bluuebunny/crossref_metadata_2025_split huggingface dataset:
0021 Bimodality prevalence in a sample of undergraduate students in Sao Paulo city
Authors: Vallim Julia , Tsuji Heloisa , Lima Gabriela , Xylaras Beatriz , Louzada Fernando , Tufik Sergio , D'Almeida Vânia 
Published: May 2023

Abstract

Introduction
The concept of bimodality comprises a fourth circadian phenotype, the bimodal, whose identification is made by a re-scoring of the Morningness-Eveningness Questionnaire and was first described by Martynhak and colleagues in 2010. Bimodal subjects are originally classified as “intermediate” because they answer the questions sometimes as a morning person and, some others, as an evening one. The present work aimed to describe the prevalence of the bimodal chronotype in a sample of undergraduate students in Sao Paulo city.

Methods
A web-based cross-sectional study conducted between September 2018 and March 2021, with a convenience sampling method, composed by 615 undergraduate students (82% females, mean age: 23.4±6.5 years) from two higher education institutions at Sao Paulo city.

Results
Forty-four students (7%) had positive bimodality indexes and bimodal individuals comprised 19 students (74% female; 26% male), 3% of the total sample.

Conclusion
We met our initial goal and found a bimodality prevalence of 3%. The description of bimodality prevalence in a sample of college students is interesting because they may be a maladaptive population from a sleep and rhythmicity point of view, which may be interesting for the design of academic public policies more consistent with the circadian preferences of students.

Support (if any)
Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES), Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP), Associação Fundo de Incentivo à Pesquisa (AFIP).
0023 Circadian fragmentation and stability distinguish employment status
Authors: Glickenstein David , Karamchandani Avinash , Korbut Camille , Bang Michelle , Aucoin Alexa , Callovini Leah , Lin Kevin , Haynes Patricia 
Published: May 2023

Abstract

Introduction
This analysis used data from the Assessing Daily Activity Patterns Through Occupational Transitions (ADAPT) study to compare differences in employment status on activity variability, as an indicator of circadian fragmentation and stability. Circadian fragmentation refers to frequency of alterations between rest and activity relative to the daily rhythm and stability refers to day-to-day similarity. High fragmentation and low stability have been linked to a number of negative health outcomes including depression, obesity, and heightened mortality risk.

Methods
The sample consisted of 155 participants that included 702 total cases (n = 434 employed, n = 268 unemployed) assessed over 18 months. Participants were required to have involuntarily lost their jobs in the last 90 days. Employment status was determined for all participants at each visit based on demographic surveys. Daily activity patterns were assessed via actigraphy (Actiwatch-Spectrum) at 30 second epochs for 14 days. The nonparametric measures of intradaily variability (IV) and interdaily stability (IS) were used as measures of circadian fragmentation and stability. Several subsampling intervals were considered for IV, ranging from 5 minutes to 4 hours.

Results
Unemployment was associated with higher IV (for 1 hour subsampling intervals, t=5.23, p &lt; .001) and lower IS than employment (t=2.38, p &lt;.05). The same is true with changes in the subsampling interval for IV, with highest significance at 45 minutes to 1.5 hours. Significant findings remained even for 5 minute (t=2.15, p &lt; .05) and 4 hour (t = 4.07, p&lt;.001) intervals.

Conclusion
In a sample of adults with variable employment status, employment was associated with less circadian fragmentation and more stability than unemployment. These findings suggest that circadian fragmentation and instability may be two mechanisms by which job loss increases negative health risk. Future planned studies examining prospective changes in circadian fragmentation/stability during employment transitions will help answer this question.

Support (if any)
NIH #1R01HL117995-01A1, NSF DMS 1937229, NSF CCF 1740858
0024 Decreased circadian period associated with enhanced autophagy in cereblon knockout mice
Authors: Kim Jinhong , Park Chul-Seung , Kim Tae 
Published: May 2023

Abstract

Introduction
The circadian rhythm plays a critical role in many biological processes. Recently, autophagy was suggested as a regulating factor of circadian rhythm by reducing protein trafficking. Cereblon (CRBN) is a component of the E3 ubiquitin ligase complex for ubiquitination and degradation of the target proteins. Interestingly, cereblon ubiquitinates beclin 1, a potent autophagy inducer. Accordingly, the changes in cereblon are likely to affect the circadian rhythm. However, how cereblon can regulate circadian rhythm is not understood yet. Therefore, we hypothesized that cereblon might be an important regulator of the circadian rhythm and sought to investigate the relationship.

