Optogenetic and chemogenetic approaches reveal differences in neuronal circuits that mediate initiation and maintenance of social interaction.

For social interaction to be successful, two conditions must be met: the motivation to initiate it and the ability to maintain it. This study uses both optogenetic and chemogenetic approaches to reveal the specific neural pathways that selectively influence those two social interaction components.

Astrocytic ß-catenin signaling via TCF7L2 regulates synapse development and social behavior.

The Wnt/ß-catenin pathway contains multiple high-confidence risk genes that are linked to neurodevelopmental disorders, including autism spectrum disorder. However, its ubiquitous roles across brain cell types and developmental stages have made it challenging to define its impact on neural circuit development and behavior. Here, we show that TCF7L2, which is a key transcriptional effector of the Wnt/ß-catenin pathway, plays a cell-autonomous role in postnatal astrocyte maturation and impacts adult social behavior. TCF7L2 was the dominant Wnt effector that was expressed in both mouse and human astrocytes, with a peak during astrocyte maturation. The conditional knockout of Tcf7l2 in postnatal astrocytes led to an enlargement of astrocytes with defective tiling and gap junction coupling. These mice also exhibited an increase in the number of cortical excitatory and inhibitory synapses and a marked increase in social interaction by adulthood. These data reveal an astrocytic role for developmental Wnt/ß-catenin signaling in restricting excitatory synapse numbers and regulating adult social behavior.

Social buffering diminishes fear response but does not equal improved fear extinction.

Social support during exposure-based psychotherapy is believed to diminish fear and improve therapy outcomes. However, some clinical trials challenge that notion. Underlying mechanisms remain unknown, hindering the understanding of benefits and pitfalls of such approach. To study social buffering during fear extinction, we developed a behavioral model in which partner's presence decreases response to fear-associated stimuli. To identify the neuronal background of this phenomenon, we combined behavioral testing with c-Fos mapping, optogenetics, and chemogenetics. We found that the presence of a partner during fear extinction training causes robust inhibition of freezing; the effect, however, disappears in subjects tested individually on the following day. It is accompanied by lowered activation of the prelimbic (PL) and anterior cingulate (ACC) but not infralimbic (IL) cortex. Accordingly, blocking of IL activity left social buffering intact. Similarly, inhibition of the ventral hippocampus-PL pathway, suppressing fear response after prolonged extinction training, did not diminish the effect. In contrast, inhibition of the ACC-central amygdala pathway, modulating social behavior, blocked social buffering. By reporting that social modulation of fear inhibition is transient and insensitive to manipulation of the fear extinction-related circuits, we show that the mechanisms underlying social buffering during extinction are different from those of individual extinction.

Social deficits in BTBR T+ Itpr3tf/J mice vary with ecological validity of the test.

Translational value of mouse models of neuropsychiatric disorders depends heavily on the accuracy with which they replicate symptoms observed in the human population. In mouse models of autism spectrum disorder (ASD) these include, among others, social affiliation, and communication deficits as well as impairments in understanding and perception of others. Most studies addressing these issues in the BTBR T+ Itpr3tf/J mouse, an idiopathic model of ASD, were based on short dyadic interactions of often non-familiar partners placed in a novel environment. In such stressful and variable conditions, the reproducibility of the phenotype was low. Here, we compared physical conditions and the degree of habituation of mice at the time of testing in the three chambered social affiliation task, as well as parameters used to measure social deficits and found that both the level of stress and human bias profoundly affect the results of the test. To minimize these effects, we tested social preference and network dynamics in mice group-housed in the Eco-HAB system. This automated recording allowed for long-lasting monitoring of differences in social repertoire (including interest in social stimuli) in BTBR T+ Itpr3tf/J and normosocial c57BL/6J mice. With these observations we further validate the BTBR T+ Itpr3tf/J mouse as a model for ASD, but at the same time emphasize the need for more ecological testing of social behavior within all constructs of the Systems for Social Processes domain (as defined by the Research Domain Criteria framework).

Distinct circuits in rat central amygdala for defensive behaviors evoked by socially signaled imminent versus remote danger.

