MAOA methylation is associated with impulsive and antisocial behaviour: dependence on allelic variation, family environment and diet.

Congenital absence of monoamine oxidase A (MAO-A) activity predisposes to antisocial impulsive behaviour, and the MAOA uVNTR low-expressing genotype (MAOA-L) together with childhood maltreatment is associated with similar phenotypes in males. A possible explanation of how family environment may lead to such behaviour involves DNA methylation. We have assessed MAOA methylation and impulsive/antisocial behaviour in 121 males from the Estonian Children Personality Behaviour and Health Study. Of the 12 CpG sites measured, methylation levels at the locus designated CpG3 were significantly lower in subjects with antisocial behaviour involving police contact. CpG3 methylation was lower in subjects with alcohol use disorder by age 25, but only in MAOA-H genotype. No correlation between MAOA CpG3 methylation levels and adaptive impulsivity was found at age 15, but in MAOA-L genotype a positive correlation appeared by age 18. By age 25, this positive correlation was no longer observed in subjects with better family relationships but had increased further with experience of adversity within the family. MAOA CpG3 methylation had different developmental dynamics in relation to maladaptive impulsivity. At age 18, a positive correlation was observed in MAOA-L genotype with inferior family relationships and a negative correlation was found in MAOA-H with superior home environment; both of these associations had disappeared by age 25. CpG3 methylation was associated with dietary intake of several micronutrients, most notable was a negative correlation with the intake of zinc, but also with calcium, potassium and vitamin E; a positive correlation was found with intake of phosphorus. In conclusion, MAOA CpG3 methylation is related to both maladaptive and adaptive impulsivity in adolescence in MAOA-L males from adverse home environment. By young adulthood, this relationship with maladaptive impulsivity had disappeared but with adaptive impulsivity strengthened. Thus, MAOA CpG3 methylation may serve as a marker for adaptive developmental neuroplasticity in MAOA-L genotype. The mechanisms involved may include dietary factors.

Lipopolysaccharide triggers exacerbated microglial activation, excessive cytokine release and behavioural disturbances in mice with truncated Fused-in-Sarcoma Protein (FUS).

CNS inflammation, including microglial activation, in response to peripheral infections are known to contribute to the pathology of both familial and sporadic neurodegenerative disease. The relationship between Fused-in-Sarcoma Protein (FUS)-mediated disease in the transgenic FUS[1-359] animals and the systemic inflammatory response have not been explored. Here, we investigated microglial activation, inflammatory gene expression and the behavioural responses to lipopolysaccharide-induced (LPS; 0.1 mg/kg) systemic inflammation in the FUS[1-359] transgenic mice. The pathology of these mice recapitulates the key features of mutant FUS-associated familial frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS). Here, pre-symptomatic 8-week-old mutant or wild type controls were challenged with LPS or with saline and sucrose intake, novel cage exploration, marble burying and swimming behaviours were analyzed. The level of pro-inflammatory gene expression was also determined, and microglial activation was evaluated. In chronic experiments, to discover whether the LPS challenge would affect the onset of ALS-like paralysis, animals were evaluated for clinical signs from 5 to 7 weeks post-injection. Compared to controls, acutely challenged FUS[1-359]-tg mice exhibited decreased sucrose intake and increased floating behaviours. The FUS[1-359]-tg mice exhibited an increase in immunoreactivity for Iba1-positive cells in the prefrontal cortex and ventral horn of the spinal cord, which was accompanied by increased expression of interleukin-1ß, tumour necrosis factor, cyclooxygenase-(COX)-1 and COX-2. However, the single LPS challenge did not alter the time to development of paralysis in the FUS[1-359]-tg mice. Thus, while the acute inflammatory response was enhanced in the FUS mutant animals, it did not have a lasting impact on disease progression.

Molecular signature of excessive female aggression: study of stressed mice with genetic inactivation of neuronal serotonin synthesis.

