The function of sleep in physiology, behaviour and cognition has become a primary focus of neuroscience. Its study inevitably includes experimental sleep deprivation designs. However, concerns exist regarding confounds like stress, increased locomotor activity levels, and decreased motivation to perform operant tasks induced by the methods employed. We here propose a novel procedure for sleep deprivation in rats and evaluate how it affects sleep, corticosterone concentration profiles, locomotor activity levels, and motivation to perform an operant task. Before, during and after 12. h of total sleep deprivation by means of gradually increasing the rotation variability and the speed of a novel automated, two-compartment sleep deprivation device, sleep-wake states were assessed by electroencephalography (n= 21), brain extracellular corticosterone concentrations using microdialysis (n= 11), locomotor activity by infrared measurements (n= 8), and operant performance using a fixed-interval-fixed-ratio task (n= 16). Sleep was effectively prevented during the procedure; rats on average slept less than 1% of the time (0.8 ± 0.2%, mean ± standard error). Brain corticosterone concentrations were mildly increased during the procedure, but did not exceed normal peak concentrations. Locomotor activity was not only increased during the procedure, but also did not exceed the peak levels found during undisturbed wakefulness. Food restriction to 12. g/rat/day prevented sleep deprivation from reducing the motivation to perform an operant task. This novel procedure can be applied to sleep deprive rats in a highly effective way, while keeping corticosterone and locomotor activity within the normal range. © 2011 Elsevier B.V.