Casein kinase 1 epsilon and circadian misalignment impact affective behaviours in mice.

Affective behaviours and mental health are profoundly affected by disturbances in circadian rhythms. Casein kinase 1 epsilon (CSNK1E) is a core component of the circadian clock. Mice with tau or null mutation of this gene have shortened and lengthened circadian period respectively. Here, we examined anxiety-like, fear, and despair behaviours in both male and female mice of these two different mutants. Compared with wild-type mice, we found reductions in fear and anxiety-like behaviours in both mutant lines and in both sexes, with the tau mutants exhibiting the greatest phenotypic changes. However, the behavioural despair had distinct phenotypic patterns, with markedly less behavioural despair in female null mutants, but not in tau mutants of either sex. To determine whether abnormal light entrainment of tau mutants to 24-h light-dark cycles contributes to these phenotypic differences, we also examined these behaviours in tau mutants on a 20-h light-dark cycle close to their endogenous circadian period. The normalized entrainment restored more wild-type-like behaviours for fear and anxiety, but it induced behavioural despair in tau mutant females. These data show that both mutations of Csnk1e broadly affect fear and anxiety-like behaviours, while the effects on behavioural despair vary with genetics, photoperiod, and sex, suggesting that the mechanisms by which Csnk1e affects fear and anxiety-like behaviours may be similar, but distinct from those affecting behavioural despair. Our study also provides experimental evidence in support of the hypothesis of beneficial outcomes from properly entrained circadian rhythms in terms of the anxiety-like and fear behaviours.

Systems Genetic Analyses Highlight a TGFß-FOXO3 Dependent Striatal Astrocyte Network Conserved across Species and Associated with Stress, Sleep, and Huntington's Disease.

Recent systems-based analyses have demonstrated that sleep and stress traits emerge from shared genetic and transcriptional networks, and clinical work has elucidated the emergence of sleep dysfunction and stress susceptibility as early symptoms of Huntington's disease. Understanding the biological bases of these early non-motor symptoms may reveal therapeutic targets that prevent disease onset or slow disease progression, but the molecular mechanisms underlying this complex clinical presentation remain largely unknown. In the present work, we specifically examine the relationship between these psychiatric traits and Huntington's disease (HD) by identifying striatal transcriptional networks shared by HD, stress, and sleep phenotypes. First, we utilize a systems-based approach to examine a large publicly available human transcriptomic dataset for HD (GSE3790 from GEO) in a novel way. We use weighted gene coexpression network analysis and differential connectivity analyses to identify transcriptional networks dysregulated in HD, and we use an unbiased ranking scheme that leverages both gene- and network-level information to identify a novel astrocyte-specific network as most relevant to HD caudate. We validate this result in an independent HD cohort. Next, we computationally predict FOXO3 as a regulator of this network, and use multiple publicly available in vitro and in vivo experimental datasets to validate that this astrocyte HD network is downstream of a signaling pathway important in adult neurogenesis (TGFß-FOXO3). We also map this HD-relevant caudate subnetwork to striatal transcriptional networks in a large (n = 100) chronically stressed (B6xA/J)F2 mouse population that has been extensively phenotyped (328 stress- and sleep-related measurements), and we show that this striatal astrocyte network is correlated to sleep and stress traits, many of which are known to be altered in HD cohorts. We identify causal regulators of this network through Bayesian network analysis, and we highlight their relevance to motor, mood, and sleep traits through multiple in silico approaches, including an examination of their protein binding partners. Finally, we show that these causal regulators may be therapeutically viable for HD because their downstream network was partially modulated by deep brain stimulation of the subthalamic nucleus, a medical intervention thought to confer some therapeutic benefit to HD patients. In conclusion, we show that an astrocyte transcriptional network is primarily associated to HD in the caudate and provide evidence for its relationship to molecular mechanisms of neural stem cell homeostasis. Furthermore, we present a unified systems-based framework for identifying gene networks that are associated with complex non-motor traits that manifest in the earliest phases of HD. By analyzing and integrating multiple independent datasets, we identify a point of molecular convergence between sleep, stress, and HD that reflects their phenotypic comorbidity and reveals a molecular pathway involved in HD progression.

