Melatonin as a hypnotic: Pro

Sleep Medicine Reviews (2005) 9, 51–65

www.elsevier.com/locate/smrv

CLINICAL REVIEW

Melatonin as a hypnotic: Pro Irina V. Zhdanova* Department of Anatomy and Neurobiology, Boston University School of Medicine, 715 Albany St, R-913, Boston, MA 02118, USA KEYWORDS Melatonin; Sleep; Insomnia; Receptors; Aging

Summary In diurnal species, nocturnal melatonin secretion coincides with the habitual hours of sleep, in contrast to nocturnal animals which are at the peak of their activity while producing melatonin. Studies in humans, diurnal non-human primates, birds and fish show that melatonin treatment can facilitate sleep initiation during the daytime or improve altered overnight sleep. Behaviorally, the sleep-promoting effects of melatonin are distinctly different from those of common hypnotics and are not associated with alterations in sleep architecture. The effects of melatonin on sleep are mediated via specific melatonin receptors and physiologic doses of the hormone, those inducing circulating levels under 200 pg/ml, are sufficient to promote sleep in diurnal species. Aging reduces responsiveness to melatonin treatment and this correlates with reduced functional potency of melatonin receptors. Since melatonin receptors are present in different tissues and organs and involved in multiple physiologic functions, using physiologically relevant doses (0.1 –0.3 mg, orally) and time of administration (at bedtime) is recommended, in order to avoid known and unknown side effects of melatonin treatment. q 2004 Elsevier Ltd. All rights reserved.

Introduction Melatonin is the principal hormone of the circadian system, secreted exclusively at night in both nocturnal and diurnal species. The way this circadian signal is interpreted may depend on the animal’s strategy of adaptation to periodic changes in the environmental illumination. Being a highly lipophylic hormone, melatonin reaches every cell of the body. The wide distribution of melatonin receptors also suggests that melatonin affects various tissues and organs and, thus, can be involved in multiple physiologic processes. Only some of these processes have been studied, one of them is sleep. It took one (unrelated) clinical trial to discover that melatonin affects sleep. It has taken decades *Tel.: þ617-6388002; fax: þ617-6384676. E-mail address: [email protected]

thereafter to confirm, detail and further explore this phenomenon. Major differences in experimental designs and populations studied often make it difficult to compare studies on the effects of melatonin on human sleep. However, the overall data from human and, lately, animal-based studies suggest that: (a) Melatonin can promote sleep in healthy humans and other diurnal animals, if administered during habitual hours of wakefulness. It can improve overnight sleep in insomniacs but does not alter sleep in healthy individuals. Melatonin can facilitate the effects of common hypnotics, thus reducing their effective dose and facilitating drug withdrawal. (b) The dose-dependence of melatonin’s effect on sleep is within physiologic or low pharmacologic range, i.e. 50 – 200 pg/ml in blood plasma.

1087-0792/$ - see front matter q 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.smrv.2004.04.003

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Increasing melatonin above these levels does not increase melatonin efficacy but may cause side effects, e.g. circadian rhythm alterations. (c) The efficacy of melatonin to promote an individual’s sleep depends on circulating melatonin levels and sensitivity of the individual’s melatonin receptors. Sensitivity to melatonin may decline with age and as a result of neurodegenerative disorders. (d) Melatonin affects sleep via specific melatonin receptors, presumably MT1, and via cAMPdependent signaling pathway. It remains to be elucidated whether one particular physiologic target or the combined response of several organs and tissues defines a behavioral picture we call the ‘sleep-promoting effect of melatonin’.

Melatonin can promote sleep The first word on melatonin affecting sleep came from the ‘father’ of melatonin, Aaron Lerner, who identified and named this substance produced by the pineal gland and linked it to previously described lightening effects of pineal extracts on amphibian skin color. Since Dr Lerner studied the human pigmentation disease, vitiligo, he administered the newly-discovered endogenous substance to his patients hoping to see an effect on altered skin pigmentation. Instead, his patients became sleepy. The report of this unexpected result1 opened a new area of research on the effects of melatonin on sleep. While Dr Lener’s observations were strictly behavioral and relied on subjects’ self-reports, further studies employed polysomnography

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to objectively document changes in sleep processes.2 – 4 The results of these early studies confirmed that humans fall asleep faster on melatonin and that after melatonin treatment their sleep is less disturbed by environmental factors, e.g. noise. Moreover, it was shown at that time that no major changes in sleep architecture or day-after side effects result from melatonin administration.

Dose-dependence of melatonin’s effects on sleep The effects of biologically-active substances are always dose-dependent, although the type of dosedependence varies. The initial studies, including those by Dr Lerner, utilized doses of melatonin in the range of 50 – 1000 mg orally or up to 200 mg iv.1 – 3 After physiologic circulating melatonin levels have been measured in humans, it became clear that in all these studies gigantic increases in circulating melatonin were induced, to an extent that is never experienced by the organism under normal circumstances (Fig. 1). Fig. 2 shows increases in circulating melatonin levels after oral administration of different melatonin doses. Most striking is the 1000-fold increase in plasma melatonin after ingestion of a 10 mg dose, when compared to peak melatonin levels normally occurring at night in a young healthy adult. The realization that the doses of melatonin used in these studies resulted in such high supraphysiologic plasma levels should have resulted in drastic reductions in the doses administered to experimental subjects, at least in human studies. That this did

Figure 1 Twenty four hour serum melatonin profiles measured from 9 to 9 a.m. in a group of six young healthy males; p the time of the onset of habitual evening sleepiness.15

