External Photoperiod Research Articles

Transgenerational effects of maternal circadian melatonin deficiency and melatonin replacement in rats during pregnancy and lactation on the energy metabolism and thermoregulation in the offspring subjected to a high-fat diet

Pineal melatonin participates in the control of numerous biological functions through its immediate effects, which result from its high presence in the systemic circulation during the dark phase of the circadian cycle or, through its prolonged effects, when its level is extremely low during the light phase. At pregnancy, maternal melatonin signals the external photoperiod to the fetus, highlighting its importance not only in synchronizing rhythms, but also in preparing the fetus to adapt the external environments. The fetus and newborns are exclusively dependent on maternal melatonin since their pineal melatonin production only occurs weeks after birth. Thus, maternal hypomelatoninemia stands out as an important factor capable of modulating the physiological systems of their descendants, demonstrating its transgenerational capacity. The present study evaluated the transgenerational influence of maternal melatonin deficiency and replacement during pregnancy on morphometric parameters, thermoregulation and energy metabolism of the offspring submitted to the normal and high-fat diets, respectively. For this, nulliparous Wistar rats at an age of 8 weeks were used and randomized into three groups: CTL (pregnant rats), PINX (pinealectomized pregnant rats), PINX + MEL (pinealectomized pregnant rats submitted to melatonin replacement). After birth, the pups were divided into three groups: (C) pups from control mothers, (P) pups from PINX mothers and (PM) pups from PINX + MEL mothers. One week after weaning, part of the animals was fed a high-fat diet (DH) and rest of them were fed a normal diet (ND) for 12 weeks. Subsequently, the animals were euthanized at ZTs 6 and 18. The results showed that maternal melatonin deficiency disrupted the energy metabolism of the offspring and melatonin replacement normalized the energy metabolism in the offspring submitted to the high-fat diet, enabling them to make functional adaptations such as reduced food consumption and greater thermoregulatory capacity, resulting in reduction in body weight gain white adipose tissue mass.

From gestational chronodisruption to noncommunicable diseases: Pathophysiological mechanisms of programming of adult diseases, and the potential therapeutic role of melatonin.

During gestation, the developing fetus relies on precise maternal circadian signals for optimal growth and preparation for extrauterine life. These signals regulate the daily delivery of oxygen, nutrients, hormones, and other biophysical factors while synchronizing fetal rhythms with the external photoperiod. However, modern lifestyle factors such as light pollution and shift work can induce gestational chronodisruption, leading to the desynchronization of maternal and fetal circadian rhythms. Such disruptions have been associated with adverse effects on cardiovascular, neurodevelopmental, metabolic, and endocrine functions in the fetus, increasing the susceptibility to noncommunicable diseases (NCDs) in adult life. This aligns with the Developmental Origins of Health and Diseasetheory, suggesting that early-life exposures can significantly influence health outcomes later in life. The consequences of gestational chronodisruption also extend into adulthood. Environmental factors like diet and stress can exacerbate the adverse effects of these disruptions, underscoring the importance of maintaining a healthy circadian rhythm across the lifespan to prevent NCDs and mitigate the impact of gestational chronodisruption on aging. Research efforts are currently aimed at identifying potential interventions to prevent or mitigate the effects of gestational chronodisruption. Melatonin supplementation during pregnancy emerges as a promising intervention, although further investigation is required to fully understand the precise mechanisms involved and to develop effective strategies for promoting health and preventing NCDs in individuals affected by gestational chronodisruption.