Animals display a rich repertoire of defensive responses adequate to the threat proximity. In social species, these reactions can be additionally influenced by the behavior of fearful conspecifics. However, the majority of neuroscientific studies on socially triggered defensive responses focuses on one type of behavior, freezing. To study a broader range of socially triggered reactions and underlying mechanisms, we directly compared two experimental paradigms, mimicking occurrence of the imminent versus remote threat. Observation of a partner currently experiencing aversive stimulation evokes passive defensive responses in the observer rats. Similar interaction with a partner that has just undergone the aversive stimulation prompts animals to increase active exploration. Although the observers display behaviors similar to those of the aversively stimulated demonstrators, their reactions are not synchronized in time, suggesting that observers' responses are caused by the change in their affective state rather than mimicry. Using opsins targeted to behaviorally activated neurons, we tagged central amygdala (CeA) cells implicated in observers' responses to either imminent or remote threat and reactivated them during the exploration of a novel environment. The manipulation revealed that the two populations of CeA cells promote passive or active defensive responses, respectively. Further experiments confirmed that the two populations of cells at least partially differ in expression of molecular markers (protein kinase C-δ [PKC-δ] and corticotropin-releasing factor [CRF]) and connectivity patterns (receiving input from the basolateral amygdala or from the anterior insula). The results are consistent with the literature on single subjects' fear conditioning, suggesting that similar neuronal circuits control defensive responses in social and non-social contexts.

Social Transfer of Fear in Rodents.

Social transfer of fear is a potent tool facilitating response to danger in animals forming social groups. With many factors influencing the transfer-such as proximity of the animal receiving information to the donor, familiarity, proximity of danger, and species-specific coping strategies-it allows studies of neuronal correlates of a variety of behavioral responses. Since both the transfer of fear and social modulation of fear responses are impaired in many neuropsychological disorders, the models described in this article could be useful in disentangling the neuronal circuitry involved in the pathogenesis of these disorders. © 2019 by John Wiley & Sons, Inc. Basic Protocol 1: Imminent threat in rats Alternate Protocol 1: Imminent threat in mice Basic Protocol 2: Remote threat in rats Alternate Protocol 2: Remote threat in mice Basic Protocol 3: Social modulation of fear extinction in rats Alternate Protocol 3: Social modulation of fear extinction in mice.

Risk assessment and serotonin: Animal models and human psychopathologies.

Risk assessment (RA) is an evolved, generally adaptive, mechanism comprising focused attention and appraisal of potential threat stimuli and situations. Initially characterized in animal models, it provides a number of behavioral and functional parallels to patterns of rumination, gaze biases, and other forms of affective cognition that appear to be disregulated in depression and anxiety. Serotonergic mechanisms are involved in these mood disorders, and an emerging body of evidence suggests that they may modulate the affective cognitive changes common to such psychopathologies. Findings of parallel effects of serotonin systems in RA would support a view that it may provide a useful behavioral endophenotype for translational research on mood disorders. This review examines the involvement of serotonergic mechanisms in both animal models of RA, and in an array of tasks focusing on affective cognitive changes in individuals with depression or anxiety. Results suggest substantial serotonin involvement in both RA behaviors measured in rats or mice, and in the "intersection of emotional and cognitive processes" [43] in people.

What can rodents teach us about empathy?

While many consider empathy an exclusively human trait, non-human animals are capable of simple forms of empathy, such as emotional contagion, as well as consolation and helping behavior. Rodent models are particularly useful for describing the neuronal background of these phenomena. They offer the possibility of employing single-cell resolution mapping of the neuronal activity as well as novel techniques for manipulation of in vivo activity, which are currently unavailable in human studies. Here, we review recent developments in the field of rodent empathy research with special emphasis on behavioral paradigms and data on neuronal correlates of emotional contagion. We hope that the use of rodent models will enhance our understanding of social deficits in neuropsychiatric disorders characterized with empathy impairments and the evolutionary continuity of the empathic trait.

Why mother rats protect their children.

The presence of the hormone oxytocin in the central amygdala makes a mother rat willing to put her life in danger in order to protect her offspring.

Matrix Metalloproteinase-9 and Synaptic Plasticity in the Central Amygdala in Control of Alcohol-Seeking Behavior.

