Forced Desynchrony Research Articles

Mathematical Analysis of Light-sensitivity Related Challenges in Assessment of the Intrinsic Period of the Human Circadian Pacemaker.

Accurate assessment of the intrinsic period of the human circadian pacemaker is essential for a quantitative understanding of how our circadian rhythms are synchronized to exposure to natural and man-made light-dark (LD) cycles. The gold standard method for assessing intrinsic period in humans is forced desynchrony (FD) which assumes that the confounding effect of lights-on assessment of intrinsic period is removed by scheduling sleep-wake and associated dim LD cycles to periods outside the range of entrainment of the circadian pacemaker. However, the observation that the mean period of free-running blind people is longer than the mean period of sighted people assessed by FD (24.50 0.17 h vs 24.15 0.20 h, p 0.001) appears inconsistent with this assertion. Here, we present a mathematical analysis using a simple parametric model of the circadian pacemaker with a sinusoidal velocity response curve (VRC) describing the effect of light on the speed of the oscillator. The analysis shows that the shorter period in FD may be explained by exquisite sensitivity of the human circadian pacemaker to low light intensities and a VRC with a larger advance region than delay region. The main implication of this analysis, which generates new and testable predictions, is that current quantitative models for predicting how light exposure affects entrainment of the human circadian system may not accurately capture the effect of dim light. The mathematical analysis generates new predictions which can be tested in laboratory experiments. These findings have implications for managing healthy entrainment of human circadian clocks in societies with abundant access to light sources with powerful biological effects.

Influence of circadian phase and extended wakefulness on glucose levels during forced desynchrony

ObjectivesCircadian misalignment and sleep deprivation often occur in tandem, and both negatively impact glucose homeostasis and metabolic health. The present study employed a forced desynchrony protocol to examine the influence of extended wakefulness and circadian misalignment on hourly glucose levels. MethodsNine healthy adults (4F/5M; 26 ± 4years) completed a 31-day in-laboratory protocol. After three 24 hour baseline days with 8 hours scheduled sleep opportunities, participants were scheduled to 14 consecutive 42.85 hour sleep-wake cycles, with 28.57 hours extended wakefulness and 14.28 hours sleep opportunities each cycle. Blood was sampled hourly across the forced desynchrony and over 600 plasma samples per participant were analyzed for glucose levels. ResultsBoth hours into the 42.85 hours forced desynchrony day and circadian phase modulated glucose levels (p < .0001). Glucose peaked after each meal during scheduled wakefulness and decreased during scheduled sleep/fasting. Glucose levels were, on average, lowest during the biological daytime and rose throughout the biological night, peaking in the biological morning. When analyzed separately for scheduled sleep vs. wakefulness, the peak timing of the circadian rhythm in glucose was later during sleep (p < .05). Glucose area under the curve levels increased rapidly from the beginning of the forced desynchrony protocol and were highest on the second forced desynchrony day (p < .01), returning towards forced desynchrony day 1 levels thereafter. ConclusionsThese findings have important implications for understanding factors contributing to altered glucose metabolism during sleep loss and circadian misalignment, and for potential physiological adaptation of metabolism in healthy adults, who are increasingly exposed to such conditions in our society.

Circadian- and wake-dependent influences on face-name memory in healthy men and women over 3 weeks of chronic sleep restriction

ObjectivesFacial recognition is one of the key functions of the human brain, and linking a face to a name is critical in many social and occupational settings. This study assessed circadian- and wake-dependent effects on face-name recognition in healthy adults. MethodsThirteen healthy adults (20-70years; 7 F) were studied in a 39-day inpatient protocol that included 3weeks of 28 hours forced desynchrony with sleep restriction (6.5:21.5 hours sleep:wake). Starting 3 hours after scheduled wake, 6 novel face-name pairs were presented every 4 waking hours; recognition was tested 2 hours later. Performance data were averaged across ∼4 hours circadian phase or time-awake bins. ResultsFace-name recognition deteriorated with increased time awake (p < .0001) and exhibited significant circadian variation (p < .0001), with worst performance shortly after the core temperature nadir. There was a significant interaction between sex and circadian phase (p = .0177), with women performing significantly better than men at all circadian phases except 60° and 120°. Women exhibited a significantly higher amplitude than men during the third week of forced desynchrony (p < .01). ConclusionsLike many other aspects of neurobehavioral performance, recalling face-name associations is impacted by both duration of time awake and circadian phase. These results have implications for face recognition testing in medical contexts, such as in testing for dementia, because performance may be impacted by sleep deficiency and the time of testing.

