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Epilepsy-associated developmental lesions, including malformations of cortical development and low-grade developmental tumors, represent a major cause of drug-resistant seizures requiring surgical intervention in children. Brain-restricted somatic mosaicism has been implicated in the genetic etiology of these lesions; however, many contributory genes remain unidentified.We enrolled 50 children who were undergoing epilepsy surgery into a translational research study. Resected tissue was divided for clinical neuropathologic evaluation and genomic analysis. We performed exome and RNA sequencing to identify somatic variation and we confirmed our findings using high-depth targeted DNA sequencing.We uncovered candidate disease-causing somatic variation affecting 28 patients (56%), as well as candidate germline variants affecting 4 patients (8%). In agreement with previous studies, we identified somatic variation affecting solute carrier family 35 member A2 (SLC35A2) and mechanistic target of rapamycin kinase (MTOR) pathway genes in patients with focal cortical dysplasia. Somatic gains of chromosome 1q were detected in 30% (3 of 10) of patients with Type I focal cortical dysplasia (FCD)s. Somatic variation in mitogen-activated protein kinase (MAPK) pathway genes (i.e., fibroblast growth factor receptor 1 [FGFR1], FGFR2, B-raf proto-oncogene, serine/threonine kinase [BRAF], and KRAS proto-oncogene, GTPase [KRAS]) was associated with low-grade epilepsy-associated developmental tumors. RNA sequencing enabled the detection of somatic structural variation that would have otherwise been missed, and which accounted for more than one-half of epilepsy-associated tumor diagnoses. Sampling across multiple anatomic regions revealed that somatic variant allele fractions vary widely within epileptogenic tissue. Finally, we identified putative disease-causing variants in genes not yet associated with focal cortical dysplasia.These results further elucidate the genetic basis of structural brain abnormalities leading to focal epilepsy in children and point to new candidate disease genes.

Variants in the AUTS2 gene are associated with a broad spectrum of neurological conditions characterized by intellectual disability, microcephaly, and congenital brain malformations. Here, we use a human cerebral organoid (CO) model to investigate the pathophysiology of a heterozygous de novo missense AUTS2 variant identified in a patient with multiple neurological impairments including primary microcephaly and profound intellectual disability. Proband COs exhibit reduced growth, deficits in neural progenitor cell (NPC) proliferation and disrupted NPC polarity within ventricular zone-like regions compared to control COs. We used CRISPR-Cas9-mediated gene editing to correct this variant and demonstrate rescue of impaired organoid growth and NPC proliferative deficits. Single-cell RNA sequencing revealed a marked reduction of G1/S transition gene expression and alterations in WNT-β-Catenin signaling within proband NPCs, uncovering a novel role for AUTS2 in NPCs during human cortical development. Collectively, these results underscore the value of COs to uncover molecular mechanisms underlying AUTS2 syndrome.

Summary Cerebral organoids (COs) are rapidly accelerating the rate of translational neuroscience based on their potential to model complex features of the developing human brain. Several studies have examined the electrophysiological and neural network features of COs; however, no study has comprehensively investigated the developmental trajectory of electrophysiological properties in whole-brain COs and correlated these properties with developmentally linked morphological and cellular features. Here, we profiled the neuroelectrical activities of COs over the span of 5 months with a multi-electrode array platform and observed the emergence and maturation of several electrophysiologic properties, including rapid firing rates and network bursting events. To complement these analyses, we characterized the complex molecular and cellular development that gives rise to these mature neuroelectrical properties with immunohistochemical and single-cell transcriptomic analyses. This integrated approach highlights the value of COs as an emerging model system of human brain development and neurological disease., Highlights • CO electrophysiology can be quantified with a multi-electrode array method • CO electrophysiological trajectories correlate with molecular and cellular development • The neurotrophin/TRK signaling pathway is active in COs by 5 months in culture, Cerebral organoids (COs) have a unique advantage in modeling human-specific features of early brain development. Here, we profile their neuroelectrical activities with an MEA platform over time and correlate these activities with their increasingly complex molecular and cellular features. This integrated approach highlights the value of COs as an emerging model system of human brain development and neurological disease.

Epilepsy-associated developmental lesions, including malformations of cortical development and low-grade developmental tumors, represent a major cause of drug-resistant seizures requiring surgical intervention in children. Brain-restricted somatic mosaicism has been implicated in the genetic etiology of these lesions; however, many contributory genes remain unidentified. We enrolled 50 children undergoing epilepsy surgery into a translational research study. We performed exome and RNA-sequencing of resected brain tissue samples to identify somatic variation. We uncovered candidate disease-causing somatic variation affecting 28 patients (56%), as well as candidate germline variants affecting 4 patients (8%). We confirmed somatic findings using high-depth targeted DNA sequencing. In agreement with previous studies, we identified somatic variation affecting SLC35A2 and MTOR pathway genes in patients with focal cortical dysplasia. Somatic gains of chromosome 1q were detected in 30% (3 of 10) Type I FCD patients. Somatic variation of MAPK pathway genes (i.e., FGFR1, FGFR2, BRAF, KRAS) was associated with low-grade epilepsy-associated developmental tumors. Somatic structural variation accounted for over one-half of epilepsy-associated tumor diagnoses. Sampling across multiple anatomic regions revealed that somatic variant allele fractions vary widely within epileptogenic tissue. Finally, we identified putative disease-causing variants in genes (EEF2, NAV2, PTPN11) not yet associated with focal cortical dysplasia. These results further elucidate the genetic basis of structural brain abnormalities leading to focal epilepsy in children and point to new candidate disease genes.

