bluuebunny/crossref_metadata_2025 huggingface dataset:
0012 Morning Misery: Circadian Timing and Negative Affect in a Sample of Adolescents
Authors: Mirchandaney Riya , Buysse Daniel , Clark Duncan , Guo Kathryn , Siegle Greg , Hasler Brant
Published: April 2024

Introduction

Adolescents with later circadian timing evidence an increased risk for depression. Depressed individuals report higher negative affect (NA), especially in the morning, which may reflect altered circadian timing in NA. Untangling the circadian influences on mood is challenging due to inconsistent operationalizations of circadian timing. This study aims to analyze the associations between three metrics of circadian timing (circadian preference, chronotype, and circadian phase) and daily levels of NA among adolescents.

Methods

This study analyzed 8 days of ecological momentary assessment (EMA) data among 119 adolescents (54.6% female; mean age 17.3 years). Participants completed sleep diaries with a visual analog scale (VAS) measuring NA (0-100; calm-tense) each morning, and the Positive and Negative Affect Scale–Short Form (PANAS-SF) approximately every 3 waking hours. NA scores were averaged across mornings, evenings, and overall. Circadian preference was assessed by the Composite Scale of Morningness (CSM), chronotype by the Munich Chronotype Questionnaire (MCTQ), and circadian phase by salivary dim light melatonin onset (DLMO; 4pg/ml threshold). We conducted multiple regression analyses to examine circadian preference, chronotype, and circadian phase predicting morning (PANAS, VAS), evening, and overall NA (PANAS), controlling for age, sex, and socioeconomic status.

Results

Correlations between circadian metrics were low (rDLMO-MCTQ=.20; rDLMO-CSM=.39; rCSM-MCTQ=.27). Greater evening preference predicted higher morning NA (PANAS: p=.026, β=-.21; VAS: p< 0.001, β=-.33) and evening NA (p=.032, β=-.20), but not overall NA (p=.106, β=-.15). These associations were not significant after isolating the Sleep Timing and Activity factor of the CSM. Later chronotype predicted higher NA on the morning VAS (VAS: p=.048, β=.18), but not on the PANAS (p=.756, β=-.03); chronotype did not predict evening NA (p=.259, β=-.11) or overall NA (p=.412, β=-.08). Circadian phase did not predict morning NA (PANAS: p=.841, β=-.02; VAS: p=.552, β=.05), evening NA (p=.625, β=.05), or overall NA (p=.391, β=.03).

Conclusion

These results highlight the importance of specificity when operationalizing circadian timing and NA, and add to the mixed literature concerning the role of circadian phase in negative mood modulation. Adolescents with later chronotype may be particularly vulnerable to negative mood in the morning. Important next steps involve modeling diurnal patterns of NA and positive affect.

Support (if any)

R01-AA025626 (Hasler)
0014 Time to Step Up: The Role of Circadian Misalignment on Adolescent Physical Activity Metrics
Authors: Crandall Gracie , Wright Isabella , Kay Daniel , Duraccio Kara
Published: April 2024

Introduction

Circadian misalignment, the discrepancy between internal circadian timing and sleep timing, impacts overall health and well-being. While past research emphasizes the connection between physical activity and sleep duration, we aim to examine differences in physical activity levels and abilities in adolescents experiencing experimentally manipulated circadian-aligned versus misaligned sleep conditions.

Methods

29 night-owl adolescents underwent both aligned (12:30am-9:30am) and misaligned sleep (9:30pm-6:30am) for five nights each, with order randomized. To determine adherence to study protocols, wrist accelerometers tracked sleep duration and timing and waist accelerometers measured physical activity. After each experimental period, participants completed the 6-minute walk test and a sit-to-stand test. Repeated measures t-tests were used to assess the impact of the circadian timing condition on minutes in sedentary behavior, minutes in moderate-to-vigorous physical activity (MVPA), 6-minute walk distance, and sit-to-stand repetitions.

Results

Participants in the aligned condition significantly outperformed the misaligned group in sit-to-stand repetitions (M=24.10, SD=8.978; M=19.72, SD=6.524, respectively; p=0.003, d=0.616). Additionally, 6-min walk results revealed a significant difference, with participants in the aligned condition walking significantly farther than the control group (M=0.2952, SD=0.13095; M=0.2331, SD=0.10674, respectively; p=0.011, d=0.503). There was no significant difference between aligned and misaligned conditions in MVPA or sedentary behavior.

Conclusion

Participants undergoing circadian alignment had significantly better performance in walking and sitting tasks compared to when they were misaligned; there was no impact of circadian timing on physical activity levels across the week. These results could have significant implications for how teen athletes train and compete. Specifically, aligning teen athlete’s school and sport training and competition schedules with their circadian rhythm might lead to better performance.

Support (if any)
0015 Feasibility of an At-Home Experimental Circadian Misalignment Induction for Adolescents
Authors: Beebe Dean , Fidler Andrea , Crowley Stephanie
Published: April 2024

Introduction

For many adolescents, the only practical way to lengthen sleep on school nights is an earlier bedtime. However, this can be misaligned with their circadian phase. The effects of circadian misalignment remain understudied. Here, we present preliminary findings from an ongoing experimental trial comparing sleep extension that is aligned vs. misaligned with adolescents’ circadian phase.