Methods
We obtained 3-month-old CRBN gene knockout (KO) and wild-type (WT) mice (n=4 and 8, respectively). The mice were housed in the wheel-based activity recording system under 12:12 light-dark (LD) condition for 1 week and constant darkness (DD) condition for the subsequent 2 weeks. Activity data were analyzed by ClockLab Analysis 6 (Actimetrics, USA). Autophagy-related proteins, including beclin1, microtubule-associated protein light chain-3 (LC3), and p63, were measured by the ELISA and western blot.

Results
We found that CRBN KO mice showed decreased circadian period during constant darkness (23.7 vs 23.4 hours, p&lt; 0.05). The latency from the light-off time to initiating activities in the dark period was shorter in CRBN KO mice and WT mice. Protein analysis assay revealed that beclin1, LC3, and p62 increased in CRBN KO mice, indicating increased autophagy.

Conclusion
Our results demonstrated that CRBN knockout mice showed shorter circadian periods and reduced autophagy-related molecules. We interpret that the lack of cereblon resulted in reduced degradation of beclin1 that triggers autophagy. As per Beesley et al (2020), autophagy-induced decrease in cytoplasmic congestion facilitates the protein trafficking for the crucial components of the molecular circadian clock. The shorter circadian period in CRBN KO mice was primarily due to the enhanced autophagy, although its implication in general health is unclear. Taken together, we suggest that CRBN is associated with circadian rhythm via controlling autophagy. Further investigations to understand how circadian amplitude was increased and the activity latency during the dark period was shortened are needed.

Support (if any)
0028 Systematic Review: Time of day differences in complete blood count values
Authors: Busza Anna , Sharma Vani , Ferguson Kendall , Fawcett Andrea , Knoll Justin , Zee Phyllis , Fishbein Anna 
Published: May 2023

Abstract

Introduction
The complete blood count (CBC) is the most commonly ordered blood test with a large range of reference values that do not consider time of day for interpretation. Clinicians are largely unfamiliar with the influence of time of day on CBC values. Our objective was to systematically review this topic to report on peak and trough timing of CBC values.

Methods
A systematic search of PubMed, Ovid, Scopus, CINAHL, Cochrane, Embase, and Web of Science was performed for studies evaluating any part of the CBC over 24 hours. The studies were screened for eligibility based on predetermined criteria including: measurement of a part of the CBC with at least 3 time points in 24 hours. In total, 164 full-text articles were screened and 32 were included in the final analysis. The aggregated data was analyzed with the Cosinor package of R statistical environment and Stata SE to create cosinor graphs and Forest plots for each component of the CBC, respectively.

Results
Of the 32 articles, a metalysis was performed on the following CBC subsets: leukocytes, erythrocytes, hemoglobin, hematocrit, platelets, neutrophils, lymphocytes, monocytes, eosinophils, and basophils. Lymphocytes exhibited a statistically significant diurnal rhythm (p=0.014, n=10 articles) with a peak of 2280.10 cells/uL at 23:17 (CI: 1731.80, 2828.40) and trough of 1661.49 cells/uL at 08:53 (CI: 1242.64, 2080.33). Erythrocytes, hemoglobin, and hematocrit peaked in the morning; while platelets, neutrophils, monocytes, and basophils peaked in the late afternoon and early evening; and lymphocytes and eosinophils peaked late at night. High heterogeneity in the results across studies was noted, likely attributable to the difference in circadian factors controlled. Limitations include the small sample size for each component of the CBC, the limited samples included in the meta analysis, and the significant heterogeneity affecting the results.

Conclusion
There are significant time of day changes in CBC values that might be important to consider as we move towards precision medicine. Given the limited number of studies with small sample sizes, future work should evaluate large data sources to inform time-dependent interpretation of the CBC.