BACKGROUND: Dysfunction of the glutamatergic system has been implicated in alcohol addiction; however, the molecular underpinnings of this phenomenon are still poorly understood. In the current study we have investigated the possible function of matrix metalloproteinase-9 (MMP-9) in alcohol addiction because this protein has recently emerged as an important regulator of excitatory synaptic plasticity. METHODS: For long-term studies of alcohol drinking in mice we used IntelliCages. Dendritic spines were analyzed using Diolistic staining with DiI. Whole-cell patch clamp was used to assess silent synapses. Motivation for alcohol in human subjects was assessed on the basis of a Semi-Structured Assessment for the Genetics of Alcoholism interview. RESULTS: Mice devoid of MMP-9 (MMP-9 knockout) drank as much alcohol as wild-type animals; however, they were impaired in alcohol seeking during the motivation test and withdrawal. The deficit could be rescued by overexpression of exogenous MMP-9 in the central nucleus of the amygdala (CeA). Furthermore, the impaired alcohol seeking was associated with structural alterations of dendritic spines in the CeA and, moreover, whole-cell patch clamp analysis of the basal amygdala to CeA projections showed that alcohol consumption and withdrawal were associated with generation of silent synapses. These plastic changes were impaired in MMP-9 knockout mice. Finally, C/T polymorphism of MMP-9 gene at position -1562, which upregulates MMP-9 expression, correlated with increased motivation for alcohol in alcoholics. CONCLUSIONS: In aggregate, our results indicate a novel mechanism of alcohol craving that involves MMP-9-dependent synaptic plasticity in CeA.

Neuronal correlates of asocial behavior in a BTBR T (+) Itpr3(tf)/J mouse model of autism.

Autism spectrum disorder (ASD) is a neurodevelopmental disorder characterized, in part, by an inability to adequately respond to social cues. Patients diagnosed with ASD are often devoid of empathy and impaired in understanding other people's emotional perspective. The neuronal correlates of this impairment are not fully understood. Replicating such a behavioral phenotype in a mouse model of autism would allow us insight into the neuronal background of the problem. Here we tested BTBR T(+)Itpr3(tf)/J (BTBR) and c57BL/6J (B6) mice in two behavioral paradigms: the Transfer of Emotional Information test and the Social Proximity test. In both tests BTBR mice displayed asocial behavior. We analyzed c-Fos protein expression in several brain regions after each of these tests, and found that, unlike B6 mice, BTBR mice react to a stressed cagemate exposure in the Transfer of Emotional Information test with no increase of c-Fos expression in either the prefrontal cortex or the amygdala. However, after Social Proximity exposure we observed a strong increase in c-Fos expression in the CA3 field of the hippocampus and two hypothalamic regions of BTBR brains. This response was accompanied by a strong activation of periaqueductal regions related to defensiveness, which suggests that BTBR mice find unavoidable social interaction highly aversive.

A novel automated behavioral test battery assessing cognitive rigidity in two genetic mouse models of autism.

Repetitive behaviors are a key feature of many pervasive developmental disorders, such as autism. As a heterogeneous group of symptoms, repetitive behaviors are conceptualized into two main subgroups: sensory/motor (lower-order) and cognitive rigidity (higher-order). Although lower-order repetitive behaviors are measured in mouse models in several paradigms, so far there have been no high-throughput tests directly measuring cognitive rigidity. We describe a novel approach for monitoring repetitive behaviors during reversal learning in mice in the automated IntelliCage system. During the reward-motivated place preference reversal learning, designed to assess cognitive abilities of mice, visits to the previously rewarded places were recorded to measure cognitive flexibility. Thereafter, emotional flexibility was assessed by measuring conditioned fear extinction. Additionally, to look for neuronal correlates of cognitive impairments, we measured CA3-CA1 hippocampal long term potentiation (LTP). To standardize the designed tests we used C57BL/6 and BALB/c mice, representing two genetic backgrounds, for induction of autism by prenatal exposure to the sodium valproate. We found impairments of place learning related to perseveration and no LTP impairments in C57BL/6 valproate-treated mice. In contrast, BALB/c valproate-treated mice displayed severe deficits of place learning not associated with perseverative behaviors and accompanied by hippocampal LTP impairments. Alterations of cognitive flexibility observed in C57BL/6 valproate-treated mice were related to neither restricted exploration pattern nor to emotional flexibility. Altogether, we showed that the designed tests of cognitive performance and perseverative behaviors are efficient and highly replicable. Moreover, the results suggest that genetic background is crucial for the behavioral effects of prenatal valproate treatment.