Methods

Healthy 14-18-year-olds in the bottom and top quartile of reported midsleep on non-school nights (“lark” vs “owl” chronotype) underwent a 3-week protocol with three multi-night periods at home: 8-hours/night of sleep opportunity aligned with their chronotype (Week1), 6.5-hours/night centered between chronotypes (Week2), and 9.5 hours/night randomly assigned to early bedtime or late risetime conditions (Week3). Linear mixed models compared Dim Light Melatonin Onset (DLMO, collected at the end of each week) and sleep midpoint and duration (via wrist actigraphy) across week, chronotype, and random assignment during Week3 (early vs late sleep extension). Alpha was set at .01.

Results

Of 68 adolescents randomized, 57 (84%) completed the protocol (Mean age=15.9, 53% female). At Week1, larks and owls markedly differed in midsleep (03:16 vs 05:38) and DLMO (21:18 vs 23:30), ps<.001, with midsleep and DLMO correlating well (r=.76, p<.001). By design, sleep duration markedly differed across weeks (7.7 vs 6.4 vs 8.5 hours; p<.001) without any significant interactions involving chronotype or randomization. Sleep midpoint and DLMO each showed week-by-randomization interactions (ps<.001) but no 3-way interactions. By design, late risetime significantly delayed midsleep in Week3 only (p<.001). Week-by-week follow-up tests of the randomization effect on DLMO were non-significant. Sleep midpoint showed a week-by-chronotype interaction (p<.001), with later midsleep for owls pre-randomization (ps<.003), but not during Week3. Midsleep at Week3 and subsequent DLMO correlated well for adolescents who were randomized to a condition that aligned with their chronotype (r=.70, p<.001), but not those randomized to a misaligned condition (r=.26, p>.2).

Conclusion

It is feasible to experimentally induce sleep patterns that meet recommendations for healthy sleep duration but are aligned or misaligned with individual adolescents’ circadian phase. We will use this protocol to study the effect of circadian misalignment on health outcomes.

Support (if any)

US National Institutes of Health (R01HL147915)
0017 Circadian Phase Alignment and Sleep Duration in Overweight Adults: Associations with Cardiometabolic Risk Factors
Authors: Baron Kelly , Appelhans Bradley , Burgess Helen , Quinn Laurie , Carlson Steven , Allen Chelsea , Greene Tom
Published: April 2024

Introduction

Both short sleep duration and circadian rhythm misalignment are associated with increased cardiometabolic risk factors. Although experimental research has demonstrated the independent and interactive effects of these two exposures, few studies have examined the interaction between these two factors on outcomes among free-living participants. The goal of this study is to examine how sleep duration and circadian alignment predict cardiometabolic risk factors and extend prior research by examining the mechanisms of insulin resistance.

Methods

Participants included adults aged 18-60 y with BMI 25-35 kg/m2 who completed body measurements, at least 7 days of actigraphy and an in lab dim light melatonin onset assessment (DLMO). The main measure of circadian misalignment was the phase angle, or duration between DLMO and habitual sleep onset time in the past 7 days. Participants attended an outpatient visit to the clinical research unit to complete a frequently sampled intravenous glucose tolerance test (FSIVGTT) and data were analyzed using Bergman’s minimal model analysis. Multivariable linear regression models evaluated the effects of sleep duration both individually and their interaction in predicting cardiometabolic outcomes in models controlling for age, sex, race and ethnicity.

Results

Body measurement data were available for n= 92 participants and FSIVGTT were available for 87 participants (age: M= 36 SD=10 years, 47 female). Average sleep duration was 6.9 hours (SD=0.6) and average phase angle was 3.3 hours (SD=1.0). We found there was higher insulin sensitivity among those with shorter phase angle (Si, estimate= -2.58 CI= -4.63, -0.53 (mU/L)-1min-1, p=.02) and a trend among shorter sleep duration (estimate = -3.71, CI= -7.35, -0.07 (mU/L)-1min-1, p=.05). Phase angle and sleep duration were not associated with metabolic measures, BMI, body fat or hip/waist ratio.

Conclusion

Results demonstrate contrary to our hypotheses, shorter phase angle and sleep were associated with higher insulin sensitivity and unrelated to other cardiometabolic risk markers in our sample of overweight adults without diabetes.

Support (if any)

The research reported in this publication was supported in part by the National Heart Lung and Blood Institute under Award Number 1R01HL141706-01A1 and the National Center for Advancing Translational Sciences of the National Institutes of Health under Award Number UL1TR002538
0018 Extroversion Predicts Higher Evening Caloric Intake
Authors: Kennedy Kathryn , Huskey Alisa , Grandner Michael , Killgore William
Published: April 2024

Introduction

Personality has been shown to predict health-related outcomes, including eating behavior. However, the findings on extroversion and caloric intake have been inconsistent. This study sought to examine whether trait extroversion predicted total caloric intake, as well as calories from specific macronutrients, at different times of day during a sleep deprivation protocol.

Methods

Participants (n=46; 23 women) conducted a baseline personality assessment before completing a 30-hour sleep deprivation protocol a week later. The laboratory visit began at 6pm, and participants departed the following day at 2pm. While in the lab, participants completed serial neurocognitive assessments and engaged in ad libitum feeding. Caloric intake was assessed in 6-hour bins from 6pm (start of laboratory protocol) until 12am, 12am-6am, and 6am-12pm. Multivariate regression models were used with extroversion as the primary predictor (assessed via NEO Personality Inventory scores) and primary outcomes including total caloric intake, calories from total carbohydrates, sugars, protein, total fat, saturated fat, and transfats. Models were adjusted for age, sex, BMI, and chronotype (assessed using the Morningness-Eveningness Questionnaire (MEQ)).