Support (if any)
0032 ADA Functional Polymorphism Modulates REM Sleep Following Total Sleep Deprivation
Authors: Fluke Sofia , Hansen Devon , Satterfield Brieann , Van Dongen Hans 
Published: May 2023

Abstract

Introduction
Adenosine Deaminase (ADA) regulates extracellular levels of adenosine, a brain correlate of sleep homeostasis. A single nucleotide polymorphism (SNP) of the ADA gene (rs73598374) is known to be associated with increased homeostatic pressure for NREM sleep after total sleep deprivation (TSD). Whether this genotype effect also impacts REM sleep after TSD has not been systematically investigated. The limit cycle reciprocal interaction model posits that NREM/REM cycles are regulated by cholinergic and GABAergic neurotransmitter systems. While NREM/REM regulation is also thought to be mediated by sleep homeostatic state, evidence of an association with adenosine levels has not been reported. Here we investigate the influence of ADA on REM sleep at baseline and following TSD.

Methods
N=48 healthy, normal sleepers (ages 27.7 + 5.3, 23 females) participated in one of three in-laboratory TSD studies. Following 10h baseline sleep (22:00–08:00), participants underwent 38h TSD, with subsequent 10h recovery sleep (22:00–08:00). Sleep periods were recorded polysomnographically and scored visually according to AASM criteria. Genomic DNA was extracted from whole blood, and the ADA SNP was assayed using real-time PCR. Sleep stages were analyzed using mixed-effect ANOVA with fixed effects of ADA genotype, night (baseline vs. recovery), and their interaction, controlling for study, with a random effect over subject on the intercept.

Results
The frequency of genotypes (G/G: 42, A/G: 6) did not vary from Hardy-Weinberg equilibrium. There was no genotype effect on total sleep time and no genotype by night interaction (p&gt;0.7). However, A allele carriers spent more time in REM during baseline and less during recovery compared to G/G homozygotes (genotype by night interaction: F[1,46]=8.13, p=0.007). Conversely, while not statistically significant (p&gt;0.1), A allele carriers spent less time in NREM during baseline and more during recovery compared to G/G homozygotes.

Conclusion
ADA genotype was associated with a shift in balance between REM and NREM at baseline, as well as changes therein after TSD, without concomitant effect on total sleep time. This suggests a role for adenosine in mediating REM sleep regulation, possibly through a sleep homeostatic effect on the reciprocal interaction between NREM and REM.

Support (if any)
CDMRP W81XWH-05-1-0099; ONR N00014-13-C-0063; NIH R21CA16769; ARO W911NF2210223.
0035 Melanopsin genetic variants and delayed sleep phenotype – whole genome sequencing analysis
Authors: Smieszek Sandra 
Published: May 2023

Abstract

Introduction
Melanopsin (OPN4) is a blue light-sensitive opsin-type G-protein coupled receptor. It is highly expressed in photosensitive retinal ganglion cells which mediate responses to light, including regulation of sleep, circadian photoentrainment, and pupillary light response. Mutations in OPN4 were shown to affect responses to light, ultimately affecting the regulation of circadian rhythms and sleep. Previously, we describe a male carrier of the OPN4 missense variant diagnosed with delayed sleep-wake phase disorder (DSWPD), with a consistent recurrent pattern of delayed sleep onset The rs143641898 [NM_033282.4:c.502C&gt;T p.(Arg168Cys)] variant in the OPN4 gene that was shown in a functional study to render the OPN4 protein non-functional.

Methods
The analysis was conducted on whole genome sequencing samples obtained from patients with a clinical diagnosis of DSWPD. Age and sex-matched samples were used as controls.

Results
Here we report an enrichment of OPN4 rare coding variants, MAF&lt; 1, in a set of 117 DSWPD samples as compared to 315 healthy controls. This significant enrichment likely implies that other rare pLOFs can similarly contribute to the delayed sleep phenotype. Moreover, we report a significant association of rs rs1079610, a common variant in OPN4 – with delayed bedtime in DSWPD patients. This variant has been previously reported in association with altered pupillary responses. This effect is not seen in a large set of healthy controls without DSWPD diagnosis.