Aggression is a complex social behavior, critically involving brain serotonin (5-HT) function. The neurobiology of female aggression remains elusive, while the incidence of its manifestations has been increasing. Yet, animal models of female aggression are scarce. We previously proposed a paradigm of female aggression in the context of gene x environment interaction where mice with partial genetic inactivation of tryptophan hydroxylase-2 (Tph2+/- mice), a key enzyme of neuronal 5-HT synthesis, are subjected to predation stress resulting in pathological aggression. Using deep sequencing and the EBSeq method, we studied the transcriptomic signature of excessive aggression in the prefrontal cortex of female Tph2+/- mice subjected to rat exposure stress and food deprivation. Challenged mutants, but not other groups, displayed marked aggressive behaviors. We found 26 genes with altered expression in the opposite direction between stressed groups of both Tph2 genotypes. We identified several molecular markers, including Dgkh, Arfgef3, Kcnh7, Grin2a, Tenm1 and Epha6, implicated in neurodevelopmental deficits and psychiatric conditions featuring impaired cognition and emotional dysregulation. Moreover, while 17 regulons, including several relevant to neural plasticity and function, were significantly altered in stressed mutants, no alteration in regulons was detected in stressed wildtype mice. An interplay of the uncovered pathways likely mediates partial Tph2 inactivation in interaction with severe stress experience, thus resulting in excessive female aggression.

Serotonin Transporter (SERT) Expression Modulates the Composition of the Western-Diet-Induced Microbiota in Aged Female Mice.

Background. The serotonin transporter (SERT), highly expressed in the gut and brain, is implicated in metabolic processes. A genetic variant of the upstream regulatory region of the SLC6A4 gene encoding SERT, the so-called short (s) allele, in comparison with the long (l) allele, results in the decreased function of this transporter, altered serotonergic regulation, an increased risk of psychiatric pathology and type-2 diabetes and obesity, especially in older women. Aged female mice with the complete (Sert-/-: KO) or partial (Sert+/-: HET) loss of SERT exhibit more pronounced negative effects following their exposure to a Western diet in comparison to wild-type (Sert+/+: WT) animals. Aims. We hypothesized that these effects might be mediated by an altered gut microbiota, which has been shown to influence serotonin metabolism. We performed V4 16S rRNA sequencing of the gut microbiota in 12-month-old WT, KO and HET female mice that were housed on a control or Western diet for three weeks. Results. The relative abundance of 11 genera was increased, and the abundance of 6 genera was decreased in the Western-diet-housed mice compared to the controls. There were correlations between the abundance of Streptococcus and Ruminococcaceae_UCG-014 and the expression of the pro-inflammatory marker Toll-like-Receptor 4 (Tlr4) in the dorsal raphe, as well as the expression of the mitochondrial activity marker perixome-proliferator-activated-receptor-cofactor-1b (Ppargc1b) in the prefrontal cortex. Although there was no significant impact of genotype on the microbiota in animals fed with the Control diet, there were significant interactions between diet and genotype. Following FDR correction, the Western diet increased the relative abundance of Intestinimonas and Atopostipes in the KO animals, which was not observed in the other groups. Erysipelatoclostridium abundance was increased by the Western diet in the WT group but not in HET or KO animals. Conclusions. The enhanced effects of a challenge with a Western diet in SERT-deficient mice include the altered representation of several gut genera, such as Intestinimonas, Atopostipes and Erysipelatoclostridium, which are also implicated in serotonergic and lipid metabolism. The manipulation of these genera may prove useful in individuals with the short SERT allele.

The long and the short of it: 5-HTTLPR and moral judgement.

Differences in moral sentiments are widespread. Increasingly, their biological correlates are investigated to elucidate potential sources of divergent moral attitudes and choices. Serotonin is one such potential modulator. We investigated the effects of a functional serotonergic polymorphism, 5-HTTLPR, which was previously linked to moral choices albeit with inconsistent findings. N = 157 healthy young adults completed a set of congruent and incongruent moral dilemmas. In addition to the traditional moral response score, this set allows by using a process dissociation (PD) approach an estimation a deontological and a utilitarian parameter. While there was no main effect of 5-HTTLPR on any of the three moral judgement parameters, there was an interaction effect between 5-HTTLPR and endocrine status on PD parameters, which was mainly due to the deontological but not the utilitarian parameter. In men and free cycling women, LL homozygotes showed reduced deontological tendencies compared to S allele carriers. Contrariwise, in women using oral contraceptives, LL homozygotes had increased deontology parameter scores. Furthermore, LL genotypes in general reported less difficulty in making harmful choices, which were in addition associated with less negative emotions. The findings suggest that 5-HTTLPR might be involved in modulating cognitive and emotional processes contributing to moral decisions.