Results

The fitted regression model with extroversion as the primary predictor and food variables as outcomes was statistically significant for total carbohydrates (F(4,40) = 3.153; R2 = 0.164; p = 0.024) and total calories (F(4,40) = 3.179; R2 = 0.165; p = 0.023) consumed between 6pm and midnight, adjusted for age, sex, and BMI. Extroversion was positively associated with greater total carbohydrates (β (3.148) = p < 0.0174) and greater total calories (β (22.088) = p < 0.0057) consumed between 6pm and midnight. Chronotype did not predict food intake in models further adjusted for MEQ score.

Conclusion

Trait extroversion may be associated a greater propensity for hedonic eating behavior, thus leading to higher caloric intake. However, it may also be that in a laboratory setting where social interactions are limited, extroversion predicts the consumption of foods as a self-soothing strategy in the event of loneliness. Further investigation is needed to understand this association in real world settings.

Support (if any)

DARPA AWARD #: D12AP00241
0020 Heart Rate Variability Changes During Sleep in Postmenopausal Women
Authors: Pérez-Medina-Carballo Rafael , Kosmadopoulos Anastasi , Boudreau Philippe , Boivin Diane B
Published: April 2024

Introduction

Young women present a lower risk of cardiovascular disease (CVD) than age-matched men, but this risk steeply increases at menopause. Heart rate variability (HRV) has been proposed as a marker of cardiovascular health, which changes with circadian phase and menopause. We aim to understand the changes in the circadian variation of HRV that occurs in postmenopausal women.

Methods

Eight healthy PMW (54.8±3.4 years, one taking hormones) and 12 healthy young women (YW; 25.8±3.4 years) in their mid-follicular phase were enrolled. Participants were healthy sleepers. After a regular 8-h sleep schedule at home for ≥2 weeks, participants entered the laboratory. After a baseline nocturnal sleep period, they underwent a 48-h (PMW) or 72-h (YW) ultradian sleep-wake cycle procedure (USW) consisting of alternating 60-min wake and nap opportunities. Sleep was polysomnographically recorded. EKG recordings were used to calculate HRV parameters: heart rate (HR), SDNN, RMSSD, HF. Circadian parameters (mesor, amplitude, phase) were calculated by fitting a cosinor to HRV parameters during the first 24 naps of the USW. Linear mixed-effects models were used for between-group comparisons.

Results

During the hourly analysis of the baseline sleep period, PMW presented lower SDNN, RMSSD and HF compared to YW (p≤0.025), with no differences in HR. PMW also had lower SDNN, RMSSD, and HF throughout N2, N3, REM sleep, and wake epochs (p≤0.013). During the USW procedure, YW showed a significant circadian variation of HR during NREM sleep and wake, and HF during NREM sleep (p≤0.010), whereas these variations were not significant in PMW. Both groups presented a significant circadian variation of SDNN in NREM sleep and wake (p≤0.037), with no differences in amplitude or phase. PMW also presented a lower mesor of SDNN, RMSSD and HF (p≤0.024).

Conclusion

The lower HRV (SDNN) and parasympathetic activity (RMSSD and HF) during the sleep of PMW is consistent with the increased risk of developing CVD after menopause. Furthermore, the dampened circadian variation of HR and HF in PMW suggest a weakened circadian regulation of the cardiovascular system. This could be a mechanism involved in the increased CV risk after menopause, although this would require further investigation.

Support (if any)

Canadian Institutes of Health Research.
0023 Light- and Meal-induced Phase Resetting of Circadian Lipid Rhythms in Humans
Authors: Grant Leilah , Chung Joon , Garbazza Corrado , McKenzie Lauren , Nogueiras Montserrat Perez , Gotto Alexander , Hilaire Melissa St , Lockley Steven , Rahman Shadab
Published: April 2024

Introduction

We have characterized endogenous circadian rhythms in total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C) and triglycerides (TG) and shown that they can be reset according to a Type I phase response curve (PRC) by a combined stimulus of light exposure and meals. The resulting PRCs are different in phase and amplitude than that describing melatonin rhythm resetting and therefore we hypothesized that lipid rhythms may be reset by meal timing rather than light. In the current study, we are constructing three PRCs that systematically examine the contribution of light and/or meal timing on the circadian resetting response of lipids.

Methods

To date, 15 young healthy adults (mean age±SD= 25.9±3.6 yrs, 7F) have been randomized to one of three stimulus conditions: Light + Meals (n=5): 6.5-h ~10,000 lux white light exposure with a 12-h meal window; Meals only (n=4): dim light (< 3 lux) with a 12-h meal window; or Time-restricted Meals (n=6): dim light with a 6.5-h meal window. Individuals are randomized to one of 16 stimulus times distributed every 90-minutes (~22.5◦) across the 24-hour day. Cosinor analysis was used to assess 24-h rhythms in lipids measured during constant routines conducted before and after stimulus administration. Phase shifts were calculated as the difference between pre- and post-stimulus acrophase.