The Circadian Clock Mutation Promotes Intestinal Dysbiosis.

BACKGROUND: Circadian rhythm disruption is a prevalent feature of modern day society that is associated with an increase in pro-inflammatory diseases, and there is a clear need for a better understanding of the mechanism(s) underlying this phenomenon. We have previously demonstrated that both environmental and genetic circadian rhythm disruption causes intestinal hyperpermeability and exacerbates alcohol-induced intestinal hyperpermeability and liver pathology. The intestinal microbiota can influence intestinal barrier integrity and impact immune system function; thus, in this study, we sought to determine whether genetic alteration of the core circadian clock gene, Clock, altered the intestinal microbiota community. METHODS: Male Clock(Δ19) -mutant mice (mice homozygous for a dominant-negative-mutant allele) or littermate wild-type mice were fed 1 of 3 experimental diets: (i) a standard chow diet, (ii) an alcohol-containing diet, or (iii) an alcohol-control diet in which the alcohol calories were replaced with dextrose. Stool microbiota was assessed with 16S ribosomal RNA gene amplicon sequencing. RESULTS: The fecal microbial community of Clock-mutant mice had lower taxonomic diversity, relative to wild-type mice, and the Clock(Δ19) mutation was associated with intestinal dysbiosis when mice were fed either the alcohol-containing or the control diet. We found that alcohol consumption significantly altered the intestinal microbiota in both wild-type and Clock-mutant mice. CONCLUSIONS: Our data support a model by which circadian rhythm disruption by the Clock(Δ19) mutation perturbs normal intestinal microbial communities, and this trend was exacerbated in the context of a secondary dietary intestinal stressor.

Disruption of the clock components CLOCK and BMAL1 leads to hypoinsulinaemia and diabetes.

The molecular clock maintains energy constancy by producing circadian oscillations of rate-limiting enzymes involved in tissue metabolism across the day and night. During periods of feeding, pancreatic islets secrete insulin to maintain glucose homeostasis, and although rhythmic control of insulin release is recognized to be dysregulated in humans with diabetes, it is not known how the circadian clock may affect this process. Here we show that pancreatic islets possess self-sustained circadian gene and protein oscillations of the transcription factors CLOCK and BMAL1. The phase of oscillation of islet genes involved in growth, glucose metabolism and insulin signalling is delayed in circadian mutant mice, and both Clock and Bmal1 (also called Arntl) mutants show impaired glucose tolerance, reduced insulin secretion and defects in size and proliferation of pancreatic islets that worsen with age. Clock disruption leads to transcriptome-wide alterations in the expression of islet genes involved in growth, survival and synaptic vesicle assembly. Notably, conditional ablation of the pancreatic clock causes diabetes mellitus due to defective beta-cell function at the very latest stage of stimulus-secretion coupling. These results demonstrate a role for the beta-cell clock in coordinating insulin secretion with the sleep-wake cycle, and reveal that ablation of the pancreatic clock can trigger the onset of diabetes mellitus.

Chronic Alcohol Exposure and the Circadian Clock Mutation Exert Tissue-Specific Effects on Gene Expression in Mouse Hippocampus, Liver, and Proximal Colon.

BACKGROUND: Chronic alcohol exposure exerts numerous adverse effects, although the specific mechanisms underlying these negative effects on different tissues are not completely understood. Alcohol also affects core properties of the circadian clock system, and it has been shown that disruption of circadian rhythms confers vulnerability to alcohol-induced pathology of the gastrointestinal barrier and liver. Despite these findings, little is known of the molecular interactions between alcohol and the circadian clock system, especially regarding implications for tissue-specific susceptibility to alcohol pathologies. The aim of this study was to identify changes in expression of genes relevant to alcohol pathologies and circadian clock function in different tissues in response to chronic alcohol intake. METHODS: Wild-type and circadian Clock(Δ19) mutant mice were subjected to a 10-week chronic alcohol protocol, after which hippocampal, liver, and proximal colon tissues were harvested for gene expression analysis using a custom-designed multiplex magnetic bead hybridization assay that provided quantitative assessment of 80 mRNA targets of interest, including 5 housekeeping genes and a predetermined set of 75 genes relevant for alcohol pathology and circadian clock function. RESULTS: Significant alterations in expression levels attributable to genotype, alcohol, and/or a genotype by alcohol interaction were observed in all 3 tissues, with distinct patterns of expression changes observed in each. Of particular interest was the finding that a high proportion of genes involved in inflammation and metabolism on the array was significantly affected by alcohol and the Clock(Δ19) mutation in the hippocampus, suggesting a suite of molecular changes that may contribute to pathological change. CONCLUSIONS: These results reveal the tissue-specific nature of gene expression responses to chronic alcohol exposure and the Clock(Δ19) mutation and identify specific expression profiles that may contribute to tissue-specific vulnerability to alcohol-induced injury in the brain, colon, and liver.

