Data from animal studies suggest that serotonin release promotes wakefulness and suppresses REM sleep, but there are dangers in extrapolating these findings to humans. Binding of the radioligand [18F]MPPF to 5HT1A receptors is sensitive to levels of endogenous serotonin. In this study, we aimed to demonstrate changes in serotonin receptor availability in the human brain in wakefulness and sleep using [18F]MPPF and positron emission tomography. 14 subjects with narcolepsy cataplexy underwent [18F]MPPF PET scans in wakefulness and in sleep. Subjects who used the stimulant methylphenidate took their normal medication for the wake scan but omitted it prior to the sleep scan. The change in binding potential (BP) between the sleep and wake scans was examined using paired t test. Methylphenidate is thought to have little or no effect on serotonergic neurotransmission, and in order to confirm the absence of an effect on [18F]MPPF binding, a concurrent study was performed using a beta-microprobe technique to examine the effect of methylphenidate administration on [18F]MPPF binding in Sprague-Dawley rats. The human study showed a significant increase in [18F]MPPF binding in sleep compared to wakefulness in the whole brain and all regions of interest examined (temporal cortex, mesial temporal region and cingulate cortex). The beta-microprobe study confirmed that methylphenidate administration had no effect on [18F]MPPF binding. These findings indicate that serotonin receptor availability is increased in sleep compared to wakefulness in narcoleptic humans.