Melatonin as a hypnotic: Pro

Figure 2 Mean serum melatonin profiles of 20 subjects sampled at intervals after ingesting 0.1, 0.3, 1.0, and 10 mg of melatonin or placebo at 11:45 h.13

not occur for quite a while was due mainly to two factors: melatonin has very low toxicity and melatonin’s effects on sleep are subtle. Indeed, no major abnormalities in behavior or physiologic functions have been documented even after ingestion of extremely high melatonin doses. Consistent with this, induction of high circulating melatonin levels did not cause uncontrollable sleep initiation or general anesthesia, unlike the effects produced by high doses of common hypnotics (e.g. benzodiazepines). Due to these two factors, human studies continued to employ pharmacologic melatonin doses, in the hope that a higher dose would increase melatonin’s efficiency. While the majority of these studies reported a hypnotic effect of daytime melatonin administration,4 – 11 others did not,12 in spite of the similarities in the pharmacologic melatonin doses used (2 – 10 mg). These discrepancies in the results obtained are not easy to explain, since study designs and melatonin preparations employed were quite variable and individual circulating melatonin levels following treatment were rarely documented. Only a few of the studies aimed at establishing the dose-dependence of the behavioral effects of melatonin and did not find such within the 1 – 80 mg range of doses tested.7,10 This led some investigators to believe that sleep-related effects of melatonin reflect a pharmacologic side effect of the excessive doses of the hormone used, rather than indicating that melatonin plays a role in physiologic sleep regulation in humans. In 1994, we reported the results of a study comparing the effects of a wide range of melatonin doses (0.1 – 10 mg), administered during the day to young healthy subjects.13 The results showed that

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‘physiologic doses’, i.e. those resulting in circulating (blood serum or plasma) melatonin levels within normal nocturnal range, can promote sleep onset in young healthy volunteers. Physiologic circulating melatonin levels (under 200 pg/ml) were achieved after ingestion of 0.1 –0.3 mg preparations. This finding was followed by a series of polysomnographic studies on the effects of physiologic doses of melatonin administered to healthy young individuals at different times of day, though assuring that bedtime was scheduled at least 2 h prior to the subjects’ habitual bedtime.14,15 In all of these studies melatonin significantly promoted sleep onset, after being ingested at noon, or in the early or late afternoon. In these healthy subjects, normal overnight sleep duration or sleep architecture was not significantly altered by such treatment, with minor increases observed in the duration of stage 2 sleep.15 Overall, these studies showed that in young healthy adults physiologic circulating melatonin levels can accelerate sleep onset, if induced prior to habitual hours of sleep. Importantly, these studies established that melatonin efficacy does not significantly differ between physiologic and pharmacologic doses of melatonin (Fig. 3). These results provided the first evidence that normal

Figure 3 Effects of melatonin (0.3 or 1.0 mg, p.o.) on average (^SEM) latency to: (A) sleep onset (B) stage 2 sleep, relative to placebo ðn ¼ 11Þ: Treatment administered at the time of low sleep propensity, 2–4 h before habitual bedtime; *p , 0:005. Reprinted with permission from Ref. 15.

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nighttime melatonin production may indeed play an important role in normal sleep regulation. Similar results of physiologic and low pharmacologic melatonin doses were later reported as a result of other polysomnographic studies.16,17 A comparison of the effects of 0.1, 0.5, 1, 5 and 10 mg doses administered either in the evening, prior to a nap, or before nighttime sleep, confirmed that all of these doses can promote sleep onset when administered during the day but do not further improve normal sleep in healthy volunteers.17 This study also confirmed our observation on the lack of dose response over the range of 0.5 – 10 mg doses of melatonin. The studies on the effects of low melatonin doses underscored some peculiar characteristics of melatonin dose-dependence. While doses in the range of 0.3 – 10 mg produced effects of similar magnitude, a decline in melatonin efficacy became noticeable with a lower dose, 0.1 mg, which tended to affect a smaller number of subjects. However, in those who responded to this minimal dose, the effect was not different from that induced by higher doses. Such results suggested that the dosedependence of melatonin effects on sleep is either confined to a narrow range of low doses and saturates at physiologic levels of the hormone, or it may reflect a threshold phenomenon. If the latter were true, each individual could be sensitive to a specific circulating melatonin level within a physiologic range, with higher concentrations causing no increase in melatonin efficacy. Furthermore, such a threshold level could be constant for each individual or vary depending on the functional state of the organism. To add to the complexity of the phenomenon, some young healthy subjects tested repeatedly did not show any sleep-promoting effect of melatonin, independent of the dose used, suggesting that there are non-responders to melatonin’s effects on sleep. As will be discussed below, such an overall picture might be explained by variations in an individual’s melatonin receptor sensitivity and with receptor saturation at nearphysiologic levels. Overall, human studies and, as discussed below, those conducted in animal models, demonstrate that the effect of melatonin on sleep has dosedependence within a narrow range of effective concentrations, corresponding to physiologic or low pharmacologic circulating levels of the hormone. While using a term ‘physiologic doses’ or ‘physiologic circulating melatonin levels’, we compare melatonin levels induced by a particular dose to the endogenous nocturnal melatonin concentrations observed in blood circulation. Those are lower than the nocturnal melatonin levels in the cerebrospinal