Results

Endogenous circadian rhythms were observed in 73%, 86% and 100% of participants for TC, HDL-C and TG, respectively, during the first CR. These percentages were similar for the second CR: 80% for TC and HDL-C and 93% for TG. Consistent with our previous work, TC and HDL-C peaked in the afternoon (~15:30), whereas TG peaked in the early morning (~03:30). TC, HDL-C and TG demonstrated both advance (up to 7.1 hours) and delay (up to 8.9 hours) phase shifts depending on the circadian time of stimulus administration that were broadly consistent with our published PRCs.

Conclusion

Once completed, these PRCs will differentiate the role of light versus meals in resetting lipid rhythms, and the impact of meal window duration. This work is essential for providing evidence-based advice to shift workers and others with circadian misalignment to reduce their risk of lipid rhythm desynchrony.

Support (if any)

R01HL159207
0028 Chronotype and Circadian Timing in Day and Chronic Night-shift Workers
Authors: McAuliffe Kathryn , Shafer Brooke , McHill Andrew
Published: April 2024

Introduction

In populations who live and work typical day-shift work hours, chronotype (i.e., morning lark or night owl) is tightly associated with dim-light melatonin onset (DLMO) timing, the gold standard marker of circadian phase. The association between these markers is less clear in populations chronically working fixed, overnight shift work schedules which may be vital for making population-based recommendations. We therefore examined the relationship between chronotype and DLMO in chronic, fixed overnight shift working nurses compared to fixed day-shift working nurses.

Methods

Sixteen volunteers (15 female; aged [average ± SD] 31.8±7.7yr) working either a fixed day (n=8) or nightshift schedule (n=8; 19:00-07:00 shift; duration of night-shift work 8.6 ± 7.2yr) participated in an ~8h evening in-laboratory stay in dim-lighting (< 5 lux). Beginning at 17:00, saliva was collected every 30 minutes to calculate circadian phase via salivary DLMO (3pg/ml threshold). Chronotype was determined via daily sleep/wake diaries collected over 7-days and defined as mid-sleep time on free days corrected for sleep loss on participant-identified work days (MSFsc). Pearson Correlations were used to determine associations between DLMO and MSFsc within each work-schedule group and circadian timing between groups was compared using Independent T-Test. DLMO was not captured in two night-shift working participants, thus they were not included in DLMO analysis.

Results

The timing of MSFsc and DLMO were significantly correlated such that later mid-sleep timing was correlated with later DLMO timing in day-shift (r=0.77, p< 0.025), but not night-shift (r=0.005, p=0.99) workers. Moreover, the average MSFsc (01:25± 0:45 vs 11:16±01:35; p< 0.001) and DLMO timing (19:13±01:04 vs 21:12±01:01; p=0.005) of the day-shift nurses were significantly earlier than the night-shift nurses.

Conclusion

Working and living on a day-shift schedule results in an association between chronotype and circadian timing, while a stable night-shift schedule results in no discernable relationship between the measures, despite a later DLMO timing. Understanding the connection between these sleep and circadian measurements could have implications for interventions to improve sleep, health, and performance in shift-working populations.

Support (if any)

OHSU School of Nursing PhD Student Scholarship Award, T32HL083808, K01HL146992
0029 Bimodality in Undergraduate Students Is Associated with Negative Health and Sleep Outcomes
Authors: Vallim Julia , Tsuji Heloisa , Lima Gabriela , Xylaras Beatriz , Louzada Fernando , Tufik Sergio , D'Almeida Vania
Published: April 2024

Introduction

The bimodal chronotype is a fourth circadian phenotype proposed by re-scoring the Morningness-Eveningness Questionnaire as first described by Martynhak and colleagues in 2010. Bimodal subjects are classified as “intermediate”, but they answer some of the questions as a morning person and others as an evening person. The present work aimed to describe the prevalence of the bimodal chronotype in a sample of undergraduate students in the city of Sao Paulo and to characterize the bimodal type in terms of their health and sleep-related outcomes.

Methods

A web-based cross-sectional study conducted between September 2018 and March 2021, using a convenience sampling method. The sample was composed of undergraduate students from two higher education institutions in Sao Paulo, who completed an electronic form that included the Morningness and Eveningness Questionnaire, the Pittsburgh Sleep Quality Index, the Self-Compassion Scale, the Epworth Sleepiness Scale, the Hospital Anxiety and Depression Scale, and the World Health Organization Subjective Well-Being Index. The final sample consisted of 615 students (82% female, mean age: 23.4±6.5 years).

Results

Of the 615 students, 108 (18%) had positive bimodality indexes. Bimodal subjects comprised 48 students (79% female, 19% male, 2% other gender), 8% of the total sample. Bimodal subjects had poorer subjective sleep quality (F(3,598)=20.0 p< 0.001), more daytime sleepiness (F(3,611)=7.1 p< 0.001), lower subjective well-being (F(3,611)=14.0 p< 0.001), greater anxiety (F(3,611)=9.6 p< 0.001) and depression symptoms (F(3,611)=11.8 p< 0.001), and lower self-compassion (F(3,611)=10.1 p< 0.001) than morning and/or intermediate types. However, they did not differ from evening types.