Serotonergic modulation of normal and abnormal brain dynamics: The genetic influence of the TPH2 G-703T genotype and DNA methylation on wavelet variance in children and adolescents with and without ADHD.

Attention-deficit/hyperactivity disorder (ADHD) is a neurodevelopmental disorder that often persists into adulthood. Core symptoms of ADHD, such as impulsivity, are caused by an interaction of genetic and environmental factors. Epigenetic modifications of DNA, such as DNA methylation, are thought to mediate the interplay of these factors. Tryptophan hydroxylase 2 (TPH2) is the rate-limiting enzyme in brain serotonin synthesis. The TPH2 gene has frequently been investigated in relation to ADHD, e.g., showing that TPH2 G-703T (rs4570625) polymorphism influences response control and prefrontal signaling in ADHD patients. In this (epi)genetic imaging study we examined 144 children and adolescents (74 patients, 14 females) using fMRI at rest and during performing a waiting impulsivity (WI) paradigm. Both, TPH2 G-703T (rs4570625) genotype and DNA methylation in the 5' untranslated region (5'UTR) of TPH2 were associated with wavelet variance in fronto-parietal regions and behavioral performance, taking TPH2 genotype into account. In detail, comparisons between genotypes of patients and controls revealed highest wavelet variance and longest reaction times in patients carrying the T allele [indicative for a gene-dosage effect, i.e., the WI phenotype is a direct result of the cumulative effect of ADHD and TPH2 variation]. Regressions revealed a significant effect on one specific DNA methylation site in ADHD patients but not controls, in terms of a significant prediction of wavelet variance in fronto-parietal regions as well as premature responses. By the example of the TPH2 G-703T (rs4570625) polymorphism, we provide insight into how interactive genetic and DNA methylation affect the ADHD and/or impulsive endophenotype.

Genome-wide DNA methylation analysis of aggressive behaviour: a longitudinal population-based study.

BACKGROUND: Human aggression is influenced by an interplay between genetic predisposition and experience across the life span. This interaction is thought to occur through epigenetic mechanisms, inducing differential gene expression, thereby moderating neuronal cell and circuit function, and thus shaping aggressive behaviour. METHODS: Genome-wide DNA methylation (DNAm) levels were measured in peripheral blood obtained from 95 individuals participating in the Estonian Children Personality Behaviours and Health Study (ECPBHS) at 15 and 25 years of age. We examined the association between aggressive behaviour, as measured by Life History of Aggression (LHA) total score and DNAm levels both assessed at age 25. We further examined the pleiotropic effect of genetic variants regulating LHA-associated differentially methylated positions (DMPs) and multiple traits related to aggressive behaviours. Lastly, we tested whether the DNA methylomic loci identified in association with LHA at age 25 were also present at age 15. RESULTS: We found one differentially methylated position (DMP) (cg17815886; p = 1.12 × 10-8 ) and five differentially methylated regions (DMRs) associated with LHA after multiple testing adjustments. The DMP annotated to the PDLIM5 gene, and DMRs resided in the vicinity of four protein-encoding genes (TRIM10, GTF2H4, SLC45A4, B3GALT4) and a long intergenic non-coding RNA (LINC02068). We observed evidence for the colocalization of genetic variants associated with top DMPs and general cognitive function, educational attainment and cholesterol levels. Notably, a subset of the DMPs associated with LHA at age 25 also displayed altered DNAm patterns at age 15 with high accuracy in predicting aggression. CONCLUSIONS: Our findings highlight the potential role of DNAm in the development of aggressive behaviours. We observed pleiotropic genetic variants associated with identified DMPs, and various traits previously established to be relevant in shaping aggression in humans. The concordance of DNAm signatures in adolescents and young adults may have predictive value for inappropriate and maladaptive aggression later in life.

