Search

Endogenous retroviruses (ERVs) are ancestorial retroviral elements that were integrated into the mammalian genome through germline infections and insertions during evolution. While increased ERV expression has been repeatedly implicated in psychiatric and neurodevelopmental disorders, recent evidence suggests that aberrant endogenous retroviral activity may contribute to biologically defined subgroups of psychotic disorders with persisting immunological dysfunctions. Here, we explored whether ERV expression is altered in a mouse model of maternal immune activation (MIA), a transdiagnostic environmental risk factor of psychiatric and neurodevelopmental disorders. MIA was induced by maternal administration of poly(I:C) on gestation day 12 in C57BL/6N mice. Murine ERV transcripts were quantified in the placentae and fetal brains shortly after poly(I:C)-induced MIA, as well as in adult offspring that were stratified according to their behavioral profiles. We found that MIA increased and reduced levels of class II ERVs and syncytins, respectively, in placentae and fetal brain tissue. We also revealed abnormal ERV expression in MIA-exposed offspring depending on whether they displayed overt behavioral anomalies or not. Taken together, our findings provide a proof of concept that an inflammatory stimulus, even when initiated in prenatal life, has the potential of altering ERV expression across fetal to adult stages of development. Moreover, our data highlight that susceptibility and resilience to MIA are associated with differential ERV expression, suggesting that early-life exposure to inflammatory factors may play a role in determining disease susceptibility by inducing persistent alterations in the expression of endogenous retroviral elements., Brain, Behavior, and Immunity, 107, ISSN:0889-1591, ISSN:1090-2139

Theoretical and empirical evidence indicates that low external validity due to rigorous standardization of study populations is a cause of poor replicability in animal research. Here, we report a multi-laboratory study to test whether heterogenization of study populations by using animals from different breeders increases the replicability of results from single-laboratory studies. We found that heterogenization by breeder was not potent enough to increase external validity sufficiently to improve replicability.

Viral infections during pregnancy are associated with increased incidence of psychiatric disorders in offspring. The pathological outcomes of viral infection appear to be caused by the deleterious effects of innate immune response-associated factors on development of the fetus, which predispose the offspring to pathological conditions in adulthood. The negative impact of viral infections varies substantially between pregnancies. Here, we explored whether differential stress sensitivity underlies the high heterogeneity of immune reactivity and whether this may influence the pathological consequences of maternal immune activation. Using mouse models of social dominance (Dom) and submissiveness (Sub), which possess innate features of stress resilience and vulnerability, respectively, we identified differential immune reactivity to the synthetic analogue of viral double-stranded RNA, Poly(I:C), in Sub and Dom nulliparous and pregnant females. More specifically, we found that Sub females showed an exacerbated pro- and anti-inflammatory cytokine response to Poly(I:C) as compared with Dom females. Sub offspring born to Sub mothers (stress sensitive offspring) showed enhanced locomotory response to the non-competitive NMDA antagonist, MK-801, which was potentiated by prenatal Poly(I:C) exposure. Our findings suggest that inherited stress sensitivity may lead to functional changes in glutamatergic signaling, which in turn is further exacerbated by prenatal exposure to viral-like infection. The maternal immunome seems to play a crucial role in these observed phenomena.

Human endogenous retroviruses (ERVs) are ancestorial retroviral elements that were integrated into our genome through germline infections and insertions during evolution. They have repeatedly been implicated in the aetiology and pathophysiology of numerous human disorders, particularly in those that affect the central nervous system. In addition to the known association of ERVs with multiple sclerosis and amyotrophic lateral sclerosis, a growing number of studies links the induction and expression of these retroviral elements with the onset and severity of neurodevelopmental and psychiatric disorders. Although these disorders differ in terms of overall disease pathology and causalities, a certain degree of (subclinical) chronic inflammation can be identified in all of them. Based on these commonalities, we discuss the bidirectional relationship between ERV expression and inflammation and highlight that numerous entry points to this reciprocal sequence of events exist, including initial infections with ERV-activating pathogens, exposure to non-infectious inflammatory stimuli, and conditions in which epigenetic silencing of ERV elements is disrupted., Brain, Behavior, and Immunity, 107, ISSN:0889-1591, ISSN:1090-2139

The prefrontal cortex (PFC) provides executive top-down control of a variety of cognitive processes. A distinctive feature of the PFC is its protracted structural and functional maturation throughout adolescence to early adulthood, which is necessary for acquiring mature cognitive abilities. Using a mouse model of cell-specific, transient and local depletion of microglia, which is based on intracerebral injection of clodronate disodium salt (CDS) into the PFC of adolescent male mice, we recently demonstrated that microglia contribute to the functional and structural maturation of the PFC in males. Because microglia biology and cortical maturation are partly sexually dimorphic, the main objective of the present study was to examine whether microglia similarly regulate this maturational process in female mice as well. Here, we show that a single, bilateral intra-PFC injection of CDS in adolescent (6-week-old) female mice induces a local and transient depletion (70 to 80% decrease from controls) of prefrontal microglia during a restricted window of adolescence without affecting neuronal or astrocytic cell populations. This transient microglia deficiency was sufficient to disrupt PFC-associated cognitive functions and synaptic structures at adult age. Inducing transient prefrontal microglia depletion in adult female mice did not cause these deficits, demonstrating that the adult PFC, unlike the adolescent PFC, is resilient to transient microglia deficiency in terms of lasting cognitive and synaptic maladaptations. Together with our previous findings in males, the present findings suggest that microglia contribute to the maturation of the female PFC in a similar way as to the prefrontal maturation occurring in males., Brain, Behavior, and Immunity, 111, ISSN:0889-1591, ISSN:1090-2139

