to PIS. Finally, this striking increase in synaptic connectivity was accompanied by enhanced anxiety-like behavior in the elevated plus-maze. Thus, we did not detect any obvious morphological correlate of adaptive changes within the BLA that may have been activated by prolonged and repeated application of the same stressor for 21 days. These findings add to accumulating evidence that structural encoding of aversive experiences, through enhanced availability of postsynaptic dendritic surface and synaptic inputs on principal neurons of the BLA, may contribute to the affective symptoms of stress disorders.

Recovery after chronic stress fails to reverse amygdaloid neuronal hypertrophy and enhanced anxiety-like behavior.

The hippocampus and amygdala are important components of the neural circuitry mediating stress responses. While structural plasticity in the hippocampus may mediate cognitive aspects of behavioral impairments caused by severe stress, changes in the amygdala are more likely to contribute to the affective aspects of stress disorders. Recent reports have identified cellular and molecular correlates of stress-induced amygdaloid plasticity that may underlie anxiety. Hence, we examined the impact of chronic stress, in terms of its duration, at the cellular and behavioral levels in rats. We found that, even after 21 days of stress-free recovery, animals exposed to chronic immobilization stress (CIS) continue to exhibit enhanced anxiety, as manifested by a significant reduction in open-arm exploration and risk-assessment behavior in the elevated plus-maze. At the cellular level, we tested if CIS-induced dendritic remodeling in the amygdala is also as long-lasting as enhanced anxiety after 21 days of recovery. Indeed, long-lasting facilitation of CIS-induced anxiety is accompanied by a persistent increase in dendritic arborization of basolateral amygdala (BLA) spiny neurons. Moreover, CIS-induced BLA hypertrophy is distinct from hippocampal CA3 atrophy, which is reversible within the same period of stress-free recovery. These findings on persistent dendritic remodeling in the amygdala, in addition to highlighting important differences with hippocampal structural plasticity, may provide a cellular basis for examining anxiety and mood disorders triggered by chronic stress.

Forebrain-specific calcineurin knockout selectively impairs bidirectional synaptic plasticity and working/episodic-like memory.

Calcineurin is a calcium-dependent protein phosphatase that has been implicated in various aspects of synaptic plasticity. By using conditional gene-targeting techniques, we created mice in which calcineurin activity is disrupted specifically in the adult forebrain. At hippocampal Schaffer collateral-CA1 synapses, LTD was significantly diminished, and there was a significant shift in the LTD/LTP modification threshold in mutant mice. Strikingly, although performance was normal in hippocampus-dependent reference memory tasks, including contextual fear conditioning and the Morris water maze, the mutant mice were impaired in hippocampus-dependent working and episodic-like memory tasks, including the delayed matching-to-place task and the radial maze task. Our results define a critical role for calcineurin in bidirectional synaptic plasticity and suggest a novel mechanistic distinction between working/episodic-like memory and reference memory.

2-Amino-3-phosphonopropionic acid, an inhibitor of glutamate-stimulated phosphoinositide turnover, blocks induction of homosynaptic long-term depression, but not potentiation, in rat hippocampus.

Recently, we have reported an associative long-term depression (LTD) of synaptic strength in hippocampal field CA1 that is produced when a low-frequency test input is negatively correlated in time with a high-frequency conditioning input. We have also found that pairing of synaptic activity with postsynaptic hyperpolarization is sufficient to induce LTD. We report here that 2-amino-3-phosphonopropionate (AP3), a selective inhibitor of phosphoinositide (PI) turnover mediated by the metabotropic glutamate receptor, blocks the induction of associative LTD in hippocampal field CA1, but does not impair the induction of LTP. Our data suggest that metabotropic glutamate receptor activation is involved in the induction of LTD.

Commissural synapses, but not mossy fiber synapses, in hippocampal field CA3 exhibit associative long-term potentiation and depression.

When CA3 commissural afferents received low-frequency (weak) stimuli synchronized with a train of mossy fiber bursts (strong), associative long-term potentiation (LTP) was induced at mixed commissural and associational synapses on hippocampal CA3 pyramidal cells in vitro. In contrast, a weak mossy fiber input did not potentiate when given in phase with commissural/associational bursts. Furthermore, commissural/associational synapses receiving low-frequency stimuli out-of-phase with strong rhythmic mossy fiber input showed associative long-term depression (LTD), whereas mossy fiber synaptic strengths were not depressed when they received weak inputs out-of-phase with a strong commissural/associational input. Thus, both associative LTP and associative LTD can be induced at commissural/associational synapses, but not at mossy fiber synapses.