Generation of N-ethyl-N-nitrosourea (ENU) diabetes models in mice demonstrates genotype-specific action of glucokinase activators.

We performed genome-wide mutagenesis in C57BL/6J mice using N-ethyl-N-nitrosourea to identify mutations causing high blood glucose early in life and to produce new animal models of diabetes. Of a total of 13 new lines confirmed by heritability testing, we identified two semi-dominant pedigrees with novel missense mutations (Gck(K140E) and Gck(P417R)) in the gene encoding glucokinase (Gck), the mammalian glucose sensor that is mutated in human maturity onset diabetes of the young type 2 and the target of emerging anti-hyperglycemic agents that function as glucokinase activators (GKAs). Diabetes phenotype corresponded with genotype (mild-to-severe: Gck(+/+) < Gck(P417R/+), Gck(K140E)(/+) < Gck(P417R/P417R), Gck(P417R/K140E), and Gck(K140E/K140E)) and with the level of expression of GCK in liver. Each mutant was produced as the recombinant enzyme in Escherichia coli, and analysis of k(cat) and tryptophan fluorescence (I(320/360)) during thermal shift unfolding revealed a correlation between thermostability and the severity of hyperglycemia in the whole animal. Disruption of the glucokinase regulatory protein-binding site (GCK(K140E)), but not the ATP binding cassette (GCK(P417R)), prevented inhibition of enzyme activity by glucokinase regulatory protein and corresponded with reduced responsiveness to the GKA drug. Surprisingly, extracts from liver of diabetic GCK mutants inhibited activity of the recombinant enzyme, a property that was also observed in liver extracts from mice with streptozotocin-induced diabetes. These results indicate a relationship between genotype, phenotype, and GKA efficacy. The integration of forward genetic screening and biochemical profiling opens a pathway for preclinical development of mechanism-based diabetes therapies.

A Prebiotic Diet Alters the Fecal Microbiome and Improves Sleep in Response to Sleep Disruption in Rats.

Sleep disruption is a challenging and exceedingly common physiological state that contributes to a wide range of biochemical and molecular perturbations and has been linked to numerous adverse health outcomes. Modern society exerts significant pressure on the sleep/wake cycle via myriad factors, including exposure to electric light, psychological stressors, technological interconnection, jet travel, shift work, and widespread use of sleep-affecting compounds. Interestingly, recent research has identified a link between the microbiome and the regulation of sleep, suggesting that interventions targeting the microbiome may offer unique therapeutic approaches to challenges posed by sleep disruption. In this study, we test the hypothesis that administration of a prebiotic diet containing galactooligosaccharides (GOS) and polydextrose (PDX) in adult male rats improves sleep in response to repeated sleep disruption and during recovery sleep. We found that animals fed the GOS/PDX prebiotic diet for 4 weeks exhibit increased non-rapid eye movement (NREM) and rapid eye movement (REM) sleep during 5 days of sleep disruption and increased total sleep time during 24 h of recovery from sleep disruption compared to animals fed a control diet, despite similar baseline sleep characteristics. Further, the GOS/PDX prebiotic diet led to significant changes in the fecal microbiome. Consistent with previous reports, the prebiotic diet increased the relative abundance of the species Parabacteroides distasonis, which positively correlated with sleep parameters during recovery sleep. Taken together, these findings suggest that the GOS/PDX prebiotic diet may offer an approach to improve resilience to the physiologic challenge of sleep disruption, in part through impacts on the microbiome.