Melatonin reduced colon inflammation but had no effect on energy metabolism in ageing Mongolian gerbils (Meriones unguiculatus)

In photoperiod-sensitive wild animals, the secretion of melatonin (MT) is modulated by external photoperiod, and MT affects inflammation and the ageing process. The beneficial effects of MT in delaying the progress of ageing have been reported in laboratory mice and rats. However, little is known about MT in wild mammals. In the current study, we investigated energy metabolism, microbial community structure and colon homeostasis in ageing Mongolian gerbils (Meriones unguiculatus) through exogenous supplementation of MT to test the hypothesis that MT has beneficial effects on gut homeostasis in ageing gerbils. Exogenous MT supplementation had no effect on energy metabolism in Mongolian gerbils but reduced the levels of circulating tumor necrosis factor-α (TNF-α), immune globulin G (IgG) and corticosterone (CORT). The increase in the level of inflammation in ageing animals was related to changes in the structure and diversity of the gut microbiota. At the genus level, the relative abundance of Prevotella, Treponema, Corynebacterium, and Sphingomonas was increased in ageing animals and decreased significantly by the treatment of MT. Christensenella and Lactobacillus were attenuated in ageing animals, and tended to be enhanced by MT treatment. Functions related to glycosphingolipid biosynthesis-ganglio series and lipopolysaccharide biosynthesis (metabolisms of cofactors, vitamins and glycan) were increased in ageing animals and decreased significantly by the treatment of MT. Our data suggest that a supplement of MT could improve colon homeostasis through changing the composition of gut microbiota and reducing inflammation in ageing gerbils.

Epigenetic Regulation of miR-25 and Lnc107153 on Expression of Seasonal Estrus Key Gene CHGA in Sheep

Pituitary pars tuberalis (PT) plays an important role as the transmission center in the seasonal reproduction of animals. It helps convert external photoperiod signals into intrinsic seasonal reproduction signals. In sheep PT, specific expression patterns of several genes (including short photoperiod-induced gene CHGA and long photoperiod genes EYA3 and TSHβ) under different photoperiods are crucial characteristics during this signal transduction. Recent studies have revealed the role of epigenetics in regulating the expression of seasonal reproductive key genes. Therefore, we explored whether microRNAs and LncRNAs regulated the expressions of the above key genes. Firstly, the expression of miR-25 and CHGA showed a significant negative correlation in sheep PT. Results of the dual luciferase reporter assay and miR-25 overexpression indicated that miR-25 could inhibit the expression of CHGA by specifically binding to its 3'UTR region in pituitary cells. Then, expression negative correlation and dual luciferase reporter analyses were used to screen and identify the candidate LncRNA (Lnc107153) targeted by miR-25. Finally, the results of fluorescence in situ hybridization and Lnc107153 overexpression suggested that Lnc107153 and miR-25 were involved in the epigenetic regulation of CHGA expression. However, the expressions of EYA3 and TSHβ were not regulated by miRNAs. These results will provide new insights into the epigenetic regulatory network of key genes in sheep seasonal reproduction.

Plant Melatonin: Regulatory and Protective Role

Melatonin is an antioxidant that is widely distributed in plants and animals. It is a conservative molecule. In early studies, scientists often used isolation and identification techniques to observe whether the endogenous melatonin cycle in plants was related to the external photoperiod, plant growth, and development cycles, including seed germination, plant rooting, and floral induction. With the development of isolation and identification technology, there is ample evidence that plants possess a variety of melatonin-synthesis pathways. The comprehensive application of molecular biology, genomics, and computational biology has also led to a comprehensive understanding of the physiological functions of plant melatonin. In this paper, we not only highlight the candidate genes from Arabidopsis thaliana and Oryza sativa that might be contributing to increasing plant endogenous melatonin but also elucidate and characterize the role of melatonin in plant growth and development in response to biotic and abiotic stresses.

Seasonality of prolactin in birds and mammals.

