Can you remember silence? Epigenetic memory and reversibility as a site of intervention.

Just over 20 years ago, molecular biologists Leonie Ringrose and Renato Paro published an article with a provocative title, "Remembering Silence", in BioEssays. The article focused on how epigenetic elements could return to their silent state, operationally defined as their epigenetic status before their modulation by experimental or environmental factors. Though Ringrose and Paro's article was on fruit flies and factors affecting embryological growth, the article asked a question of considerable importance to rapidly expanding research in neuroepigenetics on the correlation between trauma and neuropsychiatric risk: If you experience a traumatic event and, as a result, acquire an epigenetic trait that is considered pathological, can you free yourself of that trait? Ultimately, we are interested in how a return to silence is envisioned in neuroepigenetics research, how interventions purported to bring about that silence might function, and what this might mean for people who live in the aftermath of trauma.

Functional genomic mechanisms of opioid action and opioid use disorder: a systematic review of animal models and human studies.

In the past two decades, over-prescription of opioids for pain management has driven a steep increase in opioid use disorder (OUD) and death by overdose, exerting a dramatic toll on western countries. OUD is a chronic relapsing disease associated with a lifetime struggle to control drug consumption, suggesting that opioids trigger long-lasting brain adaptations, notably through functional genomic and epigenomic mechanisms. Current understanding of these processes, however, remain scarce, and have not been previously reviewed systematically. To do so, the goal of the present work was to synthesize current knowledge on genome-wide transcriptomic and epigenetic mechanisms of opioid action, in primate and rodent species. Using a prospectively registered methodology, comprehensive literature searches were completed in PubMed, Embase, and Web of Science. Of the 2709 articles identified, 73 met our inclusion criteria and were considered for qualitative analysis. Focusing on the 5 most studied nervous system structures (nucleus accumbens, frontal cortex, whole striatum, dorsal striatum, spinal cord; 44 articles), we also conducted a quantitative analysis of differentially expressed genes, in an effort to identify a putative core transcriptional signature of opioids. Only one gene, Cdkn1a, was consistently identified in eleven studies, and globally, our results unveil surprisingly low consistency across published work, even when considering most recent single-cell approaches. Analysis of sources of variability detected significant contributions from species, brain structure, duration of opioid exposure, strain, time-point of analysis, and batch effects, but not type of opioid. To go beyond those limitations, we leveraged threshold-free methods to illustrate how genome-wide comparisons may generate new findings and hypotheses. Finally, we discuss current methodological development in the field, and their implication for future research and, ultimately, better care.

Homeorhesis: envisaging the logic of life trajectories in molecular research on trauma and its effects.

What sets someone on a life trajectory? This question is at the heart of studies of 21st-century neurosciences that build on scientific models developed over the last 150 years that attempt to link psychopathology risk and human development. Historically, this research has documented persistent effects of singular, negative life experiences on people's subsequent development. More recently, studies have documented neuromolecular effects of early life adversity on life trajectories, resulting in models that frame lives as disproportionately affected by early negative experiences. This view is dominant, despite little evidence of the stability of the presumably early-developed molecular traits and their potential effects on phenotypes. We argue that in the context of gaps in knowledge and the need for scientists to reason across molecular and phenotypic scales, as well as time spans that can extend beyond an individual's life, specific interpretative frameworks shape the ways in which individual scientific findings are assessed. In the process, scientific reasoning oscillates between understandings of cellular homeostasis and organisms' homeorhesis, or life trajectory. Biologist and historian François Jacob described this framework as the "attitude" that researchers bring to bear on their "objects" of study. Through an analysis of, first, historical and contemporary scientific literature and then ethnographic research with neuroscientists, we consider how early life trauma came to be associated with specific psychological and neurobiological effects grounded in understandings of life trajectories. We conclude with a consideration of the conceptual, ontological, and ethical implications of interpreting life trajectories as the result of the persistence of long-embodied biological traits, persistent life environments, or both.

Plasticity of the epigenome during early-life stress.