Variants in the AUTS2 gene are associated with a broad spectrum of neurological conditions characterized by intellectual disability, microcephaly, and congenital brain malformations. Here, we use a human cerebral organoid model to investigate the pathophysiology of a heterozygous de novo missense AUTS2 variant identified in a patient with multiple neurological impairments including primary microcephaly and profound intellectual disability. Proband cerebral organoids exhibit reduced growth, deficits in neural progenitor cell (NPC) proliferation and disrupted NPC polarity within ventricular zone-like regions compared to control cerebral organoids. We used CRISPR-Cas9-mediated gene editing to correct this variant and demonstrate rescue of impaired organoid growth and NPC proliferative deficits. Single-cell RNA sequencing revealed a marked reduction of G1/S transition gene expression and alterations in WNT-β-catenin signalling within proband NPCs, uncovering a novel role for AUTS2 in NPCs during human cortical development. Collectively, these results underscore the value of cerebral organoids to investigate molecular mechanisms underlying AUTS2 syndrome.

Transposable elements (TEs) are mobile sequences that engender widespread mutations and thus are a major hazard that must be silenced. The most abundant active class of TEs in mammalian genomes is long interspersed element class 1 (LINE1). Here, we report that LINE1 transposition is suppressed in the male germline by transcription factors encoded by a rapidly evolving X-linked homeobox gene cluster. LINE1 transposition is repressed by many members of this RHOX transcription factor family, including those with different patterns of expression during spermatogenesis. One family member—RHOX10—suppresses LINE1 transposition during fetal development in vivo when the germline would otherwise be susceptible to LINE1 activation because of epigenetic reprogramming. We provide evidence that RHOX10 suppresses LINE transposition by inducing Piwil2, which encodes a key component in the Piwi-interacting RNA pathway that protects against TEs. The ability of RHOX transcription factors to suppress LINE1 is conserved in humans but is lost in RHOXF2 mutants from several infertile human patients, raising the possibility that loss of RHOXF2 causes human infertility by allowing uncontrolled LINE1 expression in the germline. Together, our results support a model in which the Rhox gene cluster is in an evolutionary arms race with TEs, resulting in expansion of the Rhox gene cluster to suppress TEs in different biological contexts.

Transposable elements (TEs) are mobile sequences that engender widespread mutations and thus are a major hazard that must be silenced. The most abundant active class of TEs in mammalian genomes is long interspersed element class 1 (LINE1). Here, we report that LINE1 transposition is suppressed in the male germline by transcription factors encoded by a rapidly evolving X-linked homeobox gene cluster. LINE1 transposition is repressed by many members of this RHOX transcription factor family, including those with different patterns of expression during spermatogenesis. One family member-RHOX10-suppresses LINE1 transposition during fetal development in vivo when the germline would otherwise be susceptible to LINE1 activation because of epigenetic reprogramming. We provide evidence that RHOX10 suppresses LINE transposition by inducing Piwil2, which encodes a key component in the Piwi-interacting RNA pathway that protects against TEs. The ability of RHOX transcription factors to suppress LINE1 is conserved in humans but is lost in RHOXF2 mutants from several infertile human patients, raising the possibility that loss of RHOXF2 causes human infertility by allowing uncontrolled LINE1 expression in the germline. Together, our results support a model in which the Rhox gene cluster is in an evolutionary arms race with TEs, resulting in expansion of the Rhox gene cluster to suppress TEs in different biological contexts.

Phosphatase and tensin homologue (PTEN) regulates cell growth and survival through inhibition of the mammalian target of rapamycin (MTOR) signalling pathway. Germline genetic variation of PTEN is associated with autism, macrocephaly and PTEN hamartoma tumour syndromes. The effect of developmental PTEN somatic mutations on nervous system phenotypes is not well understood, although brain somatic mosaicism of MTOR pathway genes is an emerging cause of cortical dysplasia and epilepsy in the paediatric population. Here we report two somatic variants of PTEN affecting a single patient presenting with intractable epilepsy and hemimegalencephaly that varied in clinical severity throughout the left cerebral hemisphere. High-throughput sequencing analysis of affected brain tissue identified two somatic variants in PTEN. The first variant was present in multiple cell lineages throughout the entire hemisphere and associated with mild cerebral overgrowth. The second variant was restricted to posterior brain regions and affected the opposite PTEN allele, resulting in a segmental region of more severe malformation, and the only neurons in which it was found by single-nuclei RNA-sequencing had a unique disease-related expression profile. This study reveals brain mosaicism of PTEN as a disease mechanism of hemimegalencephaly and furthermore demonstrates the varying effects of single- or bi-allelic disruption of PTEN on cortical phenotypes.

Gastroblastomas are rare tumors with a biphasic epithelioid/spindle cell morphology that typically present in early adulthood and have recurrent MALAT1-GLI1 fusions. We describe an adolescent patient with Wiskott-Aldrich syndrome who presented with a large submucosal gastric tumor with biphasic morphology. Despite histologic features consistent with gastroblastoma, a MALAT1-GLI1 fusion was not found in this patient's tumor; instead, comprehensive molecular profiling identified a novel EWSR1-CTBP1 fusion and no other significant genetic alterations. The tumor also overexpressed NOTCH and FGFR by RNA profiling. The novel fusion and expression profile suggest a role for epithelial-mesenchymal transition in this tumor, with potential implications for the pathogenesis of biphasic gastric tumors such as gastroblastoma.

