Using an experimental design we photographed the faces of 23 adults (mean age 23, range 18-31 years, 11 women) between 14.00 and 15.00 under two conditions in a balanced design: after a normal night’s sleep (at least eight hours of sleep between 23.00-07.00 and seven hours of wakefulness) and after sleep deprivation (sleep 02.00-07.00 and 31 hours of wakefulness). We advertised for participants at four universities in the Stockholm area. Twenty of 44 potentially eligible people were excluded. Reasons for exclusion were reported sleep disturbances, abnormal sleep requirements (for example, sleep need out of the 7-9 hour range), health problems, or availability on study days (the main reason). We also excluded smokers and those who had consumed alcohol within two days of the protocol. One woman failed to participate in both conditions. Overall, we enrolled 12 women and 12 men.
The participants slept in their own homes. Sleep times were confirmed with sleep diaries and text messages. The sleep diaries (Karolinska sleep diary) included information on sleep latency, quality, duration, and sleepiness. Participants sent a text message to the research assistant by mobile phone (SMS) at bedtime and when they got up on the night before sleep deprivation. They had been instructed not to nap. During the normal sleep condition the participants’ mean duration of sleep, estimated from sleep diaries, was 8.45 (SE 0.20) hours. The sleep deprivation condition started with a restriction of sleep to five hours in bed; the participants sent text messages (SMS) when they went to sleep and when they woke up. The mean duration of sleep during this night, estimated from sleep diaries and text messages, was 5.06 (SE 0.04) hours. For the following night of total sleep deprivation, the participants were monitored in the sleep laboratory at all times. Thus, for the sleep deprivation condition, participants came to the laboratory at 22.00 (after 15 hours of wakefulness) to be monitored, and stayed awake for a further 16 hours. We therefore did not observe the participants during the first 15 hours of wakefulness, when they had had a slightly restricted sleep, but had good control over the last 16 hours of wakefulness when sleepiness increased in magnitude. For the sleep condition, participants came to the laboratory at 12.00 (after five hours of wakefulness). They were kept indoors two hours before being photographed to avoid the effects of exposure to sunlight and the weather. We had a series of five or six photographs (resolution 3872×2592 pixels) taken in a well lit room, with a constant white balance (×900l; colour temperature 4200 K, Nikon D80; Nikon, Tokyo). The white balance was differently set during the two days of the study and affected seven photographs (four taken during sleep deprivation and three during a normal night’s sleep). Removing these participants from the analyses did not affect the results. The distance from camera to head was fixed, as was the focal length, within 14 mm (between 44 and 58 mm). To ensure a fixed surface area of each face on the photograph, the focal length was adapted to the head size of each participant.
For the photo shoot, participants wore no makeup, had their hair loose (combed backwards if long), underwent similar cleaning or shaving procedures for both conditions, and were instructed to “sit with a straight back and look straight into the camera with a neutral, relaxed facial expression.” Although the photographer was not blinded to the sleep conditions, she followed a highly standardised procedure during each photo shoot, including minimal interaction with the participants. A blinded rater chose the most typical photograph from each series of photographs. This process resulted in 46 photographs; two (one from each sleep condition) of each of the 23 participants. This part of the study took place between June and September 2007.
In October 2007 the photographs were presented at a fixed interval of six seconds in a randomised order to 65 observers (mainly students at the Karolinska Institute, mean age 30 (range 18-61) years, 40 women), who were unaware of the conditions of the study. They rated the faces for attractiveness (very unattractive to very attractive), health (very sick to very healthy), and tiredness (not at all tired to very tired) on a 100 mm visual analogue scale. After every 23 photographs a brief intermission was allowed, including a working memory task lasting 23 seconds to prevent the faces being memorised. To ensure that the observers were not primed to tiredness when rating health and attractiveness they rated the photographs for attractiveness and health in the first two sessions and tiredness in the last. To avoid the influence of possible order effects we presented the photographs in a balanced order between conditions for each session.
Data were analysed using multilevel mixed effects linear regression, with two crossed independent random effects accounting for random variation between observers and participants using the xtmixed procedure in Stata 9.2. We present the effect of condition as a percentage of change from the baseline condition as the reference using the absolute value in millimetres (rated on the visual analogue scale). No data were missing in the analyses.