Effect of long-term exposure to constant dim light on the circadian system of rats

The newly discovered multi-oscillatory nature of the mammalian circadian clock system and the cloning of the genes involved in the molecular mechanism that generates circadian rhythmicity have opened new approaches for understanding how mammals are temporally organized and how the mammalian circadian system reacts to the lack of normal synchronization cues. In the present study we investigated the effects of long-term exposure to constant red dim light on the pattern of the expression of Period 1 in the suprachiasmatic nuclei of the hypothalamus and of Arylalkylamine N-acetyltransferase(Aa-nat) in the retina and pineal gland. Our data demonstrate that Period 1 mRNA expression in the suprachiasmatic nuclei of the hypothalamus was not affected by exposure to constant red dim light for 60 days, whereas Aa-nat mRNA expression in the retina and in the pineal gland was significantly affected, since in some animals (20-30%) Aa-nat mRNA levels were found to be higher during the subjective day. A circadian rhythm of serum melatonin and locomotor activity was present in all the animals tested. In 4 animals serum melatonin levels were high during the subjective day. Our data suggest that long-term exposure to constant red dim light may induce desynchronization between the circadian rhythm of locomotor activity and serum melatonin levels.     

肾上腺糖皮质激素与生物钟基因表达调控的相关研究进展

由生物体内源性生物钟所产生的昼夜节律是近年来生命科学的研究热点之一。哺乳动物中的昼夜节律系统由位于下丘脑SCN核内的主钟和位于多数外周细胞中的子钟组成。生物钟基因及其编码的蛋白质组成反馈回路,维持振荡系统持续进行并与环境周期保持同步。光照和食物是生物钟重要的授时因子, 光照刺激能引起肾上腺中基因表达变化以及糖皮质激素的分泌, 而肾上腺糖皮质激素能减缓由食物因子引起的外周生物钟时相的移动。可见, 肾上腺糖皮质激素与生物钟有着非常密切的关系。文章综述了两者的相互影响并对今后的研究方向做了展望。     

Programming of Mice Circadian Photic Responses by Postnatal Light Environment

Early life programming has important consequences for future health and wellbeing. A key new aspect is the impact of perinatal light on the circadian system. Postnatal light environment will program circadian behavior, together with cell morphology and clock gene function within the suprachiasmatic nucleus (SCN) of the hypothalamus, the principal circadian clock in mammals. Nevertheless, it is still not clear whether the observed changes reflect a processing of an altered photic input from the retina, rather than an imprinting of the intrinsic molecular clock mechanisms. Here, we addressed the issue by systematically probing the mouse circadian system at various levels. Firstly, we used electroretinography, pupillometry and histology protocols to show that gross retinal function and morphology in the adult are largely independent of postnatal light experiences that modulate circadian photosensitivity. Secondly, we used circadian activity protocols to show that only the animal''s behavioral responses to chronic light exposure, but not to constant darkness or the acute responses to a light stimulus depend on postnatal light experience. Thirdly, we used real-time PER2::LUC rhythm recording to show long-term changes in clock gene expression in the SCN, but also heart, lung and spleen. The data showed that perinatal light mainly targets the long-term adaptive responses of the circadian clock to environmental light, rather than the retina or intrinsic clock mechanisms. Finally, we found long-term effects on circadian peripheral clocks, suggesting far-reaching consequences for the animal''s overall physiology.     

Prenatal ethanol exposure alters the expression of period genes governing the circadian function of beta-endorphin neurons in the hypothalamus

Sleep-wake disturbances and stress hyper-responsiveness have been observed in human neonates, children and adolescents who were exposed to alcohol during the prenatal period. Using the laboratory rat as an animal model, we investigated whether fetal ethanol exposure during gestational days 10-21 affects the circadian function of the stress-axis regulatory beta-endorphin neurons in the hypothalamus. Fetal ethanol-exposed rats showed abnormality in the circadian expression of proopiomelanocortin (POMC) mRNA encoding the peptide beta-endorphin in the arcuate nucleus of the hypothalamus during the adult period. These rats also showed altered circadian expression of the clock governing Period genes rPer1, rPer2 and rPer3, in the arcuate nucleus, and rPer1 and rPer 2 mRNA levels in the suprachiasmatic nucleus. Laser captured microdissection analysis identified constitutive expression of rPer1, rPer2 and rPer3 genes in beta-endorphin-containing neurons. These data suggest for the first time that fetal exposure to ethanol significantly alters the clock mechanisms governing the circadian function of beta-endorphin neurons.     