Generation of a ST3GAL3 null mutant induced pluripotent stem cell (iPSC) line (UKWMPi002-A-3) by CRISPR/Cas9 genome editing.

Fibroblasts isolated from a skin biopsy of a healthy individual were infected with Sendai virus containing the Yamanaka factors to produce transgene-free human induced pluripotent stem cells (iPSCs). CRISPR/Cas9 was used to generate an isogenic cell line carrying an inactivation of ST3GAL3, a risk gene associated with neurodevelopmental and psychiatric disorders. This ST3GAL3 null mutant (ST3GAL3-/-) iPSC line, which displays the expression of pluripotency-associated markers, the ability to differentiate into cells of the three germ layers in vitro, and a normal karyotype, is a powerful tool to investigate the impact of deficient sialylation of glycoproteins in neural development and plasticity.

Understanding the Role of Oxidative Stress, Neuroinflammation and Abnormal Myelination in Excessive Aggression Associated with Depression: Recent Input from Mechanistic Studies.

Aggression and deficient cognitive control problems are widespread in psychiatric disorders, including major depressive disorder (MDD). These abnormalities are known to contribute significantly to the accompanying functional impairment and the global burden of disease. Progress in the development of targeted treatments of excessive aggression and accompanying symptoms has been limited, and there exists a major unmet need to develop more efficacious treatments for depressed patients. Due to the complex nature and the clinical heterogeneity of MDD and the lack of precise knowledge regarding its pathophysiology, effective management is challenging. Nonetheless, the aetiology and pathophysiology of MDD has been the subject of extensive research and there is a vast body of the latest literature that points to new mechanisms for this disorder. Here, we overview the key mechanisms, which include neuroinflammation, oxidative stress, insulin receptor signalling and abnormal myelination. We discuss the hypotheses that have been proposed to unify these processes, as many of these pathways are integrated for the neurobiology of MDD. We also describe the current translational approaches in modelling depression, including the recent advances in stress models of MDD, and emerging novel therapies, including novel approaches to management of excessive aggression, such as anti-diabetic drugs, antioxidant treatment and herbal compositions.

The translational genetics of ADHD and related phenotypes in model organisms.

Attention-deficit/hyperactivity disorder (ADHD) is a highly prevalent neurodevelopmental disorder resulting from the interaction between genetic and environmental risk factors. It is well known that ADHD co-occurs frequently with other psychiatric disorders due, in part, to shared genetics factors. Although many studies have contributed to delineate the genetic landscape of psychiatric disorders, their specific molecular underpinnings are still not fully understood. The use of animal models can help us to understand the role of specific genes and environmental stimuli-induced epigenetic modifications in the pathogenesis of ADHD and its comorbidities. The aim of this review is to provide an overview on the functional work performed in rodents, zebrafish and fruit fly and highlight the generated insights into the biology of ADHD, with a special focus on genetics and epigenetics. We also describe the behavioral tests that are available to study ADHD-relevant phenotypes and comorbid traits in these models. Furthermore, we have searched for new models to study ADHD and its comorbidities, which can be useful to test potential pharmacological treatments.

Immunomodulatory effects of new phytotherapy on human macrophages and TLR4- and TLR7/8-mediated viral-like inflammation in mice.

