Bright light exposure of a large skin area does not affect melatonin or bilirubin levels in humans.

BACKGROUND: Light treatment through the eyes is effective in alleviating the symptoms of some psychiatric disorders. A recent report suggested that skin light exposure can affect human circadian rhythms. Bilirubin can serve as a hypothetical blood-borne mediator of skin illumination into the brain. We studied whether bright light directed to a large body area could suppress the pineal melatonin secretion or decrease serum total bilirubin in conditions that could be used for therapeutic purposes. METHODS: Seven healthy volunteers participated in two consecutive overnight sessions that were identical except for a light exposure on the chest and abdomen in the second night from 12:00 AM to 6:00 AM (10,000-lux, 32 W/m(2) cool white for six subjects and 3000-lux, 15 W/m(2) blue light for one subject). Hourly blood samples were collected from 7:00 PM to 7:00 AM for melatonin radioimmunoassays. Bilirubin was measured by a modified diazo method in blood samples taken at 12:00 AM and 6:00 AM and in urine samples collected from 7:00 PM to 11:00 PM and from 11:00 PM to 7:00 AM. RESULTS: The skin light exposure did not cause any significant changes in serum melatonin or bilirubin levels. The excretion of bilirubin in urine was also the same in both sessions. CONCLUSIONS: Significant melatonin suppression by extraocular light does not occur in humans. Robust concentration changes of serum total bilirubin do not have a role in mediating light information from the skin to the central nervous system.

No evidence for extraocular light induced phase shifting of human melatonin, cortisol and thyrotropin rhythms.

The view that light affects the mammalian circadian clock only through the eyes was recently challenged by a study in which the phases of human circadian rhythms were shifted by extraocular light exposure. This finding has not been confirmed, however. We studied the effects of light exposure (3 h, broad spectrum fluorescent white light, 13000 lux) on abdomen and chest on the circadian rhythms of serum melatonin, cortisol and thyrotropin in six subjects. The protocol consisted of two 3-day sessions in a dimly lit (< 10 lux) experimental unit. In both sessions hourly serum samples were collected for hormone analysis on days 1 and 3. The skin light exposure was delivered on day 2 from 22.00 to 01.00h in one of the two sessions in a randomized order. In both sessions all three rhythms tended to delay, presumably due to the endogenous circadian cycle length being slightly longer than 24 h. However, the phase shifts did not differ significantly between the sessions. Thus, the present study does not support the existence of extraocular photic regulation of the circadian rhythms in humans.

Evidence for central alpha(2)-adrenergic modulation of rat pineal melatonin synthesis.

This study was performed to distinguish central and peripheral alpha(2)-adrenoceptors in the inhibition of rat pineal melatonin synthesis. The rats received lipo- or hydrophilic alpha(2)-adrenoceptor ligand injections at middark; after 1 or 2 h the pineal melatonin contents were measured. The lipophilic agonist medetomidine (100 microg/kg s.c.) suppressed the melatonin contents significantly, while the hydrophilic agonists ST-91 and p-aminoclonidine (10 or 100 microg/kg i.v.) did not. The suppression by medetomidine was counteracted by the lipophilic antagonist yohimbine (0.3-3.0 mg/kg i.p.) but not by the hydrophilic antagonist L-659,066 (1-10 mg/kg i.v.). In conclusion, the suppression of nocturnal melatonin synthesis by alpha(2)-adrenoceptor agonists is mainly of central origin.

Suppression of melatonin by 2000-lux light in humans with closed eyelids.

BACKGROUND: In order to clarify the role of light in regulating body functions in sleeping humans, we studied whether the light-sensitive pineal hormone melatonin can be suppressed by facial light exposure in subjects with closed eyelids. METHODS: Eight healthy volunteers participated in 3 nightly sessions: a dim-light control session (< 10 lux) and two light-exposure sessions (2000 lux, 60 min between 2400 and 0200 h). One light exposure occurred with eyes open and the other with eyes closed. Saliva samples were collected at least every hour from 1900 to 0300 h. Melatonin concentrations were measured by radioimmunoassay. RESULTS: Salivary melatonin concentrations decreased only in 2 of the 8 volunteers during light-exposure sessions with eyes closed. On average, light exposure did not decrease the salivary melatonin concentration. CONCLUSIONS: Because indoor illuminance is usually much lower than 2000 lux, light is probably ineffective in regulating the neuroendocrine hypothalamic functions in people during their sleep. Nevertheless, the possibility remains that higher illuminances, often used for therapeutic purposes, can inhibit the secretion of melatonin even in sleeping patients.

Evidence against alpha2-adrenoceptor involvement in the regulation of rat melatonin synthesis by ambient lighting.

