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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.

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.

Background Parvalbumin inhibitory interneurons (PVIs) are crucial for maintaining proper excitatory/inhibitory balance and high-frequency neuronal synchronization. Their activity supports critical developmental trajectories, sensory and cognitive processing, and social behavior. Despite heterogeneity in the etiology across schizophrenia and autism spectrum disorder, PVI circuits are altered in these psychiatric disorders. Identifying mechanism(s) underlying PVI deficits is essential to establish treatments targeting in particular cognition. Based on our previous publications and new data, we propose oxidative stress as a common pathological mechanism leading to PVI impairment in schizophrenia and some forms of autism. Methods Using immunohistochemistry technique and confocal imaging analysis, we assessed the relationship between oxidative stress (as revealed by 8-oxo-DG immunolabeling) and PVI and their perineuronal net (PNN) in twelve established animal models relevant to autism (i.e., the fmr1 KO and CNV 15q13.3) and schizophrenia (CNV: 22q11, 15q13.3, 1q21, serine racemase (SR) KO, GRIN2A KO, Gclm KO) with or without additional insult (e.g., environmental: Gclm KO + GBR12909, GRIN2A KO + GBR12909, neonatal ventral hippocampal lesion (NVHL), methylazoxymethanol acetate developmental rodent model (MAM) and poly:IC). Results When PVI deficits in the anterior cingulate cortex were found in these animal models carrying genetic and/or environmental risks relevant to diverse etiological aspects of these disorders, oxidative stress was always present. Specifically, oxidative stress was negatively correlated with the integrity of PVIs and the extracellular perineuronal net enwrapping these interneurons. Oxidative stress may result from dysregulation of systems typically affected in schizophrenia, including glutamatergic, dopaminergic, immune, and antioxidant signaling. As convergent endpoint, redox dysregulation has successfully been targeted to protect PVIs with antioxidants/redox regulators across several animal models (e.g., Gclm KO, NVHL rats, GRIN2A KO and SR KO mice). D-serine, an allosteric modulator of brain NMDA receptor also protected PVIs and PNN against oxidative stress in SR KO mice. Discussion In view of the fact that the established pathophysiological processes dopamine excess, immune dysregulation and NMDA receptor hypofunction could all induce oxidative stress and are potentiated by additional oxidative insults, this mechanism could be central to damage of the highly metabolically active PVIs and the PNN surrounding them. Antioxidant systems are therefore potential therapeutic targets, assuming that redox regulators could be applied early, during environmental impacts, long before the clinical emergence of the disease.

Neuronal dysfunctions in the cortical GABAergic system have been widely documented in neuropsychiatric disorders with prenatal infectious etiologies, including schizophrenia. At least some of these abnormalities may stem from transcriptional impairments in the GABAergic transcriptome. However, the extent to which prenatal exposure to immune challenge can induce long-term alterations in GABAergic gene transcription remains largely elusive. Here, we use an established mouse model of prenatal immune activation induced by maternal gestational administration of the viral mimetic poly(I:C) (= polyriboinosinic-polyribocytidilic acid) to demonstrate that prenatal immune activation causes maturation-dependent alterations in prefrontal GABAergic gene expression. The spectrum of abnormalities included altered mRNA expression levels of enzymes regulating γ-aminobutyric acid (GABA) biosynthesis (glutamic acid decarboxylase 65-kDa [GAD65] and GAD67), vesicular GABA transporter (VGAT), alpha-subunits of the GABA(A) receptor (α2, α3, α4, and α5), and the chloride transporters sodium-potassium-chloride cotransporter 1 and potassium-chloride cotransporter 2. Additional western blot analyses confirmed the deficits in prefrontal GAD65/GAD67 and VGAT expression at the protein level. Intriguingly, the prefrontal GABAergic transcriptome was found to be more strongly affected in adult compared with peripubertal offspring born to immune-challenged mothers, and these age-dependent changes in GABAergic gene expression were paralleled by an adult onset of working memory deficiency. Collectively, our data emphasize a critical impact of prenatal immune-related insults on long-term GABAergic changes relevant to neuropsychiatric disorders with prenatal infectious etiologies, especially for those with delayed onset in early adulthood.