Spontaneous attentional failures reflect multiplicative interactions of chronic sleep loss with acute sleep loss and circadian misalignment

ObjectivesAcute and chronic sleep loss and circadian timing interact such that, depending on their combination, small or very large performance decrements are observed in tasks of attention. Here, we tested whether such nonlinear interactions extend to a physiological measure of spontaneous visual attentional failures, indicating a fundamental principle of sleep-wake regulation. MethodsNine healthy volunteers completed an in-laboratory 3-week forced desynchrony protocol consisting of 12 consecutive 42.85-hour cycles with a sleep-wake ratio of 1:3.3. The protocol induced increasing chronic sleep loss, while extended wake (32.85 hours) and sleep episodes (10 hours) occurred at multiple circadian phases. Attentional failure rate was quantified from continuous electrooculograms (number of 30-second epochs with slow eye movements/h of wakefulness) as a function of time since scheduled wake (acute sleep loss), week of study (chronic sleep loss), and circadian (melatonin) phase. ResultsDuring the first ∼8 hours awake, attentional failure rate was low, irrespective of the week. During the following wake hours, attentional failure rate increased steadily but at a faster rate in weeks 2 and 3 compared to week 1. The effects of acute and chronic sleep loss on attentional failure rate were magnified during the biological night compared to the biological day. ConclusionsA single extended sleep episode can only temporarily reverse attentional impairment associated with chronic sleep loss. Multiplicative effects of acute and chronic sleep loss—further amplified during the biological night—substantiate the interaction of 2 homeostatic response mechanisms and caution against underestimating their disproportionate combined impact on performance, health, and safety.

0008 The endogenous circadian system provides for enhanced coronary microvascular function in the morning

Abstract Introduction Major adverse cardiovascular events occur most commonly in the morning compared to other times of the day or night. Evaluation of coronary microvascular reserve perfusion can be used to reflect the anatomic and functional status of the coronary vascular system, both of which contribute to major adverse cardiovascular events. Prior work suggests that coronary microvascular function is attenuated in the morning, but whether the circadian system contributes to this attenuation is unknown. Methods To determine if the circadian system drives the morning attenuation in coronary microvascular function, we studied 13 healthy participants (8 females, 5 males; 42+/-13 years [mean+/-SD]) in a 5-day forced desynchrony protocol in dim light, where all behaviors and measurements were distributed across the 24-hour cycle. Coronary microvascular function was evaluated by vasodilator myocardial contrast echocardiography, sequentially 16 hours apart, corresponding to each participant’s morning, evening and night. We analyzed time-intensity data to calculate microvascular flux rate (beta) at rest and during hyperemia for each measurement. Circadian phases were calculated based on individual dim-light melatonin onsets. We grouped data into three ~8-hour circadian bins corresponding to each participant’s morning, evening and night. Mixed model analyses were used to determine if there were significant differences between data collected in the circadian morning compared to the other times while controlling for the measurement schedule and between-subject variability. Results There were no significant time-dependent differences in echocardiographic indices of ventricular function, including stroke volume, cardiac output, or myocardial work. Contrary to our hypothesis, hyperemic perfusion was lowest in the evening rather than the morning (2.49 /s vs. 2.82 /s, p=0.042) but not different between morning and night (also 2.82 /s, p=0.96). Conclusion In healthy people, there is no circadian-system-driven morning attenuation in coronary microvascular function: indeed, the circadian system facilitates coronary vascular flow in the morning, possibly to accommodate the anticipated behaviorally driven increases in myocardial demand after awakening. Support (if any) NIH KL2TR002370, R01HL163232, R35HL155681, R01HL140577, R01HL130046, R01HL078610, R01HL156948, K01HL146992, K01HL151745, ULTR00128, Sleep Research Society Foundation, Oregon Institute of Occupational Health Sciences.

Desynchronizing the sleep---wake cycle from circadian timing to assess their separate contributions to physiology and behaviour and to estimate intrinsic circadian period.