I.V. Zhdanova fluid (CSF), as shown in both animal18 and human19 studies. Such difference reflects a parallel release of melatonin from the pineal gland into the bloodstream and CSF and its greater dilution in the blood. Differences observed in blood and CSF melatonin levels may lead to another question. If sleeppromoting effects of melatonin were mediated via central structures, would it be possible that ‘CSFlike physiologic melatonin levels’, rather than ‘physiologic blood melatonin levels’, are needed to produce this effect? Based on the experimental evidence, this would be unlikely. Oral administration of low dose melatonin, which has been shown to promote sleep while increasing plasma melatonin levels within their normal nocturnal range, could hardly increase CSF levels above those observed in the blood. This would require a mechanism for active accumulation of melatonin in CSF, which has not been shown. Higher melatonin doses, which would result in ‘CSF-like’ melatonin levels in the blood and, presumably, in CSF, do not appear to be more powerful, as discussed above. Thus, unless only some peripheral effects of melatonin mediate its sleep-related actions, relatively low melatonin levels are sufficient to affect the central structures. The latter is also suggested by the low dose melatonin being able to produce a circadian entrainment, presumably via its receptors located in the hypothalamus. Rapid increase in CSF melatonin levels, however, may provide a prompt response of the brain structures to the initiation of nocturnal activity of the pineal gland. Further studies are needed to address these issues.

Acute versus circadian effects of melatonin on sleep It has been well established that melatonin can shift the circadian phase of suprachiasmatic nuclei of the hypothalamus (SCN) activity or entrain a freerunning circadian rhythm to a period of melatonin administration. This action is likely to underlie the effects of melatonin on the circadian phase of sleep initiation, helpful to blind individuals with freerunning rhythms or to those suffering from jet-lag. It is, however, important to distinguish the circadian from the acute sleep-promoting effects of melatonin treatment. The circadian effect largely depends on the time of melatonin administration and can produce opposite effects. Morning administration of melatonin can delay the onset of evening sleepiness by delaying the phase of the circadian rhythms, while evening melatonin

Melatonin as a hypnotic: Pro

treatment can advance the circadian rhythms, including time of sleep onset.20,21 The latter effect, for some time, has confused the researchers, since an earlier onset of sleep after late afternoon melatonin treatment could be attributed to both acute and circadian effects. The distinction came with the appreciation that melatonin can promote sleep within 30 – 60 min after morning or early afternoon treatment.13 – 15 In contrast, the circadian effect of a single dose of melatonin is powerful enough to induce around a 1 h phase advance, at most. Thus, the sleeppromoting effect of melatonin manifesting at noon13 cannot be attributed to an instant 9 h shift in the circadian clock. Importantly, doses required for both the acute sleep-promoting and circadian effect of melatonin are similar and are within the physiologic or low pharmacologic range.13 – 15,22 Using higher melatonin doses to entrain circadian rhythms in blind individuals proved to be counterproductive.22 Part of the reason could be an alteration in the circadian pattern of circulating melatonin after administering a high dose. Increased levels at both morning and evening hours can produce opposite circadian shifts, thus reducing an overall efficacy of such treatment. We suggest that yet another reason might be desensitization of melatonin receptors during prolonged elevation of circulating melatonin to supraphysiologic levels.

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Effects of melatonin on sleep in animal models

there are several important similarities between humans and diurnal non-human primates, favoring the use of these animals to model normal and pathological sleep-related processes. Those include: (1) Similar temporal patterns of activation of the major circadian pacemaker, the SCN, relative to the rest-activity cycle in both species, i.e. high activity of the SCN neurons during the day correlates with these species’ daytime activity, in contrast to nocturnal animals whose SCN is active during their daytime rest period; (2) Similar temporal patterns of melatonin production, occurring during habitual nighttime sleep period; (3) A consolidated nocturnal sleep episode, with similar sleep architecture, in contrast to the majority of nocturnal or diurnal species which tend to have a polyphasic sleep pattern. In all three species of diurnal macaques studied, the sleep process showed high sensitivity to daytime melatonin administration.24 Sleep initiation was significantly promoted by a wide range of melatonin doses used and, as in humans, showed a lack of dose dependence of the effect, once the dose (5 – 20 mg/kg, orally, Fig. 4) was sufficient to induce physiologic circulating levels of the hormone (above 50 pg/ml). Lower doses failed to promote sleep in the macaques studied. Another diurnal species we are using to investigate the effects of melatonin on sleep is the zebrafish, a favorite model for studying vertebrate genetics and development. After characterizing sleep-like behavior in this diurnal vertebrate, since it has not been done before, we compared the effects of melatonin and other hypnotic agents in zebrafish.25 The results showed an already

Technical and ethical issues preclude many types of studies that could help to elucidate the mechanisms of melatonin action in humans. Initial animal-based studies on the effects of melatonin on sleep produced inconsistent results, only some being able to show that very high pharmacologic doses of melatonin produce sedative effects. However, animals used to study the effects of melatonin on sleep at that time were either those with crepuscular (cats) or nocturnal (rats) activity pattern. Unlike humans, nocturnal animals are sleeping during the day, when their melatonin levels are low, and are at the peak of their activity at night, at the time of melatonin production by the pineal gland. Thus, melatonin is an unlikely candidate for being a sleep-promoting agent in nocturnally-active species. To explore the nature of sleep-promoting effects of melatonin, we have initiated studies in diurnal macaques.23 In addition to phylogenetic proximity,

Figure 4 Effects of a long-term melatonin treatment with escalating melatonin doses. Mean (SEM) sleep onset times in six monkeys (M. nemestrina and M. mulatta) during administration of escalating melatonin doses (5 –320 mg/kg, 3 days each dose; black bars), compared to periods of placebo treatment (white bars). PLC1, basal placebo treatment; PLC2, washout placebo treatment.

