Circadian Rhythms and Time Perception in Healthy Adults During Constant Wakefulness

Background and Rationale Daily experience of time does not always match clock time: under some conditions time seems to pass faster or slower, and people may misjudge short intervals or deadlines. At the same time, human physiology and cognition are strongly regulated by circadian rhythms, which interact with the build-up of sleep pressure during extended wakefulness. However, it is not well understood how the internal circadian phase and increasing sleepiness jointly shape time perception and related cognitive functions.

The "constant routine" (CR) protocol is a well-established method in circadian research to unmask endogenous rhythmicity. By keeping environmental and behavioural factors as constant as possible while participants remain awake in a semi-recumbent position, the protocol minimises external masking and allows changes over time to be attributed primarily to the underlying circadian system and homeostatic sleep pressure.

Objectives The primary objective is to characterise how time perception (e.g., subjective passage of time and estimation/production of short intervals) varies across the circadian cycle during 36 hours of constant wakefulness. Secondary objectives are to examine how these changes relate to: 1) Biological markers of circadian phase (salivary melatonin), 2) Subjective sleepiness and mood, 3) Objective alertness and vigilance (reaction time), and 4) Simple cognitive decisions and perceptual judgements (e.g., responses to numerical information and colour adjustments).

Study Design This is a single-centre, interventional, basic science study using a within-subject, single-group assignment. Healthy adults attend a screening and briefing visit, followed by one in-laboratory constant-routine session of approximately 36 hours of continuous wakefulness. During the CR, environmental conditions (light, temperature, noise) and posture are controlled, and participants receive small, isocaloric snacks at regular intervals. No drugs, devices, or randomised treatment arms are used; the "intervention" is prolonged wakefulness under constant-routine conditions.

Procedures After eligibility screening and consent, participants complete questionnaires on sleep habits and general health and are instructed to maintain a regular sleep-wake schedule prior to the laboratory visit. For the main CR session, participants arrive at the sleep laboratory in the morning. From that point onward, they remain awake in a controlled setting for about 36 hours. Light is kept at a low, constant level, physical activity is restricted, and posture is standardised as far as feasible.

Every two hours, participants perform a structured test block lasting approximately 45-60 minutes. This block includes: 1) Subjective ratings of sleepiness and mood, 2) A psychomotor vigilance task (reaction-time task), 3) Brief tasks assessing subjective passage of time and the ability to estimate or produce short time intervals, 4) Simple decision-making and estimation tasks, and 5) Perceptual judgements such as colour adjustments.

At regular intervals across the CR, saliva samples are collected to measure melatonin, providing an estimate of individual circadian phase. Vital signs and adverse events are monitored throughout. After the CR, participants are debriefed and provided with recovery sleep arrangements according to local safety procedures.

Outcomes and Analysis Primary outcomes are measures of time perception (e.g., ratings of how fast or slow time seems to pass and accuracy of time interval estimation/production) modelled as a function of circadian phase and hours awake. Secondary outcomes include reaction-time performance, subjective sleepiness and mood ratings, decision-making measures, and colour perception indices. Salivary melatonin profiles are used to determine dim-light melatonin onset (DLMO) and circadian phase position.

Analyses will examine rhythmic changes across the 36-hour protocol and test whether distortions in time perception align with circadian night and with rising sleep pressure. The study is exploratory/basic science in nature and is not designed to evaluate a clinical treatment. Findings are expected to advance understanding of how biological time and psychological time interact, with potential implications for scheduling of shift work, transportation, and other situations requiring sustained wakefulness.

Inclusion criteria

• Age 23 to 45 years.
• Able and willing to provide written informed consent.
• Fluent in Danish and able to understand study procedures and instructions.
• Generally healthy, as assessed by medical history, screening questionnaires, and basic clinical measures (e.g., blood pressure, heart rate).
• Self-reported regular sleep-wake schedule for at least 4 weeks prior to the laboratory visit (typically 6.5-9 hours of sleep per night, with usual sleep period between approximately 22:00-01:00 and 06:00-09:00).
• Body mass index (BMI) within a non-extreme range (for example, approximately 18.5-30 kg/m²), if required by the study physician.
• No regular night work or rotating shift work during the 3 months before participation.
• No travel across more than 2 time zones in the 2 months before the constant-routine session.
• Willing to abstain from caffeine, nicotine, alcohol, and recreational drugs for the specified washout periods before and during the 36-hour constant-routine session.
• For participants who can become pregnant: negative pregnancy test at screening/arrival and agreement to use reliable contraception for the duration of participation.

Exclusion criteria

• Any known or suspected major sleep disorder (e.g., insomnia disorder, obstructive sleep apnoea, restless legs syndrome, narcolepsy), based on self-report or prior diagnosis.
• Current or past major psychiatric or neurological disorders (e.g., major depressive disorder, bipolar disorder, psychotic disorders, epilepsy), unless considered mild and stable and explicitly approved by the study physician.
• Chronic medical conditions that could be worsened by prolonged wakefulness or that might confound outcome measures, such as significant cardiovascular disease, uncontrolled hypertension, diabetes mellitus, severe respiratory disease, or other serious systemic illness.
• Regular use of medications or supplements that may affect sleep, circadian rhythms, melatonin secretion, alertness, or mood (e.g., hypnotics, sedative-hypnotics, melatonin, stimulants, certain antidepressants or beta-blockers), unless a safe washout is possible and approved by the study physician.
• High habitual caffeine intake (for example, >400 mg/day) or nicotine dependence if the participant is unable or unwilling to abstain for the required washout periods.
• Current harmful alcohol use or substance use disorder, or frequent use of recreational drugs.
• Pregnancy or breastfeeding.
• Previous severe adverse reaction to sleep deprivation, extended wakefulness, or similar laboratory protocols.
• Claustrophobia or inability to tolerate prolonged stays in a controlled laboratory environment.
• Any condition or circumstance that, in the judgement of the investigators, would make participation unsafe, interfere with the 36-hour wakefulness protocol, or compromise data quality (e.g., inability to remain awake despite support, strong fear of needles or saliva sampling, or inability to comply with study restrictions).