Circadian clocks drive cyclic variations in many aspects of physiology, but some daily variations are evoked by periodic changes in the environment or sleep-wake state and associated behaviors, such as changes in posture, light levels, fasting or eating, rest or activity and social interactions; thus, it is often important to quantify the relative contributions of these factors. Yet, circadian rhythms and these evoked effects cannot be separated under typical 24-h day conditions, because circadian phase and the length of time awake or asleep co-vary. Nathaniel Kleitman's forced desynchrony (FD) protocol was designed to assess endogenous circadian rhythmicity and to separate circadian from evoked components of daily rhythms in multiple parameters. Under FD protocol conditions, light intensity is kept low to minimize its impact on the circadian pacemaker, and participants have sleep-wake state and associated behaviors scheduled to an imposed non-24-h cycle. The period of this imposed cycle, Τ, is chosen so that the circadian pacemaker cannot entrain to it and therefore continues to oscillate at its intrinsic period (τ, ~24.15 h), ensuring circadian components are separated from evoked components of daily rhythms. Here we provide detailed instructions and troubleshooting techniques on how to design, implement and analyze the data from an FD protocol. We provide two procedures: one with general guidance for designing an FD study and another with more precise instructions for replicating one of our previous FD studies. We discuss estimating circadian parameters and quantifying the separate contributions of circadian rhythmicity and the sleep-wake cycle, including statistical analysis procedures and an R package for conducting the non-orthogonal spectral analysis method that enables an accurate estimation of period, amplitude and phase.

Contrasting Effects of Sleep Restriction, Total Sleep Deprivation, and Sleep Timing on Positive and Negative Affect.

Laboratory-based sleep manipulations show asymmetries between positive and negative affect, but say little about how more specific moods might change. We report extensive analyzes of items from the Positive and Negative Affect Scale (PANAS) during days following nights of chronic sleep restriction (6 h sleep opportunity), during 40 h of acute sleep deprivation under constant routine conditions, and during a week-long forced desynchrony protocol in which participants lived on a 28-h day. Living in the laboratory resulted in medium effects sizes on all positive moods (Attentiveness, General Positive Affect, Joviality, Assuredness), with a general deterioration as the days wore on. These effects were not found with negative moods. Sleep restriction reduced some positive moods, particularly Attentiveness (also General Positive), and increased Hostility. A burden of chronic sleep loss also led to lower positive moods when participants confronted the acute sleep loss challenge, and all positive moods, as well as Fearfulness, General Negative Affect and Hostility were affected. Sleeping at atypical circadian phases resulted in mood changes: all positive moods reduced, Hostility and General Negative Affect increased. Deteriorations increased the further participants slept from their typical nocturnal sleep. In most cases the changes induced by chronic or acute sleep loss or mistimed sleep waxed or waned across the waking day, with linear or various non-linear trends best fitting these time-awake-based changes. While extended laboratory stays do not emulate the fluctuating emotional demands of everyday living, these findings demonstrate that even in controlled settings mood changes systematically as sleep is shortened or mistimed.

Bright Light During Wakefulness Improves Sleep Quality in Healthy Men: A Forced Desynchrony Study Under Dim and Bright Light (III).

Under real-life conditions, increased light exposure during wakefulness seems associated with improved sleep quality, quantified as reduced time awake during bed time, increased time spent in non-rapid eye movement (NREM) sleep, or increased power of the electroencephalogram delta band (0.5-4 Hz). The causality of these important relationships and their dependency on circadian phase and/or time awake has not been studied in depth. To disentangle possible circadian and homeostatic interactions, we employed a forced desynchrony protocol under dim light (6 lux) and under bright light (1300 lux) during wakefulness. Our protocol consisted of a fast cycling sleep-wake schedule (13 h wakefulness—5 h sleep; 4 cycles), followed by 3 h recovery sleep in a within-subject cross-over design. Individuals (8 men) were equipped with 10 polysomnography electrodes. Subjective sleep quality was measured immediately after wakening with a questionnaire. Results indicated that circadian variation in delta power was only detected under dim light. Circadian variation in time in rapid eye movement (REM) sleep and wakefulness were uninfluenced by light. Prior light exposure increased accumulation of delta power and time in NREM sleep, while it decreased wakefulness, especially during the circadian wake phase (biological day). Subjective sleep quality scores showed that participants rated their sleep quality better after bright light exposure while sleeping when the circadian system promoted wakefulness. These results suggest that high environmental light intensity either increases sleep pressure buildup during wakefulness or prevents the occurrence of micro-sleep, leading to improved quality of subsequent sleep.