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I.V. Zhdanova

Figure 5 Melatonin and conventional sedatives promote rest behavior in larval zebrafish. Melatonin, diazepam and sodium pentobarbital (barbital) significantly and dose-dependently reduced zebrafish locomotor activity ((a), (c), (e)) and increased arousal threshold ((b), (d), (f)). Each data point represents mean ^SEM group changes in a 2 h locomotor activity relative to basal activity, measured in each treatment or control group for 2 h prior to treatment administration. Arousal threshold data are expressed as the mean ^ SEM group number of stimuli necessary to initiate locomotion in a resting fish. Closed diamond-treatment, open square-vehicle control; N ¼ 20; each group. Reprinted with permission from Ref. 25.

familiar picture of a ‘sleep’-promoting effect of melatonin with relatively flat dose-dependence, in contrast to barbiturates or benzodiazepines (Fig. 5). Moreover, the dose-dependence shown in Fig. 5 largely reflects more rapid onset of the effect after administration of high melatonin doses, presumably due to differences in the bioavailability of melatonin administered into the medium. Locomotor activity measured starting an hour after melatonin treatment significantly differed only within a narrow range of 1 – 100 nM of melatonin. These results in diurnal monkeys and fish are in accord with observations in diurnal birds. Melatonin administration reduces locomotor activity in the house sparrow and the Japanese quail.26 In pigeons, melatonin induces daytime sleep and increases EEG slow wave activity, without altering subsequent nighttime sleep.27 In contrast, melatonin does not change locomotor activity in the nocturnally active owl.26 Combined, these results obtained in animal models illustrate that the acute sleep-promoting effect of melatonin is not unique to humans but is typical of the effect seen in other diurnal species. Importantly, this allows studying the mechanisms of melatonin effects on sleep in more detail.

Behavioral effects of melatonin differ from those of common hypnotics The nature of melatonin’s effect on sleep is such that it typically does not produce a rapid increase in subjective sleepiness, an uncontrollable urge to fall asleep or major impairments in cognitive performance.1 – 17 In contrast to common hypnotics, and we have repeatedly underscored this, melatonin induces a behavioral state that resembles quiet wakefulness, which normally predisposes to normal sleep initiation, rather than sleepiness or drowsiness. These differences in the behavioral effects of melatonin and common hypnotics are, at least in part, responsible for some doubts about melatonin’s efficacy to affect sleep. On the one hand, physicians or sleep researchers are used to robust dose-dependent effects of the pharmacologic hypnotics, ranging from minor sedation to general anesthesia and resulting in significant changes in sleep architecture. On the other hand, patients expect to feel sleepy, drowsy or fatigued soon after taking a ‘sleeping pill’. Apparently, some of these effects of common hypnotics and the underlying

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Figure 6 Effect of melatonin and temazepam on subjective sleepiness, using a visual analogue scale. W represents the placebo condition, B represents the melatonin condition, K represents the temazepam condition. Each point represents mean ^ SEM. Administration of 5 mg melatonin, 10 mg temazepam or placebo was at 1200 h (dashed line). *** represents a significant ðp # 0:05Þ difference between the placebo and melatonin conditions; ** represents a significant ðp , 0:05Þ difference between the placebo and temazepam conditions; * represents a significant ðp , 0:05Þ difference between the melatonin and temazepam conditions. Reprinted with permission from Ref. 28.

mechanisms involved are not related to normal processes of sleep regulation, judging from specific changes in EEG patterns following their administration. Melatonin administered during the day can potentiate sleep onset, if the environmental conditions are appropriate for sleep initiation. Stimuli that would interfere with normal unmedicated sleep, e.g. turning lights on, changing to an upright position or the necessity of conducting a task requiring a high level of attention or motivation, temporarily interfere with the effect of melatonin on sleep. Importantly, under such circumstances, a person would typically feel quite alert, rather than sedated, in spite of high circulating melatonin levels. Once the interference is removed, the ability to fall asleep faster is restored. It should be noted however that in some individuals high pharmacologic doses of melatonin may induce behavioral alterations typical of sedatives. These sedative-like effects may reflect direct or indirect interaction of melatonin with other, e.g. GABA, receptors. These differences in the effects of melatonin and common hypnotics were well documented by a recent study, comparing the effects of melatonin (5 mg) and temazepam (10 mg) ingestion on sleepiness and cognitive performance.28 While temazepam caused an abrupt but short-lived increase in subjective sleepiness, melatonin gradually

increased sleepiness and maintained it for a significantly longer period of time (Fig. 6). In contrast, a decrease in performance following temazepam treatment was greater than after melatonin administration. While this and other studies show moderate changes in performance following administration of pharmacologic melatonin doses, our dose-dependence studies in humans, monkeys and zebrafish13 – 15,24,25 suggest that even these alterations in performance can be avoided by using physiologic melatonin doses, without jeopardizing melatonin’s sleep-promoting effects. Principal differences in the effects of melatonin and common hypnotics are further underscored by the absence of substantial changes in sleep architecture following melatonin administration, whichever melatonin doses are used. However, high dose melatonin, e.g. 5 mg administered orally, has been reported to enhance EEG power density in non-REM sleep in the sleep spindles frequency range (13.75 – 14.0 Hz bin) and reduce activity in the 15.25 – 16.5 Hz band, suggesting that melatonin, at least in supraphysiologic levels, could influences thalamocortical oscillations.29 The distinctively subtle nature of melatonin’s effect on sleep appears to be evolutionary conserved, manifesting in other diurnal species, including non-human primates. Our current studies in zebrafish, involving high-speed camera recordings (1000 frames/s), allow precise measurement of