Early life adversity remains a significant risk factor for the development of a host of negative behavioural and pathological outcomes in adulthood long after the stressor is over. Recent evidence indicates that these lasting effects of ELS may occur via alterations in the epigenetic landscape. Here, we review the main findings of the effects of early life adversity on DNA methylation, histone post-translational modification, and non-coding RNAs in the context of psychiatric disease in animal models and human cohorts. We specifically explore how early life adversity alters epigenetic patterns in both a global manner, and in specific candidate genes that play a role in relevant systems such as the hypothalamic-pituitary-adrenal axis, as well as neurotransmitter and neuroendocrine signalling. We also discuss how individual factors, such as genetics, sex, and age, as well as the type, and timing of early life adversity, can create differential susceptibility and significantly moderate outcomes. Although challenges remain in deciphering the complexity of how the early environment interacts with individual factors to determine epigenetic patterns, as well as how to translate these mechanistic findings into clinically relevant populations, the reviewed literature sheds light on the potential of the field to identify effective interventions for vulnerable individuals.

The opioid system and the social brain: implications for depression and suicide.

In the past decade, considerable attention has been drawn to social interactions and behaviors as sources of pleasurable (social reward) and painful (social pain) emotional states. While the role of the opioid system in the regulation of reward and pain processes has long been recognized, it has more recently been investigated and characterized in the specific context of social experiences across several mammalian species. Accordingly, the present narrative review provides a comprehensive summary of studies detailing how the opioid system controls social reward and social pain. From a translational and pathophysiological perspective, we further discuss how opioid-dependent regulation of social behaviors may contribute to depressive illness and suicidal behaviors, and ultimately provide innovative therapeutic opportunities.

Suicide and suicide behaviors: A review of transcriptomics and multiomics studies in psychiatric disorders.

Aberrant gene expression can contribute to brain and nervous system dysfunction that causes many psychiatric illnesses. Here, we review how transcriptomic approaches have deepened our understanding of the neurobiological underpinnings of psychiatric disorders and how they have to the identification of biomarkers for these disorders, with a focus on their relevance to suicide and suicide behaviors. We begin by providing an overview of the genetic, transcriptomic, and epigenetic factors (including non-coding RNAs) implicated in suicide and their roles in the regulation of gene expression. Then, we highlight the unique benefits and limitations of using either postmortem brain or peripheral tissues in transcriptomic analyses. We examine the current shift from candidate gene to genome-wide approaches in psychiatric research, which are concurrently emerging with the increased consideration of the Research Domain Criteria (RDoC) framework for classifying mental illnesses. Furthermore, we discuss the use of transcription networks and how they can be integrated into multiomic analyses. Finally, we end by highlighting recent findings of peripheral markers of suicide risk identified through the use of transcriptomic tools. Technological advancements and increased accessibility of these technologies are drastically shaping the current research landscape. We present an overview of the significant changes currently taking place to usher in a new era of psychiatric research.

Hyperactivity of Anterior Cingulate Cortex Areas 24a/24b Drives Chronic Pain-Induced Anxiodepressive-like Consequences.

Pain associates both sensory and emotional aversive components, and often leads to anxiety and depression when it becomes chronic. Here, we characterized, in a mouse model, the long-term development of these sensory and aversive components as well as anxiodepressive-like consequences of neuropathic pain and determined their electrophysiological impact on the anterior cingulate cortex (ACC, cortical areas 24a/24b). We show that these symptoms of neuropathic pain evolve and recover in different time courses following nerve injury in male mice. In vivo electrophysiological recordings evidence an increased firing rate and bursting activity within the ACC when anxiodepressive-like consequences developed, and this hyperactivity persists beyond the period of mechanical hypersensitivity. Whole-cell patch-clamp recordings also support ACC hyperactivity, as shown by increased excitatory postsynaptic transmission and contribution of NMDA receptors. Optogenetic inhibition of the ACC hyperactivity was sufficient to alleviate the aversive and anxiodepressive-like consequences of neuropathic pain, indicating that these consequences are underpinned by ACC hyperactivity.SIGNIFICANCE STATEMENT Chronic pain is frequently comorbid with mood disorders, such as anxiety and depression. It has been shown that it is possible to model this comorbidity in animal models by taking into consideration the time factor. In this study, we aimed at determining the dynamic of different components and consequences of chronic pain, and correlated them with electrophysiological alterations. By combining electrophysiological, optogenetic, and behavioral analyses in a mouse model of neuropathic pain, we show that the mechanical hypersensitivity, ongoing pain, anxiodepressive consequences, and their recoveries do not necessarily exhibit temporal synchrony during chronic pain processing, and that the hyperactivity of the anterior cingulate cortex is essential for driving the emotional impact of neuropathic pain.