BackgroundPediatric cancers typically have a distinct genomic landscape when compared to adult cancers and frequently carry somatic gene fusion events that alter gene expression and drive tumorigenesis. Sensitive and specific detection of gene fusions through the analysis of next-generation-based RNA sequencing (RNA-Seq) data is computationally challenging and may be confounded by low tumor cellularity or underlying genomic complexity. Furthermore, numerous computational tools are available to identify fusions from supporting RNA-Seq reads, yet each algorithm demonstrates unique variability in sensitivity and precision, and no clearly superior approach currently exists. To overcome these challenges, we have developed an ensemble fusion calling approach to increase the accuracy of identifying fusions.ResultsOur ensemble fusion detection approach utilizes seven fusion calling algorithms: Arriba, CICERO, FusionMap, FusionCatcher, JAFFA, MapSplice, and STAR-Fusion, which are packaged as a fully automated pipeline using Docker and AWS serverless technology. This method uses paired end RNA-Seq sequence reads as input, and the output from each algorithm is examined to identify fusions detected by a consensus of at least three algorithms. These consensus fusion results are filtered by comparison to an internal database to remove likely artifactual fusions occurring at high frequencies in our internal cohort, while a “known fusion list” prevents failure to report known pathogenic events. We have employed the ensemble fusion-calling pipeline on RNA-Seq data from 229 patients with pediatric cancer or blood disorders studied under an IRB-approved protocol. The samples consist of 138 central nervous system tumors, 73 solid tumors, and 18 hematologic malignancies or disorders. The combination of an ensemble fusion-calling pipeline and a knowledge-based filtering strategy identified 67 clinically relevant fusions among our cohort (diagnostic yield of 29.3%), including RBPMS-MET, BCAN-NTRK1, and TRIM22-BRAF fusions. Following clinical confirmation and reporting in the patient’s medical record, both known and novel fusions provided medically meaningful information.ConclusionsOur ensemble fusion detection pipeline offers a streamlined approach to discover fusions in cancer, at higher levels of sensitivity and accuracy than single algorithm methods. Furthermore, this method accurately identifies driver fusions in pediatric cancer, providing clinical impact by contributing evidence to diagnosis and, when appropriate, indicating targeted therapies.

ObjectiveMany genetic studies of intractable epilepsy in pediatric patients primarily focus on inherited, constitutional genetic deficiencies identified in patient blood. Recently, studies have revealed somatic mosaicism associated with epilepsy in which genetic variants are present only in a subset of brain cells. We hypothesize that tissue-specific, somatic mosaicism represents an important genetic etiology in epilepsy and aim to discover somatic alterations in epilepsy-affected brain tissue.MethodsWe have pursued a research study to identify brain somatic mosaicism, using next-generation sequencing (NGS) technologies, in patients with treatment refractory epilepsy who have undergone surgical resection of affected brain tissue.ResultsWe used an integrated combination of NGS techniques and conventional approaches (radiology, histopathology, and electrophysiology) to comprehensively characterize multiple brain regions from a single patient with intractable epilepsy. We present a 3-year-old male patient with West syndrome and intractable tonic seizures in whom we identified a pathogenic frameshift somatic variant in SLC35A2, present at a range of variant allele fractions (4.2%–19.5%) in 12 different brain tissues detected by targeted sequencing. The proportion of the SLC35A2 variant correlated with severity and location of neurophysiology and neuroimaging abnormalities for each tissue.ConclusionsOur findings support the importance of tissue-based sequencing and highlight a correlation in our patient between SLC35A2 variant allele fractions and the severity of epileptogenic phenotypes in different brain tissues obtained from a grid-based resection of clinically defined epileptogenic regions.

Wilson disease is a medically actionable rare autosomal recessive disorder of defective copper excretion caused by mutations in ATP7B, one of two highly evolutionarily conserved copper-transporting ATPases. Hundreds of disease-causing variants in ATP7B have been reported to public databases; more than half of these are missense changes, and a significant proportion are presumed unequivocal loss-of-function variants (nonsense, frameshift, and canonical splice site). Current molecular genetic testing includes sequencing all coding exons (±10 bp) as well as deletion/duplication testing, with reported sensitivity of >98%. We report a proband from a consanguineous family with a biochemical phenotype consistent with early-onset Wilson disease who tested negative on conventional molecular genetic testing. Using a combination of whole-genome sequencing and transcriptome sequencing, we found that the proband's disease is due to skipping of exons 6–7 of the ATP7B gene associated with a novel intronic variant (NM_000053.4:c.1947-19T > A) that alters a putative splicing enhancer element. This variant was also homozygous in the proband's younger sister, whose subsequent clinical evaluations revealed biochemical evidence of Wilson disease. Our work adds to emerging evidence that ATP7B exon skipping from deep intronic variants outside typical splice junctions is an important mechanism of Wilson disease; the variants responsible may elude standard genetic testing.

Genomic plasticity during brain development Mice genomes contain many mobile retrotransposons. Bedrosian et al. analyzed DNA from the mouse hippocampus during development (see the Perspective by Song and Gleeson). They found that the amount of maternal care in the first few weeks of a mouse pup's life affected the number of copies of the L1 retrotransposon. The experience of maternal care was thus “recorded” in the DNA of these mice pups during a time when the brain was still actively developing. Science , this issue p. 1395 ; see also p. 1330

Nighttime lighting is one of the great conveniences of modernization; however, there is mounting evidence that inopportune light exposure can disrupt physiological and behavioral functions. Hospital patients may be particularly vulnerable to the consequences of light at night due to their compromised physiological state. Cardiac arrest/cardiopulmonary resuscitation (CA) was used to test the hypothesis in mice that exposure to dim light at night impairs central nervous system (CNS) recovery from a major pathological insult. Mice exposed to dim light at night (5 lx) had higher mortality in the week following cardiac arrest compared to mice housed in dark nights (0 lx). Neuronal damage was significantly greater in surviving mice exposed to dim light at night after CA versus those housed in dark nights. Dim light at night may have elevated neuronal damage by amplifying pro-inflammatory pathways in the CNS; Iba1 immunoreactivity (an indication of microglia activation) and pro-inflammatory cytokine expression were elevated in mice exposed to dim light at night post-CA. Furthermore, selective inhibition of IL-1β or TNFα ameliorated damage in mice exposed to dim light at night. The effects of light at night on CA outcomes were also prevented by using a wavelength of nighttime light that has minimal impact on the endogenous circadian clock, suggesting that replacing broad-spectrum nighttime light with specific circadian-inert wavelengths could be protective. Together, these data indicate that exposure to dim light at night after global cerebral ischemia increases neuroinflammation, in turn exacerbating neurological damage and potential for mortality.