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the reaction time to stimuli of different intensity. While melatonin-treated zebrafish larvae show a significant reduction in spontaneous locomotor activity and increased duration of inactivity, medium or high intensity stimuli induce locomotor responses that are quantitatively and qualitatively indistinguishable from those in control animals. Furthermore, independent of the melatonin dose administered at daytime, arousal thresholds are maintained close to those observed normally at night.25 In appreciation of the distinct qualities of melatonin’s effect on sleep, compared to those of common hypnotics, we chose to call melatonin a ‘sleep-promoting’ hormone, rather than ‘sedative’, ‘hypnotic’ or ‘soporific’, since all the latter terms imply that the substance induces sleepiness or sleep. Whatever linguistic term to be used, the distinct pattern of melatonin’s effect on sleep awaits a more biological definition, which would also require a better understanding of the physiologic nature and function of the sleep process per se.

Circulating melatonin levels and insomnia Recognition that the pineal hormone produced every night can promote sleep suggested that melatonin deficiency may underlie some sleep disorders. One of the first questions to answer was whether insomniacs have lower melatonin levels than individuals with normal sleep. Typically, melatonin secretion undergoes a predictable change over the life span. Newborn babies do not secrete melatonin for approximately the first 3 months of life. The onset of melatonin secretion appears to coincide with the overall increase in circadian rhythmicity and development of the major nocturnal sleep episode. Young children are reported to have the highest circulating melatonin levels, which start declining around the time of puberty. Mean circulating melatonin levels in aged individuals are significantly lower than in young healthy adults.30 – 33 It should be noted, however, that there is a substantial inter-individual variability in melatonin levels in all age groups (Fig. 7) and healthy elderly may preserve melatonin levels typical of young adults. While some studies documented a significant correlation between low melatonin production and insomnia,32 others reported either a tendency34 or no correlation between these two variables.35 Does that mean that a decline in circulating melatonin

I.V. Zhdanova

Figure 7 Individual peak endogenous serum melatonin levels in people of different ages ðn ¼ 36Þ.

with age has nothing to do with the development of insomnia? Not necessarily. All of the studies addressing possible links between circulating melatonin levels and incidence of insomnia were crosssectional, rather than longitudinal. They have documented an absolute melatonin level at a particular age but not the degree of change that has occurred over the years. What if melatonin deficiency is a relative term? For example, subject A has maintained 40 pg/ml peak melatonin level from age 20 to 70 and, thus, does not experience melatonin deficiency. In contrast, in Subject B, during the same period of time, circulating melatonin levels dropped from 120 to 40 pg/ml, causing relative melatonin deficiency. In such a case, unless we knew of the ‘pre-existing condition’, we would not be able to suggest that insomnia in subject B could be due to melatonin deficiency. This theoretical scenario is supported by inter-individual variability in circulating melatonin levels in all age groups and the substantial overlap in melatonin levels observed in young and aged adults (Fig. 7).33 Thus, so far, it can be concluded that peak circulating melatonin level might not be a decisive factor in an individual’s sleep control. However, melatonin deficiency might be part of the problem. Consequently, alterations in melatonin production resulting from pathological conditions, medications or aging may play a significant role in sleep pathology. These issues can be studied in humans and in animal models. One of the approaches would be to see if increasing circulating melatonin levels could improve altered sleep.

Effects of melatonin treatment in insomniacs Observations of sleep-promoting effects of daytime melatonin administration in healthy individuals suggested that melatonin might help insomniacs

Melatonin as a hypnotic: Pro

to sleep better. In order to test this hypothesis, several research groups have conducted studies treating insomniacs with different melatonin doses. The results of these studies have varied, with some documenting positive effects of melatonin treatment, while others report no effect. Among possible explanations for differences in the results obtained is variability in the subjects’ age, origin of insomnia, concurrent medications and different study designs used, including the melatonin doses administered. The majority of melatonin studies in insomniacs were conducted in ‘extreme’ age groups, children and the elderly. Children appear to be quite sensitive to melatonin treatment. In contrast, high variability in melatonin efficacy was found among the elderly. For this reason, we will discuss the results relevant to each of these age groups separately.

Children-insomniacs are sensitive to the effects of melatonin on sleep Studies in children with severe insomnia, resulting from multiple neurological disorders, showed that administration of pharmacologic melatonin doses could substantially improve their sleep patterns and increase sleep duration.36 – 39 Similarly, our study in children with Angelman syndrome (AS), a rare genetic disorder characterized by severe mental retardation, hyperactivity, and disturbed sleep, showed that timely administration of a low melatonin dose (0.3 mg) can promote nighttime sleep and, in some cases, improve daytime performance in these children.40 Children with normal development, but suffering from chronic insomnia, also showed significantly increased total sleep time following melatonin treatment.41 By documenting the circadian patterns of plasma melatonin in Angelman syndrome children before and after treatment period, we could also see that the majority of them were well-entrained to the environmental light – dark cycle. In such children with entrained rhythms, bedtime melatonin treatment improved sleep without altering the circadian rhythmicity. In contrast, in few children with circadian phase delay, melatonin entrained the rhythm and synchronized it with the time of melatonin administration. These data illustrated that acute and circadian effects of melatonin could occur jointly or separately, both contributing to the therapeutic effect observed. Overall, studies conducted in children suggest that this age group is remarkably sensitive to the effects of melatonin on sleep. Similar conclusions can be derived from our animal-based studies,

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showing that younger animals, both primates and zebrafish, are more responsive to the effects of melatonin on sleep than older ones. This notion might help us understand the mechanisms of melatonin action and inter-individual variability in melatonin efficacy, as discussed below.