Neural adrenergic/cyclic AMP regulation of the immunoglobulin E receptor alpha-subunit expression in the mammalian pinealocyte: a neuroendocrine/immune response link?

The high affinity immunoglobulin E receptor (FcepsilonRI) complex is dedicated to immunoglobulin E-mediated allergic responses. Expression of the FcepsilonRI receptor is thought to be relatively stable and limited to mast cells, basophils, eosinophils, monocytes, Langerhans cells, platelets, and neutrophils. We now report that the FcepsilonRIalpha and FcepsilonRIgamma polypeptides are expressed in the pinealocyte, the melatonin-secreting cell of the pineal gland. Moreover, Fcer1a mRNA levels increased approximately 100-fold at night to levels that were higher than in other tissues examined. Pineal FcepsilonRIalpha protein also increased markedly at night from nearly undetectable daytime levels. Our studies indicate that pineal Fcer1a mRNA levels are controlled by a well described neural pathway that controls pineal function. This pathway includes the master circadian oscillator in the suprachiasmatic nucleus and passes through central and peripheral structures. The circadian expression of FcepsilonRIalpha in the pineal gland is driven by this neural circuit via an adrenergic/cyclic AMP mechanism. Pineal FcepsilonRIalpha and FcepsilonRIgamma may represent a previously unrealized molecular link between the neuroendocrine and immune systems.     

Chronic ethanol consumption impairs the circadian rhythm of pro-opiomelanocortin and period genes mRNA expression in the hypothalamus of the male rat

Certain psychiatric disorders are known to alter the body's biological rhythms. However, currently, very little information is known about the effect of chronic ethanol administration on the circadian clock or the rhythm of beta-endorphin-containing neurons that participate in the control of the reward and reinforcement of alcohol drinking. Here, we report that administration of ethanol, via a liquid diet paradigm for a period of 2 weeks, abolishes the circadian rhythm of pro-opiomelanocortin mRNA expression of beta-endorphin neurons in the arcuate nucleus of the hypothalamus. The circadian expression of the clock governing rat period genes (rPeriod1 mRNA and rPeriod2 mRNA) in the arcuate nucleus was significantly altered, suggesting that ethanol administration disrupted the internal clock. Moreover, ethanol consumption altered the circadian rhythms of rPeriod2 and rPeriod3 mRNA levels in the suprachiasmatic nucleus, suggesting that ethanol also affected the function of the central pacemaker. Our findings identified the vulnerability of the body's clock machinery and its opioidergic system to chronic alcohol drinking.     

Enhanced effect of daytime restricted feeding on the circadian rhythm of streptozotocin-induced type 2 diabetic rats

There is increasing awareness of the link between impaired circadian clocks and multiple metabolic diseases. However, the impairment of the circadian clock by type 2 diabetes has not been fully elucidated. To understand whether and how the function of circadian clock is impaired under the diabetic condition, we examined not only the expression of circadian genes in the heart and pineal gland but also the behavioral rhythm of type 2 diabetic and control rats in both the nighttime restricted feeding (NRF) and daytime restricted feeding (DRF) conditions. In the NRF condition, the circadian expression of clock genes in the heart and pineal gland was conserved in the diabetic rats, being similar to that in the control rats. DRF shifted the circadian phases of peripheral clock genes more efficiently in the diabetic rats than those in the control rats. Moreover, the activity rhythm of rats in the diabetic group was completely shifted from the dark phase to the light phase after 5 days of DRF treatment, whereas the activity rhythm of rats in the control group was still under the control of the suprachiasmatic nucleus (SCN) after the same DRF treatment. Furthermore, the serum glucose rhythm of type 2 diabetic rats was also shifted and controlled by the external feeding schedule, ignoring the SCN rhythm. Therefore, DRF shows stronger effect on the reentrainment of circadian rhythm in the type 2 diabetic rats, suggesting that the circadian system in diabetes is unstable and more easily shifted by feeding stimuli.     