Environmental light-dark cycles play an important role in behavioural and physiological processes. It is essential that laboratory vivaria be designed to properly control the light conditions in which laboratory mice are housed; however, this is not universally the case. Some laboratory vivarium doors are designed with windows, which allow light from the hallways to leak into the housing space during the rodents’ dark phase. Personnel entering and exiting the housing space during the dark phase can also create excessive light leak from brightly illuminated hallways. In this study, we investigated the hypothesis that exposure to dim light at night, as commonly experienced in many laboratory rodent housing spaces, alters mouse (Mus musculus) behaviour. We specifically analysed patterns of locomotor activity, anxiety- and depressive-like responses. Exposure to dim (5 lux) light at night altered home-cage locomotor activity and increased anxiety and some depressive responses among laboratory mice. These results suggest that light conditions can alter mouse behaviour and potentially influence experimental outcomes. Increased care should be taken to properly control light-dark conditions for laboratory animals.

Humans and other organisms have adapted to a consistent and predictable 24-h solar cycle, but over the past ~130 years the widespread adoption of electric light has transformed our environment. Instead of aligning behavioral and physiological processes to the natural solar cycle, individuals respond to artificial light cycles created by social and work schedules. Urban light pollution, night shift work, transmeridian travel, televisions and computers have dramatically altered the timing of light used to entrain biological rhythms. In humans and other mammals, light is detected by the retina and intrinsically photosensitive retinal ganglion cells project this information both to the circadian system and limbic brain regions. Therefore, it is possible that exposure to light at night, which has become pervasive, may disrupt both circadian timing and mood. Notably, the rate of major depression has increased in recent decades, in parallel with increasing exposure to light at night. Strong evidence already links circadian disruption to major depression and other mood disorders. Emerging evidence from the past few years suggests that exposure to light at night also negatively influences mood. In this review, we discuss evidence from recent human and rodent studies supporting the novel hypothesis that nighttime exposure to light disrupts circadian organization and contributes to depressed mood.

Both normal aging and dementia are associated with altered circadian regulation of physiology and behavior. Elderly individuals and dementia patients commonly experience disrupted sleep-wake cycles, which may lead to psychomotor agitation, confusion, and wandering. These behaviors are disruptive to both patients and caregivers. Sundowning syndrome, which encompasses many of these behaviors, is characterized by a temporal pattern in the severity of symptoms, usually expressed as worse during the late afternoon or evening. Other than antipsychotic medications, off-label medications, and restraint, few treatment options are available. The aim of this paper is to review mouse studies of circadian behavioral disturbances relevant to sundowning, in order to determine potential models for studying the mechanisms of sundowning syndrome. The emergence of a useful mouse model should facilitate the development of novel therapeutic approaches.

The healthy human brain is a mosaic of varied genomes. Using a single cell sequencing approach targeting L1 elements, the authors show that the contribution of L1 to somatic mosaicism goes beyond retrotransposition and includes deletion of genomic regions associated with L1. The healthy human brain is a mosaic of varied genomes. Long interspersed element-1 (LINE-1 or L1) retrotransposition is known to create mosaicism by inserting L1 sequences into new locations of somatic cell genomes. Using a machine learning-based, single-cell sequencing approach, we discovered that somatic L1-associated variants (SLAVs) are composed of two classes: L1 retrotransposition insertions and retrotransposition-independent L1-associated variants. We demonstrate that a subset of SLAVs comprises somatic deletions generated by L1 endonuclease cutting activity. Retrotransposition-independent rearrangements in inherited L1s resulted in the deletion of proximal genomic regions. These rearrangements were resolved by microhomology-mediated repair, which suggests that L1-associated genomic regions are hotspots for somatic copy number variants in the brain and therefore a heritable genetic contributor to somatic mosaicism. We demonstrate that SLAVs are present in crucial neural genes, such as DLG2 (also called PSD93), and affect 44–63% of cells of the cells in the healthy brain.

The prevalence of major depression has increased in recent decades and women are twice as likely as men to develop the disorder. Recent environmental changes almost certainly have a role in this phenomenon, but a complete set of contributors remains unspecified. Exposure to artificial light at night (LAN) has surged in prevalence during the past 50 years, coinciding with rising rates of depression. Chronic exposure to LAN is linked to increased risk of breast cancer, obesity and mood disorders, although the relationship to mood is not well characterized. In this study, we investigated the effects of chronic exposure to 5 lux LAN on depression-like behaviors in female hamsters. Using this model, we also characterized hippocampal brain-derived neurotrophic factor expression and hippocampal dendritic morphology, and investigated the reversibility of these changes 1, 2 or 4 weeks following elimination of LAN. Furthermore, we explored the mechanism of action, focusing on hippocampal proinflammatory cytokines given their dual role in synaptic plasticity and the pathogenesis of depression. Using reverse transcription-quantitative PCR, we identified a reversible increase in hippocampal tumor necrosis factor (TNF), but not interleukin-1β, mRNA expression in hamsters exposed to LAN. Direct intracerebroventricular infusion of a dominant-negative inhibitor of soluble TNF, XPro1595, prevented the development of depression-like behavior under LAN, but had no effect on dendritic spine density in the hippocampus. These results indicate a partial role for TNF in the reversible depression-like phenotype observed under chronic dim LAN. Recent environmental changes, such as LAN exposure, may warrant more attention as possible contributors to rising rates of mood disorders.

