Entrainment of the circadian rhythm in the rat pineal N-acetyltransferase activity by melatonin is photoperiod dependent

Entraining effect of melatonin on the circadian rhythm in rat pineal N-acetyltransferase (NAT) activity was studied under various photoperiods. Melatonin administration prior to dark onset for 5 successive days phase-advanced the evening NAT rise under the light:dark (LD) cycle of either LD 10:14 or LD 8:16, but not under LD 12:12. It is assumed that under the latter regime, the end of a light period exhibited a phase-delaying effect on the NAT rise. The light exposure appeared to be a stronger Zeitgeber than melatonin itself. Data show that melatonin applied in the late light period advances the evening NAT rise under a short photoperiod only; under a longer photoperiod, the phase-advancing effect of melatonin may conflict with a phase-delaying effect of the end of a light period, and the effect of light exposure overrides that of melatonin.     

Phase-shifting effects of light on the circadian rhythms of 5-methoxytryptophol and melatonin in the chick pineal gland

In the chick pineal gland, 5-methoxytryptophol and melatonin concentrations fluctuate in a rhythmic manner. These rhythms are circadian in nature persisting in constant darkness and have opposite phases. Acute exposure of chicks to white light (30 lux for 5, 10, 20, and 30 min) at night increased the amount of pineal 5-methoxytryptophol and decreased pineal melatonin content. A 6 hr pulse of light (100 lux) applied early in the subjective night (CT12-CT18) caused a delay in the phase of the circadian rhythms of 5-methoxytryptophol and melatonin by 3.7 and 4.5 h, respectively, compared to untreated controls. When the 6 hr light pulse was given during the late subjective night (C18 CT24) it advanced the phase of the 5-methoxytryptophol and melatonin rhythms by 8.1 and 11.9 h, respectively. In the chick pineal the phase-advancing effects of light on the circadian rhythms of 5-methoxytryptophol and melatonin were more pronounced than the phase-delaying effects. Our results provide the first evidence that light is capable of phase shifting the 5-methoxytryptophol rhythm in a manner similar to its action on the melatonin rhythm.     

The human pineal gland and melatonin in aging and Alzheimer's disease

The pineal gland is a central structure in the circadian system which produces melatonin under the control of the central clock, the suprachiasmatic nucleus (SCN). The SCN and the output of the pineal gland, i.e. melatonin, are synchronized to the 24-hr day by environmental light, received by the retina and transmitted to the SCN via the retinohypothalamic tract. Melatonin not only plays an important role in the regulation of circadian rhythms, but also acts as antioxidant and neuroprotector that may be of importance in aging and Alzheimer's disease (AD). Circadian disorders, such as sleep-wake cycle disturbances, are associated with aging, and even more pronounced in AD. Many studies have reported disrupted melatonin production and rhythms in aging and in AD that, as we showed, are taking place as early as in the very first preclinical AD stages (neuropathological Braak stage I-II). Degeneration of the retina-SCN-pineal axis may underlie these changes. Our recent studies indicate that a dysfunction of the sympathetic regulation of pineal melatonin synthesis by the SCN is responsible for melatonin changes during the early AD stages. Reactivation of the circadian system (retina-SCN-pineal pathway) by means of light therapy and melatonin supplementation, to restore the circadian rhythm and to relieve the clinical circadian disturbances, has shown promising positive results.     

Perinatal development of circadian melatonin production in domestic chicks

In contrast to the situation in mammals, in which circadian melatonin production by the pineal gland does not begin until some time after birth, the development of pineal gland rhythmicity is an embryonic event in the precocial domestic fowl. A distinct melatonin rhythm was found in 19-d-old chick embryos maintained under light:dark (LD) 16:8. No significant variation in melatonin levels was detected in embryos exposed to LD 8:16. The melatonin rhythm in the pineal gland and plasma of chick embryos incubated for 18 d in LD 12:12 persisted for 2 d in constant darkness indicating that melatonin production is under circadian control at least from the end of embryonic life. A 1-d exposure to a LD cycle during the first postembryonic day was sufficient to entrain the melatonin rhythm, and previous embryonic exposure to either LD or constant darkness (DD) neither modified this rapid synchronization nor did it affect the melatonin pattern during the two subsequent days in DD. It is suggested that, in contrast to the situation in mammals, the avian embryo has evolved its own early circadian melatonin-producing system because, as a consequence of its extrauterine development, it cannot use the system of its mother.     