The BTBR T+ tf/J mouse model for autism spectrum disorders-in search of biomarkers.

Autism spectrum disorders (ASD) form a common group of neurodevelopmental disorders appearing to be under polygenic control, but also strongly influenced by multiple environmental factors. The brain mechanisms responsible for ASD are not understood and animal models paralleling related emotional and cognitive impairments may prove helpful in unraveling them. BTBR T+ tf/J (BTBR) mice display behaviors consistent with the three diagnostic categories for ASD. They show impaired social interaction and communication as well as increased repetitive behaviors. This review covers much of the data available to date on BTBR behavior, neuroanatomy and physiology in search for candidate biomarkers, which could both serve as diagnostic tools and help to design effective treatments for the behavioral symptoms of ASD.

Local blockade of NMDA receptors in the rat prefrontal cortex increases c-Fos expression in multiple subcortical regions.

Ketamine, phencyclidine and MK801 are uncompetitive NMDA receptor (NMDAR) antagonists which are used widely to model certain features of schizophrenia in rats. Systemic administration of NMDAR antagonists, in addition to provoking an increase in c-Fos expression, leads to important neurochemical and electrophysiological changes within the medial prefrontal cortex (mPFC). Since the mPFC is considered to exert a top-down regulatory control of subcortical brain regions, we examined the effects of local infusion of the NMDAR antagonist, MK801, into the mPFC on the expression of c-Fos protein (widely used marker of neuronal activation) in several subcortical structures. The experiment was performed on freely moving rats, bilaterally implanted with guide cannulae in the prelimbic mPFC, infused with MK801 or saline. Bilateral administration of MK801 to the mPFC produced changes in the behavior (increased stereotypy and decreased sleep-like behavior) and complex changes in c-Fos protein expression with significant increases observed in the nucleus accumbens (core and shell), amygdala (basolateral and central nuclei), the CA1 field of the hippocampus, and mediodorsal and paraventricular thalamic nuclei, as compared to the saline group. Together, we demonstrate that blockade of NMDA receptors in the mPFC is sufficient to lead to behavioral abnormalities and increased c-Fos expression in many, but not all, of the subcortical structures examined. Our findings suggest that some of the behavioral abnormalities produced by uncompetitive NMDAR antagonists may result from aberrant activity in cortico-subcortical pathways. These data support an increasing body of literature, suggesting that the mPFC is an important site mediating the effects of NMDAR antagonists.

Absence of social conditioned place preference in BTBR T+tf/J mice: relevance for social motivation testing in rodent models of autism.

A major goal of translation research in autism is to characterize the physiological and psychological processes underlying behavioral abnormalities. Since autism reflects impairments in social motivation, we modified the mouse three-chamber social approach apparatus for use as a social conditioned place preference arena. We paired one of two unique contexts with social interactions in juvenile mice for five or ten conditioning sessions in BTBR T+tf/J mice and a control strain with normal approach behaviors (C57BL/6J) since the BTBR T+tf/J inbred mouse strain displays a variety of behavioral alterations analogous to symptoms of autism spectrum disorders. While C57BL/6J mice formed a conditioned place preference to the context associated with social interactions, particularly those receiving ten days of conditioning, BTBR T+tf/J mice did not. Neither absence of social proximity nor avoidance due to high rates of autogrooming appeared to underlie the impaired positive incentive value of the unconditioned social stimulus in the BTBR T+tf/J strain. These data contribute to a growing body of evidence suggesting that the BTBR T+tf/J strain shows impairments in all diagnostic domains of autism including social motivation. Additionally, social conditioning testing might provide an important social motivation measure in other rodent models of neuropsychiatric disorders characterized by social abnormalities.

Reduced sulfate plasma concentrations in the BTBR T+tf/J mouse model of autism.

Clinical studies have shown that children diagnosed with autism show abnormal sulfate chemistry, which is critical for cellular and metabolic processes. To determine if the inbred BTBR T+tf/J mouse shows autism-relevant aberrations in sulfate chemistry, the present study examined plasma sulfate concentrations in BTBR T+tf/J, inbred C57BL/6J, and outbred CD-1 mice. Results showed that the BTBR T+tf/J mouse exhibits significantly lower plasma sulfate concentrations in comparison to both C57BL/6J and CD-1 mice. These results suggest that the BTBR mouse shows autism-relevant abnormalities in sulfate chemistry and may serve additional utility in examining the role of sulfate and sulfate-dependent systems in relation to autism-relevant behavioral aberrations.