The elderly have increased variability in melatonin efficacy A number of research groups have investigated the effects of melatonin administration in aged humans, focusing on those suffering from agerelated insomnia. Melatonin, with its low toxicity, and with no apparent side effects typical of pharmacologic hypnotic agents, e.g. locomotor and memory impairments or morning grogginess, could be an ideal medication for aged insomniacs, the main consumers of hypnotic drugs and the main sufferers from their side effects. Moreover, in addition to the typical reduction in melatonin levels in normal aging, patients with neurodegenerative disorders, such as Alzheimer or Parkinson disease, were reported to have dramatically impaired melatonin production. These diseases are associated with severe insomnia, further suggesting that lack of melatonin’s sleep-promoting effect could be one of the factors altering normal sleep process in these patients. However, studies on melatonin administration to aged individuals suffering from insomnia produced mixed results. Some found melatonin treatment (0.1 – 6 mg, orally) to facilitate overnight sleep in elderly insomniacs. 34,42 – 44 In contrast, other studies failed to document significant changes in elderly patients’ sleep after melatonin (1 – 5 mg) administration.35,45 Differences in the results obtained could not be attributed to melatonin doses used, since their range largely overlapped in these studies. Only two of melatonin-related studies in aged insomniacs used polysomnography to assess sleep,34,45 others fully relied on actigraphy and/or subjective reports. One of these polysomnographic studies45 reported a reduction in latency to sleep onset after both physiologic and pharmacologic doses were administered, but no change in sleep efficiency. In contrast, in our patients34 physiologic and pharmacologic doses of melatonin did not cause significant changes in latency to sleep (which was rather short at baseline) but improved sleep efficiency (Fig. 8A), especially in the middle portion of the night. In both of these studies, induced individual melatonin levels or concomitant changes

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reports. Although the reason for not conducting PSG studies in this population is fully appreciated, this might add to the variability in the results obtained. More importantly, and that brings us to the question of the mechanisms of melatonin action discussed below, there is no information on individual functional sensitivity of melatonin receptors to endogenous or exogenous melatonin in aged individuals studied. This parameter could be a critical determinant of the degree of an individual’s response to melatonin treatment, since it is subject to age-related alteration and possibly further deterioration due to neurodegenerative disorders.

Mechanisms of melatonin’s sleep-promoting effect There are several critical questions regarding the mechanisms of the behavioral effects of melatonin. Do specific melatonin receptors mediate sleep-promoting effect of the hormone? If yes, which melatonin receptor subtype is responsible for the effects observed and where are these receptors localized? Which other receptors melatonin might act upon? Figure 8 Sleep efficiency (A) and core body temperature profiles (B) in adults over 50 years of age following melatonin or placebo treatment. V, placebo; A, 0.1 mg; O, 0.3 mg, W, 3 mg. *p , 0:05. Reprinted from The Journal of Clinical Endocrinology and Metabolism, vol. 86, pp. 4727–4730, Copyright 2001, The Endocrine Society.

in core body temperature did not correlate with melatonin efficacy. Similarly, conflicting results were obtained in studies on the effects of melatonin in Alzheimer Disease (AD) patients, some demonstrating improvement in patients’ sleep after melatonin administration,46 – 48 others reporting the absence of a significant effect.49,50 There are many factors that could contribute to the inconsistent results obtained, including different subject selection criteria, the melatonin doses and preparations used, time of treatment, circulating melatonin levels induced, co-medications, food intake around the treatment time, the environmental conditions in a laboratory, home or institution. Importantly, only a few studies measured endogenous and treatment-induced circulating melatonin levels, leaving unanswered questions regarding differences in baseline melatonin levels and drug metabolism in these elderly patients. Unfortunately, studies in AD patients had to rely on actigrahy, rather than polysomnography (PSG), and on care-takers’

Does melatonin affect sleep via melatonin receptors? Studying the effects of melatonin receptors in humans is a challenge, since melatonin receptor agonists or antagonists are not approved for human use. However, using a diurnal animal model, we showed that the effect of melatonin on sleep is indeed mediated via specific melatonin receptors.25 Specific melatonin receptor antagonists, including luzindole, can attenuate or block sleep-promoting effect of melatonin (Fig. 9). Comparing the relative ability of melatonin receptor ligands to simulate or counteract the effects of melatonin on zebrafish sleep (Zhdanova, unpublished data), also suggests that the MT1 receptor subtype is likely to be responsible for the effects of melatonin on sleep. However, the lack of MT1 subtype selective ligands precludes a more definitive conclusion. Future availability of such ligands and/or studies in diurnal animals lacking specific melatonin receptor subtypes could fully clarify this issue. Zebrafish-based studies might also explain interindividual variability in melatonin’s efficacy to promote sleep, repeatedly documented in human studies. Our preliminary data suggest that the promotion of sleep by melatonin correlates with