Context Prenatal infection and traumatizing experiences have both been linked with schizophrenia, but none of these factors seem sufficient to cause the disorder. However, recent evidence suggests that these environmental insults act in synergy to increase schizophrenia risk. Objective To estimate the independent and joint effects of exposure to prenatal infection and peripubertal psychological trauma on the risk of schizophrenia. Design Danish nationwide registers were linked in this prospective cohort study. We used survival analysis to report incidence rate ratios (IRRs) and corresponding 95% confidence intervals (95% CIs). Analyses were adjusted for age and calendar period and stratified by sex. Participants A total of 979701 persons born between 1980 and 1998 were followed up from January 1, 1995 through December 31, 2013, with 9656 having a hospital contact for schizophrenia. Results Females exposed to prenatal infection had a significantly increased risk of schizophrenia (IRR: 1.61, 95% CI: 1.30–2.00), but not males (IRR: 0.99, 95% CI: 0.77–1.28). Peripubertal trauma was associated with increased risk in both sexes. Males, however, had a significantly higher risk of schizophrenia after exposure to both prenatal infection and peripubertal psychological trauma (IRR: 2.85, 95% CI: 2.32–3.51), with significant interaction between infection and peripubertal trauma on the multiplicative scale (P = .007). Conclusions Our study demonstrated for the first time that prenatal infection and psychological trauma in peripubertal life can act in synergy to increase the risk of schizophrenia, with a potentially stronger susceptibility in males.

Background Successful development of high affinity radioligands for the translocator protein 18 KDa (TSPO) has contributed to a rapid rise in their use with positron emission tomography (PET) imaging to quantitatively detect the higher density of TSPO in neuropsychiatric conditions with putative microglial activation or reactive gliosis in vivo. [11C]PK11195 has been widely used to study TSPO in many neurological and psychiatric diseases, but the quality of quantified binding estimates using this first-generation radioligand is hampered by low signal to noise ratio, which also limits the sensitivity to detect group differences in binding. Second-generation radiotracers for TSPO such as [11C]DPA-713, [11C]PBR28, or [18F]FEPPA have superior specificity for the target and improved brain penetrance. However, in spite of these promising newer generation radioligands, their use with PET neuroimaging to study the immune response in psychotic diseases like schizophrenia has yielded inconsistent results of low, unchanged, or even tendency toward decreased binding to TSPO compared to data from controls. Methods In this presentation, we will provide necessary biological and methodological perspective to help interpret better the recent results from imaging TSPO in psychosis. We will first review its expression by many cell types including activated microglia, the resident immune cells in the brain, and its diverse functional roles including TSPO as a biomarker of classic neuroinflammatory processes. We will then present optimized methods for estimating useful binding outcomes that go beyond correction for TSPO genotype to minimize effects of factors, including those related to the diverse roles of TSPO, which otherwise introduce limiting, inter-individual variability in binding. Results These methods include the reporting of relative binding in one tissue to another, where such global factors are cancelled out by their appearance in the numerator and denominator of the outcome ratio. Use of this ratio approach may decrease inter-individual variability in binding measures and improve the sensitivity and statistical power to detect differences between cohorts. In contrast, use of a relative outcome measure may limit the utility of TSPO imaging since a difference between cohorts or within a subject over time may reflect either abnormal TSPO density or a mere shift in possibly normal, relative distribution between the two tissue regions. The most useful pseudoreference region is therefore one in which the true regional density of TSPO is unchanged in the study population, and is yet unidentified in schizophrenia. Building on this biology and methodology, we discuss misconceptions about imaging TSPO in psychosis and cautiously remind the field that this technique should not be equated with ‘imaging microglial activation’ or ‘imaging neuroinflammation.’ Indeed, PET-based TSPO estimates have not correlated with increased peripheral or central pro-inflammatory cytokine levels in schizophrenia or other psychiatric diseases like major depressive disorder. Discussion Together, a less simplified approach to imaging TSPO may inform its utility in studying other biological processes captured by its use in psychosis, and may guide future, complimentary research in vitro and in vivo to enhance our understanding of altered neuroimmune processes in psychosis.