Background: While all efforts have been undertaken to propagate the vaccination and develop remedies against SARS-CoV-2, no satisfactory management of this infection is available yet. Moreover, poor availability of any preventive and treatment measures of SARS-CoV-2 in economically disadvantageous communities aggravates the course of the pandemic. Here, we studied a new immunomodulatory phytotherapy (IP), an extract of blackberry, chamomile, garlic, cloves, and elderberry as a potential low-cost solution for these problems given the reported efficacy of herbal medicine during the previous SARS virus outbreak. Methods: The key feature of SARS-CoV-2 infection, excessive inflammation, was studied in in vitro and in vivo assays under the application of the IP. First, changes in tumor-necrosis factor (TNF) and lnteurleukin-1 beta (IL-1ß) concentrations were measured in a culture of human macrophages following the lipopolysaccharide (LPS) challenge and treatment with IP or prednisolone. Second, chronically IP-pre-treated CD-1 mice received an agonist of Toll-like receptors (TLR)-7/8 resiquimod and were examined for lung and spleen expression of pro-inflammatory cytokines and blood formula. Finally, chronically IP-pre-treated mice challenged with LPS injection were studied for "sickness" behavior. Additionally, the IP was analyzed using high-potency-liquid chromatography (HPLC)-high-resolution-mass-spectrometry (HRMS). Results: LPS-induced in vitro release of TNF and IL-1ß was reduced by both treatments. The IP-treated mice displayed blunted over-expression of SAA-2, ACE-2, CXCL1, and CXCL10 and decreased changes in blood formula in response to an injection with resiquimod. The IP-treated mice injected with LPS showed normalized locomotion, anxiety, and exploration behaviors but not abnormal forced swimming. Isoquercitrin, choline, leucine, chlorogenic acid, and other constituents were identified by HPLC-HRMS and likely underlie the IP immunomodulatory effects. Conclusions: Herbal IP-therapy decreases inflammation and, partly, "sickness behavior," suggesting its potency to combat SARS-CoV-2 infection first of all via its preventive effects.

Genetic architecture of 11 major psychiatric disorders at biobehavioral, functional genomic and molecular genetic levels of analysis.

We interrogate the joint genetic architecture of 11 major psychiatric disorders at biobehavioral, functional genomic and molecular genetic levels of analysis. We identify four broad factors (neurodevelopmental, compulsive, psychotic and internalizing) that underlie genetic correlations among the disorders and test whether these factors adequately explain their genetic correlations with biobehavioral traits. We introduce stratified genomic structural equation modeling, which we use to identify gene sets that disproportionately contribute to genetic risk sharing. This includes protein-truncating variant-intolerant genes expressed in excitatory and GABAergic brain cells that are enriched for genetic overlap across disorders with psychotic features. Multivariate association analyses detect 152 (20 new) independent loci that act on the individual factors and identify nine loci that act heterogeneously across disorders within a factor. Despite moderate-to-high genetic correlations across all 11 disorders, we find little utility of a single dimension of genetic risk across psychiatric disorders either at the level of biobehavioral correlates or at the level of individual variants.

Predation Stress Causes Excessive Aggression in Female Mice with Partial Genetic Inactivation of Tryptophan Hydroxylase-2: Evidence for Altered Myelination-Related Processes.

The interaction between brain serotonin (5-HT) deficiency and environmental adversity may predispose females to excessive aggression. Specifically, complete inactivation of the gene encoding tryptophan hydroxylase-2 (Tph2) results in the absence of neuronal 5-HT synthesis and excessive aggressiveness in both male and female null mutant (Tph2-/-) mice. In heterozygous male mice (Tph2+/-), there is a moderate reduction in brain 5-HT levels, and when they are exposed to stress, they exhibit increased aggression. Here, we exposed female Tph2+/- mice to a five-day rat predation stress paradigm and assessed their emotionality and social interaction/aggression-like behaviors. Tph2+/- females exhibited excessive aggression and increased dominant behavior. Stressed mutants displayed altered gene expression of the 5-HT receptors Htr1a and Htr2a, glycogen synthase kinase-3 ß (GSK-3ß), and c-fos as well as myelination-related transcripts in the prefrontal cortex: myelin basic protein (Mbp), proteolipid protein 1 (Plp1), myelin-associated glycoprotein (Mag), and myelin oligodendrocyte glycoprotein (Mog). The expression of the plasticity markers synaptophysin (Syp) and cAMP response element binding protein (Creb), but not AMPA receptor subunit A2 (GluA2), were affected by genotype. Moreover, in a separate experiment, naïve female Tph2+/- mice showed signs of enhanced stress resilience in the modified swim test with repeated swimming sessions. Taken together, the combination of a moderate reduction in brain 5-HT with environmental challenges results in behavioral changes in female mice that resemble the aggression-related behavior and resilience seen in stressed male mutants; additionally, the combination is comparable to the phenotype of null mutants lacking neuronal 5-HT. Changes in myelination-associated processes are suspected to underpin the molecular mechanisms leading to aggressive behavior.