Bright Light Decreases Peripheral Skin Temperature in Healthy Men: A Forced Desynchrony Study Under Dim and Bright Light (II).

Human thermoregulation is strictly regulated by the preoptic area of the hypothalamus, which is directly influenced by the suprachiasmatic nucleus (SCN). The main input pathway of the SCN is light. Here, thermoregulatory effects of light were assessed in humans in a forced desynchrony (FD) design. The FD experiment was performed in dim light (DL, 6 lux) and bright white light (BL, 1300 lux) in 8 men in a semi-randomized within-subject design. A 4 × 18 h FD protocol (5 h sleep, 13 h wake) was applied, with continuous core body temperature (CBT) and skin temperature measurements at the forehead, clavicles, navel, palms, foot soles and toes. Skin temperature parameters indicated sleep-wake modulations as well as internal clock variations. All distal skin temperature parameters increased during sleep, when CBT decreased. Light significantly affected temperature levels during the wake phase, with decreased temperature measured at the forehead and toes and increased navel and clavicular skin temperatures. These effects persisted when the lights were turned off for sleep. Circadian amplitude of CBT and all skin temperature parameters decreased significantly during BL exposure. Circadian proximal skin temperatures cycled in phase with CBT, while distal skin temperatures cycled in anti-phase, confirming the idea that distal skin regions reflect heat dissipation and proximal regions approximate CBT. In general, we find that increased light intensity exposure may have decreased heat loss in humans, especially at times when the circadian system promotes sleep.

Bright Light Increases Alertness and Not Cortisol in Healthy Men: A Forced Desynchrony Study Under Dim and Bright Light (I).

Light-induced improvements in alertness are more prominent during nighttime than during the day, suggesting that alerting effects of light may depend on internal clock time or wake duration. Relative contributions of both factors can be quantified using a forced desynchrony (FD) designs. FD designs have only been conducted under dim light conditions (<10 lux) since light above this amount can induce non-uniform phase progression of the circadian pacemaker (also called relative coordination). This complicates the mathematical separation of circadian clock phase from homeostatic sleep pressure effects. Here we investigate alerting effects of light in a novel 4 × 18 h FD protocol (5 h sleep, 13 h wake) under dim (6 lux) and bright light (1300 lux) conditions. Hourly saliva samples (melatonin and cortisol assessment) and 2-hourly test sessions were used to assess effects of bright light on subjective and objective alertness (electroencephalography and performance). Results reveal (1) stable free-running cortisol rhythms with uniform phase progression under both light conditions, suggesting that FD designs can be conducted under bright light conditions (1300 lux), (2) subjective alerting effects of light depend on elapsed time awake but not circadian clock phase, while (3) light consistently improves objective alertness independent of time awake or circadian clock phase. Reconstructing the daily time course by combining circadian clock phase and wake duration effects indicates that performance is improved during daytime, while subjective alertness remains unchanged. This suggests that high-intensity indoor lighting during the regular day might be beneficial for mental performance, even though this may not be perceived as such.