A Dual Noradrenergic Mechanism for the Relief of Neuropathic Allodynia by the Antidepressant Drugs Duloxetine and Amitriptyline.

In addition to treating depression, antidepressant drugs are also a first-line treatment for neuropathic pain, which is pain secondary to lesion or pathology of the nervous system. Despite the widespread use of these drugs, the mechanism underlying their therapeutic action in this pain context remains partly elusive. The present study combined data collected in male and female mice from a model of neuropathic pain and data from the clinical setting to understand how antidepressant drugs act. We show two distinct mechanisms by which the selective inhibitor of serotonin and noradrenaline reuptake duloxetine and the tricyclic antidepressant amitriptyline relieve neuropathic allodynia. One of these mechanisms is acute, central, and requires descending noradrenergic inhibitory controls and α2A adrenoceptors, as well as the mu and delta opioid receptors. The second mechanism is delayed, peripheral, and requires noradrenaline from peripheral sympathetic endings and ß2 adrenoceptors, as well as the delta opioid receptors. We then conducted a transcriptomic analysis in dorsal root ganglia, which suggested that the peripheral component of duloxetine action involves the inhibition of neuroimmune mechanisms accompanying nerve injury, including the downregulation of the TNF-α-NF-κB signaling pathway. Accordingly, immunotherapies against either TNF-α or Toll-like receptor 2 (TLR2) provided allodynia relief. We also compared duloxetine plasma levels in the animal model and in patients and we observed that patients' drug concentrations were compatible with those measured in animals under chronic treatment involving the peripheral mechanism. Our study highlights a peripheral neuroimmune component of antidepressant drugs that is relevant to their delayed therapeutic action against neuropathic pain.SIGNIFICANCE STATEMENT In addition to treating depression, antidepressant drugs are also a first-line treatment for neuropathic pain, which is pain secondary to lesion or pathology of the nervous system. However, the mechanism by which antidepressant drugs can relieve neuropathic pain remained in part elusive. Indeed, preclinical studies led to contradictions concerning the anatomical and molecular substrates of this action. In the present work, we overcame these apparent contradictions by highlighting the existence of two independent mechanisms. One is rapid and centrally mediated by descending controls from the brain to the spinal cord and the other is delayed, peripheral, and relies on the anti-neuroimmune action of chronic antidepressant treatment.

Medium throughput bisulfite sequencing for accurate detection of 5-methylcytosine and 5-hydroxymethylcytosine.

BACKGROUND: Epigenetic modifications of DNA, such as 5-methylcytosine and 5-hydroxymethycytosine, play important roles in development and disease. Here, we present a cost-effective and versatile methodology for the analysis of DNA methylation in targeted genomic regions, which comprises multiplexed, PCR-based preparation of bisulfite DNA libraries followed by customized MiSeq sequencing. RESULTS: Using bisulfite and oxidative bisulfite conversion of DNA, we have performed multiplexed targeted sequencing to analyse several kilobases of genomic DNA in up to 478 samples, and achieved high coverage data of 5-methylcytosine and 5-hydroxymethycytosine at single-base resolution. Our results demonstrate the ability of this methodology to detect all levels of cytosine modifications at greater than 100× coverage in large sample sets at low cost compared to other targeted methods. CONCLUSIONS: This approach can be applied to multiple settings, from candidate gene to clinical studies, and is especially useful for validation of differentially methylated or hydroxymethylated regions following whole-genome analyses.

BisQC: an operational pipeline for multiplexed bisulfite sequencing.