Conclusion

We found a bimodality prevalence of 8% and show that this profile is associated with negative health and sleep outcomes. The description of bimodality prevalence in a sample of college students is interesting because they are often an at-risk population for abnormal sleep and circadian disruption, which may be of interest for the design of academic public policies more consistent with the circadian reality 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).
0034 Sleep Is Associated with Telomere Shortening: A Population-Based Longitudinal Study
Authors: Tempaku Priscila , D'Almeida Vania , Andersen Monica , Tufik Sergio
Published: April 2024

Introduction

Evidence has shown that as the chronological age of an organism increases, there is a decrease in the telomere length (TL) via oxidative stress and inflammation pathways. Epidemiological studies have measured this parameter as the mean leukocyte TL and several associations between TL and age-related diseases have been described. Since the major pathophysiological factors related to inadequate sleep (including sleep complaints and sleep disorders) contribute to the exacerbation of inflammation and oxidative stress, the association of sleep and TL has been proposed in many cross-sectional studies. Thus, the aim of this study was to evaluate the association between sleep-related variables with TL in a population-based longitudinal framework.

Methods

We used data derived from the São Paulo Epidemiologic Sleep Study cohort, which was followed over 8 years. All individuals answered sleep-related questionnaires, underwent a full-night polysomnography (PSG) and had their blood collected for DNA extraction. TL was measured through quantitative real time polymerase chain reaction. For analysis in a single point in time, the Generalized Linear Model was used. The Generalized Estimating Equations test was applied for the comparison between baseline and follow-up, considering age, sex and body mass index (BMI) as covariables; and Spearman correlations were applied to compare the delta values between baseline and follow-up.

Results

Of the 1,042 individuals in the EPISONO cohort, 68.3% accepted to participate in the follow-up study (n=712). We have found that baseline SpO2 (ß=0.007, p=0.013), medium SpO2 (ß=0.009, p=0.004) and total sleep time < 90% (ß=-0.142, p=0.037) had an effect on TL from the follow-up. The 8-year TL attrition was inversely associated with sleep quality, sleepiness, total sleep time, sleep efficiency, sleep architecture variables, wake after sleep onset, arousal index, oxygen-related variables baseline and the presence of obstructive sleep apnea (OSA).

Conclusion

Individuals with worse sleep quality, alterations in sleep architecture and OSA had greater TL attrition over the 8 years. Using a longitudinal approach, these findings confirm previous cross-sectional evidence linking sleep with accelerated biological aging.

Support (if any)

This work was supported by grants from AFIP, CNPq and FAPESP.
0035 Characterization of Metabolites in a Randomized, Controlled Trial of Clarithromycin for Pathological Sleepiness
Authors: Saini Prabhjyot , Keating Glenda , Rye David , Bliwise Donald , Ortlund Eric , Trotti Lynn Marie
Published: April 2024

Introduction

Pathologic sleepiness in CDH is associated with the presence of an endogenous, cerebrospinal fluid peptide that can positively, allosterically modulate GABA-A receptors. The macrolide antibiotic clarithromycin has been shown to benefit patients with hypersomnolence that is refractory to standard of care agents and is a negative allosteric modulator of GABA-A receptors. One explanation of its wake-promoting effects is the alteration of gastrointestinal microbiota composition. Here we characterize the metabolomics of serum and stool samples collected as part of a randomized, controlled trial of clarithromycin.

Methods

Ten patients and 10 Healthy controls had serum and stool samples taken on baseline study day and after a 2 week trial of clarithromycin dosed at 500mg bid, A total of 60 serum and stool samples were analyzed using Mass Spectrometry with a metabolomics panel. A total of 214 metabolites were detected, 122 in serum and 148 in stool with 56 overlapping metabolites.

Results

Samples were normalized by QC samples, log transformed and scaled separately for serum and stool. When comparing the 3 groups at the metabolite level, univariate analyses revealed 7 in serum and 4 in stool that had different levels between at nominal significance (p< 0.05). The 4 in stool were 2-Deoxyadenosine, 3-Hydroxybenzaldehyde, Imidazole acetic acid and N-Acetyl aspartate. The first 2 were driven by lower values in the Treatment group vs Healthy Controls, and the latter 2 were the opposite. The 7 in serum were (R)-2,3-Dihydroxy-isovalerate, Homoserine, L-Allothreonine, L-Threonine, O-Acetyl-L-Serine, Palmitoyl carnitine, Riboflavin. Paired log2 fold-change analysis in serum between treatment and baseline revealed nominally significant decrease in the expression of Theobromine (P=0.4). In stool, the analysis revealed changes in 3-Hydroxybutyric acid, 2-Deoxyadenosine,3-2-Hydroxyethylindole,3-Hydroxy-2-Methylpyridine, Glycerol 2-phosphate, N-Acetylaspartate, Nicotinic Acid, Mononucleotide, Riboflavin, and Theobromine. PCA loadings of serum and stool samples, overall and in each group, revealed the first 2 PC’s consisting of 3-Hydroxybutyric acid and (R)-2,3-Dihydroxy-isovalerate

Conclusion

Despite similar profiles in serum and stool in patients, before and after treatment and healthy controls; subtle differences in particular metabolites may need to be further studied. Alternative pathways not currently captured in metabolomics in this study may be better suited to capture evidence of alternative mechanisms of clarithromycin efficacy.

Support (if any)

NS111280
0036 Exploring Common Genetic Factors Between Obstructive Sleep Apnea and Asthma: Insights into Inflammatory Pathways
Authors: Souza Maingredy , Moysés-Oliveira Mariana , Porcacchia Allan , Rosa Daniela , Tufik Sergio , Andersen Monica
Published: April 2024

Introduction

Obstructive sleep apnea (OSA) and asthma are globally prevalent respiratory disorders that share risk factors, symptoms and pathophysiological mechanisms. Emerging evidence suggests a bidirectional relationship, with each condition having a negative impact on the clinical course of the other. Although both diseases have a strong hereditary component, research assessing the genetic basis of their co-occurrence remains limited. Thus, the aim of this study was to analyze the interaction of risk genes and common biological pathways between OSA and asthma.