This study was carried out to clarify the role of alpha2-adrenoceptors in the regulation of pineal melatonin synthesis. Medetomidine, a selective alpha2-adrenoceptor agonist, was previously found to be a potent suppressor of nocturnal melatonin levels in rats. Medetomidine and alpha2-adrenoceptor antagonists atipamezole and yohimbine were injected into rats in different conditions, and their pineal melatonin contents were measured by radioimmunoassay. Experiment 1: Blocking the alpha2-adrenoceptors and possible non-adrenergic binding sites with atipamezole did not counteract the light-induced suppression of nocturnal melatonin. These receptors are, thus, not essential for the suppression of melatonin by light. Experiment 2: Blocking the alpha2-adrenoceptors with atipamezole or yohimbine did not sensitize the pineal melatonin synthesis to daytime darkness in the light/dark-entrained rats. The binding sites are not involved in keeping the daytime melatonin levels low, even in darkness. Experiment 3: The rats were sensitized to daytime darkness by keeping them for seven days in constant light. The dark-elicited melatonin rise was suppressed by a lower dose of medetomidine than the normal nocturnal rise in light/dark-entrained rats, while atipamezole had no effect. The results showed that alpha2-adrenoceptor insufficiency is not involved in the constant light-induced pineal supersensitivity. In summary, the experiments indicated that the physiological regulation of melatonin synthesis by ambient lighting in rats does not depend on alpha2-adrenergic mechanisms.

Melatonin ineffective in neuronal ceroid lipofuscinosis patients with fragmented or normal motor activity rhythms recorded by wrist actigraphy.

Melatonin was tested as a sleeping pill in five patients with neuronal ceroid lipofuscinoses. The single-blind, placebo-controlled study consisted of motor activity recordings, sleep logs, and administration of placebo or melatonin (2.5 or 5 mg). Daily motor activity rhythms were measured by wrist actigraphy during four 7-day periods (baseline, placebo, melatonin 2.5 mg, and melatonin 5 mg). The placebo or melatonin was administered in the evenings for 3 weeks, and the recordings were made during the last week of the 3-week treatment. Sleep logs were kept by the caregivers during the recordings. Based on period analyses, the activity recordings were evaluated to display a normal (24-h) or fragmented rhythm. Three patients had normal motor activity patterns during the baseline recordings, and administration of placebo or melatonin did not affect their rest/activity rhythms. Two patients had abnormally fragmented activity rhythms during the baseline periods, and administration of placebo or melatonin did not induce synchronization. According to the actigraphic data, there were no changes in activity rhythms resulting from administration of melatonin. However, based on the observations, three families reported that melatonin slightly improved the sleep quality of the patients. These controversial findings show the difficulties involved in specifying the role of melatonin in modulating sleep. Thus, we conclude that more evidence is required before the significance of melatonin as a sleeping pill is defined.

Supersensitivity with reduced capacity for pineal melatonin synthesis in constant light-treated rats.

Melatonin is synthetized from serotonin in two steps driven by the enzymes N-acetyltransferase and hydroxyindole-O-methyltransferase. Constant light treatment reduces rat pineal hydroxyindole-O-methyltransferase activity while the activation of N-acetyltransferase becomes supersensitive to adrenergic stimulation. We studied the effect of this discrepancy on the production of melatonin. Male rats were kept under 12/ 12-h light/dark (LD) conditions or for 7 days under constant light (LL). They received subcutaneous injections of isoproterenol or methoxamine in the middle of the light period (LD-rats) or the estimated rest phase (LL-rats). A low dose of isoproterenol (0.1 mg/kg) increased pineal melatonin only marginally in LD-rats, while a maximum effect was found in LL-rats. A medium dose (0.2mg/kg) produced similar levels in both groups. A high dose (0.4 mg/kg) elevated pineal melatonin contents significantly more in normal than light-treated rats. Methoxamine (0.8 mg/kg) had no effects alone nor combined with isoproterenol. The results suggest supersensitivity with reduced capacity for melatonin formation in constant light-treated rats.

Bright light suppresses melatonin in blind patients with neuronal ceroid-lipofuscinoses.

We studied whether light information can reach the pineal glands of clinically blind patients with neuronal ceroid-lipofuscinoses. The suppression of melatonin by light was used as an indicator. Seven patients and seven control subjects were exposed to 3,000-lux light for 60 minutes at the rising phase of the melatonin synthesis. Most patients were not cooperative, and their eyelids were opened by a researcher every 2 minutes for 2 seconds. The control subjects opened and closed their eyes similarly by themselves. Light suppressed melatonin in three of seven control subjects and in all patients. The average postlight levels were 80% (control subjects) and 51% (patients) of the corresponding levels during the dim-light session. Despite degenerated retinas of the blind patients, light can penetrate their visual system to the hypothalamic and pineal levels and regulate neuroendocrine function.

Pineal melatonin in rats: suppression by the selective alpha2-adrenoceptor agonist medetomidine.