Pharmacological validation of candidate causal sleep genes identified in an N2 cross.

Despite the substantial impact of sleep disturbances on human health and the many years of study dedicated to understanding sleep pathologies, the underlying genetic mechanisms that govern sleep and wake largely remain unknown. Recently, the authors completed large-scale genetic and gene expression analyses in a segregating inbred mouse cross and identified candidate causal genes that regulate the mammalian sleep-wake cycle, across multiple traits including total sleep time, amounts of rapid eye movement (REM), non-REM, sleep bout duration, and sleep fragmentation. Here the authors describe a novel approach toward validating candidate causal genes, while also identifying potential targets for sleep-related indications. Select small-molecule antagonists and agonists were used to interrogate candidate causal gene function in rodent sleep polysomnography assays to determine impact on overall sleep architecture and to evaluate alignment with associated sleep-wake traits. Significant effects on sleep architecture were observed in validation studies using compounds targeting the muscarinic acetylcholine receptor M3 subunit (Chrm3) (wake promotion), nicotinic acetylcholine receptor alpha4 subunit (Chrna4) (wake promotion), dopamine receptor D5 subunit (Drd5) (sleep induction), serotonin 1D receptor (Htr1d) (altered REM fragmentation), glucagon-like peptide-1 receptor (Glp1r) (light sleep promotion and reduction of deep sleep), and calcium channel, voltage-dependent, T type, alpha 1I subunit (Cacna1i) (increased bout duration of slow wave sleep). Taken together, these results show the complexity of genetic components that regulate sleep-wake traits and highlight the importance of evaluating this complex behavior at a systems level. Pharmacological validation of genetically identified putative targets provides a rapid alternative to generating knock out or transgenic animal models, and may ultimately lead towards new therapeutic opportunities.

NREM delta power and AD-relevant tauopathy are associated with shared cortical gene networks.

Reduced NREM sleep in humans is associated with AD neuropathology. Recent work has demonstrated a reduction in NREM sleep in preclinical AD, pointing to its potential utility as an early marker of dementia. We test the hypothesis that reduced NREM delta power and increased tauopathy are associated with shared underlying cortical molecular networks in preclinical AD. We integrate multi-omics data from two extensive public resources, a human Alzheimer's disease cohort from the Mount Sinai Brain Bank (N = 125) reflecting AD progression and a (C57BL/6J × 129S1/SvImJ) F2 mouse population in which NREM delta power was measured (N = 98). Two cortical gene networks, including a CLOCK-dependent circadian network, are associated with NREM delta power and AD tauopathy progression. These networks were validated in independent mouse and human cohorts. Identifying gene networks related to preclinical AD elucidate possible mechanisms associated with the early disease phase and potential targets to alter the disease course.

Immunization with a heat-killed bacterium, Mycobacterium vaccae NCTC 11659, prevents the development of cortical hyperarousal and a PTSD-like sleep phenotype after sleep disruption and acute stress in mice.

STUDY OBJECTIVES: Sleep deprivation induces systemic inflammation that may contribute to stress vulnerability and other pathologies. We tested the hypothesis that immunization with heat-killed Mycobacterium vaccae NCTC 11659 (MV), an environmental bacterium with immunoregulatory and anti-inflammatory properties, prevents the negative impacts of 5 days of sleep disruption on stress-induced changes in sleep, behavior, and physiology in mice. METHODS: In a 2 × 2 × 2 experimental design, male C57BL/6N mice were given injections of either MV or vehicle on days -17, -10, and -3. On days 1-5, mice were exposed to intermittent sleep disruption, whereby sleep was disrupted for 20 h per day. Immediately following sleep disruption, mice were exposed to 1-h social defeat stress or novel cage (control) conditions. Object location memory (OLM) testing was conducted 24 h after social defeat, and tissues were collected 6 days later to measure inflammatory markers. Sleep was recorded using electroencephalography (EEG) and electromyography (EMG) throughout the experiment. RESULTS: In vehicle-treated mice, only the combination of sleep disruption followed by social defeat (double hit): (1) increased brief arousals and NREM beta (15-30 Hz) EEG power in sleep immediately post-social defeat compared to baseline; (2) induced an increase in the proportion of rapid-eye-movement (REM) sleep and number of state shifts for at least 5 days post-social defeat; and (3) induced hyperlocomotion and lack of habituation in the OLM task. Immunization with MV prevented most of these sleep and behavioral changes. CONCLUSIONS: Immunization with MV ameliorates a stress-induced sleep and behavioral phenotype that shares features with human posttraumatic stress disorder.