Methods

Two sets of genes associated with OSA (2,159 genes) and asthma (786 genes) were manually curated from significant single nucleotide polymorphisms (SNP) revealed in genome-wide association studies (GWAS). These lists were subsequently compared to identify intersecting genes, and the statistical significance of the overlap was assessed using Fisher's Exact Test. Pathway enrichment analysis was conducted utilizing the Benjamini-Hochberg test with a significance threshold set at an adjusted p-value< 0.05.

Results

There were 187 overlapping genes between OSA and asthma gene sets, indicating a significantly higher occurrence than expected by chance (p< 1.07E–29; odds ratio=2.3). The pathway overrepresentation analysis of these intersecting genes identified processes associated with immune system functions, encompassing human leucocyte antigen (HLA), antigen presentation, T cell differentiation, cell signaling, and positive regulation of inflammatory mediators.

Conclusion

The shared genetic basis between OSA and asthma suggests dysregulation of the immune response and pro-inflammatory processes as underlying pathophysiological mechanisms contributing to the susceptibility and manifestation of these comorbid conditions.

Support (if any)

Associação Fundo de Incentivo à Pesquisa (AFIP); Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP), and Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq).
0037 Investigating Common Genetic Factors Between Obstructive Sleep Apnea and Prostate Cancer
Authors: Porcacchia Allan , Moysés-Oliveira Mariana , Pires Gabriel , Andersen Monica , Tufik Sergio
Published: April 2024

Introduction

Cancer is one of the leading causes of death in the world. Studies have highlighted the possible association between obstructive sleep apnea (OSA) and an increased risk of incidence or prevalence of some types of cancer, especially prostate cancer. However, the extent to which genes that are related to both conditions contribute to their comorbidity is still unclear. The aim of this study was to evaluate the intersection of risk genes found in both OSA and prostate cancer.

Methods

Two sets of genes associated respectively with OSA and prostate cancer were manually curated and then compared to obtain an intersection gene list. Fisher’s exact test was employed to evaluate the statistical significance of the overlap, with a significance threshold of p-value< 0.05. The identification of enriched pathways among the intersection gene list was performed using the Benjamini–Hochberg test, with a significance threshold of adjusted p-value< 0.05. A functional network was generated from the intersect gene list which was used as an input in a protein–protein interaction analysis conducted using the String database.

Results

Sixty-eight genes overlapped between prostate cancer risk genes and OSA-associated genes, indicating more overlap than expected by chance (odds ratio=2.665; p=2.606E–10). This intersect list was associated mainly with hypoxia, apoptosis, oxidative stress, and cell cycle, proliferation, or cell damage. The hypoxia-inducible factor (HIF)-1 signaling pathway stands out from the list of enriched pathways reported in both conditions (adjusted p-value< 0.0006). A 17-node network was retrieved from the overlapping gene list and included key proteins from the aforementioned enriched pathways, which comprised TNF, STAT3, BCL-2, TGF, and ATM.

Conclusion

The genetic basis of prostate cancer and OSA was significantly shared, indicating common molecular etiology in respect of this comorbidity. Relevant proliferation, apoptosis, and hypoxia pathways were significantly enriched among the genes associated with both conditions, suggesting their role in pathophysiological processes.

Support (if any)

Associação Fundo de Incentivo à Pesquisa (AFIP); Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP), and Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq).
0038 Kinesin Binding as a Shared Pathway Underlying the Genetic Basis of Male Infertility and Insomnia
Authors: Adami Luana , Moyses-Oliveira Mariana , Tufik Sergio , Andersen Monica
Published: April 2024

Introduction

Fertility problems are becoming increasingly common, with issues relating to men accounting for half of the cases. In addition to all the known causes of male infertility, some complaints are of idiopathic origin, and lifestyle, including sleep habits, might be an important underlying factor. Sleep disorders such as insomnia, which are prevalent in the population, might interfere in male fertility. Therefore, we aimed to perform an in silico study to test whether male infertility and insomnia share genetic risk variants, and to identify any molecular, cellular, and biological interactions between them.

Methods

Two lists of gene sets were manually curated through a literature review using PUBMED, 1 associated with male infertility (454 genes) and the other with insomnia (921 genes). The statistical significance of the gene list overlap was performed by the Fisher Exact test using Nematode Bioinformatics and Molbiotools online tools. This analysis considered a total of 21,196 genes in the human genome with a statistical threshold of P-value< 0.05. The enriched terms over-represented in the intersect gene list were acquired from the Benjamini-Hochberg and the protein-protein interaction (PPI) analysis was performed via String database.

Results

Among these genes, 28 were common for both lists, representing more overlap than expected by chance (Fisher's exact test, p-value=0.041). The biological function of the genes contained in the intersection gene list suggested that pathways related to kinesin binding were commonly found in both conditions (p-value=0.038). A protein-protein interaction (PPI) analysis using the intersection list as input retrieved 25 nodes and indicated that 2 of them were kinesin-related proteins (PLEKHM2 and KCL1).

Conclusion

One of the major physiological mechanisms supporting the crosstalk between the sperm and the neuron through kinesis-related processes is mitochondria recycling. The intersecting gene list and the enriched pathways retrieved from our analysis might reflect how the rhythm control of neural circuits may affect spermatogenesis and male fertility. These results might serve as preliminary findings for future clinical trials focused on the identification of pharmacological interventions tailored to this comorbidity.