In most animals, annual rhythms in environmental cues and internal programs regulate seasonal physiology and behavior. Prolactin, an evolutionarily ancient hormone, serves as a molecular correlate of seasonal timing in most species. Prolactin is highly pleiotropic with a wide variety of well-documented physiological effects; in a seasonal context prolactin is known to regulate annual changes in pelage and molt. While short-term homeostatic variation of prolactin secretion is under the control of the hypothalamus, long-term seasonal rhythms of prolactin are programmed by endogenous timers that reside in the pituitary gland. The molecular basis of these rhythms is generally understood to be melatonin dependent in mammals. Prolactin rhythmicity persists for several years in many species, in the absence of hypothalamic signaling. Such evidence in mammals has supported the hypothesis that seasonal rhythms in prolactin derive from an endogenous timer within the pituitary gland that is entrained by external photoperiod. In this review, we describe the conserved nature of prolactin signaling in birds and mammals and highlight its role in regulating multiple diverse physiological systems. The review will cover the current understanding of the molecular control of prolactin seasonality and propose a mechanism by which long-term rhythms may be generated in amniotes.

Relationships between endogenous circadian period, physiological and cognitive parameters and sex in aged gray mouse lemurs (Microcebus murinus)

ABSTRACT The biological clock generates circadian rhythms, with an endogenous period tau close to 24 h. The circadian resonance theory proposes that lifespan is reduced when endogenous period goes far from 24 h. It has been suggested that daily resetting of the circadian clock to the 24 h external photoperiod might induce marginal costs that would accumulate over time and forward accelerate aging and affect fitness. In this study, we aimed to evaluate the link between the endogenous period and biomarkers of aging in order to investigate the mechanisms of the circadian resonance theory. We studied 39 middle-aged and aged Microcebus murinus, a nocturnal non-human primate whose endogenous period is about 23.1 h, measuring the endogenous period of locomotor activity, as well as several physiological and behavioral parameters (rhythm fragmentation and amplitude, energetic expenditure, oxidative stress, insulin-like growth factor-1 (IGF-1) concentrations and cognitive performances) in both males and females. We found that aged males with tau far from 24 h displayed increased oxidative stress. We also demonstrated a positive correlation between tau and IGF-1 concentrations, as well as learning performances, in males and females. Together these results suggest that a great deviation of tau from 24 h leads to increased biomarkers of age-related impairments.

Melatonin and osmoregulation in fish: A focus on Atlantic salmon Salmo salar smoltification.

Part of the life cycle of several fish species includes important salinity changes, as is the case for the sea bass (Dicentrarchus labrax) or the Atlantic salmon (Salmo salar). Salmosalar juveniles migrate downstream from their spawning sites to reach seawater, where they grow and become sexually mature. The process of preparation enabling juveniles to migrate downstream and physiologically adapt to seawater is called smoltification. Daily and seasonal variations of photoperiod and temperature play a role in defining the timing of smoltification, which may take weeks to months, depending on the river length and latitude. Smoltification is characterised by a series of biochemical, physiological and behavioural changes within the neuroendocrine axis. This review discusses the current knowledge and gaps related to the neuroendocrine mechanisms that mediate the effects of light and temperature on smoltification. Studies performed in S. salar and other salmonids, as well as in other species undergoing important salinity changes, are reviewed, and a particular emphasis is given to the pineal hormone melatonin and its possible role in osmoregulation. The daily and annual variations of plasma melatonin levels reflect corresponding changes in external photoperiod and temperature, which suggests that the hormonal time-keeper melatonin might contribute to controlling smoltification. Here, we review studies on (i) the impact of pinealectomy and/or melatonin administration on smoltification; (ii) melatonin interactions with hormones involved in osmoregulation (e.g., prolactin, growth hormone and cortisol); (iii) the presence of melatonin receptors in tissues involved in osmoregulation; and (iv) the impacts of salinity changes on melatonin receptors and circulating melatonin levels. Altogether, these studies show evidence indicating that melatonin interacts with the neuroendocrine pathways controlling smoltification, although more information is needed to clearly decipher its mechanisms of action.