Hippocampal Over-Expression of Cyclooxygenase-2 (COX-2) Is Associated with Susceptibility to Stress-Induced Anhedonia in Mice.

The phenomenon of individual variability in susceptibility/resilience to stress and depression, in which the hippocampus plays a pivotal role, is attracting increasing attention. We investigated the potential role of hippocampal cyclooxygenase-2 (COX-2), which regulates plasticity, neuroimmune function, and stress responses that are all linked to this risk dichotomy. We used a four-week-long chronic mild stress (CMS) paradigm, in which mice could be stratified according to their susceptibility/resilience to anhedonia, a key feature of depression, to investigate hippocampal expression of COX-2, a marker of microglial activation Iba-1, and the proliferation marker Ki67. Rat exposure, social defeat, restraints, and tail suspension were used as stressors. We compared the effects of treatment with either the selective COX-2 inhibitor celecoxib (30 mg/kg/day) or citalopram (15 mg/kg/day). For the celecoxib and vehicle-treated mice, the Porsolt test was used. Anhedonic (susceptible) but not non-anhedonic (resilient) animals exhibited elevated COX-2 mRNA levels, increased numbers of COX-2 and Iba-1-positive cells in the dentate gyrus and the CA1 area, and decreased numbers of Ki67-positive cells in the subgranular zone of the hippocampus. Drug treatment decreased the percentage of anhedonic mice, normalized swimming activity, reduced behavioral despair, and improved conditioned fear memory. Hippocampal over-expression of COX-2 is associated with susceptibility to stress-induced anhedonia, and its pharmacological inhibition with celecoxib has antidepressant effects that are similar in size to those of citalopram.

Chronic mild stress paradigm as a rat model of depression: facts, artifacts, and future perspectives.

RATIONALE: The chronic mild stress (CMS) paradigm was first described almost 40 years ago and has become a widely used model in the search for antidepressant drugs for major depression disorder (MDD). It has resulted in the publication of almost 1700 studies in rats alone. Under the original CMS procedure, the expression of an anhedonic response, a key symptom of depression, was seen as an essential feature of both the model and a depressive state. The prolonged exposure of rodents to unpredictable/uncontrollable mild stressors leads to a reduction in the intake of palatable liquids, behavioral despair, locomotor inhibition, anxiety-like changes, and vegetative (somatic) abnormalities. Many of the CMS studies do not report these patterns of behaviors, and they often fail to include consistent molecular, neuroanatomical, and physiological phenotypes of CMS-exposed animals. OBJECTIVES: To critically review the CMS studies in rats so that conceptual and methodological flaws can be avoided in future studies. RESULTS: Analysis of the literature supports the validity of the CMS model and its impact on the field. However, further improvements could be achieved by (i) the stratification of animals into 'resilient' and 'susceptible' cohorts within the CMS animals, (ii) the use of more refined protocols in the sucrose test to mitigate physiological and physical artifacts, and (iii) the systematic evaluation of the non-specific effects of CMS and implementation of appropriate adjustments within the behavioral tests. CONCLUSIONS: We propose methodological revisions and the use of more advanced behavioral tests to refine the rat CMS paradigm, which offers a valuable tool for developing new antidepressant medications.

Consortium neuroscience of attention deficit/hyperactivity disorder and autism spectrum disorder: The ENIGMA adventure.