0226 Circadian influence on food intake among adolescents with overweight and healthy weight

Abstract Introduction Meal timing has been linked to obesity in adults and children; however, evidence for an endogenous influence of the circadian system on food intake is unknown. We measured food intake during forced desynchrony (FD) in adolescents with healthy weight (HW) or overweight (OW), hypothesizing that circadian timing would affect food intake independent of the environmental cycle and that the food intake rhythm would be delayed in adolescents with OW compared to HW. Methods Participants were 51 (29m) adolescents (12-15yr); 24 were HW and 27 OW determined by CDC norms. Participants completed seven, 28h-FD cycles; 6 meals occurred at fixed times each cycle with foods selected ~1h before meals and weighed before and after. Each meal’s energy intake was computed relative to total energy consumed in that FD cycle. Endogenous circadian period was determined using salivary dim-light melatonin onset (DLMO) phases (DLMO=0°). Time awake effect was assessed using mixed effect models and circadian phase with multilevel cosinor that included time awake as a categorical covariate. Results Circadian period was not significantly different between OW and HW (mean, StDev; HW =24.19h,0.22; OW=24.22h,0.14). Overall, participants consumed more calories on the cycles’ first meals (22.0% [21.3;22.6]) compared to the last meal (12.4% [11.8;13.0]). Adolescents with OW vs. HW consumed a higher proportion of calories later in the wake episode (F(5,2081)=2.63,p=.02). As hypothesized, circadian phase influenced caloric intake with an amplitude of 2.66% [2.19;3.14] percent daily calories and an acrophase of 301°[291;310], equivalent to ~17:52 in this population. The circadian influence differed by weight category (likelihood ratio test of both amplitude and acrophase; χ2(2)=10.7,p&amp;lt;.01), with those with OW showing a lower amplitude (OW = 2.11%[1.40;2.82], HW=3.53%[2.94;4.12] and later acrophase (OW=301° [287;314], HW=290° [278;302]). Conclusion Results show for the first time an independent influence of the endogenous circadian timing system on caloric intake of humans that differed as a function of body weight: caloric intake for adolescents with OW had a circadian rhythm with blunted amplitude and delayed peak phase. These observations show circadian control of food intake that may be stronger in HW than OW adolescents. Support (If Any) P20GM139743; R01DK101046; R01HL153969

High dose melatonin increases sleep duration during nighttime and daytime sleep episodes in older adults.

Aging is associated with changes in sleep, and improving sleep may have important consequences for the health, cognition, and quality of life of older adults. Many prescription sleep aids increase the risk of nighttime falls, have adverse effects on next-day cognition, and are associated with increased mortality. Melatonin, a hormone secreted at night, increases sleep duration in young adults but only when administered during the day when endogenous levels are low. In a month-long cross-over study, we randomized 24 healthy older (age >55, mean 64.2 ± 6.3 years) participants to receive 2 weeks of placebo and 2 weeks of either a low (0.3 mg) or high (5.0 mg) dose of melatonin 30 min before lights out. Sleep was polysomnographically recorded and was scheduled during both the biological day and night using a forced desynchrony design. Although0.3 mg melatonin had a trend towards increasing sleep efficiency (SE) overall, this was due to its effects on sleep during the biological day. In contrast, 5 mg melatonin significantly increased SE during both biological day and night, mainly by increasing the duration of Stage 2 non-rapid eye movement sleepand slightly shortening awakenings. Melatonin should be further explored as a sleep aid for older adults.

Proof-of-principle demonstration of endogenous circadian system and circadian misalignment effects on human oral microbiota.

Circadian misalignment-the misalignment between the central circadian "clock" and behavioral and environmental cycles (including sleep/wake, fasting/eating, dark/light)-results in adverse cardiovascular and metabolic effects. Potential underlying mechanisms for these adverse effects include alterations in the orogastrointestinal microbiota. However, it remains unknown whether human oral microbiota has endogenous circadian rhythms (i.e., independent of sleep/wake, fasting/eating, and dark/light cycles) and whether circadian misalignment influences oral microbiota community composition. Healthy young individuals [27.3±2.3years (18-35years), 4 men and 2 women, body-mass index range: 18-28kg/m2 ] were enrolled in a stringently controlled 14-day circadian laboratory protocol. This included a 32-h constant routine (CR) protocol (endogenous circadian baseline assessment), a forced desynchrony protocol with four 28-h "days" under ~3lx to induce circadian misalignment, and a post-misalignment 40-h CR protocol. Microbiota assessments were performed on saliva samples collected every 4h throughout both CR protocols. Total DNA was extracted and processed using high-throughput 16S ribosomal RNA gene amplicon sequencing. The relative abundance of specific oral microbiota populations, i.e., one of the five dominant phyla, and three of the fourteen dominant genera, exhibited significant endogenous circadian rhythms. Importantly, circadian misalignment dramatically altered the oral microbiota landscape, such that four of the five dominant phyla and eight of the fourteen dominant genera exhibited significant circadian misalignment effects. Moreover, circadian misalignment significantly affected the metagenome functional content of oral microbiota (inferred gene content analysis), as indicated by changes in specific functional pathways associated with metabolic control and immunity. Collectively, our proof-of-concept study provides evidence for endogenous circadian rhythms in human oral microbiota and show that even relatively short-term experimental circadian misalignment can dramatically affect microbiota community composition and functional pathways involved in metabolism and immune function. These proof-of-principle findings have translational relevance to individuals typically exposed to circadian misalignment, including night shift workers and frequent flyers.