In most parts of the world, environmental conditions vary in a predictable seasonal manner. Thus, seasonal variation in reproductive timing and immune function has emerged in some species to cope with disparate seasonal demands. During the long days of spring and summer when food availability is high and thermoregulatory demands low, Siberian hamsters invest in reproduction, whereas during the harsh short days of winter hamsters divert energy away from reproductive activities and modify immune capabilities. Many seasonal adaptations can be recapitulated in a laboratory setting by adjusting day length (photoperiod). Early-life photoperiods are important sources of seasonal information and can establish an individual’s developmental trajectory. Siberian hamsters housed under short days (SD; 8 h light/day) recover more rapidly than long-day (LD; 16 h light/day) hamsters from immune activation with lipopolysaccharide (LPS). SD hamsters attenuate fever response, reduce cytokine production, and abrogate behavioral responses following LPS injection. The mechanism by which SD Siberian hamsters attenuate febrile response remains unspecified. It is possible that periphery-to-brain communication of inflammatory signals is altered by exposure to photoperiod. Rather than testing photoperiod effects on each of the multiple routes by which immunological cues are communicated to the CNS, we administered LPS intracerebroventricularly (i.c.v.) following adolescent exposure to either 6 weeks of SD or LD. Injection of LPS i.c.v. led to a similar immune reaction in SD hamsters as previously reported with intraperitoneal injection. Short days attenuated the response to LPS with diminished fever spike and duration, as well as decreased locomotor inactivity. Furthermore, only LD hamsters demonstrated anhedonic-like behavior following LPS injection as evaluated by decreased preference for a milk solution. These results suggest that photoperiodic differences in response to infection are due in part to changes in central immune activation.

Many nontropical species undergo physiological and behavioral adaptations in response to seasonal changes in photoperiod, or day length. In most rodent species, short winter photoperiods reduce testosterone concentrations, which provoke gonadal regression and reduce testosterone-dependent behaviors such as mating and aggression. Seasonally-breeding Siberian hamsters, however, are paradoxically more aggressive in short-days, despite much reduced reproductive activity and testosterone concentrations. Nitric oxide (NO) signaling has been proposed as part of an alternate mechanism underlying this phenomenon. A reduction in neuronal nitric oxide synthase (nNOS), the enzyme responsible for synthesizing NO in the brain, is associated with increased aggression in male short-day hamsters. In the present study, we hypothesized that pharmacological inhibition of nNOS would increase aggressive behavior in long days, but not in short days because nNOS is already reduced. Adult male Siberian hamsters were housed in either long (LD 16:8 h) or short (LD 8:16 h) photoperiods for 8 weeks, then treated with either the selective nNOS inhibitor, 3-bromo-7-nitroindazole (3BrN) or oil vehicle, and subsequently tested for aggression in a resident-intruder test. Treatment with 3BrN increased attack frequency and duration in long days, but had no effect in short days. Short days also reduced testosterone concentrations, without any effect of treatment. These data provide further evidence linking reduced nNOS to elevated short-day aggression and support a role for NO signaling in this phenomenon.

Aggression frequently coincides with specific dimensions of emotionality, such as impulsivity, risk-taking, and drug abuse. Serotonergic (5-HTergic) neurotransmission contributes to the regulation of numerous neurobiological functions, and is thought to play a key role in modulating aggressive responses. The current study uses selectively-bred High (bHR) and Low (bLR) Responder rats that exhibit differences in emotionality and behavioral control, with bHRs exhibiting heightened novelty-induced exploration, impulsivity, and increased sensitivity to drugs of abuse, and with bLRs characterized by exaggerated depressive- and anxiety-like behaviors. Based on this behavioral profile we hypothesized that bHR rats exhibit increased aggression along with changes in testosterone and corticosterone secretion characteristic of aggression, and that these changes are accompanied by alterations in the expression of key genes that regulate 5-HTergic neurotransmission (Tph2 and Sert) as well as in the activation of 5-HTergic cell groups following aggressive encounter. Our data demonstrate that when compared to bLR rats, bHRs express increased baseline Tph2 and Sert in select brainstem nuclei, and when tested on the resident-intruder test they exhibited: 1) increased aggressive behavior; 2) potentiated corticosterone and testosterone secretion; and 3) diminished intrusion-induced c-fos expression in select 5-HTergic brainstem cell groups. The most prominent gene expression differences occurred in the B9 cell group, pontomesencephalic reticular formation, median raphe, and the gigantocellular nucleus pars α. These data are consistent with the notion that altered 5-HT neurotransmission contributes to bHRs' heightened aggression. Furthermore, they indicate that a specific subset of brainstem 5-HTergic cell groups contributes to the regulation of intrusion-elicited behavioral responses.

Both normal aging and dementia are associated with dysregulation of the biological clock, which contributes to disrupted circadian organization of physiology and behavior. Diminished circadian organization in conjunction with the loss of cholinergic input to the cortex likely contributes to impaired cognition and behavior. One especially notable and relatively common circadian disturbance among the aged is “sundowning syndrome,” which is characterized by exacerbated anxiety, agitation, locomotor activity, and delirium during the hours before bedtime. Sundowning has been reported in both dementia patients and cognitively intact elderly individuals living in institutions; however, little is known about temporal patterns in anxiety and agitation, and the neurobiological basis of these rhythms remains unspecified. In the present study, we explored the diurnal pattern of anxiety-like behavior in aged and amyloid precursor protein (APP) transgenic mice. We then attempted to treat the observed behavioral disturbances in the aged mice using chronic nightly melatonin treatment. Finally, we tested the hypothesis that time-of-day differences in acetylcholinesterase and choline acetyltransferase expression and general neuronal activation (i.e., c-Fos expression) coincide with the behavioral symptoms. Our results show a temporal pattern of anxiety-like behavior that emerges in elderly mice. This behavioral pattern coincides with elevated locomotor activity relative to adult mice near the end of the dark phase, and with time-dependent changes in basal forebrain acetylcholinesterase expression. Transgenic APP mice show a similar behavioral phenomenon that is not observed among age-matched wild-type mice. These results may have useful applications to the study and treatment of age- and dementia-related circadian behavioral disturbances, namely, sundowning syndrome.