The epidemiological literature reporting increased risk for neurodevelopmental and psychiatric disorders after prenatal exposure to maternal immune activation (MIA) is still evolving, and so are the attempts to model this association in animals. Epidemiological studies of MIA offer the advantage of directly evaluating human populations but are often limited in their ability to uncover pathogenic mechanisms. Animal models, on the other hand, are limited in their generalizability to psychiatric disorders but have made significant strides toward discovering causal relationships and biological pathways between MIA and neurobiological phenotypes. Like in any other model system, both planned and unplanned sources of variability exist in animal models of MIA. Therefore, the design, implementation, and interpretation of MIA models warrant a careful consideration of these sources, so that appropriate strategies can be developed to handle them satisfactorily. While every research group may have its own strategy to this aim, it is essential to report the methodological details of the chosen MIA model in order to enhance the transparency and comparability of models across research laboratories. Even though it poses a challenge for attempts to compare experimental findings across laboratories, variability does not undermine the utility of MIA models for translational research. In fact, variability and heterogenous outcomes in MIA models offer unique opportunities for new discoveries and developments in this field, including the identification of disease pathways and molecular mechanisms determining susceptibility and resilience to MIA. This review summarizes the most important sources of variability in animal models of MIA and discusses how model variability can be used to investigate neurobiological and immunological factors causing phenotypic heterogeneity in offspring exposed to MIA.

Circular RNAs (circRNAs) are a large class of relatively stable RNA molecules that are highly expressed in animal brains. Many circRNAs have been associated with CNS disorders accompanied by an aberrant wake-sleep cycle. However, the regulation of circRNAs in brain homeostasis over daily light-dark (LD) cycles has not been characterized. Here, we aim to quantify the daily expression changes of circRNAs in physiological conditions in healthy adult animals. Using newly generated and public RNA-Seq data, we monitored circRNA expression throughout the 12:12 h LD cycle in various mouse brain regions. We identified that Cdr1as, a conserved circRNA that regulates synaptic transmission, is highly expressed in the suprachiasmatic nucleus (SCN), the master circadian pacemaker. Despite its high stability, Cdr1as has a very dynamic expression in the SCN throughout the LD cycle, as well as a significant regulation in the hippocampus following the entry into the dark phase. Computational integration of different public datasets predicted that Cdr1as is important for regulating light entrainment in the SCN. We hypothesize that the expression changes of Cdr1as in the SCN, particularly during the dark phase, are associated with light-induced phase shifts. Importantly, our work revises the current beliefs about natural circRNA stability and suggests that the time component must be considered when studying circRNA regulation.

Circular RNAs (circRNAs) are a large class of relatively stable RNA molecules that are highly expressed in animal brains. Many circRNAs have been associated with CNS disorders accompanied by an aberrant wake-sleep cycle. However, the regulation of circRNAs in brain homeostasis over daily light-dark (LD) cycles has not been characterized. Here, we aim to quantify the daily expression changes of circRNAs in physiological conditions in healthy adult animals. Using newly generated and public RNA-Seq data, we monitored circRNA expression throughout the 12:12 h LD cycle in various mouse brain regions. We identified that

The designer receptor exclusively activated by designer drugs (DREADD) system is one of the most widely used chemogenetic techniques to modulate the activity of cell populations in the brains of behaving animals. DREADDs are activated by acute or chronic administration of their ligand, clozapine-N-oxide (CNO). There is, however, a current lack of a non-invasive CNO administration technique that can control for drug timing and dosing without inducing substantial distress for the animals. Here, we evaluated whether the recently developed micropipette-guided drug administration (MDA) method, which has been used as a non-invasive and minimally stressful alternative to oral gavages, may be applied to administer CNO orally to activate DREADDs in a dosing- and timing-controlled manner. Unlike standard intraperitoneal injections, administration of vehicle substances via MDA did not elevate plasma levels of the major stress hormone, corticosterone, and did not attenuate exploratory activity in the open field test. At the same time, however, administration of CNO via MDA or intraperitoneally was equally efficient in activating hM3DGq-expressing neurons in the medial prefrontal cortex, as evident by time-dependent increases in mRNA levels of neuronal immediate early genes (cFos, Arc and Zif268) and cFos-immunoreactive neurons. Compared to vehicle given via MDA, oral administration of CNO via MDA was also found to potently increase locomotor activity in mice that express hM3DGq in prefrontal neurons. Taken together, our study confirms the effectiveness of CNO given orally via MDA and provides a novel method for non-stressful, yet well controllable CNO treatments in mouse DREADD systems.

Infectious or noninfectious maternal immune activation (MIA) is an environmental risk factor for psychiatric and neurological disorders with neurodevelopmental etiologies. Whilst there is increasing evidence for significant health consequences, the effects of MIA on the offspring appear to be variable. Here, we aimed to identify and characterize subgroups of isogenic mouse offspring exposed to identical MIA, which was induced in C57BL6/N mice by administration of the viral mimetic, poly(I:C), on gestation day 12. Cluster analysis of behavioral data obtained from a first cohort containing >150 MIA and control offspring revealed that MIA offspring could be stratified into distinct subgroups that were characterized by the presence or absence of multiple behavioral dysfunctions. The two subgroups also differed in terms of their transcriptional profiles in cortical and subcortical brain regions and brain networks of structural covariance, as measured by ex vivo structural magnetic resonance imaging (MRI). In a second, independent cohort containing 50 MIA and control offspring, we identified a subgroup of MIA offspring that displayed elevated peripheral production of innate inflammatory cytokines, including IL-1β, IL-6, and TNF-α, in adulthood. This subgroup also showed significant impairments in social approach behavior and sensorimotor gating, whereas MIA offspring with a low inflammatory cytokine status did not. Taken together, our results highlight the existence of subgroups of MIA-exposed offspring that show dissociable behavioral, transcriptional, brain network, and immunological profiles even under conditions of genetic homogeneity. These data have relevance for advancing our understanding of the variable neurodevelopmental effects induced by MIA and for biomarker-guided approaches in preclinical psychiatric research.