Intracellular Ca2+ signaling pathway is involved in light-induced phase advance, but may not be in phase delay, of the circadian melatonin rhythm in chick pineal cell

Chick pineal cells have photoreceptive, circadian clock and melatonin synthetic capacities, and express circadian oscillation of melatonin release in vitro. Light pulses cause phase-dependent phase shift of the melatonin rhythm. The purpose of this study was to address the questions whether intracellular calcium is involved in both light-induced phase advance and delay. Thapsigargin and cyclopiazonic acid, which deplete the intracellular calcium stores, blocked the light-induced phase advance in a dose-dependent manner. The pulses of ryanodine receptor antagonist (dantrolene sodium or ruthenium red) also blocked the light-induced phase advance. Most agents did not cause a significant phase shift by themselves. On the other hand, all the agents used, failed to block the light-induced phase delay, even if the magnitude of phase delay was decreased using low intensity light. An antagonist of nitric oxide synthase blocked neither light-induced phase advance nor phase delay. These results indicate the following possibilities: (1) the mechanism of light-induced phase advance and delay may be different in chick pineal cells, or (2) if intracellular calcium is involved in both light-induced phase advance and delay, the sensitivity to light and/or agents used in this study may differ according to Zeitgeber time.     

Effect of the pineal gland on circadian rhythmicity of colony forming units for granulocytes and macrophages (CFU-GM) from rat bone marrow cell cultures

The present study provides evidence that the pineal gland has a physiological role in the proliferation of colony forming units for granulocytes and macrophages (CFU-GM). A biphasic circadian rhythm of CFU-GM proliferation in rat bone marrow cell cultures (BMC) from intact animals peaking at 0600 and 1800 was observed. Pinealectomy (Px) at 1600 obliterated the circadian rhythm patterns of CFU-GM. Afternoon injections of melatonin (1630, 20 micrograms/per day for 10 days) to Px animals restored the rhythmicity. When pinealectomy was done at 0800, the morning peak remained unaltered and the colony number at 1800 was higher than that found in the afternoon Px animals. In conclusion, the pineal gland or its main hormone melatonin seems to have a regulatory role in the proliferation of CFU-GM in rat BMC. Further, the expression of the activity of CFU-GM in rat BMC depends on the time when pinealectomy is done or melatonin is substituted.     

Putting cancer to sleep at night: the neuroendocrine/circadian melatonin signal

Physiological and pharmacological blood concentrations of melatonin inhibit tumorigenesis in a variety of in vivo and in vitro experimental models of neoplasia. Evidence indicates that melatonin’s anticancer effects are exerted via inhibition of cell proliferation and a stimulation of differentiation and apoptosis. A new mechanism by which physiological and pharmacological blood levels of melatonin inhibit cancer growth in vivo is via a melatonin-induced suppression of tumor linoleic acid (LA) uptake and its metabolism to the important mitogenic signaling molecule 13-hydroxyoctadecadienoic acid (13-HODE). Melatonin suppresses cAMP formation and inhibits tumor uptake of LA and its metabolism to 13-HODE via a melatonin receptor-mediated mechanism in both tissue-isolated rat hepatoma 7288 CTC and human breast cancer xenografts. It has been postulated that in industrialized societies, light at night, by suppressing melatonin production, poses a new risk for the development of breast cancer and, perhaps, other cancers as well. In support of this hypothesis, light during darkness suppresses nocturnal melatonin production and stimulates the LA metabolism and growth of rat hepatoma and human breast cancer xenografts. Nocturnal dietary supplementation with melatonin, at levels contained in a melatonin-rich diet, inhibits rat hepatoma growth via the mechanisms described above. The nocturnal melatonin signal organizes tumor metabolism and growth within circadian time structure that can be further reinforced by appropriately timed melatonin supplementation. Dietary melatonin supplementation working in concert with the endogenous melatonin signal has the potential to be a new preventive/therapeutic strategy to optimize the host/cancer balance in favor of host survival and quality of life.     

Roles of nocturnal melatonin and the pineal gland in modulation of water-immersion restraint stress-induced gastric mucosal lesions in rats

The roles of melatonin and the pineal gland in the circadian variation of water-immersion restraint stress-induced gastric mucosal lesions in rats were investigated. Fasted rats were subjected to water-immersion restraint stress during both the diurnal and nocturnal phases of a light:dark cycle. Pinealectomized and sham-operated rats were also subjected to water-immersion restraint stress at night. The lesion area after 4 hr of stress during the dark phase was significantly lower than in light-phase controls. Pinealectomy increased the lesion area in the dark phase, compared to the sham operation, but this effect was counteracted by intracisternal melatonin preadministration at a dose of 100 ng/rat. Melatonin concentrations in control rats during the light phase were significantly increased 4 hr after water-immersion restraint stress. In contrast, melatonin concentrations 4 hr after water-immersion restraint stress in the dark phase were significantly depressed compared with the control levels at the corresponding time. Melatonin levels after stress exposure were markedly decreased in pinealectomized rats as compared with sham-operated rats. These results suggest that circadian rhythm has an important role in the formation of stress-induced gastric mucosal lesions in rats and that melatonin responses to water-immersion restraint stress differ between day and night. The pineal gland modulates the stress response and melatonin contributes to gastric protection via a mechanism involving the central nervous system.     