Nondestructive evaluation of oviposition behavior of the bamboo powderpost beetle, Dinoderus minutus, using X-ray computed tomography and acoustic emission

In this study, X-ray computed tomography (CT) and acoustic emission (AE) monitoring were used to nondestructively analyze the oviposition behavior of the bamboo powderpost beetle, Dinoderus minutus, which takes place inside bamboo culms. Newly mated adult females of D. minutus were singly exposed to a bamboo piece, obtained from culms of madake (Phyllostachys bambusoides), and were kept at a temperature of 28 °C and a relative humidity of 65% with a photoperiod of L12:D12. The bamboo pieces were subjected to X-ray CT scanning every 2–5 days, and they were also monitored continuously using AE. The behavior of males after mating was also analyzed in a similar manner. Oviposition tunnels bored across the bamboo vascular bundles and eggs deposited in the metaxylem vessels were visualized in the CT images. The females lived for 75 ± 17 days (mean ± SD) after mating, and they created tunnels that were 116.4 ± 52.1 mm in length and 177.2 ± 78.9 mm3 in volume and laid 253.1 ± 125.3 eggs in total. Using AE monitoring, the tunneling activity of the females was detected. The AE data indicated that continuous tunneling activity of the females was separated by regular short pauses, lasting approximately 1 h, occurring at intervals of around 2 days. The tunneling activity of the females did not exhibit clear circadian rhythms, suggesting that it was unaffected by external photoperiod. The males tunneled more slowly than the females and had no preference to the boring direction, and their activities were in synchronization with the photoperiod.

Melatonin and the development of circadian and seasonal rhythmicity

We have investigated whether the maternal melatonin rhythm provides the fetus with either a circadian or seasonal 'signal' during development. Our findings provide evidence that melatonin can generate and entrain the early evening peak in the daily rhythm of fetal breathing movements. In contrast, daily variations in maternal and fetal prolactin concentrations are present in pinealectomized ewes, are altered by changes in the time of onset of darkness, but are unaltered by changes in the phase of the daily melatonin rhythm. The mechanisms that generate and control the daily prolactin rhythm before and after birth are therefore unknown. It is clear from a number of studies that the duration of the nocturnal melatonin signal provides the adult and fetal sheep with photoperiodic information. We investigated whether there are differences in the fetal plasma concentrations of prolactin in ewes held in long and short photoperiods after surgical disconnection of the fetal hypothalamus and pituitary and demonstrated that there was a fetal prolactin response to the external photoperiod in sheep fetuses in which the hypothalamo-pituitary axis was either intact or surgically disconnected. We have suggested that one potential extrahypothalamic site of action of maternal melatonin is at the pars tuberalis of the fetal pituitary. It appears therefore that there are a number of different mechanisms for the neuroendocrine transmission of information about the time of day and duration of the external photoperiod to the sheep fetus throughout late gestation.

Scotostimulation of reproductive neural pathways and gonadal maturation are not correlated with hypothalamic expression of deiodinases in subtropical spotted munia.

Circannual rhythm regulates the annual timing of reproduction in spotted munia, with sex differences in its relationship with the external photoperiod environment. Interestingly, munia show an atypical photosensitivity and exhibit gonadal maturation when acutely exposed to an unnatural short photoperiod (eg 3hours of light per day; ie a long scotoperiod). The proximate mechanisms regulating scotoperiod-induced hypothalamic-pituitary-gonadal (HPG) activation are unclear. Because thyroid hormone signalling plays a central role in photoperiodic induction, we hypothesised the involvement of similar mechanism, comprising alterations in hypothalamic deiodinases, under long scotoperiod-induced HPG activation. To test this, several endpoints of cellular and molecular correlates were assayed in male and female munias after 1 and 4 weeks of exposure to an 3:21hour light/dark cycle (3L:21D), with controls on a 21:3hour light/dark cycle (21L:3D). We measured the hypothalamic expression of mRNA and protein of light-sensitive (neuropsin, OPN5) and reproductive (vasoactive intestinal peptide [VIP], neuropeptide Y [NPY], gonadotrophin-releasing hormone [GnRH], gonadotrophin-inhibiting hormone [GnIH]) neuropeptides by quantitative polymerase chain reaction (PCR) and immunohistochemistry, respectively. In addition, we also measured mRNA expression of types 2 (DIO2) and 3 (DIO3) deiodinases that regulate triiodothyronine-mediated GnRH release and gonadal maturation in photoperiodic species. The quantitative PCR and immunohistochemistry results were consistent. Higher OPN5 levels under 21L:3D than under 3L:21D suggested its role in sensing the length of the light period. Similarly, low VIP and high NPY expression under 3L:21D than under 21L:3D were consistent with their roles as cellular correlates of photic and nonphotic environment, respectively. High GnRH-I/low GnIH levels and gonadal recrudescence under 3L:21D, and an inverse pattern under 21L:3D, confirmed the scotostimulation of HPG axis in spotted munia. However, DIO2 and DIO3 mRNA levels did not differ between 2 scotoperiods, in contrast to their reciprocal expression pattern found during long-day photostimulation. We demonstrate for the first time sex-dependent scotostimulation of reproductive neural pathways and suggest the involvement of molecules other than hypothalamic deiodinases in the regulation of gonad development cycle in 'nonphotoperiodic' seasonally breeding vertebrates.