Ehd4 is required to attain normal prepubertal testis size but dispensable for fertility in male mice.

The four highly homologous members of the C-terminal EH domain-containing (EHD) protein family (EHD1-4) regulate endocytic recycling. To delineate the role of EHD4 in normal physiology and development, mice with a conditional knockout of the Ehd4 gene were generated. PCR of genomic DNA and Western blotting of organ lysates from Ehd4(-/-) mice confirmed EHD4 deletion. Ehd4(-/-) mice were viable and born at expected Mendelian ratios; however, males showed a 50% reduction in testis weight, obvious from postnatal day 31. An early (Day 10) increase in germ cell proliferation and apoptosis and a later increase in apoptosis (Day 31) were seen in the Ehd4(-/-) testis. Other defects included a progressive reduction in seminiferous tubule diameter, dysregulation of seminiferous epithelium, and head abnormalities in elongated spermatids. As a consequence, lower sperm counts and reduced fertility were observed in Ehd4(-/-) males. Interestingly, EHD protein expression was seen to be temporally regulated in the testis and EHD4 levels peaked between days 10 and 15. In the adult testis, EHD4 was highly expressed in primary spermatocytes and EHD4 deletion altered the levels of other EHD proteins in an age-dependent manner. We conclude that high levels of EHD1 in the adult Ehd4(-/-) testis functionally compensate for lack of EHD4 and prevents the development of severe fertility defects. Our results suggest a role for EHD4 in the proper development of postmitotic and postmeiotic germ cells and implicate EHD protein-mediated endocytic recycling as an important process in germ cell development and testis function.

Repeated sleep disruption in mice leads to persistent shifts in the fecal microbiome and metabolome.

It has been established in recent years that the gut microbiome plays a role in health and disease, potentially via alterations in metabolites that influence host physiology. Although sleep disruption and gut dysbiosis have been associated with many of the same diseases, studies investigating the gut microbiome in the context of sleep disruption have yielded inconsistent results, and have not assessed the fecal metabolome. We exposed mice to five days of sleep disruption followed by four days of ad libitum recovery sleep, and assessed the fecal microbiome and fecal metabolome at multiple timepoints using 16S rRNA gene amplicons and untargeted LC-MS/MS mass spectrometry. We found global shifts in both the microbiome and metabolome in the sleep-disrupted group on the second day of recovery sleep, when most sleep parameters had recovered to baseline levels. We observed an increase in the Firmicutes:Bacteroidetes ratio, along with decreases in the genus Lactobacillus, phylum Actinobacteria, and genus Bifidobacterium in sleep-disrupted mice compared to control mice. The latter two taxa remained low at the fourth day post-sleep disruption. We also identified multiple classes of fecal metabolites that were differentially abundant in sleep-disrupted mice, some of which are physiologically relevant and commonly influenced by the microbiome. This included bile acids, and inference of microbial functional gene content suggested reduced levels of the microbial bile salt hydrolase gene in sleep-disrupted mice. Overall, this study adds to the evidence base linking disrupted sleep to the gut microbiome and expands it to the fecal metabolome, identifying sleep disruption-sensitive bacterial taxa and classes of metabolites that may serve as therapeutic targets to improve health after poor sleep.

Effects of Immunization With the Soil-Derived Bacterium Mycobacterium vaccae on Stress Coping Behaviors and Cognitive Performance in a "Two Hit" Stressor Model.