Chronic Sleep Restriction While Minimizing Circadian Disruption Does Not Adversely Affect Glucose Tolerance

Insufficient sleep, which has been shown to adversely affect metabolism, is generally associated with prolonged exposure to artificial light at night, a known circadian disruptor. There is growing evidence suggesting that circadian disruption adversely affects metabolism, yet few studies have attempted to evaluate the adverse metabolic effects of insufficient sleep while controlling for circadian disruption. We assessed postprandial glucose and insulin responses to a standard breakfast meal in healthy adults (n = 9) who underwent 3 weeks of chronic sleep restriction (CSR) in a 37-day inpatient study while minimizing circadian disruption by maintaining the same duration of light exposure each study day. We compared these results to findings from an earlier inpatient study which used a forced desynchrony (FD) protocol to assess the influence of 3 weeks of CSR combined with recurrent circadian disruption (RCD) on glycemic control in healthy adults (n = 21). CSR combined with RCD resulted in significantly elevated postprandial plasma glucose levels (p < 0.0001), while CSR with minimized circadian disruption had no adverse glycemic effects after 3 weeks of exposure (EXP). These results suggest that one mechanism by which sleep restriction impacts metabolism may be via concurrent circadian disruption.

Behaviorally and environmentally induced non-24-hour sleep-wake rhythm disorder in sighted patients.

To determine whether there was evidence of circadian or sleep-regulatory dysfunction in sighted individuals with non-24-hour sleep-wake rhythm disorder. Three sighted individuals with signs and/or symptoms of non-24-hour sleep-wake rhythm disorder were studied. Thirty-five- to 332-day laboratory and home-based assessments of sleep-wake and circadian timing, endogenous circadian period, photic input to the circadian pacemaker, and/or circadian and sleep-wake-dependent regulation of sleep were conducted. No evidence of circadian dysfunction was found in these individuals. Instead, sleep-wake timing appeared to dissociate from the circadian timing system, and/or self-selected sleep-wake and associated light/dark timing shifted the circadian pacemaker later, rather than the circadian pacemaker determining sleep-wake timing. These findings suggest that the etiology of this disorder may be light- and/or behaviorally induced in some sighted people, which has implications for the successful treatment of this disorder. Emens JS, St Hilaire MA, Klerman EB, etal. Behaviorally and environmentally induced non-24-hour sleep-wake rhythm disorder in sighted patients. J Clin Sleep Med. 2022;18(2):453-459.

A classification approach to estimating human circadian phase under circadian alignment from actigraphy and photometry data.

The time of dim light melatonin onset (DLMO) is the gold standard for circadian phase assessment in humans, but collection of samples for DLMO is time and resource-intensive. Numerous studies have attempted to estimate circadian phase from actigraphy data, but most of these studies have involved individuals on controlled and stable sleep-wake schedules, with mean errors reported between 0.5 and 1hour. We found that such algorithms are less successful in estimating DLMO in a population of college students with more irregular schedules: Mean errors in estimating the time of DLMO are approximately 1.5-1.6hours. We reframed the problem as a classification problem and estimated whether an individual's current phase was before or after DLMO. Using a neural network, we found high classification accuracy of about 90%, which decreased the mean error in DLMO estimation-identifying the time at which the switch in classification occurs-to approximately 1.3hours. To test whether this classification approach was valid when activity and circadian rhythms are decoupled, we applied the same neural network to data from inpatient forced desynchrony studies in which participants are scheduled to sleep and wake at all circadian phases (rather than their habitual schedules). In participants on forced desynchrony protocols, overall classification accuracy dropped to 55%-65% with a range of 20%-80% for a given day; this accuracy was highly dependent upon the phase angle (ie, time) between DLMO and sleep onset, with the highest accuracy at phase angles associated with nighttime sleep. Circadian patterns in activity, therefore, should be included when developing and testing actigraphy-based approaches to circadian phase estimation. Our novel algorithm may be a promising approach for estimating the onset of melatonin in some conditions and could be generalized to other hormones.