Perinatal Programming of Circadian Clock-Stress Crosstalk.

An intact communication between circadian clocks and the stress system is important for maintaining physiological homeostasis under resting conditions and in response to external stimuli. There is accumulating evidence for a reciprocal interaction between both—from the systemic to the molecular level. Disruption of this interaction by external factors such as shiftwork, jetlag, or chronic stress increases the risk of developing metabolic, immune, or mood disorders. From experiments in rodents, we know that both systems maturate during the perinatal period. During that time, exogenous factors such as stress or alterations in the external photoperiod may critically affect—or program—physiological functions later in life. This developmental programming process has been attributed to maternal stress signals reaching the embryo, which lastingly change gene expression through the induction of epigenetic mechanisms. Despite the well-known function of the adult circadian system in temporal coordination of physiology and behavior, the role of maternal and embryonic circadian clocks during pregnancy and postnatal development is still poorly defined. A better understanding of the circadian-stress crosstalk at different periods of development may help to improve stress resistance and devise preventive and therapeutic strategies against chronic stress-associated disorders.

Phospholipase C beta 4 in mouse hepatocytes: Rhythmic expression and cellular distribution

BackgroundCircadian regulated physiological processes have been well documented in the mammalian liver. Phospholipases are important mediators of both cytoplasmic and nuclear signaling mechanisms in hepatocytes, and despite a potentially critical role for these enzymes in regulating the temporal aspect of hepatic physiology, their involvement in the circadian liver clock has not been the subject of much investigation. The phospholipase C β4 (PLCβ4) enzyme is of particular interest as it has been linked to circadian clock function. In general, there is no knowledge of the role of the PLCβ4 isozyme in mammalian hepatocytes as this is the first report of its expression in the mammalian liver.ResultsWe found that in the liver of mice housed on a light:dark cycle, PLCβ4 protein underwent a significant circadian rhythm with a peak occurring during the early night. In constant darkness, the protein rhythm was more robust and peaked around dusk. We also observed a significant oscillation in plcβ4 gene expression in the livers of mice housed in both photoperiodic and constant dark conditions. The cellular distribution of the protein in hepatocytes varied over the course of the circadian day with PLCβ4 primarily cytoplasmic around dusk and nuclear at dawn.ConclusionOur results indicate that PLCβ4 gene and protein expression is regulated by a circadian clock in the mouse liver and is not dependent on the external photoperiod. A light-independent daily translocation of PLCβ4 implies that it may play a key role in nuclear signaling in hepatocytes and serve as a daily temporal cue for physiological processes in the liver.