SummaryCdr1as is a conserved circular RNA (circRNA) enriched in the CNS and important for maintaining brain homeostasis. The loss of Cdr1as results in aberrant synaptic transmission and deregulation of stress response and circadian clock genes. However, it is not known whether the expression of Cdr1as or circRNAs, in general, follows a circadian pattern in different tissues. Here, using newly generated and public RNA-Seq data, we monitor circRNA expression throughout circadian rhythm in various mouse brain regions. We demonstrate that Cdr1as, despite its stable character, has a highly dynamic expression during the circadian cycle in the mouse suprachiasmatic nucleus (SCN). Cdr1as is one of the highest expressed RNAs in a cluster associated with light-induced synaptic transmission and phase shift in the SCN. Further, we identified that another brain enriched circRNA, mbl, is also substantially deregulated upon light induction in the fly head. Our study highlights the potential impact of abundant and conserved circRNAs on maintaining a healthy circadian cycle across species.

The prefrontal cortex (PFC) is a cortical brain region that regulates various cognitive functions. One distinctive feature of the PFC is its protracted adolescent maturation, which is necessary for acquiring mature cognitive abilities in adulthood. Here, we show that microglia, the brain's resident immune cells, contribute to this maturational process. We find that transient and cell-specific deficiency of prefrontal microglia in adolescence is sufficient to induce an adult emergence of PFC-associated impairments in cognitive functions, dendritic complexity, and synaptic structures. While prefrontal microglia deficiency in adolescence also altered the excitatory-inhibitory balance in adult prefrontal circuits, there were no cognitive sequelae when prefrontal microglia were depleted in adulthood. Thus, our findings identify adolescence as a sensitive period for prefrontal microglia to act on cognitive development.

Prenatal exposure to infectious or noninfectious immune activation is an environmental risk factor for neurodevelopmental disorders and mental illnesses. Recent research using animal models suggests that maternal immune activation (MIA) during early to middle stages of pregnancy can induce transgenerational effects on brain and behavior, likely via inducing stable epigenetic modifications across generations. Using a mouse model of viral-like MIA, which is based on gestational treatment with poly(I:C), the present study explored whether transgenerational effects can also emerge when MIA occurs in late pregnancy. Our findings demonstrate that the direct descendants born to poly(I:C)-treated mothers display deficits in temporal order memory, which are similarly present in second- and third-generation offspring. These transgenerational effects were mediated via both the maternal and paternal lineages and were accompanied by transient changes in maternal care. In addition to the cognitive effects, late prenatal immune activation induced generation-spanning effects on the prefrontal expression of gamma-aminobutyric acid (GABA)ergic genes, including parvalbumin and distinct alpha-subunits of the GABAA receptor. Together, our results suggest that MIA in late pregnancy has the potential to affect cognitive functions and prefrontal gene expression patterns in multiple generations, highlighting its role in shaping disease risk across generations.

Background There is growing evidence for a role of abnormal gut-brain signaling in disorders involving altered mood and affect, including depression. Studies using vagus nerve stimulation (VNS) suggest that the disruption of vagal afferent signaling may contribute to these abnormalities. To test this hypothesis, we used a rat model of subdiaphragmatic vagal deafferentation (SDA), the most complete and selective vagal deafferentation method existing to date, to study the consequences of complete disconnection of abdominal vagal afferents on affective behaviors. Methods SDA- and Sham-operated male rats were subjected to several tests that are commonly used in preclinical rodent models to assess the presence of anhedonic behavior, namely the novel object-induced exploration test, the novelty-suppressed eating test, and the sucrose preference test. In addition, we compared SDA and Sham rats in a social interaction test and the forced swim test to assess sociability and behavioral despair, respectively. Results Compared to Sham controls, SDA rats consistently displayed signs of anhedonic behavior in all test settings used. SDA rats also showed increased immobility and reduced swimming in the forced swim test, whereas they did not differ from Sham controls with regards to social approach behavior. Limitations This study was conducted in male rats only. Hence, possible sex-specific effects of SDA on affective behaviors remained unexamined. Conclusions Our findings demonstrate that hedonic behavior and behavioral despair are subject to visceral modulation through abdominal vagal afferents. These data are compatible with preclinical models and clinical trials showing beneficial effects of VNS on depression-like and affective behaviors.

BI 409306, a phosphodiesterase-9 inhibitor under development for treatment of schizophrenia and attenuated psychosis syndrome (APS), promotes synaptic plasticity and cognition. Here, we explored the effects of BI 409306 treatment in the polyriboinosinic-polyribocytidilic acid (poly[I:C])-based mouse model of maternal immune activation (MIA), which is relevant to schizophrenia and APS. In Study 1, adult offspring received BI 409306 0.2, 0.5, or 1 mg/kg or vehicle to establish an active dose. In Study 2, adult offspring received BI 409306 1 mg/kg and/or risperidone 0.025 mg/kg, risperidone 0.05 mg/kg, or vehicle, to evaluate BI 409306 as add-on to standard therapy for schizophrenia. In Study 3, offspring received BI 409306 1 mg/kg during adolescence only, or continually into adulthood to evaluate preventive effects of BI 409306. We found that BI 409306 significantly mitigated MIA-induced social interaction deficits and amphetamine-induced hyperlocomotion, but not prepulse inhibition impairments, in a dose-dependent manner (Study 1). Furthermore, BI 409306 1 mg/kg alone or in combination with risperidone 0.025 mg/kg significantly reversed social interaction deficits and attenuated amphetamine-induced hyperlocomotion in MIA offspring (Study 2). Finally, we revealed that BI 409306 1 mg/kg treatment restricted to adolescence prevented adult deficits in social interaction, whereas continued treatment into adulthood also significantly reduced amphetamine-induced hyperlocomotion (Study 3). Taken together, our findings suggest that symptomatic treatment with BI 409306 can restore social interaction deficits and dopaminergic dysfunctions in a MIA model of neurodevelopmental disruption, lending preclinical support to current clinical trials of BI 409306 in patients with schizophrenia. Moreover, BI 409306 given during adolescence has preventive effects on adult social interaction deficits in this model, supporting its use in people with APS., Neuropsychopharmacology, 46 (8), ISSN:0893-133X, ISSN:1740-634X