Support (if any)

AFIP, FAPESP, CNPq.
0041 Drug Screening and CRISPR/Cas9 Screening of HCN Channels
Authors: Smieszek Sandra , Doldur-Balli Fusun , Keenan Brendan , Zimmerman Amber , Veatch Olivia , Polymeropoulos Christos , Birznieks Gunther , Polymeropoulos Mihael
Published: April 2024

Introduction

A growing body of evidence suggests a role for HCN (hyperpolarization-activated cyclic nucleotide-gated) channels in regulation of sleep. We first tested effects of three HCN channel blockers (Corlanor, Zatebradine Hydrochloride and ZD7288) on sleep/wake behavior in wild type zebrafish compared to DMSO control. Then, to more definitively investigate roles of HCN channels, we knocked out genes encoding for HCN channel subunits in zebrafish by utilizing CRISPR/Cas9 technique and we assessed sleep/wake behavior in the HCN channel mutants.

Methods

The compounds were tested at six concentrations varying between 0.1-30 ◻M (.i.e., 0.1 μM, 0.3 μM, 1.0 μM, 4.5 μM, 10 μM and 30 μM) in zebrafish larvae in our drug screening study. Control groups were administered DMSO. In CRISPR screening, single cell-stage zebrafish embryos were injected with preformed ribonucleoprotein complexes containing Cas9 protein and two crRNAs targeting each homologous gene of human HCN1, HCN2, HCN3, and HCN4. Control groups were negative control crRNA injected embryos. Sleep phenotyping was performed by placing individual larva into the wells of 96 well plates and by recording their activity via commercially available video monitoring equipment.

Results

Drug screening study showed shorter latency to sleep at 0.1 μM dose of Ivabradine, moderate reductions in average activity at 30 μM dose of Zatebradine Hydrochloride, and increased sleep at 4.5 μM dose of ZD7288. Our CRISPR/Cas9 screening resulted in decreased activity in hcn1 crispants, fewer sleep bouts in daytime in hcn2 crispants, and no difference in sleep parameters in hcn3 crispants. hcn4 crispants had shorter sleep in both day and nighttime, hyperactivity and shorter sleep bouts at night and fewer sleep bouts in daytime.

Conclusion

Blocking HCN channels decreased wakefulness in our drug screening study. Genetically targeting hcn1 and hcn2 genes resulted in the same direction with the results of drug screening study and knocking out hcn3 didn’t change sleep parameters. Change in sleep parameters were in the opposite direction in hcn4 crispants. HCN channel blockers might be selecting HCN1 and HCN2 however further work is required to confirm this suggestion.

Support (if any)
0043 The Role of Median Preoptic Nucleus Astrocytes in Mediating E2’s Effects on Sleep-Wake Behavior
Authors: Onwukwe Chimdiya , Viechweg Shaun , Byrd Carissa , Mong Jessica
Published: April 2024

Introduction

Although 50-70 million Americans suffer from sleep disorders, women are 2x as likely as men to experience sleep disruptions. This discrepancy emerges at puberty and is strongly associated with fluctuations in estrogen, suggesting a role for estrogens in sleep disorders. Our previous work has identified the median preoptic nucleus (MnPO) as a key site regulating estradiol (E2) effects on sleep-wake behavior. We have also shown that E2 increases extracellular adenosine in the MnPO. Astrocytes are a major source of adenosine in the central nervous system, and cortical astrocytes have been shown to regulate sleep homeostasis. Thus, we hypothesize that MnPO astrocytes play a key role in the estrogenic mechanisms regulating sleep-wake behaviors.

Methods

To clarify the role of astrocytes in E2- modulation of sleep, we expressed Gi- or Gq-DREADDs in MnPO astrocytes of ovariectomized rats. Rats were treated with subcutaneous Oil injections (baseline) followed by injections of low (5ug) and high (10ug) dose E2, 24 hours apart. Rats also received Vehicle or Clozapine-N-oxide (CNO; 1.7mg/kg) at the time of their Oil/E2 injections. EEG/EMG recordings were acquired throughout treatment. To inhibit Ca2+signaling, and, ultimately, gliotransmission in astrocytes, we used a viral approach to express Pleckstrin Homology domain of Phospholipase C (PLC)-like protein p130 (p130PH) in MnPO astrocytes. p130PH is an IP3 buffer that prevents IP3 from initiating intracellular Ca2+ release and was previously shown to reduce astrocytic Ca2+signaling and gliotransmission (PMID: 20736051). Experimental (p130PH) and Control (tdTomato) animals were then treated with the Oil/E2 paradigm described above.

Results

In Oil-treated animals, CNO activation of Gq-DREADD increased wake and decreased sleep, mimicking E2’s effects on Sleep-Wake behavior. Unexpectedly, CNO activation of Gi-DREADD produced similar effects, increasing wake and decreasing sleep. These observations may support literature findings that Gi-DREADDs, while inhibitory in neurons, may enhance activity in astrocytes. Thus, Gi-DREADDs may not be an ideal candidate for inhibiting astrocytic activity. Animals expressing p130PH (n=3) exhibited less wake and more NREM sleep than animals injected with Control virus (n=3).

Conclusion

These preliminary findings suggest that E2 may increase wake by enhancing activity of MnPO astrocytes and highlight a need for further investigation.