BACKGROUND: Bisulfite sequencing is the most efficient single nucleotide resolution method for analysis of methylation status at whole genome scale, but improved quality control metrics are needed to better standardize experiments. RESULTS: We describe BisQC, a step-by-step method for multiplexed bisulfite-converted DNA library construction, pooling, spike-in content, and bioinformatics. We demonstrate technical improvements for library preparation and bioinformatic analyses that can be done in standard laboratories. We find that decoupling amplification of bisulfite converted (bis) DNA from the indexing reaction is an advantage, specifically in reducing total PCR cycle number and pre-selecting high quality bis-libraries. We also introduce a progressive PCR method for optimal library amplification and size-selection. At the sequencing stage, we thoroughly test the benefits of pooling non-bis DNA library with bis-libraries and find that BisSeq libraries can be pooled with a high proportion of non-bis DNA libraries with minimal impact on BisSeq output. For informatics analysis, we propose a series of optimization steps including the utilization of the mitochondrial genome as a QC standard, and we assess the validity of using duplicate reads for coverage statistics. CONCLUSION: We demonstrate several quality control checkpoints at the library preparation, pre-sequencing, post-sequencing, and post-alignment stages, which should prove useful in determining sample and processing quality. We also determine that including a significant portion of non-bisulfite converted DNA with bisulfite converted DNA has a minimal impact on usable bisulfite read output.

Multiple serotonergic paths to antidepressant efficacy.

Depression is a leading cause of disability worldwide. Brain mechanisms underlying the clinical antidepressant efficacy of selective serotonin reuptake inhibitors (SSRI), currently the first-line treatment, remain poorly understood. Recent animal studies have implicated multiple serotonin receptor subtypes in SSRI response, opening new therapeutic perspectives.

Mu Opioid Receptor-Positive Neurons in the Dorsal Raphe Nucleus Are Impaired by Morphine Abstinence.

BACKGROUND: Chronic opioid exposure leads to hedonic deficits and enhanced vulnerability to addiction, which are observed and even strengthen after a period of abstinence, but the underlying circuit mechanisms are poorly understood. In this study, using both molecular and behavioral approaches, we tested the hypothesis that neurons expressing mu opioid receptors (MORs) in the dorsal raphe nucleus (DRN) are involved in addiction vulnerability associated with morphine abstinence. METHODS: MOR-Cre mice were exposed to chronic morphine and then went through spontaneous withdrawal for 4 weeks, a well-established mouse model of morphine abstinence. We studied DRN-MOR neurons of abstinent mice using 1) viral translating ribosome affinity for transcriptome profiling, 2) fiber photometry to measure neuronal activity, and 3) an opto-intracranial self-stimulation paradigm applied to DRN-MOR neurons to assess responses related to addiction vulnerability including persistence to respond, motivation to obtain the stimulation, self-stimulation despite punishment, and cue-induced reinstatement. RESULTS: DRN-MOR neurons of abstinent animals showed a downregulation of genes involved in ion conductance and MOR-mediated signaling, as well as altered responding to acute morphine. Opto-intracranial self-stimulation data showed that abstinent animals executed more impulsive-like and persistent responses during acquisition and scored higher on addiction-like criteria. CONCLUSIONS: Our data suggest that protracted abstinence to chronic morphine leads to reduced MOR function in DRN-MOR neurons and abnormal self-stimulation of these neurons. We propose that DRN-MOR neurons have partially lost their reward-facilitating properties, which in turn may lead to increased propensity to perform addiction-related behaviors.

Accelerated aging in bipolar disorders: An exploratory study of six epigenetic clocks.

Bipolar disorder (BD) is a chronic and severe psychiatric disorder associated with significant medical morbidity and reduced life expectancy. In this study, we assessed accelerated epigenetic aging in individuals with BD using various DNA methylation (DNAm)-based markers. For this purpose, we used five epigenetic clocks (Horvath, Hannum, EN, PhenoAge, and GrimAge) and a DNAm-based telomere length clock (DNAmTL). DNAm profiles were obtained using Infinium MethylationEPIC Arrays from whole-blood samples of 184 individuals with BD. We also estimated blood cell counts based on DNAm levels for adjustment. Significant correlations between chronological age and each epigenetic age estimated using the six different clocks were observed. Following adjustment for blood cell counts, we found that the six epigenetic AgeAccels (age accelerations) were significantly associated with the body mass index. GrimAge AgeAccel was significantly associated with male sex, smoking status and childhood maltreatment. DNAmTL AgeAccel was significantly associated with smoking status. Overall, this study showed that distinct epigenetic clocks are sensitive to different aspects of aging process in BD. Further investigations with comprehensive epigenetic clock analyses and large samples are required to confirm our findings of potential determinants of an accelerated epigenetic aging in BD.