Species have been adapted to specific niches optimizing survival and reproduction; however, urbanization by humans has dramatically altered natural habitats. Artificial light at night (LAN), termed ‘light pollution’, is an often overlooked, yet increasing disruptor of habitats, which perturbs physiological processes that rely on precise light information. For example, LAN alters the timing of reproduction and activity in some species, which decreases the odds of successful breeding and increases the threat of predation for these individuals, leading to reduced fitness. LAN also suppresses immune function, an important proxy for survival. To investigate the impact of LAN in a species naive to light pollution in its native habitat, immune function was examined in Siberian hamsters derived from wild-caught stock. After four weeks exposure to dim LAN, immune responses to three different challenges were assessed: (i) delayed-type hypersensitivity (DTH), (ii) lipopolysaccharide-induced fever, and (iii) bactericide activity of blood. LAN suppressed DTH response and reduced bactericide activity of blood after lipopolysaccharide treatment, in addition to altering daily patterns of locomotor activity, suggesting that human encroachment on habitats via night-time lighting may inadvertently compromise immune function and ultimately fitness.

In the version of this article initially published, NIH grant U01 MH106882 to F.H.G. was missing from the Acknowledgments. The error has been corrected in the HTML and PDF versions of the article.

Disruption of circadian rhythms, provoked by artificial lighting at night, inconsistent sleep-wake schedules, and transmeridian air travel, is increasingly prevalent in modern society. Desynchrony of biological rhythms from environmental light cycles has dramatic consequences for human health. In particular, disrupting homeostatic oscillations in endocrine tissues and the hormones that these tissues regulate can have cascading effects on physiology and behavior. Accumulating evidence suggests that chronic disruption of circadian organization of endocrine function may lead to metabolic, reproductive, sleep, and mood disorders. This review discusses circadian control of endocrine systems and the consequences of distorting rhythmicity of these systems.

Melatonin is the principle hormonal product of the pineal gland. It is secreted with a robust daily rhythm, peaking near the middle of the night. During the daytime, concentrations remain very low, as exposure to light robustly suppresses its secretion. The regulation of melatonin by light is well-characterized, but an interesting feature of the daily melatonin rhythm is that its peak occurs near the middle of the night and then levels begin to drop hours before morning light exposure. The mechanism underlying the light-independent drop in melatonin during late night remains unspecified. Feedback control is one mechanism of hormone regulation, but no studies thus far have explored the possibility of such regulation in the pineal of white-footed mice (Peromyscus leucopus). The pineal gland and SCN express melatonin receptors, and melatonin regulates its own receptor density in the brain. We investigated the possibility of feedback control of melatonin by administering melatonin receptor antagonists to female white-footed mice and then measuring plasma melatonin concentrations. In the first experiment, we observed that luzindole, a dual MT1/MT2 receptor antagonist administered 1h after lights off, caused an increase in plasma melatonin both 1 and 2h later. In a second experiment, we did not observe a change in melatonin concentrations following injection of an antagonist specific for the MT2 subtype. These results suggest the possibility of feedback control of melatonin release, occurring preferentially through the MT1 receptor subtype.

Nat. Neurosci. 19, 1583–1591 (2016); published online 12 September 2016; corrected after print 13 July 2017 In the version of this article initially published, NIH grant T32 CA009370 to F.H.G. was missing from the Acknowledgments. The error has been corrected in the HTML and PDF versions of the article.

Temporal organization of physiology is critical for human health. In the past, humans experienced predictable periods of daily light and dark driven by the solar day, which allowed for entrainment of intrinsic circadian rhythms to the environmental light–dark cycles. Since the adoption of electric light, however, pervasive exposure to nighttime lighting has blurred the boundaries of day and night, making it more difficult to synchronize biological processes. Many systems are under circadian control, including sleep–wake behavior, hormone secretion, cellular function and gene expression. Circadian disruption by nighttime light perturbs those processes and is associated with increasing incidence of certain cancers, metabolic dysfunction and mood disorders. This review focuses on the role of artificial light at night in mood regulation, including mechanisms through which aberrant light exposure affects the brain. Converging evidence suggests that circadian disruption alters the function of brain regions involved in emotion and mood regulation. This occurs through direct neural input from the clock or indirect effects, including altered neuroplasticity, neurotransmission and clock gene expression. Recently, the aberrant light exposure has been recognized for its health effects. This review summarizes the evidence linking aberrant light exposure to mood.

Violence is a significant public health problem worldwide. Neurobiological research on violence and aggression attempts to elucidate the cellular and molecular pathways that increase the propensity toward this behavior. Research over the past 40 years has implicated several brain regions and neurotransmitters in aggression, mainly using rodent models. Perhaps the strongest association is the link between serotonin and aggression, which has compelling interactions with the nitric oxide system. Recently, new insights into these relationships have been added as modern techniques allow more sophisticated analyses. This chapter will discuss current developments implicating serotonin and nitric oxide in aggressive behavior. Recently developed high-resolution methods for examining the neurobiological basis of aggression will be considered, with emphasis on future directions for the field.

Life on earth is entrained to a 24 h solar cycle that synchronizes circadian rhythms in physiology and behavior; light is the most potent entraining cue. In mammals, light is detected by (1) rods and cones, which mediate visual function, and (2) intrinsically photosensitive retinal ganglion cells (ipRGCs), which primarily project to the suprachiasmatic nucleus (SCN) in the hypothalamus to regulate circadian rhythms. Recent evidence, however, demonstrates that ipRGCs also project to limbic brain regions, suggesting that, through this pathway, light may have a role in cognition and mood. Therefore, it follows that unnatural exposure to light may have negative consequences for mood or behavior. Modern environmental lighting conditions have led to excessive exposure to light at night (LAN), and particularly to blue wavelength lights. We hypothesized that nocturnal light exposure (i.e., dim LAN) would induce depressive responses and alter neuronal structure in hamsters (Phodopus sungorus). If this effect is mediated by ipRGCs, which have reduced sensitivity to red wavelength light, then we predicted that red LAN would have limited effects on brain and behavior compared with shorter wavelengths. Additionally, red LAN would not induce c-Fos activation in the SCN. Our results demonstrate that exposure to LAN influences behavior and neuronal plasticity and that this effect is likely mediated by ipRGCs. Modern sources of LAN that contain blue wavelengths may be particularly disruptive to the circadian system, potentially contributing to altered mood regulation.