Support (if any)
0044 Sleep Need-dependent Plasticity of a Thalamic Circuit Generates Persistence of Sleep Drive
Authors: Lee Sang Soo , Liu Qiang , Kim Thomas , Palmer Isabelle , Park Kristen , Muenzberg Heike , Blackshaw Seth , Wu Mark
Published: April 2024

Introduction

Prolonged wakefulness leads to persistent, deep recovery sleep. However, the neuronal circuits mediating this process remain elusive. Here, starting from a large circuit screen, we identify and characterize a group of thalamic nucleus reuniens (RE) neurons activated by sleep pressure and required for sleep homeostasis.

Methods

To identify upstream excitatory neuronal populations inducing NREM sleep, we performed rabies virus-mediated screening in mice. Sleep/wake states were assayed using tethered EEG/EMG recordings. Activation or inhibition of neuronal activity was performed using chemogenetic and/or optogenetic techniques. Behavior during optogenetic RE activation was assessed using video recordings. To genetically access the sleep pressure-activated RE neurons, Targeted Recombination in Active Populations (TRAP2) mice were used. Viral expression of caspase3 was used for selective ablation of cells. For projection analyses, virally expressed Synaptophysin was used as a presynaptic marker. To visually examine synaptic connectivity, we performed enhanced Green fluorescent protein Reconstruction Across Synaptic Partners (eGRASP) analysis. Functional connectivity between RE to downstream neurons was tested by slice patch-clamp recordings.

Results

From a circuit screen in mice, we identified a group of excitatory thalamic RE neurons activated by sleep pressure and required for sleep homeostasis. Optogenetic activation of these neurons leads to an unusual phenotype: sleep-preparatory behaviors (e.g., nesting) followed by prolonged, intense sleep resembling recovery sleep. The activity of RE neurons (assessed by Fos) is increased by sleep deprivation (SD), and ablation of these Fos+ RE cells markedly impairs recovery sleep following SD. Moreover, inhibiting RE activity during SD impairs subsequent recovery sleep, suggesting that these neurons signal sleep need. RE neurons act upstream of sleep-promoting zona incerta (ZI) cells, and, remarkably, SD triggers plasticity of this circuit to strengthen its connectivity, as assessed by eGRASP analysis and patch-clamp recordings. Finally, our results indicate that CaMKII signaling is required for morphological plasticity of the RE-ZI circuit and persistence of homeostatic NREM sleep.

Conclusion

A subset of RE neurons encode sleep pressure and generate persistent NREM sleep, via a projection to sleep-promoting ZI neurons. Increased sleep need triggers plasticity of the RE-ZI projection which enhances functional connectivity facilitating the transmission of homeostatic sleep drive.

Support (if any)

NIH grants: R01NS094571, R35NS122181, S10OD023548, NS05027
0045 Lateral Hypothalamic Neurotensin Neurons Are Necessary for Wake Maintenance
Authors: Khanday Mudasir , Naganuma Fumito , Hasan Whidul , Bandaru Sathyajit , Ibrahim Nazifa , Nandigam Shaili , Vetrivelan Ramalingam
Published: April 2024

Introduction

The lateral Hypothalamic area (LH) has been established as a critical site for regulating wakefulness. However, the exact neural elements within this region that promote and stabilize wakefulness are not completely understood. While the orexin neurons in the LH are well-known, other subpopulations within this area may also contribute to arousal control. In this regard, we have recently identified a subset of LH neurons expressing neurotensin (Nts) promoting wakefulness. Optogenetic activation of LH-Nts neurons rapidly induces arousal from non-rapid eye movement (NREM) sleep, while chemoactivation induces up to five hours of uninterrupted wakefulness. However, the precise role of these neurons in initiating and maintaining spontaneous wakefulness remains unknown. To address this question, we investigated 1) the activity dynamics of LH-Nts neurons across sleep-wake behavior and 2) the impact of their ablation on sleep-wake amounts and architecture.

Methods

Experiment 1 – LH-Nts neuron activity dynamics: Nts-Cre mice (expressing Cre recombinase specifically in Nts neurons) were injected with an adeno-associated viral (AAV) vector coding for a Cre-dependent GCaMPS (genetically encoded calcium sensors) into the LH. They were also implanted with optical fibers targeting 50µm above the injection site and EEG/EMG electrodes to assess sleep-wake. After three weeks, fiber photometry with concurrent sleep-wake recordings was performed for two hours during the light period in all mice. Experiment 2 – sleep-wake effects of LH-Nts neuron ablation: Nts-Cre mice were injected with an AAV coding for diphtheria toxin-A subunit (AAV-flex-DTA) or a control AAV coding for a red fluorescent protein (AAV-flex-mCherry) into the LH and were implanted with transmitters for the telemetric recording of EEG/EMG. Sleep-wake recordings were performed 24 hours at four weeks after surgery. All mice were then perfused, and injection/lesion sites were verified histologically.

Results

LH-Nts neurons were more active during Wakefulness and REM sleep when compared to NREM sleep. The loss of these neurons reduced the total amounts of Wake and the average duration of Wake bouts but increased the 0.5-4 Hz delta power in Wake EEG.

Conclusion

LH-Nts neurons are necessary for maintaining spontaneous wakefulness, and their loss leads to chronic sleepiness in mice.

Support (if any)

NIH R01-NS119223 (to R.V)