This study was done to clarify the role of alpha2-adrenoceptors in the regulation of pineal melatonin synthesis. A selective alpha2-adrenoceptor agonist, medetomidine, and antagonist, atipamezole, were injected subcutaneously into rats and their pineal melatonin contents were measured by radioimmunoassay. Medetomidine (120 microg/kg) suppressed melatonin at night to a similar extent during the rising and descending phase of melatonin synthesis, but it did not affect the daytime level. A dose of 12 microg/kg was ineffective; doses of 30-180 microg/kg suppressed nocturnal melatonin levels close to the daytime levels. Significant suppression was reached within 15 min and the effect started to fade 3 h after the injection (120 microg/kg). At midday, medetomidine did not inhibit isoproterenol-stimulated synthesis of melatonin. Atipamezole (0.4 or 1.2 mg/kg) had no effect alone, but it counteracted the medetomidine-induced suppression. The effects of alpha2-adrenoceptor ligands on melatonin synthesis depend on the time of day and/or on the activity of the pineal sympathetic nerves.

Alpha2-adrenoceptor agonist medetomidine affects the melatonin rhythm in rats.

We investigated whether alpha2-adrenergic mechanisms participate in the regulation of the daily melatonin rhythm. Female Wistar rats, living under 12:12 h light-dark conditions, received a subcutaneous injection of saline or medetomidine (alpha2-adrenoceptor agonist; 100 microg/kg) 1 h after lights off. Thereafter they were kept in continuous darkness. Pineal glands were collected for melatonin measurements at 2-h intervals during the first and second subjective nights. During both nights, a significant elevation of melatonin levels in medetomidine-injected rats was found 2 h later than in control rats. We interpret the first-night delay to be a sign of medetomidine's suppressive effect on melatonin synthesis, and the second-night delay a medetomidine-induced resetting of the circadian clock controlling the melatonin onset.

Ontogeny of pineal melatonin rhythm in rats under 12:12-hr and 14:14-hr light:dark conditions.

The aim of the study was to determine whether a discrepancy between the genetically determined endogenous circadian period and an abnormally long Zeitgeber period disturbs the development of melatonin synthesis. Breeding pairs of rats were kept under 12:12- or 14:14-hr light:dark (LD) conditions. Pineal melatonin contents in the offspring were measured by radioimmunoassay. At 2 weeks of age high melatonin contents were found from lights-off to lights-on in both conditions suggesting dominance of the photic regulation. At 3 weeks of age the signs of the circadian regulation in the melatonin profiles were evident: a lag period after the light offset in control conditions and a significant decline before the light onset in both conditions. However, in 14:14-hr LD conditions the melatonin content did not decrease to daytime levels until the lights were on. This could suggest incomplete maturation of the circadian system. The phase relationships between the melatonin peak and LD cycle were different in the two conditions. A statistically significant LD difference was first found at the age of 8-10 days in male pups and at 14 days in female pups under both lighting. The results suggest that the abnormally long LD cycle did not cause any major disorders in the development of photic or circadian regulation of the melatonin synthesis.

Exogenous melatonin fails to counteract the light-induced phase delay of human melatonin rhythm.

Salivary melatonin levels were measured in 6 healthy volunteers in order to determine whether the phase shift caused by a single 60-min light pulse of 2000 lux might be inhibited by maintaining high melatonin concentration. In the control sessions, the samples were collected at 60-min intervals under lighting of < 10 lux from 18.00 to 11.00 h. In the light-exposure sessions, placebo or 0.5 mg melatonin was administered orally 60 min prior to the light pulse, timed at the rising phase of the melatonin synthesis. The after-light sessions, one day after the light exposure, were like the control sessions. The average delays of the melatonin half-rise and half-decline times were equal (about 0.7 h) in the placebo and melatonin replacement experiments. The maintenance of high melatonin levels during the light exposure did not counteract the influence of bright light on the melatonin rhythm. Thus, in the adjustment of the melatonin rhythm, light is a stronger regulator than melatonin itself.

Daytime dark exposure increases pineal melatonin in rat pups.

Photic regulation of the pineal melatonin synthesis was studied in 3- to 21-day-old rat pups by exposing the animals to light at night (30-40 min) or to darkness during the day (30-240 min). The pineal melatonin contents were measured by radioimmunoassay. A significant day/night difference in the melatonin content and the nocturnal light-induced decrease were not found until second postnatal week. A novel finding was that at the age of 13-17 days a daytime dark exposure elevated the pineal melatonin content; it was twofold as compared with the normal daytime level and about half of the nocturnal peak level. In 21-day-old rats the response had disappeared, while the nocturnal suppression by light persisted. The dark-induced increase of the melatonin synthesis was independent of the opening of the eyelids which occurs in pups at the age of two weeks, but it was greater in maternally isolated than non-isolated pups. The results suggest that one component of the circadian regulatory system matures at the end of the third postnatal week. This mechanism inhibits the elevation of the melatonin synthesis by darkness during the daytime.