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Figure 9 Melatonin and diazepam affect locomotor activity in zebrafish via specific membrane receptors. Pretreatment with specific antagonist for melatonin receptors, luzindole, blocked the decline in locomotor activity induced by (a) melatonin, but not by (b) diazepam or (c) pentobarbital. Pretreatment with specific benzodiazepine receptor antagonist, flumazenil, blocked reduction in locomotor activity following (b) diazepam, but not (a) melatonin or (c) pentobarbital treatment. Control solutions are vehicles for each treatment used. Data are expressed as mean ^ SEM group changes (%) in daytime locomotor activity, measured for 2 h after treatment, relative to basal activity. N ¼ 30; each group; **p , 0:01. Reprinted with permission from Ref. 25.

reductions in stimulated brain cAMP levels. Moreover, the ability of melatonin to promote sleep in zebrafish and to counteract cAMP increase in the brain declines with age. Such age-related changes in melatonin receptor sensitivity and/or density51 might underlie increased inter-individual variability in melatonin efficacy among elderly and patients suffering from neurodegenerative disorders.

Does melatonin affect sleep via GABA receptors? Early studies in nocturnal rodents showed that very high pharmacologic doses of melatonin can induce sedation in rats.52,53 They also suggested that the effect is mediated via the GABAergic system.53 In contrast, a human study, in which subjects received melatonin (3 mg, i.e. , 50 mg/kg) with or without a benzodiazepine receptor antagonist, showed that co-administration of flumazenil is not sufficient to disrupt the sleep-promoting effect of melatonin in humans.54 Our studies in non-human primates and zebrafish support the latter finding, showing that pretreatment with flumazenil fails to alter the effect of melatonin on sleep (Fig. 9).25 It should be noted, however, that similar to benzodiazepines, melatonin can exert anxiolytic effects in humans and in animal models. These anxiolytic effects of melatonin are explored much less than the effects on sleep but might contribute to its sleep-promoting property. It is yet to be defined whether melatonin can affect anxiety of different origin and whether individuals susceptible to the sleep-promoting effects of melatonin have increased anxiety levels.

Combined, these data suggest that although the effects of melatonin on sleep are mediated via specific melatonin receptors, the direct or indirect effects of melatonin on GABA or other receptors affecting anxiety state cannot be excluded at this point.

Is SCN the site of melatonin effects on sleep? In terms of localization of receptors responsible for the sleep-promoting effects of melatonin, the evidence remains ‘circumstantial’. The SCN was the first candidate to be considered due to the high density of melatonin receptors in this structure and its critical role in the regulation of circadian rhythms. SCN neurons are active during the day and relatively quiet at night. Exposure of the SCN to melatonin leads to acute inhibition of SCN metabolic activity and neuronal firing rate.55 Thus, it has been hypothesized that melatonin may promote sleepiness by inhibiting SCN activity. The caveat, however, is that SCN activity has a similar temporal pattern in nocturnal animals, i.e. their SCN is at the peak of activity while these animals are sleeping during the day. Perhaps, the SCN activity per se has no direct alerting effect. Instead, the processes downstream of SCN could translate its signal into either ‘be alert’ or ‘fall asleep’ command, depending on the species-specific strategy of adaptation to a periodically changing environment. So far, no experimental data have fully clarified this issue and we are currently addressing it in our polysomnographic studies on the effects of melatonin in SCNlesioned rhesus monkeys. If melatonin’s acute effect on sleep is mediated via specific receptors located in the SCN, the SCN lesion would be

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predicted to block the sleep-promoting effects of the pineal hormone.

Do systems involved in thermoregulation mediate the effects of melatonin on sleep? In addition to the brain, melatonin receptors are expressed in a variety of tissues, including cardiac and peripheral vessels, retina, kidneys and other organs. Thus, it is important to understand which peripheral melatonin targets might be involved in its sleep-promoting effect. Sleep is normally associated with a decline in core body temperature and elevation of skin temperature due to a combination of increased heat loss and reduced heat production. Melatonin treatment is often associated with a reduction in core body temperature and/or an increase in skin temperature.10,56 This suggested that the effects of melatonin on cardio-vascular system and/or thermoregulatory centers may underlie the effect of melatonin on sleep.57 Our finding58 that melatonin treatment causes an increase in daytime plasma cGMP levels, typical of vasorelaxation and normally observed at night,59 and that this effect correlates with the sleep-promoting effect of melatonin, may also support this hypothesis. However, the main question pertains to the causality of these two events. Does sleepiness or sleep promoted by melatonin drive vasorelaxation, cGMP increase, heat loss and decline in core body temperature typical for the sleep state or it is the other way around? If the hypothermic effect of melatonin mediates its ability to promote sleep, then sleep effect should follow the hypothermic effect and have a similar dose-dependence. This, however, does not seem to be the case. Temporal patterns of sleep- and temperaturerelated effects of melatonin were found to be different, with hypothermia manifesting only after the onset of the sleep-promoting effect.6 Comparison of the effects of physiologic (0.3 mg) and pharmacologic (3 mg) melatonin doses show similar effects of these doses on sleep, but a significantly more prominent decline in core body temperature after administration of the pharmacologic dose34 (Fig. 8B). In the latter study, skin temperature was also measured and, while showing the typical increase at sleep onset, did not reveal significant dose-dependence. Several human studies documenting significant sleep-promoting effects of melatonin on sleep8,60 did not find changes in body temperature, even after treatment with a 10 mg dose of melatonin. In contrast, a study in aged insomniacs showing significant changes in body temperature following

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melatonin treatment found no significant change in sleep efficiency.45 Important insights came from studies on the effects of melatonin in birds. Inhibition of locomotor activity following melatonin treatment was associated with a reduction in body temperature in diurnal birds, house sparrow and Japanese quail.26 In contrast, in the nocturnal owl, body temperature was reduced but activity levels were preserved after melatonin treatment. Furthermore, administration of melatonin directly into the brain structures of Japanese quail showed that melatonin-induced decreases in body temperature and locomotor activity result from melatonin acting at discrete brain areas.61 A decrease in body temperature alone was observed after melatonin administration to the thalamus and hypothalamus, while only locomotor activity was decreased when melatonin was injected close to the nuclei septalis medialis and septalis lateralis. Injections in some brain areas produced both physiologic effects. These studies further suggest that, although melatonin can affect both sleep and thermoregulation and these two processes can interact, the mechanisms of melatonin effects on these systems might be independent.