A Morning-Specific Phytohormone Gene Expression Program underlying Rhythmic Plant Growth

Most organisms use daily light/dark cycles as timing cues to control many essential physiological processes. In plants, growth rates of the embryonic stem (hypocotyl) are maximal at different times of day, depending on external photoperiod and the internal circadian clock. However, the interactions between light signaling, the circadian clock, and growth-promoting hormone pathways in growth control remain poorly understood. At the molecular level, such growth rhythms could be attributed to several different layers of time-specific control such as phasing of transcription, signaling, or protein abundance. To determine the transcriptional component associated with the rhythmic control of growth, we applied temporal analysis of the Arabidopsis thaliana seedling transcriptome under multiple growth conditions and mutant backgrounds using DNA microarrays. We show that a group of plant hormone-associated genes are coexpressed at the time of day when hypocotyl growth rate is maximal. This expression correlates with overrepresentation of a cis-acting element (CACATG) in phytohormone gene promoters, which is sufficient to confer the predicted diurnal and circadian expression patterns in vivo. Using circadian clock and light signaling mutants, we show that both internal coincidence of phytohormone signaling capacity and external coincidence with darkness are required to coordinate wild-type growth. From these data, we argue that the circadian clock indirectly controls growth by permissive gating of light-mediated phytohormone transcript levels to the proper time of day. This temporal integration of hormone pathways allows plants to fine tune phytohormone responses for seasonal and shade-appropriate growth regulation.

Arabidopsis Clock-Associated Pseudo-Response Regulators PRR9, PRR7 and PRR5 Coordinately and Positively Regulate Flowering Time Through the Canonical CONSTANS-Dependent Photoperiodic Pathway

Photoperiodism allows organisms to measure daylength, or external photoperiod, and to anticipate coming seasons. Daylength measurement requires the integration of light signal and temporal information by the circadian clock. In the long-day plant Arabidopsis thaliana, CONSTANS (CO) plays a crucial role in integrating the circadian rhythm and environmental light signals into the photoperiodic flowering pathway. Nevertheless, the molecular mechanism by which the circadian clock modulates the cyclic expression profile of CO is poorly understood. Here, we first showed that the clock-associated genes PSEUDO-RESPONSE REGULATOR (PRR) PRR9, PRR7 and PRR5 are involved in activation of CO expression during the daytime. Then, extensive genetic studies using CIRCADIAN CLOCK-ASSOCIATED1 (CCA1)/LATE ELONGATED HYPOCOTYL (LHY) double mutants (cca1/lhy) and prr7/prr5 were conducted. The results suggested that PRR genes act coordinately in a manner parallel with and antagonistic to CCA/LHY, upstream of the canonical CO-FLOWERING LOCUS T (FT) photoperiodic flowering pathway. Finally, we provided evidence to propose a model, in which CCA1/LHY repress CO through GIGANTEA (GI), while PRR9, PRR7 and PRR5 activate CO predominantly by repressing CYCLING DOF FACTOR1 (CDF1) encoding a DNA-binding transcriptional repressor.

The role of endogenous and exogenous factors in regulation of synchronous emergence of Trichogramma embryophagum Hartig and T. principium Sug. et Sor. (Hymenoptera, Trichogrammatidae)

The interaction of exogenous and endogenous factors affecting the daily rhythm of emergence of adult parasitoids Trichogramma embryophagum and T. princupium from laboratory strains was studied. Under a constant temperature of 20°C and different photoperiods (photophase of 12, 16, or 20 h), both species demonstrated a distinct rhythm; the majority of adults emerged during 4 first hours after light-on. The fraction of individuals emerged during this period depended on the photoperiod and was different in both the species studied. Experiments with non-periodic regimes (constant light or constant darkness, 20°C) demonstrated that the rhythm is based on the endogenous circadian oscillations that are synchronized with the external photoperiod. The time of emergence however, was not rigidly “programmed” and could be modified by the direct influence of the light and temperature Light-on and/or temperature increase (from 20° up to 30° C) induced the emergence of a certain percentage of individuals virtually at any time of the 24-h-long cycle. The magnitude of the response, estimated as the fraction of adults emerged during 2 h after the stimulation, changed daily under the control of the circadian oscillation. Thus, the plasticity of the daily dynamics of emergence was controlled (to a certain extent) by the endogenous rhythm of sensitivity to the stimuli. The interspecific differences in the relative sensitivity to light-on and temperature increase were revealed. When light and temperature stimuli were applied simultaneously, they caused a synergistic effect, exceeding a simple sum of separate impacts. These results demonstrate an important role of exogenous factors in control of the emergence rhythm in Trichogramma and can also be used in elaboration of methods of regulation and synchronization of emergence in these parasitoids.