Circadian rhythm studies in neuronal ceroid-lipofuscinosis (NCL).

Sleep disorders are common in NCL patients. The patients have problems such as frequent awakenings, difficulties with sleep onset, nightmares, and night terrors. The aim of the study was to examine whether the sleep disturbance in NCL can be explained on the basis of desynchronised circadian rhythms. Therefore we studied diurnal patterns of melatonin, cortisol, body temperature, and motor activity of 14 patients. The group consisted of 8 JNCL patients, 5 INCL children, and one boy with Jansky-Bielschowsky disease of the variant type. There were healthy age- and sex-matched control subjects. The blood samples for serum melatonin and cortisol were collected every 2 hours during 24-hour periods. Body temperature was recorded continuously for a 24-hour period by a polygraph. Diurnal motor activity was measured by wrist actigraphy for 5 days. In most of our patients sleep was fragmented and the sleep phase was irregular. Disturbances in the daily hormonal rhythms occurred only in the minority of the patients and only at an advanced stage of the disease. Although disturbances in the body temperature rhythm were found in about half of the patients, a general failure in the circadian regulatory system does not explain the frequent disturbances of the sleep-wake cycle of the NCL patients.

Uncoupling of the pineal melatonin synthesis of rats from the circadian regulation.

We show that the pineal melatonin synthesis of rats can be uncoupled from the circadian regulation by exposing the animals to abnormally long light periods. Male rats were kept 7 days under 22.5/1.5-h light/dark conditions and then exposed to darkness at different times of the day. After a 60-min dark exposure, the melatonin synthesis increased independently of the time of the day (Expt. 1). During 22 h in darkness, the mean melatonin content did not return to the low daytime level (Expt. 2). A dark-induced, time-independent increase of melatonin was also found after the rats had been 3-7 days under constant light (Expts. 3 and 4).

The adjustment of the melatonin rhythm of rats by 90-min dark pulses.

The pineal melatonin patterns were measured in rats kept 1, 2, 3, 4, 6, 8, 16, or 28 days under the lighting conditions with a dark period of 90 min only (from 18:00 to 19:30 h). The samples were collected at 30-min intervals, and melatonin was measured by radioimmunoassay. During the first and second dark pulse the melatonin levels were low, but thereafter they were significantly higher than the usual daytime levels. The increase of the synthesis started within 15 min after the light offset. The results suggest a stable adjustment of the melatonin rhythm by the dark pulses.

One-hour exposure to moderate illuminance (500 lux) shifts the human melatonin rhythm.

Salivary melatonin levels were measured in 12 healthy volunteers in order to determine whether a moderate light intensity, which suppresses the nocturnal rise of melatonin, was able to shift the melatonin rhythm. The samples were collected at 1-hr intervals under lighting of < 100 lux (experiment 1) or < 10 lux (experiment 2). The control melatonin profiles were determined during the first night. In the second night the subjects were exposed to light of 500 lux for 60 min during the rising phase of melatonin synthesis. The third series of samples was collected during the third night. The mean decrease of melatonin levels by the exposure to light was 56% of the prelight concentrations. The melatonin onset times were delayed significantly (about 30 min) the night after the exposure to light. The melatonin offset times tended to be delayed in experiment 2. The shifts of the melatonin offset correlated positively with the amount of the melatonin suppression. The results suggest that a relatively small and short lasting light-induced interruption of melatonin synthesis may affect the melatonin rhythm in humans.

Melatonin, cortisol and body temperature rhythms in Lennox-Gastaut patients with or without circadian rhythm sleep disorders.

The daily rhythms of melatonin, cortisol and body temperature were studied in 16 institutionalized subjects with the Lennox-Gastaut syndrome. The results of 9 subjects with normal daily rhythms of sleep and wakefulness (group 1) were compared with those of 7 subjects with disordered sleep (group 2). Salivary samples were collected and axillary temperature was measured every 2 h during two or three separate 26-h periods. The hormones were measured by radioimmunoassays. The rhythms were characterized with single cosinor analysis. Two subjects in group 1 and six subjects in group 2 had abnormalities in their rhythms of temperature, cortisol or melatonin. All three rhythms were disrupted in two subjects of group 2. These two subjects were the only ones with disrupted cortisol rhythm. The diversity of rhythm pathologies suggested partly separate regulatory mechanisms for each rhythm. The co-occurrence of circadian rhythm sleep disorders with the deteriorated melatonin rhythm raised the question as to whether the sleep disorders of these subjects, like those of subjects with healthy brains, could be relieved by the induction of normal melatonin rhythm.