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The heritability of common psychiatric disorders has motivated global efforts to identify risk-associated genetic variants and elucidate molecular pathways connecting DNA sequence to disease-associated brain dysfunction. The overrepresentation of risk variants among gene regulatory loci instead of protein-coding loci, however, poses a unique challenge in discerning which among the many thousands of variants identified contribute functionally to disease etiology. Defined broadly, psychiatric epigenomics seeks to understand the effects of disease-associated genetic variation on functional readouts of chromatin in an effort to prioritize variants in terms of their impact on gene expression in the brain. Here, we provide an overview of epigenomic mapping in the human brain and highlight findings of particular relevance to psychiatric genetics. Computational methods, including convolutional neuronal networks, and other machine learning approaches hold great promise for elucidating the functional impact of both common and rare genetic variants, thereby refining the epigenomic architecture of psychiatric disorders and enabling integrative analyses of regulatory noncoding variants in the context of large population-level genome and phenome databases., (Copyright © 2020 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.)

Background: Exposure to drugs of abuse alters the epigenetic landscape of the brain's reward regions, such as the nucleus accumbens. We investigated how combinations of chromatin modifications affect genes that regulate responses to cocaine. We focused on Auts2, a gene linked to human evolution and cognitive disorders, which displays strong clustering of cocaine-induced chromatin modifications in this brain region., Methods: We combined chromosome conformation capture, circularized chromosome conformation capture, and related approaches with behavioral paradigms relevant to cocaine phenotypes. Cell type-specific functions were assessed by fluorescence-activated cell sorting and viral-mediated overexpression in Cre-dependent mouse lines., Results: We observed that Auts2 gene expression is increased by repeated cocaine administration specifically in D 2 -type medium spiny neurons in the nucleus accumbens, an effect seen in male but not female mice. Auts2 messenger RNA expression was also upregulated postmortem in the nucleus accumbens of male human cocaine addicts. We obtained evidence that chromosomal looping, bypassing 1524 kb of linear genome, connects Auts2 to the Caln1 gene locus under baseline conditions. This looping was disrupted after repeated cocaine exposure, resulting in increased expression of both genes in D 2 -type medium spiny neurons. Cocaine exposure reduces binding of CCCTC-binding factor, a chromosomal scaffolding protein, and increases histone and DNA methylation at the Auts-Caln1 loop base in the nucleus accumbens. Cell type-specific overexpression of Auts2 or Caln1 in D 2 -type medium spiny neurons demonstrated that both genes promote cocaine reward., Conclusions: These findings suggest that cocaine-induced alterations of neuronal three-dimensional genome organization destabilize higher order chromatin at specific loci that regulate responses to the drug., (Copyright © 2017 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.)

Background: The nervous system may include more than 100 residue-specific posttranslational modifications of histones forming the nucleosome core that are often regulated in cell-type-specific manner. On a genome-wide scale, some of the histone posttranslational modification landscapes show significant overlap with the genetic risk architecture for several psychiatric disorders, fueling PsychENCODE and other large-scale efforts to comprehensively map neuronal and nonneuronal epigenomes in hundreds of specimens. However, practical guidelines for efficient generation of histone chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) datasets from postmortem brains are needed., Methods: Protocols and quality controls are given for the following: 1) extraction, purification, and NeuN neuronal marker immunotagging of nuclei from adult human cerebral cortex; 2) fluorescence-activated nuclei sorting; 3) preparation of chromatin by micrococcal nuclease digest; 4) ChIP for open chromatin-associated histone methylation and acetylation; and 5) generation and sequencing of ChIP-seq libraries., Results: We present a ChIP-seq pipeline for epigenome mapping in the neuronal and nonneuronal nuclei from the postmortem brain. This includes a stepwise system of quality controls and user-friendly data presentation platforms., Conclusions: Our practical guidelines will be useful for projects aimed at histone posttranslational modification mapping in chromatin extracted from hundreds of postmortem brain samples in cell-type-specific manner., Competing Interests: The authors report no biomedical financial interests or potential conflicts of interest., (Copyright © 2016 Society of Biological Psychiatry. All rights reserved.)

Background: Early childhood malnutrition affects 113 million children worldwide, impacting health and increasing vulnerability for cognitive and behavioral disorders later in life. Molecular signatures after childhood malnutrition, including the potential for intergenerational transmission, remain unexplored., Methods: We surveyed blood DNA methylomes (~483,000 individual CpG sites) in 168 subjects across two generations, including 50 generation 1 individuals hospitalized during the first year of life for moderate to severe protein-energy malnutrition, then followed up to 48 years in the Barbados Nutrition Study. Attention deficits and cognitive performance were evaluated with the Connors Adult Attention Rating Scale and Wechsler Abbreviated Scale of Intelligence. Expression of nutrition-sensitive genes was explored by quantitative reverse transcriptase polymerase chain reaction in rat prefrontal cortex., Results: We identified 134 nutrition-sensitive, differentially methylated genomic regions, with most (87%) specific for generation 1. Multiple neuropsychiatric risk genes, including COMT, IFNG, MIR200B, SYNGAP1, and VIPR2 showed associations of specific methyl-CpGs with attention and IQ. IFNG expression was decreased in prefrontal cortex of rats showing attention deficits after developmental malnutrition., Conclusions: Early childhood malnutrition entails long-lasting epigenetic signatures associated with liability for attention and cognition, and limited potential for intergenerational transmission., Competing Interests: Competing Financial Interests: The authors report no biomedical financial interests or potential conflicts of interest., (Copyright © 2016 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.)