A single injection of adrenergic agonists enhances pineal melatonin production in Turkish hamsters

Abstract: The purpose of this study was to determine whether the pineal gland of Turkish hamsters (Mesocricetus brandti) responds to adrenergic agonists with an increase in melatonin production, and, if it does, whether the sensitivity of the pineal gland to agonists would differ throughout the dark phase. Adult Turkish hamsters weighing 110–210 g received a subcutaneous injection of isoproterenol (ISO, 1 mg/kg B.W.) or norepinephrine (NE, 1 mg/kg B.W.) at different times of night. Animals exposed to LD 16:8 responded to ISO or NE with increased pineal melatonin content only when injected at dawn, when endogenous melatonin is at basal or near-basal levels. When the 8 hr scotophase was entirely replaced with light, the responsiveness to ISO injections at dawn disappeared. In animals exposed to light from 30 min prior to injection to the time of sacrifice, ISO injections increased pineal melatonin content (P < 0.005, three-way ANOVA), which varied, depending on the specific time of injection (effect of time of night, P < 0.05, three-way ANOVA). These results demonstrate that (1) adrenergic agonists enhance the production of pineal melatonin in Turkish hamsters, (2) this stimulatory effect takes place late, but not early in the 8 hr scotophase, and (3) the adrenergic induction of pineal melatonin production in Turkish hamsters requires priming by darkness during the appropriate circadian phase.     

Physiological levels of melatonin contribute to the antioxidant capacity of human serum

This work evaluates whether physiological concentrations of the pineal secretory product melatonin contribute to the total antioxidant status (TAS) of human serum. Day and nighttime serum samples were collected from healthy volunteers ranging from 2 to 89 years of age and used to measure melatonin and TAS. Results showed that both melatonin and TAS in human serum exhibited 24 hr variations with nocturnal peak values at 01:00 hr. Moreover, exposure of volunteers to light at night resulted in clear decreases of both TAS and melatonin. Furthermore, when melatonin was removed from sera collected at night, the TAS value of the sample was reduced to basal daytime values. In aging studies, it was found that nocturnal serum values of TAS and melatonin exhibited maximal values during the first four decades; thereafter, these values decreased as age advanced. In 60-year-old individuals, day/night differences in serum melatonin and TAS levels were clearly diminished, by more than 80%, with these differences being completely abolished in older individuals. Our results suggest that melatonin contributes to the total antioxidative capability of human serum. This antioxidant contribution of melatonin is reduced as age advances correlating with the age-related reduction of melatonin.     

Evidence for differential photic regulation of pineal melatonin synthesis in teleosts

The aim of this study was to compare the circadian control of melatonin production in teleosts. To do so, the effects of ophthalmectomy on circulating melatonin rhythms were studied along with ex vivo pineal culture in six different teleosts. Results strongly suggested that the circadian control of melatonin production could have dramatically changed with at least three different systems being present in teleosts when one considers the photic regulation of pineal melatonin production. First, salmonids presented a decentralized system in which the pineal gland responds directly to light independently of the eyes. Then, in seabass and cod both the eyes and the pineal gland are required to sustain full night-time melatonin production. Finally, a third type of circadian control of melatonin production is proposed in tilapia and catfish in which the pineal gland would not be light sensitive (or only slightly) and required the eyes to perceive light and inhibit melatonin synthesis. Further studies (anatomical, ultrastructural, retinal projections) are needed to confirm these results. Ex vivo experiments indirectly confirmed these results, as while the pineal gland responded normally to day-night rhythms in salmonids, seabass and cod, only very low levels were obtained at night in tilapia and no melatonin could be measured from isolated pineal glands in catfish. Together, these findings suggest that mechanisms involved in the perception of light and the transduction of this signal through the circadian axis has changed in teleosts possibly as a reflection of the photic environment in which they have evolved in.     