The mitochondrial protein, translocator protein (TSPO), is a widely used biomarker of neuroinflammation, but its non-selective cellular expression pattern implies roles beyond inflammatory processes. In the present study, we investigated whether neuronal activity modifies TSPO levels in the adult central nervous system. First, we used single-cell RNA sequencing to generate a cellular landscape of basal TSPO gene expression in the hippocampus of adult (12 weeks old) C57BL6/N mice, followed by confocal laser scanning microscopy to verify TSPO protein in neuronal and non-neuronal cell populations. We then quantified TSPO mRNA and protein levels after stimulating neuronal activity with distinct stimuli, including designer receptors exclusively activated by designer drugs (DREADDs), exposure to a novel environment and acute treatment with the psychostimulant drug, amphetamine. Single-cell RNA sequencing demonstrated a non-selective and multi-cellular gene expression pattern of TSPO at basal conditions in the adult mouse hippocampus. Confocal laser scanning microscopy confirmed that TSPO protein is present in neuronal and non-neuronal (astrocytes, microglia, vascular endothelial cells) cells of cortical (medial prefrontal cortex) and subcortical (hippocampus) brain regions. Stimulating neuronal activity through chemogenetic (DREADDs), physiological (novel environment exposure) or psychopharmacological (amphetamine treatment) approaches led to consistent increases in TSPO gene and protein levels in neurons, but not in microglia or astrocytes. Taken together, our findings show that neuronal activity has the potential to modify TSPO levels in the adult central nervous system. These findings challenge the general assumption that altered TSPO expression or binding unequivocally mirrors ongoing neuroinflammation and emphasize the need to consider non-inflammatory interpretations in some physiological or pathological contexts.

BACKGROUND: Maternal immune activation (MIA) is a proposed risk factor for multiple neuropsychiatric disorders, including schizophrenia. However, the molecular mechanisms through which MIA imparts risk remain poorly understood. A recently developed nonhuman primate model of exposure to the viral mimic poly:ICLC during pregnancy shows abnormal social and repetitive behaviors and elevated striatal dopamine, a molecular hallmark of human psychosis, providing an unprecedented opportunity for studying underlying molecular correlates. METHODS: We performed RNA-sequencing across psychiatrically relevant brain regions (prefrontal cortex, anterior cingulate, hippocampus) and primary visual cortex for comparison from 3.5–4-year old male MIA-exposed and control offspring—an age comparable to mid adolescence in humans. RESULTS: We identify 266 unique genes differentially expressed (DE) in at least one brain region with the greatest number observed in hippocampus. Co-expression networks identified region-specific alterations in synaptic signaling and oligodendrocytes. Although we observed temporal and regional differences, transcriptomic changes were shared across 1(st) and 2(nd) trimester exposures, including for the top DE genes—PIWIL2 and MGARP. In addition to PIWIL2, several other regulators of retrotransposition and endogenous transposable elements were dysregulated following MIA, potentially connecting MIA to retrotransposition. CONCLUSIONS: Together, these results begin to elucidate the brain-level molecular processes through which MIA may impart risk for psychiatric disease.

Prenatal exposure to infectious and/or inflammatory insults is increasingly recognized to contribute to the etiology of psychiatric disorders with neurodevelopmental components. Recent research using animal models suggests that maternal immune activation (MIA) can induce transgenerational effects on brain and behavior, possibly through epigenetic mechanisms. Using a mouse model of MIA that is based on gestational treatment with the viral mimeticpoly(I:C) (= polyriboinosinic-polyribocytidilic acid), the present study explored whether the transgenerational effects of MIA are extendable to dopaminergic dysfunctions. We show that the direct descendants born to poly(I:C)-treated mothers display signs of hyperdopaminergia, as manifested by a potentiated sensitivity to the locomotor-stimulating effects of amphetamine (Amph) and increased expression of tyrosine hydroxylase (Th) in the adult ventral midbrain. In stark contrast, second- and third-generation offspring of MIA-exposed ancestors displayed blunted locomotor responses to Amph and reduced expression of Th. Furthermore, we found increased DNA methylation at the promoter region of the dopamine-specifying factor, nuclear receptor-related 1 protein (Nurr1), in the sperm of first-generation MIA offspring and in the ventral midbrain of second-generation offspring of MIA-exposed ancestors. The latter effect was further accompanied by reduced mRNA levels of Nurr1 in this brain region. Together, our results suggest that MIA has the potential to modify dopaminergic functions across multiple generations with opposite effects in the direct descendants and their progeny. The presence of altered DNA methylation in the sperm of MIA-exposed offspring highlights the possibility that epigenetic processes in the male germline play a role in the transgenerational effects of MIA., Neuropsychopharmacology, 46 (2), ISSN:0893-133X, ISSN:1740-634X

