The serotonergic system has repeatedly been discussed to be involved in the pathophysiology of fibromyalgia (FM), which is a syndrome of widespread pain and sleep disturbance. Elevated levels of substance P (SP), a mediator of nociception, have been described in FM. In this study the possible relationship between SP and serotonin (5-HT) together with its precursor tryptophan (TRP) and its metabolite 5-hydroxyindoleacetic acid (5-HIAA) was evaluated in 51 serum samples of fibromyalgia patients. These parameters were compared with clinical data such as pain intensity or sleep quality. A strong negative correlation between SP and 5-HIAA (P = .000) as well as between SP and TRP (P = .009) could be demonstrated. High serum concentrations of 5-HIAA and TRP showed a significant relation to low pain scores (5-HIAA: P = .030; TRP: P = .014). Moreover, 5-HIAA was strongly related to good quality of sleep (P = .000), while SP was related to sleep disturbance (P = .005). These data are valid to support the hypothesis of a systemic involvement of 5-HT and SP in fibromyalgia.