Conclusions Melatonin is an exceptionally interesting hormone. Its evolutionary roots can be traced to unicellular organisms, invertebrates and lower vertebrates, which produce and use this small amine for time navigation. Every night humans and other mammals secrete melatonin and it conveys a message of darkness to every cell of the body. If an organism can perceive and translate this message, it will be prepared for complex adaptation to the periodically changing environment. Would the responses to this message be the same for different organisms? This is unlikely. Night means activity for some and quietness for the others, foraging or building a shelter for nocturnal species and sleep for the diurnal ones. Thus, some effects of melatonin might be conserved between the species, some will vary. Based on the knowledge accumulated in multiple studies we can conclude that in humans and other diurnal species melatonin treatment can promote sleep during the day. It can also improve overnight sleep in some insomniacs, those that are sensitive to its effects. Melatonin promotes sleep in a way that resembles the natural onset of sleep: through quiet wakefulness to the sleeping state, while maintaining normal sleep architecture.

Melatonin as a hypnotic: Pro

Circulating melatonin levels normally observed at night are sufficient to produce an effect in young healthy individuals. Perhaps, slightly higher doses might help if an individual’s melatonin receptors are partially desensitized or if their density is reduced as a result of aging or the presence of a pathological condition. If receptor sensitivity is dramatically altered in tissues or organs mediating its effect, the presence of melatonin might not help. Overdosing does not appear to facilitate the sleep-promoting effect but can prolong the period of increased circulating melatonin levels. This, in turn, may desensitize melatonin receptors and shift the phase of the circadian rhythm. Neither effect would be beneficial for individuals well-entrained to a 24 h light – dark cycle. Children-insomniacs appear to be quite sensitive to the effects of melatonin on sleep. Animal-based data suggest that this could be a result of increased functional sensitivity of melatonin receptors at younger ages and/or their higher density. In contrast, the elderly, in whom melatonin production is typically lower, are less responsive to the effects of exogenous melatonin. Presumably, the reason for this might also be the degree of receptor sensitivity, which may decline with age. It is important to stress that these assumptions are based on the preliminary data and require further investigation. While animal-based studies show that melatonin affects sleep via specific melatonin receptors, the melatonin receptor subtype responsible for the effect remains to be identified. Similarly, the sites of the receptors mediating melatonin’s sleeppromoting effect remain to be elucidated. Although our knowledge of the role melatonin plays in sleep processes remains rudimentary, one thing is clear: melatonin is uniquely suitable to coordinate two major counterparts of sleep regulation, circadian rhythm and homeostasis. Melatonin can shift the circadian phase, advancing or delaying sleep propensity. Independent of this effect or in conjunction with it, melatonin can acutely increase the homeostatic drive for sleep and ease the transition between wakefulness and sleep. In addition to the role melatonin plays in sleep regulation, its circadian, visual, reproductive and thermoregulatory effects, and additional actions about which we have only a vague idea, call for extreme caution when using melatonin for preventive or therapeutic purposes. Administration of pharmacologic melatonin doses has been shown to disrupt circadian rhythms62 or alter semen quality in some human subjects.63 Thus, unless the goal is to shift the circadian phase, melatonin treatment should be synchronized with the time of normal

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melatonin secretion, which is initiated about 2 h prior to habitual bedtime and declines prior to morning awakening. We recommend administering melatonin in doses that would maintain melatonin levels within the normal nocturnal range (under 200 pg/ml). These physiologic doses were found to be sufficient and optimal to both promote sleep13 – 17, 34 and entrain circadian rhythms.22 Using melatonin at the right time and at the right dose in individuals deficient in melatonin production would be sufficient to provide its beneficial effects on sleep and protect from the undesirable ones. This would provide important information on overall physiologic role of melatonin and help to avoid possible side effects of melatonin treatment.

Practice points † Melatonin can promote sleep at daytime and improve altered sleep at night. † Doses that induce physiologic circulating melatonin levels (0.1 – 0.5 mg, orally) are sufficient to promote sleep, induce circadian phase shift and entrain circadian rhythms, without causing side effects. † If melatonin is used to improve nighttime sleep, it should be administered close to habitual bedtime, i.e. at the time of normal melatonin secretion. † Children are more sensitive to the effects of melatonin than the elderly. However, special care should be taken not to exceed physiologic melatonin levels and not to extend a period of increased circulating melatonin in children, in view of possible melatonin’s effects on development.

Research agenda We need to continue studying: † Mechanisms mediating the effects of melatonin on sleep. † Mechanisms involved in inter-individual variability in sensitivity to melatonin treatment and its age-dependence. † Role of melatonin in other physiologic systems and organs.

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Acknowledgements This work was supported by grants AG 17636 from the National Institute on Aging and MH 65528 from the National Institute of Mental Health.

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