Rodent models of maternal immune activation (MIA) are increasingly used as experimental tools to study neuronal and behavioral dysfunctions in relation to infection-mediated neurodevelopmental disorders. One of the most widely used MIA models is based on gestational administration of poly(I:C) (= polyriboinosinic-polyribocytdilic acid), a synthetic analog of double-stranded RNA that induces a cytokine-associated viral-like acute phase response. The effects of poly(I:C)-induced MIA on phenotypic changes in the offspring are known to be influenced by various factors, including the precise prenatal timing, genetic background, and immune stimulus intensity. Thus far, however, it has been largely ignored whether differences in the basic type of laboratory housing can similarly affect the outcomes of MIA models. Here, we examined this possibility by comparing the poly(I:C)-based MIA model in two housing systems that are commonly used in preclinical mouse research, namely the open cage (OC) and individually ventilated cage (IVC) systems. Pregnant C57BL6/N mice were kept in OCs or IVCs and treated with a low (1 mg/kg, i.v.) or high (5 mg/kg, i.v.) dose of poly(I:C), or with control vehicle solution. MIA or control treatment was induced on gestation day (GD) 9 or 12, and the resulting offspring were raised and maintained in OCs or IVCs until adulthood for behavioral testing. An additional cohort of dams was used to assess the influence of the different caging systems on poly(I:C)-induced cytokine and stress responses in the maternal plasma. Maternal poly(I:C) administration on GD9 caused a dose-dependent increase in spontaneous abortion in IVCs but not in OCs, whereas MIA in IVC systems during a later gestational time-point (GD12) did not affect pregnancy outcomes. Moreover, the precise type of caging system markedly affected maternal cytokines and chemokines at basal states and in response to poly(I:C) and further influenced the maternal levels of the stress hormone, corticosterone. The efficacy of MIA to induce deficits in working memory, social interaction, and sensorimotor gating in the adult offspring was influenced by the different housing conditions, the dosing of poly(I:C), and the precise prenatal timing. Taken together, the present study identifies the basic type of caging system as a novel factor that can confound the outcomes of MIA in mice. Our findings thus urge the need to consider and report the kind of laboratory housing systems used to implement MIA models. Providing this information seems pivotal to yield reproducible results in these models.

15q13.3 microdeletion is one of several gene copy number variants (CNVs) conferring increased risk of psychiatric and neurological disorders. This microdeletion gives rise to a variable spectrum of pathological phenotypes, ranging from asymptomatic to severe clinical outcomes. The reasons for these varying phenotypic outcomes remain unknown. Using a mouse model of hemizygous deletion of the orthologous region of 15q13.3, the present study examined whether exposure to stressful life events might interact with hemizygous 15q13.3 microdeletion in the development of behavioral dysfunctions. We show that hemizygous 15q13.3 microdeletion alone induces only limited effects on adult behaviors, but when combined with psychological stress in pubescence (postnatal days 30-40), it impairs sensorimotor gating and increases the sensitivity to the psychostimulant drug, amphetamine, at adult age. Stress exposure in adolescence (postnatal days 50-60) did not induce similar interactions with 15q13.3 microdeletion, but led to impaired emotional learning and memory and social behavior regardless of the genetic background. The present study provides the first evidence for interactive effects between hemizygous 15q13.3 microdeletion and exposure to stressful life events, and at the same time, it emphasizes an important influence of the precise timing of postnatal stress exposure in these interactions. Our findings suggest that hemizygous 15q13.3 microdeletion can act as a "disease primer" that increases the carrier's vulnerability to the detrimental effects of peripubertal stress exposure on adult behaviors.

The 2017 American College of Neuropychopharmacology (ACNP) conference hosted a Study Group on 4 December 2017, Establishing best practice guidelines to improve the rigor, reproducibility, and transparency of the maternal immune activation (MIA) animal model of neurodevelopmental abnormalities. The goals of this session were to (a) evaluate the current literature and establish a consensus on best practices to be implemented in MIA studies, (b) identify remaining research gaps warranting additional data collection and lend to the development of evidence-based best practice design, and (c) inform the MIA research community of these findings. During this session, there was a detailed discussion on the importance of validating immunogen doses and standardizing the general design (e.g., species, immunogenic compound used, housing) of our MIA models both within and across laboratories. The consensus of the study group was that data does not currently exist to support specific evidence-based model selection or methodological recommendations due to lack of consistency in reporting, and that this issue extends to other inflammatory models of neurodevelopmental abnormalities. This launched a call to establish a reporting checklist focusing on validation, implementation, and transparency modeled on the ARRIVE Guidelines and CONSORT (scientific reporting guidelines for animal and clinical research, respectively). Here we provide a summary of the discussions in addition to a suggested checklist of reporting guidelines needed to improve the rigor and reproducibility of this valuable translational model, which can be adapted and applied to other animal models as well.

Since Levine and then Barker's seminal work mid to late last century demonstrating the importance of early life environment, intensive research has revealed the plasticity, vulnerability and resilience of the developing brain to environmental challenges. In particular, early exposure to infectious pathogens and inflammatory stimuli has a lasting impact on brain and behavior. These data establish clear effects on vulnerability to later disease and neuroinflammatory injury, cognitive function and emotionality, and even responses to pain and susceptibility to metabolic disorders. They also highlight the issues with defining rodent models of complex diseases like autism spectrum disorders and schizophrenia, as well as the complexity of experimental design, for instance when deciding the appropriate allocation of subjects to experimental groups when dealing with whole-litter manipulations in rodents. The studies presented in this special issue of Brain Behavior and Immunity are a collection of the very latest advances in the science of perinatal inflammation and its implications for perinatal programming of brain and behavior.

Background Epidemiological studies over the past decades have repeatedly implicated maternal immune activation (MIA) in the etiology of psychiatric illnesses, including schizophrenia and related psychotic disorders. Not all offspring exposed to MIA, however, develop overt pathologies, suggesting that some are susceptible while others are resilient to MIA. To elucidate susceptibility and resilience in MIA, we used a mouse model that is based on prenatal exposure to the viral mimic poly(I:C). Methods Poly(I:C)-based MIA was induced in C57BL6/N mice on gestation day 12. Control dams received vehicle solution only. Offspring of poly(I:C)- or vehicle-exposed dams were subjected to a comprehensive behavioral testing battery when they reached adulthood (12 weeks of age onwards). Next-generation mRNA sequencing and gene pathway analyses were conducted after behavioral testing to explore the molecular correlates of resilience and susceptibility to MIA. Results Behavioral characterization coupled with unbiased TwoStep cluster analysis of a large number offspring (N >150) revealed that offspring exposed to MIA could be stratified into susceptible and resilient subgroups. While the former was characterized by deficits in social interaction, sensorimotor gating, and working memory, the behavioral profile of the latter was indistinguishable from control offspring. Susceptible and resilient MIA offspring were also dissociable by the presence of distinct molecular profiles in cortical and subcortical brain areas. In the medial prefrontal cortex, susceptible MIA offspring displayed a more profound deregulation of genes relevant for oxidative phosphorylation and mitochondrial functions than resilient MIA offspring. In the amygdala, the susceptible and resilient offspring differed in gene transcription pertinent to opioid signaling, DARPP-32 signaling, and G protein-coupled receptor signaling. Discussion Our data show that MIA can result in substantial phenotypic and transcriptomic variability even in the context of genetic homogeneity and under identical experimental conditions. If extended further, our model system may help to explain why only a subgroup of offspring exposed to MIA develops overt neurodevelopmental sequelae.

