Serotonin (5-HT) signaling in the central nervous system (CNS) helps to regulate a variety of important cognitive and behavioral processes and it is a common therapeutic target for mood disorders. Because sleep abnormalities are frequently associated with mood disorders, there has been substantial interest in the regulatory abilities of 5-HT signaling on the sleep/wake cycle. However, to date there have been few practical and reliable ways to reversibly manipulate brain 5-HT levels without disrupting other monoaminergic signaling pathways that may be important for sleep.
In this issue of European Journal of Neuroscience, Nakamaru-Ogiso and colleagues reveal a new method for reducing brain 5-HT levels in rats, a well-established rodent model of sleep–wake architecture. Intraperitoneal injections of the hemoprotein enzyme tryptophan side chain oxidase I (TSOI) transiently reduce brain and peripheral 5-HT concentrations by reversibly depleting the rats of tryptophan, while preserving catecholeaminergic signaling. The authors report that this transient reduction of brain 5-HT abolishes the sleep/wake rhythm but has no meaningful influences on daily sleep amount. Moreover, the circadian rhythm in brain temperature is preserved in TSOI-injected rats, providing evidence that the effects of the manipulation are specific to sleep and are not caused by global effects on circadian timing. These findings suggest that in addition to its well-established regulatory influences on central circadian timing, brain 5-HT also plays a more direct role in the specific regulation of the sleep/wake rhythm.
The lack of practical methods to rapidly and reversibly manipulate brain 5-HT in mammals has been an obstacle in our understanding of the role of 5-HT signaling in sleep. Tryptophan-hydroxylase 2 (TPH2), the rate-limiting enzyme in 5-HT synthesis in the brain, has been a dependable target for brain 5-HT reduction; however, a lack of specificity of TPH2 inhibitors results in the collateral reduction of catecholamines such as the sleep/wake regulator norepinephrine, making these types of agents impractical for sleep studies. Serotonergic neurotoxins and TPH2 molecular deletions in mice have also been valuable to uncover the specific roles of 5-HT signaling, but neither manipulation is reversible, giving them limited usefulness in in vivo sleep experiments. Nakamaru-Ogiso and colleagues report that TSOI eliminates tryptophan and reduces brain 5-HT levels to 30% of controls within 12 h of treatment, with no collateral reductions in catecholeamines, other amino acids or protein synthesis. These influences of TSOI injection are no longer observed 96 h after injection. Consequently, this manipulation is faster and easier to control than the more traditional method of tryptophan depletion, the tryptophan-free diet; and it is specific to indoleamine synthesis, making it more practical for sleep experiments than manipulation with agents that non-specifically alter catecholeamines.
Another obstacle in examinations of the role of 5-HT signaling on sleep is its fundamental role in circadian timing, particularly on the entrainment of circadian rhythms by light (Ehlen et al., 2001). The mammalian circadian timing system is a primary sleep regulator and observations of 5-HT sleep regulatory properties have rarely ruled out the involvement of the central circadian pacemaker. Nakamaru-Ogiso and colleagues report that TSOI treatment temporarily eliminates the sleep–wake rhythm in rats by reducing total sleep amount during the rest phase and increasing it during the active phase. Consequently, it has no cumulative effect on 24-h total sleep amount. TSOI injection also increased sleep/wake fragmentation, which is commonly reported in manipulations that disrupt central circadian timing. This observation suggests that the disruption of the sleep/wake rhythm is a secondary effect of TSOI treatment on the central circadian pacemaker. However, the authors also report that the pacemaker-driven brain temperature rhythm remains intact, providing evidence that TSOI is acting downstream of the central circadian pacemaker. These findings are consistent with an earlier study by Kawai et al. (1994) who reported that tryptophan depletion disrupts the circadian wheel-running rhythm in rats. Taken together, these studies suggest that 5-HT may play an important role in coupling the central circadian pacemaker to behavioral rhythms.
This report fills an important gap in our understanding of the regulatory role of 5-HT on sleep, but several important questions remain. For instance, total elimination of brain 5-HT by neurotoxins and TPH2 knockout leaves sleep and behavioral rhythms intact (Morin & Blanchard, 1991; Alenina et al., 2009). The rapid reduction of 5-HT by TSOI may preclude compensatory mechanisms potentially present in non-reversible models of 5-HT depletion. The presence of sleep/wake rhythms in these non-reversible models is nonetheless paradoxical. Future studies investigating the potential role of the indoleamine melatonin, which also has sleep regulatory properties and is also tryptophan-dependent, may help to clarify these inconsistencies.