076 Bright light during wakefulness improves objective and subjective sleep quality: a forced desynchrony study

Abstract Introduction Under real life conditions, increased light exposure during wakefulness seems associated with improved sleep quality, quantified as reduced time awake during bed time, increased time spent in non-REM (NREM) sleep or increased power in the EEG delta band (0.5–4 Hz). The causality of these important relationships and their dependency on circadian clock phase and/or time awake has not been studied in depth. To establish causality of light effects during wake time on subsequent sleep, and to disentangle possible circadian and homeostatic interactions, we employed a forced desynchrony (FD) protocol under dim light (6.5 lux) and bright light (1307 lux) during wakefulness. Methods The protocol consisted of a fast cycling sleep-wake schedule (13h wakefulness – 5h sleep; 4 cycles), followed by 3h recovery sleep in a within subject cross-over design. Individuals (7 men) were equipped with 10 polysomnography electrodes. Subjective sleep quality was measured immediately after wakening. Results Results indicated that circadian variation in delta power was only detected under dim light. Circadian variation in time in rapid eye movement (REM) sleep and wakefulness were uninfluenced by light. Prior light exposure increased accumulation of delta power and time in NREM sleep, while decreasing wakefulness, especially during the circadian wake phase. Subjective sleep quality scores showed that participants were only able to assess light induced improvement of sleep quality correctly when the circadian system promoted wakefulness. Conclusion This study presents significant effects of bright light exposure on sleep architecture, leading to sleep pressure related changes in objective sleep quality. At the end of the scheduled sleep phase after increased light exposure, more delta power and NREM sleep were detected, especially when sleep occurred outside the normal sleep phase. Subjective sleep quality scores showed light-induced improvements coinciding with increased delta power and time spend in NREM sleep, suggesting that light during wakefulness may improve subsequent sleep quality. These findings may have important implications for insomnia treatment and clinical applications of light therapy. Support (if any) This research was funded by the University of Groningen Campus Fryslân (Grant No. 01110939; co-financed by Philips Drachten and Provincie Fryslân). Additional financial support was obtained from a NWO-STW Program Grant “OnTime” (project 12185).

077 Human activity levels reflect circadian influences independent of sleep/wake behavior

Abstract Introduction Actigraphy is a non-invasive method that allows long-term recordings of activity, light, and other variables in diverse environments. In real-world settings, activity usually has a 24-hour rhythm that may arise from sleep/wake-associated behavior and/or circadian rhythmicity. We tested for an independent circadian component using data from people living on non-24 hours “days” in the laboratory. Methods Data are from five inpatient studies with tightly-controlled forced desynchrony (FD) conditions. Participants (19–34 yo) were healthy by history, physical exam, laboratory tests of blood and urine, and clinical polysomnography, and did not report using prescription medicines. Caffeine-containing substances were prohibited during the study. Protocol 1: 7 participants (3 F) T-cycle (i.e., FD sleep-wake cycle duration) = 42.85h; Rest:Activity ratio 1:3.3. Protocol 2: 8 participants (3 F) T cycle =42.85h; Rest:Activity 1:2. Protocol 3: 9 participants (3 F) T cycle =28.0h; Rest:Activity 1:2. Protocol 4: 7 participants (3 F) T cycle =20.0h; Rest:Activity ratio 1:3.3. Protocol 5: 7 participants (5 F) T cycle =20.0h; Rest:Activity 1:2. At all times except during showers, participants wore an actiwatch that measured activity levels and light. Melatonin period and phase 0 (i.e., fit maximum) were computed using non-orthogonal spectral analyses. Data were analyzed relative to 3-hr Circadian Phase bins (1/8 of computed circadian period for each individual) and 3-hr Wake Duration bins. Activity data were summarized using Zero-Inflated-Poison-based statistics for each Circadian*Wake Duration bin for each individual and then across individuals within each study. Repeated measures ANOVA were conducted. Statistics were performed using SAS. Results For all protocols, there were significant differences (all p&amp;lt;0.007) by individual participant, by Circadian Phase, and by Wake Duration bin, but not by the interaction term (Circadian Phase* Wake Duration). Highest levels of activity were at Circadian Phase 7.5–10.5 (~10am–1pm) and lowest values at Circadian Phase -1.5–1.5 (~midnight–3 am). Activity values were lowest at scheduled sleep times. Conclusion Circadian rhythms independent of sleep/wake behaviors influence activity levels and may be an important component of analyses. In individuals living on non-24-hr days (e.g., some blind people and some sighted people with Non-24-hr Sleep Disorder), it may be possible to derive circadian-based metrics. Support (if any) NIH K24-HL105664, P01-AG009975, T32-HL007901, K01-HL146992.