The basolateral amygdala-anterior cingulate pathway contributes to depression-like behaviors and comorbidity with chronic pain behaviors in male mice.

While depression and chronic pain are frequently comorbid, underlying neuronal circuits and their psychopathological relevance remain poorly defined. Here we show in mice that hyperactivity of the neuronal pathway linking the basolateral amygdala to the anterior cingulate cortex is essential for chronic pain-induced depression. Moreover, activation of this pathway in naive male mice, in the absence of on-going pain, is sufficient to trigger depressive-like behaviors, as well as transcriptomic alterations that recapitulate core molecular features of depression in the human brain. These alterations notably impact gene modules related to myelination and the oligodendrocyte lineage. Among these, we show that Sema4a, which was significantly upregulated in both male mice and humans in the context of altered mood, is necessary for the emergence of emotional dysfunction. Overall, these results place the amygdalo-cingulate pathway at the core of pain and depression comorbidity, and unravel the role of Sema4a and impaired myelination in mood control.

The molecular pathophysiology of mood disorders: From the analysis of single molecular layers to multi-omic integration.

Next-generation sequencing now enables the rapid and affordable production of reliable biological data at multiple molecular levels, collectively referred to as "omics". To maximize the potential for discovery, computational biologists have created and adapted integrative multi-omic analytical methods. When applied to diseases with traceable pathophysiology such as cancer, these new algorithms and statistical approaches have enabled the discovery of clinically relevant molecular mechanisms and biomarkers. In contrast, these methods have been much less applied to the field of molecular psychiatry, although diagnostic and prognostic biomarkers are similarly needed. In the present review, we first briefly summarize main findings from two decades of studies that investigated single molecular processes in relation to mood disorders. Then, we conduct a systematic review of multi-omic strategies that have been proposed and used more recently. We also list databases and types of data available to researchers for future work. Finally, we present the newest methodologies that have been employed for multi-omics integration in other medical fields, and discuss their potential for molecular psychiatry studies.

Oxytocin receptor expression and epigenetic regulation in the anterior cingulate cortex of individuals with a history of severe childhood abuse.

Childhood abuse significantly increases the lifetime risk of negative mental health outcomes. The oxytocinergic system, which plays a role in complex social and emotional behaviors, has been shown to be sensitive to early-life experiences. While previous studies have investigated the relationship between early-life adversity and oxytocin, they did so with peripheral samples. We, therefore, aimed to characterize the relationship between early-life adversity and oxytocin receptor (OXTR) expression in the brain, using post-mortem human samples, as well as a rodent model of naturally occurring variation in early-life environment. Focusing on the dorsal anterior cingulate cortex, we compared OXTR expression and epigenetic regulation between MDD suicides with (N = 26) and without history of childhood abuse (N = 24), as well as psychiatrically healthy controls (N = 23). We also compared Oxtr expression in the cingulate cortex of adult rats raised by dams displaying high (N = 13) and low levels (N = 12) of licking and grooming (LG) behavior. Overall, our results indicate that childhood abuse associates with an upregulation of OXTR expression, and that similarly, this relationship is also observed in the cingulate cortex of adult rats raised by low-LG dams. Additionally, we found an effect of rs53576 genotype on expression, showing that carriers of the A variant also show upregulated OXTR expression. The effects of early-life adversity and rs53576 genotype on OXTR expression are, however, not explained by differences in DNA methylation within and around the MT region of the OXTR gene.

Increased functional coupling of the mu opioid receptor in the anterior insula of depressed individuals.

The mu opioid receptor (MOR) is a G protein-coupled receptor that plays an essential role in reward and hedonic processes, and that has been implicated in disorders such as depression and addiction. Over the last decade, several brain imaging studies in depressed patients have consistently found that dysregulation of MOR function occurs in particular in the anterior insular cortex, an important brain site for the perception of internal states and emotional regulation. To investigate molecular mechanisms that may underlie these effects, here we assessed genetic polymorphisms, expression, and functional G-protein coupling of MOR in a large post-mortem cohort (N = 95) composed of depressed individuals who died by suicide, and healthy controls. Results indicated that depression, but not comorbid substance use disorder or acute opiate consumption, was associated with increased MOR activity. This effect was partly explained by a specific increase in expression of the inhibitory alpha G-protein subunit GNAI2. Consistent with previous neuroimaging studies, our findings support the notion that enhanced endogenous opioidergic tone in the anterior insula may buffer negative affective states in depressed individuals, a mechanism that could potentially contribute to the antidepressant efficacy of emerging opioid-based medications.