Neuroimaging has been extensively used to study brain structure and function in individuals with attention deficit/hyperactivity disorder (ADHD) and autism spectrum disorder (ASD) over the past decades. Two of the main shortcomings of the neuroimaging literature of these disorders are the small sample sizes employed and the heterogeneity of methods used. In 2013 and 2014, the ENIGMA-ADHD and ENIGMA-ASD working groups were respectively, founded with a common goal to address these limitations. Here, we provide a narrative review of the thus far completed and still ongoing projects of these working groups. Due to an implicitly hierarchical psychiatric diagnostic classification system, the fields of ADHD and ASD have developed largely in isolation, despite the considerable overlap in the occurrence of the disorders. The collaboration between the ENIGMA-ADHD and -ASD working groups seeks to bring the neuroimaging efforts of the two disorders closer together. The outcomes of case-control studies of subcortical and cortical structures showed that subcortical volumes are similarly affected in ASD and ADHD, albeit with small effect sizes. Cortical analyses identified unique differences in each disorder, but also considerable overlap between the two, specifically in cortical thickness. Ongoing work is examining alternative research questions, such as brain laterality, prediction of case-control status, and anatomical heterogeneity. In brief, great strides have been made toward fulfilling the aims of the ENIGMA collaborations, while new ideas and follow-up analyses continue that include more imaging modalities (diffusion MRI and resting-state functional MRI), collaborations with other large databases, and samples with dual diagnoses.

Cadherin-13 is a critical regulator of GABAergic modulation in human stem-cell-derived neuronal networks.

Activity in the healthy brain relies on a concerted interplay of excitation (E) and inhibition (I) via balanced synaptic communication between glutamatergic and GABAergic neurons. A growing number of studies imply that disruption of this E/I balance is a commonality in many brain disorders; however, obtaining mechanistic insight into these disruptions, with translational value for the patient, has typically been hampered by methodological limitations. Cadherin-13 (CDH13) has been associated with autism and attention-deficit/hyperactivity disorder. CDH13 localizes at inhibitory presynapses, specifically of parvalbumin (PV) and somatostatin (SST) expressing GABAergic neurons. However, the mechanism by which CDH13 regulates the function of inhibitory synapses in human neurons remains unknown. Starting from human-induced pluripotent stem cells, we established a robust method to generate a homogenous population of SST and MEF2C (PV-precursor marker protein) expressing GABAergic neurons (iGABA) in vitro, and co-cultured these with glutamatergic neurons at defined E/I ratios on micro-electrode arrays. We identified functional network parameters that are most reliably affected by GABAergic modulation as such, and through alterations of E/I balance by reduced expression of CDH13 in iGABAs. We found that CDH13 deficiency in iGABAs decreased E/I balance by means of increased inhibition. Moreover, CDH13 interacts with Integrin-ß1 and Integrin-ß3, which play opposite roles in the regulation of inhibitory synaptic strength via this interaction. Taken together, this model allows for standardized investigation of the E/I balance in a human neuronal background and can be deployed to dissect the cell-type-specific contribution of disease genes to the E/I balance.

The Combined Effects of Amyloidosis and Serotonin Deficiency by Tryptophan Hydroxylase-2 Knockout Impacts Viability of the APP/PS1 Mouse Model of Alzheimer's Disease.

BACKGROUND: A decline of brain serotonin (5-HT) is held responsible for the changes in mood that can be observed in Alzheimer's disease (AD). However, 5-HT'ergic signaling is also suggested to reduce the production of pathogenic amyloid-ß (Aß). OBJECTIVE: To investigate the effect of targeted inactivation of tryptophan hydroxylase-2 (Tph2), which is essential for neuronal 5-HT synthesis, on amyloidosis in amyloid precursor protein (APP)swe/presenilin 1 (PS1) ΔE9 transgenic mice. METHODS: Triple-transgenic (3xTg) APP/PS1 mice with partial (+/-) or complete Tph2 knockout (-/-) were allowed to survive until 6 months old with APP/PS1, Tph2-/-, and wildtype mice. Survival and weight were recorded. Levels of Aß42/40/38, soluble APPα (sAßPPα) and sAßPPß, and cytokines were analyzed by mesoscale, neurotransmitters by mass spectrometry, and gene expression by quantitative PCR. Tph2, microglia, and Aß were visualized histologically. RESULTS: Tph2 inactivation in APP/PS1 mice significantly reduced viability, without impacting soluble and insoluble Aß42 and Aß40 in neocortex and hippocampus, and with only mild changes of soluble Aß42/Aß40. However, sAßPPα and sAßPPß in hippocampus and Aß38 and Aß40 in cerebrospinal fluid were reduced. 3xTg-/-mice were devoid of Tph2 immunopositive fibers and 5-HT. Cytokines were unaffected by genotype, as were neocortical TNF, HTR2a and HTR2b mRNA levels in Tph2-/- mice. Microglia clustered around Aß plaques regardless of genotype. CONCLUSION: The results suggest that Tph2 inactivation influences AßPP processing, at least in the hippocampus, although levels of Aß are unchanged. The reduced viability of 3xTg-/-mice could indicate that 5-HT protects against the seizures that can impact the viability of APP/PS1 mice.