Previous studies demonstrate that Mycobacterium vaccae NCTC 11659 (M. vaccae), a soil-derived bacterium with anti-inflammatory and immunoregulatory properties, is a potentially useful countermeasure against negative outcomes to stressors. Here we used male C57BL/6NCrl mice to determine if repeated immunization with M. vaccae is an effective countermeasure in a "two hit" stress exposure model of chronic disruption of rhythms (CDR) followed by acute social defeat (SD). On day -28, mice received implants of biotelemetric recording devices to monitor 24-h rhythms of locomotor activity. Mice were subsequently treated with a heat-killed preparation of M. vaccae (0.1 mg, administered subcutaneously on days -21, -14, -7, and 27) or borate-buffered saline vehicle. Mice were then exposed to 8 consecutive weeks of either stable normal 12:12 h light:dark (LD) conditions or CDR, consisting of 12-h reversals of the LD cycle every 7 days (days 0-56). Finally, mice were exposed to either a 10-min SD or a home cage control condition on day 54. All mice were exposed to object location memory testing 24 h following SD. The gut microbiome and metabolome were assessed in fecal samples collected on days -1, 48, and 62 using 16S rRNA gene sequence and LC-MS/MS spectral data, respectively; the plasma metabolome was additionally measured on day 64. Among mice exposed to normal LD conditions, immunization with M. vaccae induced a shift toward a more proactive behavioral coping response to SD as measured by increases in scouting and avoiding an approaching male CD-1 aggressor, and decreases in submissive upright defensive postures. In the object location memory test, exposure to SD increased cognitive function in CDR mice previously immunized with M. vaccae. Immunization with M. vaccae stabilized the gut microbiome, attenuating CDR-induced reductions in alpha diversity and decreasing within-group measures of beta diversity. Immunization with M. vaccae also increased the relative abundance of 1-heptadecanoyl-sn-glycero-3-phosphocholine, a lysophospholipid, in plasma. Together, these data support the hypothesis that immunization with M. vaccae stabilizes the gut microbiome, induces a shift toward a more proactive response to stress exposure, and promotes stress resilience.

Genetic analysis of daily physical activity using a mouse chromosome substitution strain.

There is considerable evidence for a genetic basis underlying individual differences in spontaneous physical activity in humans and animals. Previous publications indicate that the physical activity level and pattern vary among inbred strains of mice and identified a genomic region on chromosome 13 as quantitative trait loci (QTL) for physical activity. To confirm and further characterize the role of chromosome 13 in regulating daily physical activity level and pattern, we conducted a comprehensive phenotypic study in the chromosome 13 substitution strain (CSS-13) in which the individual chromosome 13 from the A/J strain was substituted into an otherwise complete C57BL/6J (B6) genome. The B6 and A/J parental strains exhibited pronounced differences in daily physical activity, sleep-wake structure, circadian period and body weight. Here we report that a single A/J chromosome 13 in the context of a B6 genetic background conferred a profound reduction in both total cage activity and wheel-running activity under a 14:10-h light-dark cycle, as well as in constant darkness, compared with B6 controls. Additionally, CSS-13 mice differed from B6 controls in the diurnal distribution of activity and the day-to-day variability in activity onset. We further performed a linkage analysis and mapped a significant QTL on chromosome 13 regulating the daily wheel running activity level in mice. Taken together, our findings indicate a QTL on chromosome 13 with dramatic and specific effects on daily voluntary physical activity, but not on circadian period, sleep, or other aspects of activity that are different between B6 and A/J strains.

NMDAR activation regulates the daily rhythms of sleep and mood.

STUDY OBJECTIVES: The present studies examine the effects of NMDAR activation by NYX-2925 diurnal rhythmicity of both sleep and wake as well as emotion. METHODS: Twenty-four-hour sleep EEG recordings were obtained in sleep-deprived and non-sleep-deprived rats. In addition, the day-night cycle of both activity and mood was measured using home cage ultrasonic-vocalization recordings. RESULTS: NYX-2925 significantly facilitated non-REM (NREM) sleep during the lights-on (sleep) period, and this effect persisted for 3 days following a single dose in sleep-deprived rats. Sleep-bout duration and REM latencies were increased without affecting total REM sleep, suggesting better sleep quality. In addition, delta power during wake was decreased, suggesting less drowsiness. NYX-2925 also rescued learning and memory deficits induced by sleep deprivation, measured using an NMDAR-dependent learning task. Additionally, NYX-2925 increased positive affect and decreased negative affect, primarily by facilitating the transitions from sleep to rough-and-tumble play and back to sleep. In contrast to NYX-2925, the NMDAR antagonist ketamine acutely (1-4 hours post-dosing) suppressed REM and non-REM sleep, increased delta power during wake, and blunted the amplitude of the sleep-wake activity rhythm. DISCUSSION: These data suggest that NYX-2925 could enhance behavioral plasticity via improved sleep quality as well as vigilance during wake. As such, the facilitation of sleep by NYX-2925 has the potential to both reduce symptom burden on neurological and psychiatric disorders as well as serve as a biomarker for drug effects through restoration of sleep architecture.