Background Aberrant activity of microglia, the primary innate immune cells of the brain parenchyma, may play a role in the etiology and pathophysiology of schizophrenia and related disorders. While current immunopsychiatric research indicates that microglial hyperactivity may contribute to psychotic illness in some cases, the diametrical opposite (i.e. microglial hypoactivity) may be pathologically and therapeutically relevant for others. The latter hypothesis, however, remains largely unexplored and thus warrants investigation. Methods We aimed at developing a model system in mice, in which the short- and long-term effects of selective, temporary and local depletion of microglia can be studied experimentally. This model system is based on intracerebral injection of clodronate disodium salt (CDS) into selected brain areas of interest. The behavioral and cognitive effects of temporary microglia depletion in the adolescent medial frontal cortex (mPFC) were assessed after full microglia recovery in adulthood. In addition, genome-wide transcriptional profiling was conducted during the peak of microglia depletion and after full microglia recovery in the mPFC. Results We show that a single intracerebral injection of CDS is a suitable and efficient approach to selectively deplete microglia without affecting astrocytes and neurons in-vivo, leading to a robust (~ 80% depletion) but temporary (~ 1 week) microglia deficiency in selected brain areas of interest. Using this model, we further demonstrate that CDS injection into mPFC during late adolescence (6 weeks of age) causes numerous mPFC-related cognitive dysfunctions in adulthood, that is, when microglial cells have been fully restored again. The spectrum of cognitive deficits included impairments in social recognition memory, temporal order memory and extinction of conditioned fear responses. These deficits emerged similarly in male and female animals and were paralleled by a permanent transcriptional dysregulation of genes relevant for synaptic refinement and stability. Intriguingly, CDS injections into the mPFC during early adolescence (4 weeks of age) or adulthood (12 weeks of age) did not induce similar cognitive dysfunctions in adulthood. Discussion Taken together, the present data demonstrate that temporary prefrontal microglia deficiency during adolescence leads to permanent cognitive impairments in adulthood. Our findings further highlight that distinct adolescent stages of cortical maturation show a differing sensitivity towards the long-term cognitive effects of temporary microglia hypoactivity.

Pharmacological treatments in laboratory rodents remain a cornerstone of preclinical psychopharmacological research and drug development. There are numerous ways in which acute or chronic pharmacological treatments can be implemented, with each method having certain advantages and drawbacks. Here, we describe and validate a novel treatment method in mice, which we refer to as the micropipette-guided drug administration (MDA) procedure. This administration method is based on a sweetened condensed milk solution as a vehicle for pharmacological substances, which motivates the animals to consume vehicle and/or drug solutions voluntarily in the presence of the experimenter. In a proof-of-concept study, we show that the pharmacokinetic profiles of the atypical antipsychotic drug, risperidone, were similar whether administered via the MDA procedure or via the conventional oral gavage method. Unlike the latter, however, MDA did not induce the stress hormone, corticosterone. Furthermore, we assessed the suitability and validity of the MDA method in a mouse model of maternal immune activation, which is frequently used as a model of immune-mediated neurodevelopmental disorders. Using this model, we found that chronic treatment (>4 weeks, once per day) with risperidone via MDA led to a dose-dependent mitigation of MIA-induced social interaction deficits and amphetamine hypersensitivity. Taken together, the MDA procedure described herein represents a novel pharmacological administration method for per os treatments in mice that is easy to implement, cost effective, non-invasive, and less stressful for the animals than conventional oral gavage methods., Brain, Behavior, and Immunity, 88, ISSN:0889-1591, ISSN:1090-2139

Maternal immune activation (MIA) models that are based on administration of the viral mimetic, poly(I:C), are widely used as experimental tools to study neuronal and behavioral dysfunctions in relation to immune-mediated neurodevelopmental disorders and mental illnesses. Evidence from investigations in non-pregnant rodents suggests that different poly(I:C) products can vary in terms of their immunogenicity, even if they are obtained from the same vendor. The present study aimed at extending these findings to pregnant mice, while also controlling various poly(I:C) products for potential contamination with lipopolysaccharide (LPS). We found significant variability between different batches of poly(I:C) potassium salt obtained from the same vendor (Sigma-Aldrich) in terms of the relative amount of dsRNA fragments in the high molecular weight range (1000-6000 nucleotides long) and with regards to their effects on maternal thermoregulation and immune responses in maternal plasma, placenta and fetal brain. Batches of poly(I:C) potassium salt containing larger amounts of high molecular weight fragments induced more extensive effects on thermoregulation and immune responses compared to batches with minimal amounts of high molecular weight fragments. Consistent with these findings, poly(I:C) enriched for high molecular weight dsRNA (HMW) caused larger maternal and placental immune responses compared to low molecular weight (LMW) poly(I:C). These variable effects were unrelated to possible LPS contamination. Finally, we found marked variability between different batches of the poly(I:C) potassium salt in terms of their effects on spontaneous abortion rates. This batch-to-batch variability was confirmed by three independent research groups using distinct poly(I:C) administration protocols in mice. Taken together, the present data confirm that different poly(I:C) products can induce varying immune responses and can differentially affect maternal physiology and pregnancy outcomes. It is therefore pivotal that researchers working with poly(I:C)-based MIA models ascertain and consider the precise molecular composition and immunogenicity of the product in use. We recommend the establishment of reference databases that combine phenotype data with empirically acquired quality information, which can aid the design, implementation and interpretation of poly(I:C)-based MIA models.