Chronic administration of sublethal doses of carbaryl increases pineal N-acetyltransferase and hydroxyindole-O-methyltransferase activities and serum melatonin levels

The purpose of this study was to examine the effects of chronic administration of sublethal doses of carbaryl on pineal melatonin synthesis. N-methyl 1-naphthylcarbamate (carbaryl) (8.33 mg/kg B.W. daily) was administered orally to adult male albino rats for 6 successive days. Nocturnal (0100) N-acetyltransferase and hydroxyindole-O-methyltransferase activities were increased (roughly 75% and 60%, respectively) by carbaryl administration; likewise, carbaryl augmented serum melatonin levels at 2300. Pineal tryptophan. 5-hydroxytryptophan, serotonin, and 5-hydroxindole acetic acid levels were unaffected at all three time points. The results indicate that the carbamate pesticide, i.e., carbaryl, modifies pineal melatonin synthesis in vivo.     

HPLC validation of a circadian melatonin rhythm in the pineal gland of inbred mice

Abstract: Production of melatonin in the pineal gland of inbred mice such as C57B1/6J, B ALB/c and AKR strains is still a matter of debate. In previous studies, we and other authors showed that these strains of inbred mice have a clear-cut circadian rhythm of serum melatonin and urinary 6-hydroxy-melatonin-sulphate. In contrast, other groups claimed these mice are unable to synthesize melatonin. These studies were based on RIA measurements and/or estimates of N-acetyltransferase (NAT) and hydroxyindole-O-methyltransferase (HIOMT) activities. In the present study we validate the presence of melatonin in the pineal gland of C57B1/6, BALB/c, and AKR mice by HPLC determinations. We found a short-term melatonin peak in the middle of the dark period with a pattern which mirrors that found previously in the serum. The possibility remains, although it seems unlikely, that the pineal melatonin rhythm measured here represents melatonin produced elsewhere which then was subsequently taken up by the pineal gland.     

Qualitative and quantitative changes of melatonin levels in physiological and pathological aging and in centenarians

Melatonin secretion is an endogenous synchronizer, and it may possess some anti-aging properties. Thus we examined melatonin levels in physiological aging, in extreme senescence and in senile dementia. In healthy old (age 66-94 yr) and young subjects (age 23-39 yr) and in demented patients (age 68-91 yr) plasma melatonin was measured by radioimmunoassay in eight serial blood samples. In centenarians (age 100-107 yr) melatonin levels were estimated by assaying urinary 6-hydroxymelatonin sulfate (aMT6s) in two different urine samples collected from 08:00 to 20:00 hours and from 20:00 to 08:00 hours. These data were compared with the aMT6s excretion of old and young controls. Elderly subjects, demented or not, exhibited a flattened circadian profile of plasma melatonin, because of the suppression of the nocturnal peak. An age-related decline of the circadian amplitude of the melatonin rhythm occurred in old subjects, especially in demented individuals. Furthermore, the melatonin nocturnal peak was significantly correlated with the severity of the cognitive impairment. aMT6s urinary excretion also declined with age. However, as in young controls, in centenarians the aMT6s excretion was significantly higher at night than during the day. In conclusion, pineal melatonin secretion is affected by age and by the degree of cognitive impairment. In centenarians the maintenance of the circadian organization of melatonin secretion may suggest that the amplitude of the nocturnal peak and/or the persistence of a prevalent nocturnal secretion may be an important marker of biological age and of health status.     

Sirtuins, melatonin and circadian rhythms: building a bridge between aging and cancer

Abstract:  Histone deacetylases (HDAC) have been under intense scientific investigation for a number of years. However, only recently the unique class III HDAC, sirtuins, have gained increasing investigational momentum. Originally linked to longevity in yeast, sirtuins and more specifically, SIRT1 have been implicated in numerous biological processes having both protective and/or detrimental effects. SIRT1 appears to play a critical role in the process of carcinogenesis, especially in age-related neoplasms. Similarly, alterations in circadian rhythms as well as production of the pineal hormone melatonin have been linked to aging and cancer risk. Melatonin has been found act as a differentiating agent in some cancer cells and to lower their invasive and metastatic status. In addition, melatonin synthesis and release occurs in a circadian rhythm fashion and it has been linked to the core circadian machinery genes ( Clock , Bmal1 , Periods , and Cryptochromes ). Melatonin has also been associated with chronotherapy, the timely administration of chemotherapy agents to optimize trends in biological cycles. Interestingly, a recent set of studies have linked SIRT1 to the circadian rhythm machinery through direct deacetylation activity as well as through the nicotinamide adenine dinucleotide (NAD⁺) salvage pathway. In this review, we provide evidence for a possible connection between sirtuins, melatonin, and the circadian rhythm circuitry and their implications in aging, chronomodulation, and cancer.     