Unacylated ghrelin, leptin, and appetite display diurnal rhythmicity in lean adults.

Constant routine and forced desynchrony protocols typically remove the effects of behavioral/environmental cues to examine endogenous circadian rhythms, yet this may not reflect rhythms of appetite regulation in the real world. It is therefore important to understand these rhythms within the same subjects under controlled diurnal conditions of light, sleep, and feeding. Ten healthy adults (9 M/1 F, means ±SD: age, 30 ± 10 yr; body mass index, 24.1 ± 2.7 kg·m-2) rested supine in the laboratory for 37 h. All data were collected during the final 24 h of this period (i.e., 0800-0800 h). Participants were fed hourly isocaloric liquid meal replacements alongside appetite assessments during waking before a sleep opportunity from 2200 to 0700 h. Hourly blood samples were collected throughout the 24-h period. Dim light melatonin onset occurred at 2318 ± 46 min. A diurnal rhythm in mean plasma unacylated ghrelin concentration was identified (P = 0.04), with the acrophase occurring shortly after waking (0819), falling to a nadir in the evening with a relative amplitude of 9%. Plasma leptin concentration also exhibited a diurnal rhythm (P < 0.01), with the acrophase occurring shortly after lights-out (0032 h) and the lowest concentrations at midday. The amplitude for this rhythm was 25%. Diurnal rhythms were established in all dimensions of appetite except for sweet preference (P = 0.29), with both hunger (2103 h) and prospective food consumption (1955 h) reaching their peak in the evening before falling to their nadir shortly after waking. Under controlled diurnal conditions, simultaneous measurement of leptin, unacylated ghrelin, and subjective appetite over a 24-h period revealed rhythmicity in appetite regulation in lean, healthy humans.NEW & NOTEWORTHY Simultaneous assessment of subjective appetite, unacylated ghrelin, and leptin was carried out over a continuous 37-h protocol for the first time under conditions of controlled light, sleep, and feeding in healthy, lean adults. Rhythms were observed in unacylated ghrelin, leptin, and components of subjective appetite, such as hunger, prospective consumption, and fullness. Concurrent measurement of rhythms in these variables is important to fully understand the temporal relationships between components of appetite as well as the influence of diurnal factors such as sleep, light, and feeding.

0257 Preliminary Findings: Attentional Bias for Food Cues Unrelated to Time Awake or Circadian Phase During Forced Desynchrony in Adolescents

Abstract Introduction Sleep quantity and quality influence attentional bias, and attentional bias for food cues has been associated with body weight. Whether the endogenous circadian cycle and/or time from awakening to bedtime contribute to attentional bias for food and differences in weight remain unclear. Using a 28-h forced desynchrony (FD) design, we hypothesized that adolescents with overweight (OW) and obesity (O) would have more attentional bias for food cues later in the wake episode and at a later circadian phase compared to adolescents with a healthy weight (HW). Methods 50 (28 male) adolescents (12-15yr) completed 7 FD cycles. Participants completed an attentional bias task that included three food word categories (savory, sweet, fruits/vegetables) and one non-food word category (school supplies). The task was completed at 6 fixed times each cycle: Task 1 was 1.3h after scheduled awaking, Task 2 was 2h after Task 1, and Tasks 3–6 followed at 3-h intervals. Weight categorization used body mass index (BMI) percentiles (CDC): HW (&amp;gt;5th and &amp;lt;85th; n=24), OW (85th and &amp;lt;95th; n=13), or O (≥95th; n=14). Endogenous circadian period was determined using salivary melatonin onsets (Mean: HW=23.88h; OW=24.01h; O=23.86h). Effect of circadian phase and time since scheduled awakening was assessed by mixed effects modeling using 6 circadian and 6 time-awake bins. Results We found no significant differences between weight groups in attentional bias for any of the food categories (p’s &amp;gt;.05). We also saw no significant time awake effects or circadian influence on attentional bias, nor did time awake or circadian phase moderate the associations between weight category and attentional bias (p’s &amp;gt;.05). Conclusion Weight groups did not impact food-related attentional bias across the wake episode or circadian phase. Future directions will explore whether attentional bias for food types predicts food choice and food consumption in this study. Support DK101046