Fractone-associated N-sulfated heparan sulfate shows reduced quantity in BTBR T+tf/J mice: a strong model of autism.

BTBR T+tf/J (BTBR) mice show abnormal social, communicatory, and repetitive/stereotyped behaviors paralleling many of the symptoms of autism spectrum disorders. BTBR also show agenesis of the corpus callosum (CC) suggesting major perturbations of growth or guidance factors in the dorsal forebrain [1]. Heparan sulfate (HS) is a polysaccaride found in the brain and other animal tissues. It binds to a wide variety of ligands and through these ligands modulates a number of biological processes, including cell proliferation and differentiation, migration and guidance. It is aggregated on fractal-like structures (fractones) in the subventricular zone (SVZ), that may be visualized by laminin immunoreactivity (LAM-ir), as well as by HS immunoreactivity (HS-ir). We report that the lateral ventricles of BTBR mice were drastically reduced in area compared to C57BL/6J (B6) mice while the BTBR SVZ was significantly shorter than that of B6. In addition to much smaller fractones for BTBR, both HS and LAM-ir associated with fractones were significantly reduced in BTBR, and their anterior-posterior distributions were also altered. Finally, the ratio of HS to LAM in individual fractones was significantly higher in BTBR than in B6 mice. These data, in agreement with other findings linking HS to callosal development, suggest that variations in the quantity and distribution of HS in the SVZ of the lateral ventricles may be important modulators of the brain structural abnormalities of BTBR mice, and, potentially, contribute to the behavioral pathologies of these animals.

BTBR T+tf/J mice: autism-relevant behaviors and reduced fractone-associated heparan sulfate.

BTBR T+tf/J (BTBR) mice have emerged as strong candidates to serve as models of a range of autism-relevant behaviors, showing deficiencies in social behaviors; reduced or unusual ultrasonic vocalizations in conspecific situations; and enhanced, repetitive self-grooming. Recent studies have described their behaviors in a seminatural visible burrow system (VBS); a Social Proximity Test in which avoidance of a conspecific is impossible; and in an object approach and investigation test evaluating attention to specific objects and potential stereotypies in the order of approaching/investigating objects. VBS results confirmed strong BTBR avoidance of conspecifics and in the Social Proximity Test, BTBR showed dramatic differences in several close-in behaviors, including specific avoidance of a nose-to-nose contact that may potentially be related to gaze-avoidance. Diazepam normalized social avoidance by BTBRs in a Three-Chamber Test, and some additional behaviors - but not nose to nose avoidance - in the Social Proximity Test. BTBR also showed higher levels of preference for particular objects, and higher levels of sequences investigating 3- or 4-objects in the same order. Heparan sulfate (HS) associated with fractal structures in the subventricular zone of the lateral ventricles was severely reduced in BTBR. HS may modulate the functions of a range of growth and guidance factors during development, and HS abnormalities are associated with relevant brain (callosal agenesis) and behavioral (reductions in sociality) changes; suggesting the value of examination of the dynamics of the HS system in the context of autism.

Age increases anxiety and reactivity of the fear/anxiety circuit in Lewis rats.

A growing body of data indicates that changes in emotional behavior occur with age. Young Lewis rats are known to display hypofunction of the HPA axis. With age the reactivity of this axis is thought to increase with a concomitant rise in anxiety. In the current study, we investigate how and if the pattern of neuronal activation (measured as c-Fos protein expression) in Lewis rat brains changes with age and in response to novel environments differing in aversiveness. We found that distinct parts of the fear/anxiety circuit (i.e., the amygdalar complex, hippocampus and hypothalamus) undergo diverse age-related changes in response to behavioral challenges. While in the hypothalamus an increase in responsivity to mild stressors was observed with age, no such effect was present in the hippocampus. The amygdalar complex (especially the medial and cortical nuclei) on the other hand exhibited an age-dependent decrease in neuronal activation to mild stressors. This was accompanied by a marked increase in anxiety not correlated with a decline in locomotor activity.