Orexin-A is hypothesized to promote wakefulness, and we examined whether cerebrospinal fluid (CSF) orexin-A levels are higher during the waking period in man. Within-subjects, repeated-measures design with balanced ordering of sampling at approximately 5 AM and 5 PM. Eight healthy young males. CSF orexin-A levels and standard polysomnography. Orexin-A levels during the sleep period were 4% higher than during the waking period (314.9 pg/ml versus 302.8 pg/ml, p <0.03). Sleep period orexin-A levels were negatively correlated with REM sleep as a percentage of total sleep time (p <0.05). The day and night levels of orexin-A were strongly correlated within subjects (r = 0.97; p < 0.0001) even though the samples were collected 1–2 weeks apart. Orexin-A levels in lumbar CSF are slightly higher at 5 AM than at 5 PM. Because orexin release is thought to be highest during the waking period, this observation was unexpected and may reflect a long delay between the release of orexin and its appearance in lumbar CSF. Orexin-A levels vary moderately between subjects, but are quite consistent within the same subject. Thus, for the diagnostic evaluation of narcolepsy, the time of CSF collection should have little impact.