Sex-Specific ADHD-like Behaviour, Altered Metabolic Functions, and Altered EEG Activity in Sialyltransferase ST3GAL5-Deficient Mice.

A deficiency in GM3-derived gangliosides, resulting from a lack of lactosylceramide-alpha-2,3-sialyltransferase (ST3GAL5), leads to severe neuropathology, including epilepsy and metabolic abnormalities. Disruption of ganglioside production by this enzyme may also have a role in the development of neuropsychiatric disorders. ST3Gal5 knock-out (St3gal5-/-) mice lack a-, b-, and c-series gangliosides, but exhibit no overt neuropathology, possibly owing to the production of compensatory 0-series glycosphingolipids. Here, we sought to investigate the possibility that St3gal5-/- mice might exhibit attention-deficit/hyperactivity disorder (ADHD)-like behaviours. In addition, we evaluated potential metabolic and electroencephalogram (EEG) abnormalities. St3gal5-/- mice were subjected to behavioural testing, glucose tolerance tests, and the levels of expression of brain and peripheral A and B isoforms of the insulin receptor (IR) were measured. We found that St3gal5-/- mice exhibit locomotor hyperactivity, impulsivity, neophobia, and anxiety-like behavior. The genotype also altered blood glucose levels and glucose tolerance. A sex bias was consistently found in relation to body mass and peripheral IR expression. Analysis of the EEG revealed an increase in amplitude in St3gal5-/- mice. Together, St3gal5-/- mice exhibit ADHD-like behaviours, altered metabolic and EEG measures providing a useful platform for better understanding of the contribution of brain gangliosides to ADHD and associated comorbidities.

Increased locomotor activity via regulation of GABAergic signalling in foxp2 mutant zebrafish-implications for neurodevelopmental disorders.

Recent advances in the genetics of neurodevelopmental disorders (NDDs) have identified the transcription factor FOXP2 as one of numerous risk genes, e.g. in autism spectrum disorders (ASD) and attention-deficit/hyperactivity disorder (ADHD). FOXP2 function is suggested to be involved in GABAergic signalling and numerous studies demonstrate that GABAergic function is altered in NDDs, thus disrupting the excitation/inhibition balance. Interestingly, GABAergic signalling components, including glutamate-decarboxylase 1 (Gad1) and GABA receptors, are putative transcriptional targets of FOXP2. However, the specific role of FOXP2 in the pathomechanism of NDDs remains elusive. Here we test the hypothesis that Foxp2 affects behavioural dimensions via GABAergic signalling using zebrafish as model organism. We demonstrate that foxp2 is expressed by a subset of GABAergic neurons located in brain regions involved in motor functions, including the subpallium, posterior tuberculum, thalamus and medulla oblongata. Using CRISPR/Cas9 gene-editing we generated a novel foxp2 zebrafish loss-of-function mutant that exhibits increased locomotor activity. Further, genetic and/or pharmacological disruption of Gad1 or GABA-A receptors causes increased locomotor activity, resembling the phenotype of foxp2 mutants. Application of muscimol, a GABA-A receptor agonist, rescues the hyperactive phenotype induced by the foxp2 loss-of-function. By reverse translation of the therapeutic effect on hyperactive behaviour exerted by methylphenidate, we note that application of methylphenidate evokes different responses in wildtype compared to foxp2 or gad1b loss-of-function animals. Together, our findings support the hypothesis that foxp2 regulates locomotor activity via GABAergic signalling. This provides one targetable mechanism, which may contribute to behavioural phenotypes commonly observed in NDDs.