Trait-like vulnerability of higher-order cognition and ability to maintain wakefulness during combined sleep restriction and circadian misalignment.

STUDY OBJECTIVES: Determine stability of individual differences in executive function, cognitive processing speed, selective visual attention, and maintenance of wakefulness during simulated sustained operations with combined sleep restriction and circadian misalignment. METHODS: Twenty healthy adults (eight female), aged 25.7 (±4.2 SD), body mass index (BMI) 22.3 (±2.1) kg/m2 completed an 18-day protocol twice. Participants maintained habitual self-selected 8-hour sleep schedules for 2 weeks at home prior to a 4-day laboratory visit that included one sleep opportunity per day: 8 hours on night 1, 3 hours on night 2, and 3 hours on mornings 3 and 4. After 3 days of unscheduled sleep at home, participants repeated the entire protocol. Stability and task dependency of individual differences in performance were quantified by intra-class correlation coefficients (ICC) and Kendall's Tau, respectively. RESULTS: Performance on Stroop, Visual Search, and the Maintenance of Wakefulness Test were highly consistent within individuals during combined sleep restriction and circadian misalignment. Individual differences were trait-like as indicated by ICCs (0.54-0.96) classified according to standard criteria as moderate to almost perfect. Individual differences on other performance tasks commonly reported in sleep studies showed fair to almost perfect ICCs (0.22-0.94). Kendall's rank correlations showed that individual vulnerability to sleep restriction and circadian misalignment varied by task and by metric within a task. CONCLUSIONS: Consistent vulnerability of higher-order cognition and maintenance of wakefulness to combined sleep restriction and circadian misalignment has implications for the development of precision countermeasure strategies for workers performing safety-critical tasks, e.g. military, police, health care workers and emergency responders.

Cross-species systems analysis identifies gene networks differentially altered by sleep loss and depression.

To understand the transcriptomic organization underlying sleep and affective function, we studied a population of (C57BL/6J × 129S1/SvImJ) F2 mice by measuring 283 affective and sleep phenotypes and profiling gene expression across four brain regions. We identified converging molecular bases for sleep and affective phenotypes at both the single-gene and gene-network levels. Using publicly available transcriptomic datasets collected from sleep-deprived mice and patients with major depressive disorder (MDD), we identified three cortical gene networks altered by the sleep/wake state and depression. The network-level actions of sleep loss and depression were opposite to each other, providing a mechanistic basis for the sleep disruptions commonly observed in depression, as well as the reported acute antidepressant effects of sleep deprivation. We highlight one particular network composed of circadian rhythm regulators and neuronal activity-dependent immediate-early genes. The key upstream driver of this network, Arc, may act as a nexus linking sleep and depression. Our data provide mechanistic insights into the role of sleep in affective function and MDD.

Dopamine pathway imbalance in mice lacking Magel2, a Prader-Willi syndrome candidate gene.