Less than 1.5% of the human genome encodes protein. However, vast portions of the human genome are subject to transcriptional and epigenetic regulation, and many noncoding regulatory DNA elements are thought to regulate the spatial organization of interphase chromosomes. For example, chromosomal "loopings" are pivotal for the orderly process of gene expression, by enabling distal regulatory enhancer or silencer elements to directly interact with proximal promoter and transcription start sites, potentially bypassing hundreds of kilobases of interspersed sequence on the linear genome. To date, however, epigenetic studies in the human brain are mostly limited to the exploration of DNA methylation and posttranslational modifications of the nucleosome core histones. In contrast, very little is known about the regulation of supranucleosomal structures. Here, we show that chromosome conformation capture, a widely used approach to study higher-order chromatin, is applicable to tissue collected postmortem, thereby informing about genome organization in the human brain. We introduce chromosome conformation capture protocols for brain and compare higher-order chromatin structures at the chromosome 6p22.2-22.1 schizophrenia and bipolar disorder susceptibility locus, and additional neurodevelopmental risk genes, (DPP10, MCPH1) in adult prefrontal cortex and various cell culture systems, including neurons derived from reprogrammed skin cells. We predict that the exploration of three-dimensional genome architectures and function will open up new frontiers in human brain research and psychiatric genetics and provide novel insights into the epigenetic risk architectures of regulatory noncoding DNA., (Copyright © 2014 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.)

Background: Postmortem brain studies have shown that HDAC1-a lysine deacetylase with broad activity against histones and nonhistone proteins-is frequently expressed at increased levels in prefrontal cortex (PFC) of subjects diagnosed with schizophrenia and related disease. However, it remains unclear whether upregulated expression of Hdac1 in the PFC could affect cognition and behavior., Methods: Using adeno-associated virus, an Hdac1 transgene was expressed in young adult mouse PFC, followed by behavioral assays for working and long-term memory, repetitive activity, and response to novelty. Prefrontal cortex transcriptomes were profiled by microarray. Antipsychotic drug effects were explored in mice treated for 21 days with haloperidol or clozapine., Results: Hdac1 overexpression in PFC neurons and astrocytes resulted in robust impairments in working memory, increased repetitive behaviors, and abnormal locomotor response profiles in novel environments. Long-term memory remained intact. Over 300 transcripts showed subtle but significant changes in Hdac1-overexpressing PFC. Major histocompatibility complex class II (MHC II)-related transcripts, including HLA-DQA1/H2-Aa, HLA-DQB1/H2-Ab1, and HLA-DRB1/H2-Eb1, located in the chromosome 6p21.3-22.1 schizophrenia and bipolar disorder risk locus, were among the subset of genes with a more robust (>1.5-fold) downregulation in expression. Hdac1 levels declined during the course of normal PFC development. Antipsychotic drug treatment, including the atypical clozapine, did not affect Hdac1 levels in PFC but induced expression of multiple MHC II transcripts., Conclusions: Excessive HDAC1 activity, due to developmental defects or other factors, is associated with behavioral alterations and dysregulated expression of MHC II and other gene transcripts in the PFC., (© 2013 Society of Biological Psychiatry.)

Background: Increased neuronal density in prefrontal, parietal, and temporal white matter of schizophrenia subjects is thought to reflect disordered neurodevelopment; however, it is not known if this cellular alteration affects the cingulate cortex and whether similar changes exist in bipolar disorder., Method: Eighty-two postmortem specimens (bipolar 15, schizophrenia 22, control 45) were included in this clinical study. Densities for two neuronal markers, neuron-specific nuclear protein (NeuN) and neuregulin 1 alpha (NRG), were determined in white matter up to 2.5 mm beneath the anterior cingulate cortex; density of NeuN immunopositive neurons (NeuN+) was also determined for a subset of cases in prefrontal cortex. Changes during normal development were monitored in a separate cohort of 14 brains., Results: Both the schizophrenia and bipolar cohorts demonstrated a twofold increase in NeuN+ density in cingulate white matter; this effect could be attributed to approximately 25% of cases that exceeded the second standard deviation from control subjects. Similar changes were observed in prefrontal cortex. In contrast density of NRG expressing neurons was unaltered. Cases with increased NeuN+ densities in two-dimensional (2-D) counts also showed a pronounced, > fivefold elevation in NeuN+ nuclei per cubic millimeter. Additionally, the developmental cohort demonstrated a 75% decline in NeuN+ neuronal density during the first postnatal year but was stable thereafter., Conclusions: Increased neuronal density in white matter of cingulate cortex in schizophrenia provides further evidence that this alteration occurs in multiple cortical areas. Similar changes in some cases with bipolar illness suggest that the two disorders may share a common underlying defect in late prenatal or early postnatal neurodevelopment.