Embryonic development and maternal regulation of murine circadian clock function

The importance of circadian clocks in the regulation of adult physiology in mammals is well established. In contrast, the ontogenesis of the circadian system and its role in embryonic development are still poorly understood. Although there is experimental evidence that the clock machinery is present prior to birth, data on gestational clock functionality are inconsistent. Moreover, little is known about the dependence of embryonic rhythms on maternal and environmental time cues and the role of circadian oscillations for embryonic development. The aim of this study was to test if fetal mouse tissues from early embryonic stages are capable of expressing endogenous, self-sustained circadian rhythms and their contribution to embryogenesis. Starting on embryonic day 13, we collected precursor tissues for suprachiasmatic nucleus (SCN), liver and kidney from embryos carrying the circadian reporter gene Per2::Luc and investigated rhythmicity and circadian traits of these tissues ex vivo. We found that even before the respective organs were fully developed, embryonic tissues were capable of expressing circadian rhythms. Period and amplitude of which were determined very early during development and phases of liver and kidney explants are not influenced by tissue preparation, whereas SCN explants phasing is strongly dependent on preparation time. Embryonic circadian rhythms also developed in the absence of maternal and environmental time signals. Morphological and histological comparison of offspring from matings of Clock-Δ19 mutant and wild-type mice revealed that both fetal and maternal clocks have distinct roles in embryogenesis. While genetic disruptions of maternal and embryonic clock function leads to increased fetal fat depots, abnormal ossification and organ development, Clock gene mutant newborns from mothers with a functional clock showed a larger body size compared to wild-type littermates. These data may contribute to the understanding of the ontogenesis of circadian clocks and the risk of disturbed maternal or embryonic circadian rhythms for embryonic development.     

Circadian rhythms of indoleamines and serotonin N-acetyltransferase activity in the pineal gland

Conclusion The circadian rhythm of melatonin synthesis in the pineal glands of various species has been summarized. The night-time elevation of melatonin content is in most if not all cases regulated by the change of N-acetyltransferase activity. In mammals, the N-acetyltransferase rhythm is controlled by the central nervous system, presumably by suprachiasmatic nuclei in hypothalamus through the superior cervical ganglion. In birds, the circadian oscillator that regulates the N-acetyltransferase rhythm is located in the pineal glands. The avian pineal gland may play a biological clock function to control the circadian rhythms in physiological, endocrinological and biochemical processes via pineal hormone melatonin.     

Altered behavioral rhythms and clock gene expression in mice with a targeted mutation in the Period1 gene

A group of specialized genes has been defined to govern the molecular mechanisms controlling the circadian clock in mammals. Their expression and the interactions among their products dictate circadian rhythmicity. Three genes homologous to Drosophila period exist in the mouse and are thought to be major players in the biological clock. Here we present the generation of mice in which the founding member of the family, Per1, has been inactivated by homologous recombination. These mice present rhythmicity in locomotor activity, but with a period almost 1 h shorter than wild-type littermates. Moreover, the expression of clock genes in peripheral tissues appears to be delayed in Per1 mutant animals. Importantly, light-induced phase shifting appears conserved. The oscillatory expression of clock genes and the induction of immediate-early genes in response to light in the master clock structure, the suprachiasmatic nucleus, are unaffected. Altogether, these data demonstrate that Per1 plays a distinct role within the Per family, as it may be involved predominantly in peripheral clocks and/or in the output pathways of the circadian clock.     