Non-CG methylation and multiple histone profiles associate child abuse with immune and small GTPase dysregulation.

Early-life adversity (ELA) is a major predictor of psychopathology, and is thought to increase lifetime risk by epigenetically regulating the genome. Here, focusing on the lateral amygdala, a major brain site for emotional homeostasis, we describe molecular cross-talk among multiple mechanisms of genomic regulation, including 6 histone marks and DNA methylation, and the transcriptome, in subjects with a history of ELA and controls. In the healthy brain tissue, we first uncover interactions between different histone marks and non-CG methylation in the CAC context. Additionally, we find that ELA associates with methylomic changes that are as frequent in the CAC as in the canonical CG context, while these two forms of plasticity occur in sharply distinct genomic regions, features, and chromatin states. Combining these multiple data indicates that immune-related and small GTPase signaling pathways are most consistently impaired in the amygdala of ELA individuals. Overall, this work provides insights into genomic brain regulation as a function of early-life experience.

The Role of Opiates in Social Pain and Suicidal Behavior.

With more than 800,000 deaths by suicide each year and 20 to 30 times more suicide attempts worldwide, suicide is a major public health problem. Current treatments of SB are mainly based on pharmacological treatments that are not specific of SB (e.g. antidepressants), and new therapeutic targets are urgently needed. Recent data strengthen the ancient conception pain (social, psychic, physical) that is at the core of the suicidal process and should be incorporated in the clinical assessment of suicide risk. Then, the mechanisms involved in the regulation of pain may open new avenues regarding therapeutic perspectives. Opiates appear to be a promising candidate in treatment of SB. Indeed, since the last two decades, growing evidences suggest an implication of the opioid system in the pathophysiology of SB, this conduct to the elaboration of randomized controlled trials (RCTs) using opiates in patients with SB. Results suggesting an anti-suicidal effect of buprenorphine and the potential opioidergic-related anti-suicidal effect of ketamine both contribute to the growing interest in opiates use in SB. In this review, we will summarize a large part of the evidence that leads researchers and clinicians to be interested in the use of opiates for SB treatment and discuss on new opioid pharmacological options for suicidal patients.

Neural and molecular correlates of psychological pain during major depression, and its link with suicidal ideas.

OBJECTIVES: Psychological pain increases the risk of suicidal ideas and acts, and represents a potential therapeutic target. However, the mechanisms of mental pain remain unclear. Here, we assessed the peripheral transcriptomic and central neural correlates of mental pain during a depressive episode. METHODS: 172 adult un-medicated depressed patients were recruited. Leucocytes were extracted for RNA quantification at baseline (T0) and after 8 weeks (T8) of an antidepressant treatment. Ninety-nine genes of the cortisol, immune, opioid, serotonergic, and kynurenine systems were a priori selected, and 41 were sufficiently expressed to be analyzed. At both T0 and T8, mean level of mental pain over the last 15 days was measured with a visual analog scale. A subset of 38 patients was additionally scanned with Magnetic Resonance Imaging at T0. Resting-state sequences of 4 networks (default-mode, basal ganglia, central executive, salience) were examined. RESULTS: Mean psychological pain scores significantly decreased between T0 and T8. At conservative p-corrected levels, T0 mental pain was significantly correlated with 11 brain clusters encompassing the prefrontal, parietal, and temporal cortices, the striatum, and the cerebellum. There was no direct association between peripheral gene expression and mean mental pain at any time points or in terms of temporal changes. However, expressions of 5HTR2B at p-corrected levels, and 5HTR3A, TPH1, and OPRL1 were correlated with the activity of several identified brain clusters at T0. Finally, while suicidal ideas and mental pain were correlated, the neural and molecular correlates of suicidal ideas were not the same. CONCLUSION: Our study suggests that the serotonergic and nociceptin systems are associated with the activity of a cortico-subcortical brain network underlying the perception of mental pain during depression. Mental pain may be a necessary but insufficient condition for the emergence of suicidal ideation during depression.