Background: Prefrontal deficits in gamma-aminobutyric acid (GABA)ergic gene expression, including neuropeptide Y (NPY), somatostatin (SST), and parvalbumin (PV) messenger RNAs (mRNAs), have been reported for multiple schizophrenia cohorts. Preclinical models suggest that a subset of these GABAergic markers (NPY/SST) is regulated by brain-derived neurotrophic factor (BDNF), which in turn is under the inhibitory influence of small noncoding RNAs. However, it remains unclear if these mechanisms are important determinants for dysregulated NPY and SST expression in prefrontal cortex (PFC) of subjects with schizophrenia., Methods: Using a postmortem case-control design, the association between BDNF protein, NPY/SST/PV mRNAs, and two BDNF-regulating microRNAs (miR-195 and miR-30a-5p) was determined in samples from the PFC of 20 schizophrenia and 20 control subjects. Complementary studies were conducted in cerebral cortex of mice subjected to antipsychotic treatment or a brain-specific ablation of the Bdnf gene., Results: Subjects with schizophrenia showed deficits in NPY and PV mRNAs. Within-pair differences in BDNF protein levels showed strong positive correlations with NPY and SST and a robust inverse association with miR-195 levels, which in turn were not affected by antipsychotic treatment or genetic ablation of Bdnf., Conclusions: Taken together, these results suggest that prefrontal deficits in a subset of GABAergic mRNAs, including NPY, are dependent on the regional supply of BDNF, which in turn is fine-tuned through a microRNA (miRNA)-mediated mechanism.

Alterations in RNA levels are frequently reported in brain of subjects diagnosed with autism, schizophrenia, depression, and other psychiatric diseases, but it remains unclear whether the underlying molecular pathology involves changes in gene expression, as opposed to alterations in messenger RNA processing. Pre-clinical studies have revealed that stress, drugs, and a variety of other environmental factors lead to changes in RNA levels in brain via epigenetic mechanisms, including modification of histone proteins. A number of site-specific modifications of the nucleosome core histones-including the trimethylated forms of histone H3 lysines K4, K9, and K27-are of particular interest for postmortem research, because these marks differentiate between active and inactive chromatin and seem to remain relatively stable during tissue autolysis. Therefore, histone methylation profiling at promoter regions could provide important clues about mechanisms of gene expression in human brain during development and in disease. Intriguingly, mutations within the genes encoding the H3K9-specific methyltransferase, EHMT1, and the H3K4-specific histone demethylase, JARID1C/SMCX, have been linked to mental retardation and autism, respectively. In addition, the H3K4-specific methyltransferase, MLL1, is essential for hippocampal synaptic plasticity and might be involved in cortical dysfunction of some cases of schizophrenia. Together, these findings emphasize the potential significance of histone lysine methylation for orderly brain development and also as a molecular toolbox to study chromatin function in postmortem tissue.

Background: Chromatin remodeling, including changes in histone acetylation, might play a role in the pathophysiology and treatment of depression. We investigated whether the histone deacetylase inhibitor sodium butyrate (SB) administered as single drug or in combination with the selective serotonin reuptake inhibitor (SSRI) fluoxetine exerts antidepressant-like effects in mice., Methods: Mice (C57BL/6J) received injections of SB, fluoxetine, or a combination of both drugs either acutely or chronically for a period of 28 days and were subjected to a battery of tests to measure anxiety and behavioral despair. Histone acetylation and expression of brain-derived neurotrophic factor (BDNF) were monitored in hippocampus and frontal cortex., Results: Co-treatment with SB and fluoxetine resulted in a significant 20%-40% decrease in immobility scores in the tail suspension test (TST), a measure for behavioral despair, both acutely and chronically. In contrast, decreased immobility after single drug regimens was limited either to the acute (fluoxetine) or chronic (SB) paradigm. Systemic injection of SB induced short-lasting histone hyperacetylation in hippocampus and frontal cortex. Among the four treatment paradigms that resulted in improved immobility scores in the TST, three were associated with a transient, at least 50% increase in BDNF transcript in frontal cortex, whereas changes in hippocampus were less consistent., Conclusions: The histone deacetylase inhibitor SB exerts antidepressant-like effects in the mouse. The therapeutic benefits and molecular actions of histone modifying drugs, including co-treatment with SSRIs and other newer generation antidepressant medications, warrant further exploration in experimental models.