Conclusion
The OPN4 discussed rare OPN4 variants likely increase the risk of DSWPD via its direct effect on the pathophysiology along the melanopsin axis. This study offers useful insights for the differential diagnosis and ultimately treatment of DSWPD risk in which patients carry pathogenic variants in the OPN4 gene.

Support (if any)
0038 Genioglossus muscle (GG) activity evoked by pharmacological stimulation of alpha1-adrenoceptors in pre-hypoglossal region (PHR)
Authors: Reis Leonardo Frasson , Xu Kelly , Rukhadze Irma , Fenik Victor 
Published: May 2023

Abstract

Introduction
The sleep-related withdrawal of noradrenergic excitation and increased cholinergic inhibition of hypoglossal motoneurons (HM), which control the tone of the GG, the largest protruder muscle of the tongue, play a critical role in obstructive sleep apnea pathophysiology. In an animal model of REM sleep, antagonism of alpha1 noradrenergic signaling by microinjections of prazosin into the hypoglossal nucleus (HN) strongly decreased hypoglossal nerve activity. However, a significant delay of this decrease prompted a hypothesis that prazosin must diffuse outside the HN to block alpha1-adrenoceptors that are located on interneurons, which mediate noradrenergic drive to HM. Here, we aimed to map the PHR by microinjecting alpha1 drugs for possible location of these interneurons.

Methods
Adult C57bl/6J mice (n=28) of both sexes were anesthetized with either ketamine/xylazine (120/20 mg/kg, i.p.) or 1.5-2.0% isoflurane inhaled through a nose mask. Electromyogram electrodes were implanted into GG and diaphragm to record their activity. We microinjected phenylephrine (alpha1-adrenoceptor agonist, 5-10 nl, 1 mM) in both ketamine/xylazine- and isoflurane-anesthetized mice, while prazosin (10 nl, 0.2 mM) was injected only in mice under isoflurane anesthesia. The injection sites were marked with Pontamine Skye Blue. After the completion of experiments, all animals were perfused, and brains removed for histological analysis.

Results
The activity of GG showed spontaneous phasic inspiratory modulation with minimal tonic component under both anesthesia conditions although its amplitude was markedly lager in animals anesthetized by isoflurane. Effective microinjections of phenylephrine elicited short-lasting (approximately 15 minutes) excitatory GG responses. When injected into the HN, phenylephrine strongly excited GG as compared to injections to the PHR in ketamine/xylazine-anesthetized mice. However, in isoflurane-anesthetized animals, microinjections of phenylephrine into the PHR induced greater activation of GG than injections onto the HN. Microinjections of prazosin in isoflurane-anesthetized mice produced long-lasting disfacilitation of GG activity. However, prazosin injections into the PHR elicited responses with the shortest latencies (0-120 s).

Conclusion
These results suggest that the putative interneurons that mediate noradrenergic drive to HM are located in the PHR. This finding is important for deciphering the neuronal network that controls the state-dependent activity of GG, which may provide new therapeutic targets for OSA treatment.

Support (if any)
NIHR01HL133847, NIAR01AG065233.
0039 Investigating the role of striatal dopamine in sleep and narcolepsy-cataplexy
Authors: Toth Brandon , Burgess Christian , Chang Katie 
Published: May 2023

Abstract

Introduction
Dopamine release in the striatum is known to govern reward, motor, and sleep behaviors. There also exists a link between pharmacological manipulation of dopamine receptors and cataplexy occurrence, yet the precise spatial and temporal dynamics of dopamine release in relation to cataplexy onset are not understood. In the present study, we evaluate dopamine release dynamics in orexin knockout (OX KO) mice during cataplexy and physiological behaviors.

Methods
OX KO mice were implanted with EEG/EMG electrodes, and dLight1.1 was expressed in the nucleus accumbens core (NAcc), shell (NAcSh), and dorsolateral striatum (DLS) to perform in vivo fiber photometry of dopamine release in freely-moving animals. In NAcc and NAcSh mice, ChrimsonR was retrogradely expressed in the ventral tegmental area (VTA), to allow for precise optogenetic control of midbrain dopamine projections to different regions of the striatum. We also expressed dLight1.1 in the NAcc of wildtype (WT) mice as a control. Three different stimulation parameters were used in independent recordings: phasic 20 Hz stimulation, phasic 10-40 Hz variable stimulation, and tonic 2 Hz stimulation.

