Chapter 1: A review of melatonin’s sleep-promoting role across animal models
1.10 Conclusions and future directions
4. Exogenous melatonin can entrain and phase shift circadian rhythms in birds, rodents, and humans, and may function through melatonin receptors in the SCN, which in turn regulates melatonin synthesis in the pineal gland. However, in zebrafish, rodents, and humans, there is virtually no loss-of-function evidence to support a role for endogenous melatonin in regulating circadian rhythms. Studies in birds have been interpreted as showing a requirement for melatonin for normal circadian rhythms, but experiments that (a) distinguish between effects on sleep versus circadian rhythm and (b) examine the specific loss of melatonin rather than the pineal gland are still needed.
Taken together, published studies suggest that melatonin acts downstream of the circadian clock to promote sleep in diurnal animals.
The role of melatonin in sleep seems to become less apparent (and/or more nuanced) in ‘higher’ organisms. This is evident when comparing the relatively strong effects of gain- or loss-of-function manipulations in zebrafish or birds with the relatively weak or non-existent effects in rodents or humans. What anatomical or physiological differences between these groups might explain the differences in melatonin’s effects?
Perhaps the simplest dividing factor is the complexity of their nervous systems. In an animal with a more complex nervous system, the requirement for sleep may be greater, and accordingly, the regulation of sleep might be under control of multiple redundantly acting mechanisms. In this view, melatonin’s role in sleep may be more ‘important’ in birds and fish and less so in humans or diurnal rodents, where other sleep-regulating pathways or circuits are present.
A second (non-mutually exclusive) possibility is that both the human and rodent lineages likely share a common nocturnal ancestor. The ‘nocturnal bottleneck’ hypothesis
suggests that the mammalian ancestor occupied nocturnal niches to avoid predation by diurnal dinosaurs, and not until after the extinction of dinosaurs were diurnal mammals able to flourish (Gerkema et al., 2013). Many aspects of physiology in modern mammals—
even diurnal ones—are reflective of this evolutionary period. One example may be a decreased dependence on melatonin for regulating sleep, as nocturnally secreted melatonin would be maladaptive if it promoted sleep in a nocturnal animal.
Third, as noted above, both zebrafish and birds have pineal glands that are positioned near the surface of their heads and are directly exposed to and entrained by light. In species whose circadian rhythms are so readily and directly attuned to environmental lighting, perhaps the melatonin signal is more reliable and therefore more potent, as there was no need to develop redundant or compensatory sleep-regulating circuits. Conversely, in animals whose pineal glands are buried deep within the brain (and thus not exposed to light), circadian information is received indirectly via a multi-synaptic pathway. This ‘outsourcing’ of circadian information might have led to a less reliable melatonin signal and a subsequent development of alternative, redundant mechanisms of circadian regulation of sleep.
Understanding the role of melatonin in sleep is important for evaluating and optimizing its use as a therapeutic agent. Since melatonin is a naturally-occurring sleep hormone in humans, its value in remedying sleep disorders is potentially very high. The most well-known pharmaceutical options for treating sleep disorders act by enhancing inhibitory GABAergic signaling in the brain, and their effects, while potently sedating, do not fully recapitulate natural sleep. Designing therapies around melatonin signaling, which has a key function in natural sleep regulation, could result in more natural and restorative sleep for those suffering from insomnia. Agonists or antagonists that target specific melatonin receptors with high affinity or that act on parts of the brain that mediate the
sleep-promoting role melatonin could prove useful as therapeutic agents. Optimizing melatonin’s utility as a sleep aid hinges on a greater understanding of its mode(s) of action with regards to sleep. While plenty of research has been done to uncover some of the subcellular events elicited by melatonin receptor activation, very little work has been done to identify sites of action in the brain or downstream effectors of melatonin signaling.
To advance our understanding of melatonin’s role in sleep, it seems crucial to expand melatonin research to include animal models beyond nocturnal rodents. While invertebrate research has been useful for the study of sleep, even leading to the discovery of sleep-promoting neuropeptides that play a role in vertebrate sleep (Nelson et al., 2014;
Nath et al., 2016; Lee et al., 2017), the function of melatonin in these species is questionable, and the lack of brain homology with vertebrates limits the potential for modeling melatonin’s role in human sleep. Instead, zebrafish, diurnal birds, and diurnal rodents hold great promise as model organisms to understand the functions of melatonin in diurnal vertebrates, including humans.
Zebrafish are a useful model for sleep research for the reasons described in section 1.5, and melatonin has been shown to be required for circadian regulation of sleep in this species. The genetic tractability of the zebrafish and its amenability to non-invasive whole-brain neuronal imaging makes it particularly suitable to uncover the genetic and neuronal mechanisms through which melatonin regulates sleep.
Studies using diurnal birds have been instrumental in understanding the role of the pineal gland in regulating behavior. Diurnal birds are strongly dependent on the pineal gland for normal sleep-wake cycling, but this relationship has been interpreted as an action of the pineal on the circadian rhythm, not on sleep directly. In order to determine the role of melatonin in these processes, loss-of-function manipulations that specifically target melatonin and not the entire pineal gland, perhaps using transgenesis or mutations
(Scott et al., 2010), and assays that distinguish between effects on the circadian clock versus outputs of the clock such as sleep, are needed.
Finally, a close examination of sleep (i.e., using EEG/EMG recordings) in diurnal rodents in the context of melatonin gain- and loss-of-function manipulations are necessary to determine the role of melatonin in sleep in these species. If the lack of an apparent role for melatonin in sleep in nocturnal rodents is simply a consequence of their nocturnality, then diurnal rodents could be used to validate current and future findings in nocturnal rodents—such as the role of MT2 in the reticular thalamic nucleus—and build confidence that the mechanisms uncovered in ‘lower’ organisms like birds, fish, and even worms are conserved in mammals. Studies of melatonin’s function in diurnal rodents would also aid in the development of melatonin-based therapies for human sleep disorders.
Next, in Chapter 2, I describe my efforts using zebrafish to uncover some of the mechanisms by which melatonin promotes sleep.