Background Inflammatory theories in schizophrenia have gained increasing recognition and acceptance in recent years. The evidence supporting a role of altered inflammatory processes in the etiology and pathophysiology of schizophrenia involves early-life exposure to infectious pathogens or inflammatory stimuli, increased expression of cytokines and other mediators of inflammation in the adult central nervous system (CNS) and periphery, as well as signs of glial anomalies. Given the role of dopaminergic deregulation in the pathophysiology of schizophrenia, inflammatory processes in the midbrain may contribute to dopamine abnormalities in the midbrain and its subcortical and cortical output regions. Here, we tested this hypothesis using an established neurodevelopmental mouse model with relevance to schizophrenia, namely the maternal immune activation (MIA) model. Methods Pregnant C57BL6/N mice on gestation day 17 were treated with the viral mimetic polyriboinosinic-polyribocytidilic acid (poly(I:C)) or vehicle control solution. We then quantified the gene transcripts of an array of pro-inflammatory cytokines, acute phase proteins, and dopaminergic markers in the midbrain of MIA offspring (N=32) and control offspring (N= 32) at adult age. We also assessed the cell density of microglial cells expressing Iba1 and CD68 by immunohistochemistry to ascertain whether putative inflammatory changes are accompanied by microglia anomalies. Given the large sample sizes, we performed two-step recursive cluster analyses in order to identify possible subgroups of offspring that are characterized by “high” and “low” inflammatory profiles. Results When considering the entire treatment group, MIA-exposed offspring displayed significantly increased expression of several inflammatory cytokines in the ventral midbrain, including IL-1b (p < 0.01), TNF-a (p < 0.01) and SERPINA3 (p < 0.01). These inflammatory changes occurred in the absence of overt microglia anomalies but were paralleled by changes in dopaminergic markers. The two-step cluster analyses further identified subgroups of MIA-exposed offspring that are characterized by a “high” (41 %, N = 13) and “low” (59 %, N = 19) inflammatory profiles. The “high” inflammatory subgroup of MIA-exposed offspring was defined by marked elevations of SERPINA3, IL-1β, IL-6, and TNFα mRNA levels (all p’s < 0.01). Discussion Maternal immune activation during pregnancy causes persistent signs of inflammation in the offspring’s midbrain. In agreement with post-mortem studies in schizophrenia, these inflammatory abnormalities are clearly noticeable in a subgroup of MIA-exposed offspring only. Hence, prenatal immune activation may be one of the factors inducing lasting inflammatory changes relevant to (some cases of) schizophrenia and may contribute to dopaminergic dysfunctions in this disorder.

Background 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 such as schizophrenia and autism. 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 ignored whether differences in laboratory housing systems can similarly affect the outcomes of MIA models. Here, we examined this possibility by comparing poly(I:C)-based MIA in two housing systems that are commonly used in preclinical rodent research, namely the individually ventilated cage (IVC) and open cage (OC) systems. Methods Pregnant C57BL6/N mice were kept in IVC or OC and treated with a low (1 mg/kg, i.v.) or high (5 mg/kg, i.v.) dose of poly(I:C), or with corresponding vehicle solution (pyrogen-free, sterile 0.9% NaCl; i.v.). MIA or control treatment was induced on gestation day (GD) 9 or 12, and the resulting offspring were raised and maintained in IVC or OC until adulthood for behavioral testing. An additional cohort of dams were used to assess the influence of the different caging systems on poly(I:C)-induced cytokine responses in the maternal plasma, placenta, and fetal brains 1 hr and 6 hrs post-treatment. Results Maternal administration of poly(I:C) on GD9 caused a dose-dependent increase in spontaneous abortion in IVC but not in OC system, whereas MIA in IVC systems during a later gestational time-point (GD12) did not do so. Maternal and fetal pro-inflammatory cytokine responses to poly(I:C) were markedly higher in animals kept in IVC as compared to OC systems. The efficacy of MIA to induce long-term behavioral deficits was influenced by the different housing conditions, the dosing of poly(I:C), and the precise prenatal timing. Discussion The present study identified the type of cage system as a novel factor that can confound the outcomes of MIA. Our findings thus urge the need to consider and report the kind of cages used in rodent MIA models. Providing this information seems pivotal to yield robust and reproducible results in these models.