Results
First, we show that while dopamine release in response to rewarding behaviors is muted in OX KO animals, there is no difference in dopamine release across sleep state transitions between OX KO and WT animals. Second, we demonstrate that there is a striking increase in dopamine release in both the NAcc and NAcSh, but not in the DLS, at the onset of cataplexy. This increase occurs independently of the behaviors that the animals were engaged in immediately prior to cataplexy, suggesting that the increase is characteristic of all cataplexy episodes. Finally, we find that while high frequency phasic stimulation did not affect cataplexy frequency, low frequency, tonic stimulation reduced both cataplexy and REM occurrence.

Conclusion
We show that there are differences in dopamine release in response to rewarding events, but not changes in sleep states, in OX KO and WT animals. We also identify dopamine release in the ventral striatum as a key node in cataplexy regulation, which helps to further our understanding of the limbic circuitry that underlies narcolepsy symptoms.

Support (if any)
University of Michigan Gilmore Award and Rackham Graduate Research Award.
0040 Leptin Signaling in the Dorsomedial Hypothalamus Couples Metabolism and Breathing in Obesity
Authors: Amorim Mateus , Wang Xin , Aung O , Bevans-Fonti Shannon , Anokye-Danso Frederick , Escobar Joan , Freire Carla , Pho Huy , Dergacheva Olga , Branco Luiz , Mendelowitz David , Polotsky Vsevolod 
Published: May 2023

Abstract

Introduction
Individuals with obesity hypoventilation syndrome (OHS) are unable to increase ventilation to match increased CO2 production (VCO2). Obstructive sleep apnea (OSA), defined by the closure of the upper airway during sleep, is present in 90% of OHS patients. Leptin, an adipocyte-produced hormone, acts in leptin receptor (LEPRb) positive neurons and was implicated in the pathophysiology of OSA and OHS. Diet-induced obese (DIO) C57BL/6J mice shows features of OHS and OSA. Leptin-mediated neural pathways may link metabolism and breathing in obesity. We hypothesized that LEPRb-positive neurons in the dorsomedial hypothalamus (DMH) are necessary and sufficient to match control of breathing to VCO2 and that the stimulation of these neurons relieves OHS in DIO mice.

Methods
We expressed designer receptors exclusively activated by designer drugs (DREADDs) in DMH of DIO Leprb-Cre mice and measured breathing during sleep, the hypercapnic ventilatory response (HCVR), VCO2, and O2 consumption (VO2).

Results
Activation of LEPRb-positive DMH neurons increased inspiratory flow and minute ventilation during REM sleep, without affecting sleep architecture. The effect was pronounced during inspiratory flow limitation, suggeting that upper airway obstruction during REM sleep was relieved. Activation of these neurons increased the HCVR and this response was blunted by a serotonin receptor antagonist, methysergide. Activation of these neurons augmented VCO2, VO2, and VE/VCO2, suggesting that breathing was stimulated out of proportion to increases in metabolism. In vitro experiments revealed that LEPRb-positive DMH neurons project to 5-HT neurons in the dorsal raphe (DR) and that optogenetic stimulation of these neurons evoked excitatory postsynaptic currents in 5-HT neurons of the DR. Administration of retrograde Cre-dependent AAV harboring caspase to the DR in Leprb-Cre mice deleted LEPRb-positive DMH neurons and abolished respiratory and metabolic effects of chemogenetic stimulation. Apoptosis of LEPRb-positive DMH neurons projecting to the DR also prevented the increase in ventilation during REM sleep induced by intranasal leptin.

Conclusion
Leptin increases the HCVR, improves upper airway patency during sleep, and increases VCO2 acting on LEPRb-positive DMH neurons projecting to the DR. We conclude that LEPRb-positive DMH neurons projecting to the DR couple metabolism and breathing in obesity.

Support (if any)
NIH (R01 HL128970, R01 HL133100, and R01 HL138932), AHA (#915167).