Organisation of the circadian system in melatonin-proficient C3H and melatonin-deficient C57BL mice: a comparative investigation

In all vertebrates melatonin is rhythmically synthesised in the pineal gland and functions as a hormonal message encoding for the duration of darkness. This review focuses on the role of melatonin in the circadian organisation of mammals by comparing signal transduction mechanisms in the pineal organ, the suprachiasmatic nucleus and the hypophyseal pars tuberalis in melatonin-proficient (C3H) and melatonin-deficient (C57BL) mice strains. Surprisingly, the major signal transduction cascades in the pineal organ did not differ between the two mouse strains. With regard to the suprachiasmatic nucleus, the site of the endogenous clock, it was found that melatonin at most sets the gain for clock error signals mediated via the retinohypothalamic tract, but has no effect on the rhythm generation itself or on the maintenance of the oscillation. In contrast, melatonin plays an essential role in the control of the hypophyseal pars tuberalis. Here it acts in concert with adenosine to elicit rhythms in clock gene expression. Melatonin opens a temporally restricted gate for adenosine to induce cyclic AMP (cAMP)-sensitive genes by sensitising the adenylyl cyclase. This interaction, which grants a temporally precise regulation of gene expression, may reflect the central role of melatonin, i.e. in synchronising peripheral clock cells that require unique phasing of output signals with the master clock in the brain.     

Circadian gene expression in individual fibroblasts: cell-autonomous and self-sustained oscillators pass time to daughter cells

The mammalian circadian timing system is composed of a central pacemaker in the suprachiasmatic nucleus (SCN) of the brain and subsidiary oscillators in most peripheral cell types. While oscillators in SCN neurons are known to function in a self-sustained fashion, peripheral oscillators have been thought to damp rapidly when disconnected from the control exerted by the SCN. Using two reporter systems, we monitored circadian gene expression in NIH3T3 mouse fibroblasts in real time and in individual cells. In conjunction with mathematical modeling and cell co-culture experiments, these data demonstrated that in vitro cultured fibroblasts harbor self-sustained and cell-autonomous circadian clocks similar to those operative in SCN neurons. Circadian gene expression in fibroblasts continues during cell division, and our experiments unveiled unexpected interactions between the circadian clock and the cell division clock. Specifically, the circadian oscillator gates cytokinesis to defined time windows, and mitosis elicits phase shifts in circadian cycles.     

Resetting of peripheral circadian clock by prostaglandin E2

In mammals, the master circadian pacemaker is located in the suprachiasmatic nucleus (SCN) of the hypothalamus. The SCN is thought to drive peripheral oscillators by controlling neuronal and humoral signals that can entrain the peripheral clocks. Here, we show that prostaglandin E2 (PGE2), a proinflammatory compound known to have diverse biological effects, is able to act as an in vivo clock-resetting agent. We find that in cultured NIH3T3 fibroblasts, PGE2 is able to induce transient expression of Period 1 messenger RNA and the following circadian oscillation of clock gene expression. Furthermore, we demonstrate that intraperitoneal administration of PGE2 results in the phase shift of circadian gene expression in mouse peripheral tissues in a time-dependent manner. This phase shift is also induced by the EP1/EP3 agonist sulprostone but not by the EP2 agonist butaprost. The PGE2-induced phase shift is inhibited by the EP1 antagonist SC-51322. These results suggest that PGE2 acts as an in vivo clock-resetting factor by means of the EP1 subtype of PGE receptors.     

Antibodies for assessing circadian clock proteins in the rodent suprachiasmatic nucleus

Research on the mechanisms underlying circadian rhythmicity and the response of brain and body clocks to environmental and physiological challenges requires assessing levels of circadian clock proteins. Too often, however, it is difficult to acquire antibodies that specifically and reliably label these proteins. Many of these antibodies also lack appropriate validation. The goal of this project was to generate and characterize antibodies against several circadian clock proteins. We examined mice and hamsters at peak and trough times of clock protein expression in the suprachiasmatic nucleus (SCN). In addition, we confirmed specificity by testing the antibodies on mice with targeted disruption of the relevant genes. Our results identify antibodies against PER1, PER2, BMAL1 and CLOCK that are useful for assessing circadian clock proteins in the SCN by immunocytochemistry.