The etiology of abnormal eating behaviors, including binge-eating disorder, is poorly understood. The neural circuits modulating the activities of the neurotransmitters dopamine and serotonin are proposed to be dysfunctional in individuals suffering from eating disorders. Prader-Willi syndrome is a neurodevelopmental disorder that causes extreme food seeking and binge-eating behaviors together with reduced satiety. One of the genes implicated in Prader-Willi syndrome, Magel2, is highly expressed in the regions of the brain that control appetite. Our objective was to examine behaviors relevant to feeding and the neural circuits controlling feeding in a mouse model of Prader-Willi syndrome that lacks expression of the Magel2 gene. We performed behavioral tests related to dopaminergic function, measuring cocaine-induced hyperlocomotion, binge eating, and saccharin-induced anhedonia in Magel2-deficient mice. Next, we analyzed dopaminergic neurons in various brain regions and compared these findings between genotypes. Finally, we examined biochemical markers in the brain under standard diet, high-fat diet, and withdrawal from a high-fat diet conditions. We identified abnormal behaviors and biomarkers reflecting dopaminergic dysfunction in mice lacking Magel2. Our results provide a biological framework for clinical studies of dopaminergic function in children with Prader-Willi syndrome, and may also provide insight into binge-eating disorders that occur in the general population. (PsycINFO Database Record

Altered Body Weight Regulation in CK1ε Null and tau Mutant Mice on Regular Chow and High Fat Diets.

Disruption of circadian rhythms results in metabolic dysfunction. Casein kinase 1 epsilon (CK1ε) is a canonical circadian clock gene. Null and tau mutations in CK1ε show distinct effects on circadian period. To investigate the role of CK1ε in body weight regulation under both regular chow (RC) and high fat (HF) diet conditions, we examined body weight on both RC and HF diets in CK1ε (-/-) and CK1ε (tau/tau) mice on a standard 24 hr light-dark (LD) cycle. Given the abnormal entrainment of CK1ε (tau/tau) mice on a 24 hr LD cycle, a separate set of CK1ε (tau/tau) mice were tested under both diet conditions on a 20 hr LD cycle, which more closely matches their endogenous period length. On the RC diet, both CK1ε (-/-) and CK1ε (tau/tau) mutants on a 24 hr LD cycle and CK1ε (tau/tau) mice on a 20 hr LD cycle exhibited significantly lower body weights, despite similar overall food intake and activity levels. On the HF diet, CK1ε (tau/tau) mice on a 20 hr LD cycle were protected against the development of HF diet-induced excess weight gain. These results provide additional evidence supporting a link between circadian rhythms and energy regulation at the genetic level, particularly highlighting CK1ε involved in the integration of circadian biology and metabolic physiology.

Loss of circadian photoentrainment and abnormal retinal electrophysiology in Math5 mutant mice.

PURPOSE: To determine how the absence of retinal ganglion cells (RGCs) in Math5 (Atoh7) mutant mice affects circadian behavior and retinal function. METHODS: The wheel-running behavior of wild-type and Math5 mutant mice was measured under various light-dark cycle conditions. To evaluate retinal input to the suprachiasmatic nuclei (SCN) anatomically, the retinohypothalamic tracts were labeled in vivo. To assess changes in retinal function, corneal flash electroretinograms (ERGs) from mutant and wild-type mice were compared under dark- and light-adapted conditions. Alterations in retinal neuron populations were evaluated quantitatively and with cell-type-specific markers. RESULTS: The Math5-null mice did not entrain to light and exhibited free-running circadian behavior with a mean period (23.6 +/- 0.15 hours) that was indistinguishable from that of wild-type mice (23.4 +/- 0.19 hours). The SCN showed no anterograde labeling with a horseradish peroxidase-conjugated cholera toxin B (CT-HRP) tracer. ERGs recorded from mutant mice had diminished scotopic a- and b-wave and photopic b-wave amplitudes. The scotopic b-wave was more severely affected than the a-wave. The oscillatory potentials (OPs) and scotopic threshold response (STR) were also reduced. Consistent with these ERG findings, a pan-specific reduction in the number of bipolar cells and a smaller relative decrease in the number of rods in mutant mice were observed. CONCLUSIONS: Math5-null mice are clock-blind and have no RGC projections to the SCN. RGCs are thus essential for photoentrainment in mice, but are not necessary for the development or intrinsic function of the SCN clock. RGCs are not required to generate any of the major ERG waveforms in mice, including the STR, which is produced by ganglion cells in some other species. The diminished amplitude of b-wave, OPs, and STR components in Math5 mutants is most likely caused by the decreased abundance of retinal interneurons.