Maternal immune activation (MIA), be it triggered by infectious or noninfectious stimuli, is implicated in various psychiatric and neurological disorders with developmental etiologies. Its consequences on the offspring's mental health are heterogeneous and influenced by a number of factors shaping the specificity and/or severity of pathological outcomes. There is also a substantial degree of resilience to MIA, which determines the extent to which offspring are protected from developing neurodevelopmental sequelae. This review provides a synopsis of the plausible sources that account for the heterogeneous outcomes of MIA and discusses key factors that are critical for establishing neurodevelopmental resilience and susceptibility to this early-life adversity.

BACKGROUND: Rodent models of maternal immune activation (MIA) are used as an experimental approach for neurodevelopmental disorders such as schizophrenia to study neuronal and behavioral dysfunctions of offspring. In particular, injection of the viral mimic poly(I:C) in pregnant mice causes disruption of neuronal development of the offspring leading to deficits in social behavior, cognition, sensorimotor processing and/or dopaminergic system in adulthood [1]. BI 409306 is a selective phosphodiesterase-9 (PDE9) inhibitor [2] currently under development for the prevention of relapse in patients with schizophrenia taking antipsychotic medications (NCT03351244). PDE9 inhibition is hypothesized to improve the NMDA-receptor signaling cascade by increasing cGMP levels which subsequently leads to strengthening of synaptic plasticity. Abnormalities in glutamatergic neurotransmission related to NMDA-receptor hypofunction, along with dysfunction of the dopaminergic circuitry, are believed to play a crucial role in the pathophysiology of schizophrenia. In this study, we have investigated the effect of BI 409306 on the behavioral deficits of adult offspring from the poly(I:C)-based MIA mouse model. METHODS: Pregnant C57BL6/N mice were treated with poly(I:C) (5 mg/kg, i.v.) or control (saline, i.v.) solution on gestation day 12. All male offspring (n=9/group) were administered orally once daily with BI 409306 (0.2, 0.5 or 1 mg/kg) or vehicle starting at post-natal day (PND) 70 until subjected to a battery of consecutive behavioral tests two weeks later which lasts until PND120. The order of these tests were: Y-maze spontaneous alternation for working memory performance, social interaction to assess social behavior, pre-pulse inhibition (PPI) for sensorimotor processing and amphetamine-induced hyperlocomotion to assess the dopaminergic system. Statistical analyses were done by one-way ANOVA followed by Fisher’s LSD post-hoc. RESULTS: Maternal poly(I:C)-induced immune challenge led to a significant decrease in social interaction (p

Background Prenatal exposure to infectious or inflammatory insults increases the risk of neurodevelopmental disorders. Using a well-established mouse model of prenatal viral-like immune activation, we examined whether this pathological association involves genome-wide DNA methylation differences at single nucleotide resolution. Methods Prenatal immune activation was induced by maternal treatment with the viral mimetic polyriboinosinic-polyribocytidylic acid in middle or late gestation. Following behavioral and cognitive characterization of the adult offspring ( n = 12 per group), unbiased capture array bisulfite sequencing was combined with subsequent matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and quantitative real-time polymerase chain reaction analyses to quantify DNA methylation changes and transcriptional abnormalities in the medial prefrontal cortex of immune-challenged and control offspring. Gene ontology term enrichment analysis was used to explore shared functional pathways of genes with differential DNA methylation. Results Adult offspring of immune-challenged mothers displayed hyper- and hypomethylated CpGs at numerous loci and at distinct genomic regions, including genes relevant for gamma-aminobutyric acidergic differentiation and signaling (e.g., Dlx1, Lhx5, Lhx8 ), Wnt signaling ( Wnt3, Wnt8a, Wnt7b ), and neural development (e.g., Efnb3, Mid1, Nlgn1, Nrxn2 ). Altered DNA methylation was associated with transcriptional changes of the corresponding genes. The epigenetic and transcriptional effects were dependent on the offspring's age and were markedly influenced by the precise timing of prenatal immune activation. Conclusions Prenatal viral-like immune activation is capable of inducing stable DNA methylation changes in the medial prefrontal cortex. These long-term epigenetic modifications are a plausible mechanism underlying the disruption of prefrontal gene transcription and behavioral functions in subjects with prenatal infectious histories.

Positron emission tomography (PET) imaging with radiotracers that target translocator protein 18 kDa (TSPO) has become a popular approach to assess putative neuroinflammatory processes and associated microglia activation in psychotic illnesses. It remains unclear, however, whether TSPO imaging can accurately capture low-grade inflammatory processes such as those present in schizophrenia and related disorders. Therefore, we evaluated the validity of TSPO as a disease-relevant marker of inflammation using a translational approach, which combined neurodevelopmental and neurodegenerative mouse models with PET imaging in patients with recent-onset schizophrenia and matched controls. Using an infection-mediated neurodevelopmental mouse model, we show that schizophrenia-relevant behavioral abnormalities and increased inflammatory cytokine expression are associated with reduced prefrontal TSPO levels. On the other hand, TSPO was markedly upregulated in a mouse model of acute neurodegeneration and reactive gliosis, which was induced by intrahippocampal injection of kainic acid. In both models, the changes in TSPO levels were not restricted to microglia but emerged in various cell types, including microglia, astrocytes and vascular endothelial cells. Human PET imaging using the second-generation TSPO radiotracer [(11)C]DPA-713 revealed a strong trend towards reduced TSPO binding in the middle frontal gyrus of patients with recent-onset schizophrenia, who were previously shown to display increased levels of inflammatory cytokines in peripheral and central tissues. Together, our findings challenge the common assumption that central low-grade inflammation in schizophrenia is mirrored by increased TSPO expression or ligand binding. Our study further underscores the need to interpret altered TSPO binding in schizophrenia with caution, especially when measures of TSPO are not complemented with other markers of inflammation. Unless more selective microglial markers are available for PET imaging, quantification of cytokines and other inflammatory biomarkers, along with their molecular signaling pathways, may be more accurate in attempts to characterize inflammatory profiles in schizophrenia and other mental disorders that lack robust reactive gliosis.Molecular Psychiatry advance online publication, 17 January 2017; doi:10.1038/mp.2016.248.