Several physiological and behavioral processes rely on precisely timed light information derived from the natural solar cycle. Using this information, traits have adapted to allow individuals within specific niches to optimize survival and reproduction, but urbanization by humans has significantly altered natural habitats. Nighttime light exposure alters immune function in several species, which could lead to decreased fitness or survival, particularly in the face of an environmental challenge. We exposed male Siberian hamsters (Phodopus sungorus) to five lux of light at night for four weeks, and then administered six hours of acute restraint stress. Delayed-type hypersensitivity (DTH) response was assessed immediately following stress. Acute restraint increased the DTH reaction in dark nights, but exposure to nighttime light prevented this response. Exposure to light at night prolonged the DTH response in non-stressed control hamsters. These results suggest that light pollution may significantly alter physiological responses in Siberian hamsters, particularly in response to a salient environmental challenge such as stress.

Humans and other organisms have adapted to a 24-h solar cycle in response to life on Earth. The rotation of the planet on its axis and its revolution around the sun cause predictable daily and seasonal patterns in day length. To successfully anticipate and adapt to these patterns in the environment, a variety of biological processes oscillate with a daily rhythm of approximately 24 h in length. These rhythms arise from hierarchally-coupled cellular clocks generated by positive and negative transcription factors of core circadian clock gene expression. From these endogenous cellular clocks, overt rhythms in activity and patterns in hormone secretion and other homeostatic processes emerge. These circadian rhythms in physiology and behaviour can be organised by a variety of cues, although they are most potently entrained by light. In recent history, there has been a major change from naturally-occurring light cycles set by the sun, to artificial and sometimes erratic light cycles determined by the use of electric lighting. Virtually every individual living in an industrialised country experiences light at night (LAN) but, despite its prevalence, the biological effects of such unnatural lighting have not been fully considered. Using female Siberian hamsters (Phodopus sungorus), we investigated the effects of chronic nightly exposure to dim light on daily rhythms in locomotor activity, serum cortisol concentrations and brain expression of circadian clock proteins (i.e. PER1, PER2, BMAL1). Although locomotor activity remained entrained to the light cycle, the diurnal fluctuation of cortisol concentrations was blunted and the expression patterns of clock proteins in the suprachiasmatic nucleus and hippocampus were altered. These results demonstrate that chronic exposure to dim LAN can dramatically affect fundamental cellular function and emergent physiology.

Violence is a significant public health problem worldwide. Neurobiological research on violence and aggression attempts to elucidate the cellular and molecular pathways that increase the propensity toward this behavior. Research over the past 40 years has implicated several brain regions and neurotransmitters in aggression, mainly using rodent models. Perhaps the strongest association is the link between serotonin and aggression, which has compelling interactions with the nitric oxide system. Recently, new insights into these relationships have been added as modern techniques allow more sophisticated analyses. This chapter will discuss current developments implicating serotonin and nitric oxide in aggressive behavior. Recently developed high-resolution methods for examining the neurobiological basis of aggression will be considered, with emphasis on future directions for the field.

Chronic exposure to light at night (LAN) is a circadian disruptor and may be linked to various health risks, including mood disorders. We recently demonstrated that chronic exposure to dim (5 lux) LAN provokes depressive-like behaviors and reduced hippocampal CA1 dendritic spine density in female hamsters. Whether this model is responsive to selective serotonin reuptake inhibitors remains unspecified. In this study, we exposed hamsters to 5 lux LAN and treated with citalopram to determine effects on depressive-like behavior and CA1 dendritic spine density. Female hamsters were ovariectomized at adulthood and housed in either a standard light-dark cycle (LD) or dim LAN (dLAN). After 4 weeks exposure, treatment with either citalopram or vehicle was administered for 2 weeks while hamsters remained in experimental lighting conditions. Depressive-like behavior was assayed using the forced swim test and brains were processed for Golgi-Cox staining and analyzed for dendritic spine density. Treatment with citalopram rescued behavior in the forced swim test in hamsters housed in dLAN, but had no effect on hamsters housed in LD. Dendritic spine density in CA1 was moderately improved by citalopram treatment, but not fully restored. These results validate our LAN paradigm as a depression model by showing citalopram selectively improves depressive-like behavior in dLAN conditions, but not in LD conditions. These data also suggest standard SSRI therapy may be effective for individuals experiencing depression related to circadian disruption and LAN exposure, such as shift workers.

Circadian dysfunction has long been implicated in the etiology of mood disorders. The gene Clock and related molecules (e.g. Per1, Per2) represent key regulators of circadian rhythmicity, and their targeted disruption in mutant mice produces potentiated reward drive, novelty-seeking, impulsivity, disrupted sleep, reduced depression and anxiety - a behavioral profile highly reminiscent of our selectively bred high responder (bHR) rats compared to bred low responders (bLRs). The current study evaluated potential diurnal bHR-bLR differences in behavior, gene expression, and neuroendocrinology. Relative to bHRs, bLRs showed diminished homecage locomotion during the dark (but not light) phase and a delayed corticosterone peak. In situ hybridizations in hypothalamus, amygdala, and hippocampus at Zeitgeber Time (ZT)2 and ZT14 revealed distinct bHR-bLR day-night gene expression fluctuations. bHRs exhibited altered day-night patterns of corticotrophin releasing hormone (CRH) and arginine vasopression (AVP) mRNA in the hypothalamus, and perturbed hippocampal MR:GR ratios relative to bLR rats. bHR-bLR rats showed disparate day-night Clock expression in the suprachiasmatic nucleus, a master circadian oscillator, with bHRs showing higher levels at ZT14 versus ZT2 and bLRs showing the opposite pattern. Clock, Per1 and Per2 were assessed in the substantia nigra pars compacta (SNc) and ventral tegmental area (VTA) since disruption of these genes induces "bHR-like" behavior in mutant mice. Clock and Per1 did not differ between strains, but there were robust Per2 differences, with bHRs having reduced Per2 in VTA and SNc. These findings resonate with earlier work demonstrating that perturbation of Clock and related molecules contributes to disturbances of emotional and addictive behaviors.