Maternal infection during pregnancy increases the risk of schizophrenia and other brain disorders of neurodevelopmental origin in the offspring. A multitude of infectious agents seem to be involved in this association. Therefore, it has been proposed that factors common to the immune response to a wide variety of bacterial and viral pathogens may be the critical link between prenatal infection and postnatal brain and behavioral pathology. More specifically, it has been suggested that the maternal induction of pro-inflammatory cytokines may mediate the neurodevelopmental effects of maternal infections. Here, we review recent findings from in vitro and in vivo investigations supporting this hypothesis and further emphasize the influence of enhanced anti-inflammatory cytokine signaling on early brain development. Disruption of the fetal brain balance between pro- and anti-inflammatory cytokine signaling may thus represent a key mechanism involved in the precipitation of schizophrenia-related pathology following prenatal maternal infection and innate immune imbalances.

Schizophrenia is associated with increased risk for multiple metabolic abnormalities, including altered glucose homeostasis, type-2 diabetes, obesity, and cardiovascular disease. Some of the metabolic alterations can already exist in psychosis-prone subjects prior to the onset of chronic schizophrenic disease and pharmacotherapy, indicating that they may have a developmental origin. In the present study, we tested the hypothesis that metabolic alterations pertinent to schizophrenic disease can be primed by an environmental risk factor associated with the disorder, namely prenatal exposure to immune challenge. We used a well-established mouse model of prenatal immune challenge induced by maternal gestational treatment with poly(I:C) (="polyriboinosinic-polyribocytidilic acid"), an analog of double-stranded RNA that stimulates a cytokine-associated viral-like acute phase response. Metabolic effects were studied using high-resolution computed tomography and fully automated indirect calorimetry system, along with an oral glucose tolerance test and plasma cytokine and corticosterone measurements. We found that prenatal immune activation caused altered glycemic regulation and abnormal ingestive behavior in periadolescence and led to an adult onset of excess visceral and subcutaneous fat deposition. These effects were accompanied by age-dependent changes in peripheral secretion of proinflammatory (interleukin [IL]-6 and tumor necrosis factor [TNF]-α) and T cell-related (IL-2 and interferon [IFN]-γ) cytokines and by increased release of the stress hormone corticosterone in periadolescence. Our findings show that schizophrenia-relevant metabolic and physiological abnormalities can be primed by prenatal viral-like immune activation, but at the same time, our study emphasizes that this environmental insult is unlikely to precipitate the full spectrum of metabolic and immunological changes pertinent to chronic schizophrenic disease.

Current pharmacotherapy of schizophrenia remains unsatisfactory with little hope for complete functional restoration in patients once the disease has developed. A preventive approach based on intervention in the prodromal stage of the disease aiming to preserve functional integrity by halting the progress of the disease is therefore extremely attractive. Here, we investigated the effects of preventive antipsychotic or antidepressant drug treatment in a well-established neurodevelopmental mouse model of multiple schizophrenia-related abnormalities. Pregnant mice on gestation day 9 were exposed to the viral mimic polyriboinosinic-polyribocytidylic acid (2 mg/kg, intravenously) or corresponding vehicle treatment, and the resulting offspring from both prenatal treatment conditions were subjected to chronic antipsychotic (haloperidol or clozapine), antidepressant (fluoxetine), or placebo treatment during the periadolescent stage of development. The effects of the preventive pharmacotherapy on behavioral and pharmacological functions were then investigated in adulthood using paradigms relevant to schizophrenia, namely prepulse inhibition, latent inhibition, and sensitivity to psychostimulant drugs. We show that periadolescent treatment with the reference antipsychotic and antidepressant drugs can successfully block the emergence of multiple psychosis-related behavioral and pharmacological abnormalities in subjects predisposed to adult brain pathology by exposure to prenatal immune challenge. At the same time, however, our study reveals numerous negative influences of the early pharmacological intervention on normal behavioral development in control subjects. Hence, even though preventive pharmacotherapy may be beneficial in individuals with predisposition to psychosis-related brain dysfunctions, chronic antipsychotic or antidepressant drug treatment in false-positive subjects is associated with substantial risk for long-term behavioral disturbances in adulthood.