The circadian nature of melatonin secretion in Japanese quail (Coturnix coturnix japonica)

Abstract: Plasma melatonin concentrations were measured in Japanese quail held under different photoperiods and constant darkness (<1 lux). When subjected to LD6:18 (6 hr light: 18 hr darkness), levels rose ～2 hr after lights-off, attained a peak level 8 hr after lights off, and subsequently declined to low daytime levels before the next lights-on signal. This generated a distinct daily rhythm in melatonin secretion with a duration of ～13 h. On exposing quail to a range of photoperiods, containing 6, 9, 11, 12, 13, 15, 18, or 20 hr of light per day, the onset of melatonin secretion remained essentially similar with the rise occurring soon after lights-off. However, the offset of melatonin secretion was suppressed by the light of the next day and thus a much truncated rhythm was produced under long (> 12 hr) photoperiods. Importantly, between night lengths of 4 to 18 hr (i.e., LD 20:4 to LD 6:18) a linear relationship existed between the duration of night-length and secretion of melatonin with the duration increasing by about 0.8 hr for each additional hour of darkness. If quail were released into darkness following a short (LD 6:18) or long (LD 20:4) day schedule, the rhythm persisted for at least two cycles with peaks occurring at about 24 hr intervals. In those quail coming into darkness from long days (LD 20:4), the rhythm of melatonin secretion decompressed rapidly on both sides of the peak, indicating that both the onset and offset of melatonin secretion were suppressed under long days. The endogenous nature of melatonin secretion was tested further by exposing birds to LD 6:30 for 4 cycles and then releasing into darkness. The rhythm in melatonin secretion persisted for at least three cycles before beginning to damp-out. The circadian nature of the rhythm in melatonin secretion was also examined by subjecting quail to T-cycles and then releasing into darkness. Both under the T-cycles and darkness following T-cycle treatments, the phase of the melatonin rhythm was advanced by > 3 hr under T = 27 hr cycles (LD 3:24) compared with T = 24 hr cycles (LD 3:21). This property is consistent with the melatonin oscillator being a circadian rhythm.     

大鼠心脏自噬昼夜节律的老年化改变研究

目的探讨大鼠心脏自噬昼夜节律的老年化改变及可能机制。方法随机选取健康雄性SD大鼠48只,其中6月龄大鼠24只(青年组)和26月龄大鼠24只(老年组)。观察大鼠昼夜活动习性,分别在03:00,06:00,09:00,12:00,15:00,18:00,21:00,24:00各时间点收集心脏样本,运用冷冻切片免疫荧光染色结合激光扫描共聚焦成像技术、亚细胞结构细胞核-细胞质分离技术和免疫印迹技术检测心脏自噬规律及可能调控机制。结果青年组大鼠昼伏夜出,自噬昼夜节律曲线明显,其中18:00 LC3-Ⅰ转化LC3-Ⅱ明显高于12:00,而在15:00细胞核C/EBPβ表达明显高于18:00(P<0.05);而老年组大鼠夜间及昼间均嗜睡,LC3-Ⅰ转化LC3-Ⅱ和细胞核C/EBPβ昼夜无节律性变化。结论衰老改变心脏自噬的昼夜节律,核转录因子C/EBPβ参与其调控。     

Persistence of a plasma melatonin rhythm in constant darkness and its inhibition by constant light in the sleepy lizard, Tiliqua rugosa

This study determined whether a blood plasma melatonin rhythm persists in constant photothermal environments in the sleepy lizard, Tiliqua rugosa. It builds upon an earlier investigation which provided equivocal results as to whether an in vivo melatonin rhythm persists in constant dark (DD) and light (LL) and temperature in this species. Using more frequent sampling points and new assay techniques, the present study showed that the melatonin rhythm persisted for at least 6 days at temperatures of 25 and 33 degrees C in constant dark (DD). The melatonin rhythm, however, was largely eliminated in constant light (LL) at 33 degrees C, thereby contradicting some previous findings in other species of reptiles where melatonin levels were apparently insensitive to an unexpected pulse of light at night. These results demonstrate that the sleepy lizard has a persistent, possibly circadian rhythm of melatonin in DD and constant temperature, and that the rhythm is inhibited by LL and constant temperature. Therefore, the sleepy lizard pineal gland may be an independent oscillator capable of driving the melatonin rhythm and be a transducer of the seasonally changing external photothermal environment.