Overconsumption of high-fat diets (HFDs) can critically affect synaptic and cognitive functions within telencephalic structures such as the medial prefrontal cortex (mPFC). The underlying mechanisms, however, remain largely unknown. Here we show that adolescence is a sensitive period for the emergence of prefrontal cognitive deficits in response to HFD. We establish that the synaptic modulator reelin (RELN) is a critical mediator of this vulnerability because (1) periadolescent HFD (pHFD) selectively downregulates prefrontal RELN+ cells and (2) augmenting mPFC RELN levels using transgenesis or prefrontal pharmacology prevents the pHFD-induced prefrontal cognitive deficits. We further identify N-methyl-d-aspartate-dependent long-term depression (NMDA-LTD) at prefrontal excitatory synapses as a synaptic signature of this association because pHFD abolishes NMDA-LTD, a function that is restored by RELN overexpression. We believe this study provides the first mechanistic insight into the vulnerability of the adolescent mPFC towards nutritional stress, such as HFDs. Our findings have primary relevance to obese individuals who are at an increased risk of developing neurological cognitive comorbidities, and may extend to multiple neuropsychiatric and neurological disorders in which RELN deficiency is a common feature.

Prenatal exposure to infectious or inflammatory insults can increase the risk of developing neuropsychiatric disorder in later life, including schizophrenia, bipolar disorder, and autism. These brain disorders are also characterized by pre- and postsynaptic deficits. Using a well-established mouse model of maternal exposure to the viral mimetic polyriboinosinic-polyribocytidilic acid [poly(I:C)], we examined whether prenatal immune activation might cause synaptic deficits in the hippocampal formation of pubescent and adult offspring. Based on the widely appreciated role of microglia in synaptic pruning, we further explored possible associations between synaptic deficits and microglia anomalies in offspring of poly(I:C)-exposed and control mothers. We found that prenatal immune activation induced an adult onset of presynaptic hippocampal deficits (as evaluated by synaptophysin and bassoon density). The early-life insult further caused postsynaptic hippocampal deficits in pubescence (as evaluated by PSD95 and SynGAP density), some of which persisted into adulthood. In contrast, prenatal immune activation did not change microglia (or astrocyte) density, nor did it alter their activation phenotypes. The prenatal manipulation did also not cause signs of persistent systemic inflammation. Despite the absence of overt glial anomalies or systemic inflammation, adult offspring exposed to prenatal immune activation displayed increased hippocampal IL-1β levels. Taken together, our findings demonstrate that age-dependent synaptic deficits and abnormal pro-inflammatory cytokine expression can occur during postnatal brain maturation in the absence of microglial anomalies or systemic inflammation.

Animal models of maternal immune activation (MIA) are widely used as experimental tools to study neurobehavioral and molecular dysfunctions in relation to immune-mediated neurodevelopmental disorders and mental illnesses. Both planned or intended (‘a-priori variability’) and unplanned or unintended (‘a-posteriori variability’) sources of variability exist in and are relevant to these models. The design, implementation, and interpretation of MIA models, thus, require adequate time considering both types of variability and developing appropriate strategies to handle them. While there are multiple strategies for minimizing variability in MIA models, variability per se is not an undermining characteristic of MIA models or any other model system applied to psychoneuroimmunological research. In fact, it offers unique opportunities for new (and sometimes unexpected) discoveries, granted that researchers are aware of the potential sources of variability and are open to novel methodological approaches and ways of data analysis., Current Opinion in Behavioral Sciences, 28

Background Pharmacological treatments in laboratory rodents remain a cornerstone of preclinical psychopharmacological research and drug development. There are numerous ways how acute or chronic pharmacological treatments can be implemented, with each method having certain advantages and drawbacks. To overcome the main disadvantages of conventional administration routes, we implemented and validated a novel treatment method in mice, which we refer to as the Micropipette-guided drug administration (MDA) method. Methods The MDA administration method is based on using sweetened condensed milk solution as vehicle for pharmacological substances, which motivates the animals to consume vehicle and/or drug solutions voluntarily in the presence of the experimenter. First, we measured the corticosterone and pharmacokinetic changes after the administration of a vehicle or drug (Risperidone) solution via traditional oral gavage or by MDA. We then conducted a chronic treatment paradigm in the maternal immune activation model (MIA). Offspring treatment with risperidone started on post-natal day 70 and lasted for a total of 6 weeks, with behavioral testing occurring during the last two weeks of the treatment window. Results In a proof-of-concept study, we show that the pharmacokinetic profiles of the atypical antipsychotic drug, risperidone, are similar when administered via the MDA procedure or via the conventional oral gavage method. Unlike the latter, however, MDA did not induce the stress hormone, corticosterone. Furthermore, we assessed the suitability and validity of the MDA method in a mouse model of maternal immune activation, which is frequently used as a model of immune-mediated neurodevelopmental disorders. Using this model, we found that chronic treatment (> 4 weeks, once per day) with risperidone via MDA led to a dose-dependent mitigation of MIA-induced social interaction deficits and amphetamine hypersensitivity. Discussion Taken together, the MDA procedure described represents a novel pharmacological administration method for per os treatments in mice, which is easy to implement, cost effective, non-invasive, and less stressful for the animals than conventional oral gavage methods. Moreover, it allows for a more consistent and continual control of the dosages administered to the animals compared to methods involving the dissolution of a substance in the drinking water or contained within the food of the animals.