The prevalence of major depression has increased in recent decades; however, the underlying causes of this phenomenon remain unspecified. One environmental change that has coincided with elevated rates of depression is increased exposure to artificial light at night. Shift workers and others chronically exposed to light at night are at increased risk of mood disorders, suggesting that nighttime illumination may influence brain mechanisms mediating affect. We tested the hypothesis that exposure to dim light at night may impact affective responses and alter morphology of hippocampal neurons. Ovariectomized adult female Siberian hamsters (Phodopus sungorus) were housed for 8 weeks in either a light/dark cycle (LD) or a light/dim light cycle (DM), and then behavior was assayed. DM-hamsters displayed more depression-like responses in the forced swim and the sucrose anhedonia tests compared with LD-hamsters. Conversely, in the elevated plus maze DM-hamsters reduced anxiety-like behaviors. Brains from the same animals were processed using the Golgi-Cox method and hippocampal neurons within CA1, CA3, and the dentate gyrus were analyzed for morphological characteristics. In CA1, DM-hamsters significantly reduced dendritic spine density on both apical and basilar dendrites, an effect which was not mediated by baseline cortisol, as concentrations were equivalent between groups. These results demonstrate dim light at night is sufficient to reduce synaptic spine connections to CA1. Importantly, the present results suggest that night-time low level illumination, comparable to levels that are pervasive in North America and Europe, may contribute to the increasing prevalence of mood disorders.

Selective breeding of rats exhibiting differences in novelty-induced locomotion revealed that this trait predicts several differences in emotional behavior. Bred High Responders (bHRs) show exaggerated novelty-induced locomotion, aggression, and psychostimulant self-administration, compared to bred Low Responders (bLRs), which are inhibited and prone to anxiety- and depression-like behavior. Our breeding studies highlight the heritability of the bHR/bLR phenotypes, although environmental factors like maternal care also shape some aspects of these traits. We previously reported that HR vs. LR mothers act differently, but it was unclear whether their behaviors were genetically driven or influenced by their pups. The present study (a) used cross-fostering to evaluate whether the bHR/bLR maternal styles are inherent to mothers and/or are modulated by pups; and (b) assessed oxytocin and oxytocin receptor mRNA expression to examine possible underpinnings of bHR/bLR maternal differences. While bHR dams exhibited less maternal behavior than bLRs during the dark/active phase, they were very attentive to pups during the light phase, spending greater time passive nursing and in contact with pups compared to bLRs. Cross-fostering only subtly changed bHR and bLR dams' behavior, suggesting that their distinct maternal styles are largely inherent to the mothers. We also found elevated oxytocin mRNA levels in the supraoptic nucleus of the hypothalamus in bHR versus bLR dams, which may play some role in driving their behavior differences. Overall these studies shed light on the interplay between the genetics of mothers and infants in driving differences in maternal style.

Most permanent CNS damage following global cerebral ischemia is mediated by endogenous secondary processes. The trajectory of recovery is established early, suggesting that immediate recovery environment may play a critical role in post-ischemic damage. Hospitals are stressful environments, with high noise levels, continuous disruptions by staff, and unpredictable lighting. Physiological processes such as inflammation become deranged in disruptive lighting, which led us to hypothesize that cardiac arrest (CA) outcomes are negatively affected by exposure to light at night. Following a CA and cardiopulmonary resuscitation procedure or a SHAM procedure, mice were exposed to either a standard light/dark cycle (LD) or a light/dim light cycle (dLAN). Mortality was 4-fold higher among dLAN mice compared to LD mice in the week following CA. dLAN increased hippocampal cell death 1 week after CA. Hippocampal expression of pro-inflammatory cytokines was elevated 24 h after CA. Moreover, among CA mice a single night of dLAN was sufficient to upregulate hippocampal expression of IL-1β and TNF-α compared to dark nights. dLAN-CA also increased hippocampal microglia activation and microglial pro-inflammatory cytokine expression compared to LD-CA. Post-CA corticosterone concentrations did not differ suggesting differences in outcome are not due to altered corticosterone. Blocking the inflammatory response and modulating light wavelength minimized light induced damage following CA. These results indicate that dLAN exacerbates damage following CA and may have implications for lighting in clinical settings.

Alzheimer's disease (AD) is a neurodegenerative disease that accounts for most cases of dementia. Besides progressive cognitive decline, circadian dysfunction is a prominent feature of AD. Circadian disruption is traditionally regarded as a downstream symptom of AD, but recent evidence suggests that circadian dysregulation may act to exacerbate AD pathology. A reciprocal link among sleep, circadian rhythms, and amyloid deposition has long been suspected, and data from both human and animal studies support this hypothesis. The sleep-wake cycle regulates amyloid-beta (Aβ) levels in both mice and humans. Sleep deprivation increases Aβ levels in mice, and sleep apnea and insomnia may be related to AD in humans. Furthermore, melatonin, the principal hormonal output of the circadian system, is dysregulated in AD, and this may be important because melatonin is protective in cells exposed to toxic Aβ. Initial evidence supports a reciprocal link among sleep, circadian rhythmicity, and AD. More investigation is necessary to replicate these studies and determine the extent to which the circadian system contributes to the pathogenesis of AD.

Editorial summary The identification of somatic mosaicism in the brain lends a new perspective to our understanding of the role of gene and environment interactions in psychiatric disease risk. Somatic mutations, such as retrotransposon insertions, that are precipitated by modern environmental factors may alter neuronal function and neurological traits, increasing the societal prevalence of mental disorders.