Binocular Contributions to the Responsiveness and Integrative Capacity of the Circadian Rhythm System to Light

The retinohypothalamic tract (RHT), a monosynaptic retinal projection to the SCN, is the major path by which light entrains the circadian system to the external photoperiod. The circadian system of rodents effectively integrates or counts photons, and the magnitude of the rhythm phase response is proportional to the total energy of the photic stimulus. In the present studies, responsiveness to light and integrative capacity of the circadian system were tested in hamsters after reduction of retinal photoreceptor input by 50%. At CT 19, animals in constant darkness with or without unilateral retinal occlusion were exposed to 1 of 6 irradiances of 5-min white-light pulses ranging from 0.0011 to 70 microW/cm(2) or 5 white-light pulses of 0.6 microW/cm(2) with durations ranging from 0.25 to 150.0 min. Assessment of light-induced circadian rhythm phase response and Fos expression in the SCN by these animals revealed that a 50% reduction in input from photoreceptors stimulated directly with light caused a decrease in responsiveness to the longest duration and highest irradiance pulses presented. Despite this effect, both the magnitude of Fos induction in the SCN and phase-shift response remained directly proportional to the total energy in the photic stimuli. The results support the view that a reciprocal relationship between stimulus irradiance and duration persists despite the 50% reduction in retinal photoreceptor input. The mechanism of integration neither resides in the retina nor in the RHT.

Functional analysis of the role of the median raphe as a regulator of hamster circadian system sensitivity to light

The retinohypothalamic tract, a monosynaptic retinal projection to the suprachiasmatic nucleus (SCN), is the path by which light entrains the circadian system to the external photoperiod. Serotonergic neurons in the mesencephalic median raphe nucleus (MnR) also give rise to a major SCN afferent projection. The present study was designed to determine the extent to which MnR serotonergic projections regulate sensitivity of the circadian rhythm system to light. Serotonergic neurons in the MnR were destroyed by the direct application of the neurotoxin, 5,7-dihydroxytryptamine. Animals in constant darkness were given 5-min white light pulses at circadian time 19. Light intensity varied from 0.0011 to 70 μW/cm 2. Assessment of rhythm phase response to light by lesioned and control animals revealed that animals lacking the MnR serotonergic projection are considerably more sensitive to light at high irradiances. The results are consistent with behavioral and physiological evidence implicating serotonin as an inhibitory modulator of the effects of light on circadian rhythmicity.

Photoperiod regulates clock gene rhythms in the ovine liver

To investigate the photoperiodic entrainment of peripheral rhythms in ruminants, we studied the expression of clock genes in the liver in the highly seasonal Soay sheep. Animals were kept under long (LD 16:8) or short photoperiod (LD 8:16). Daily rhythms in locomotor activity were recorded, and blood concentrations of melatonin and cortisol were measured by RIA. Per2, Bmal1, and Cry1 gene expression was determined by Northern blot analyses using ovine RNA probes in liver collected every 4 h for 24 h. Liver Per2 and Bmal1, but not Cry1, expression was rhythmic in all treatments. Under long days, peak Per2 expression occurred at end of the night with a similar timing to Bmal1, whereas, under short days the Per2 maximum was in the early night with an inverse pattern to Bmal1. There was a photoperiod × time interaction for only Per2 ( P < 0.001). The 24-h pattern in plasma cortisol matched the observed phasing of Per2 expression, suggesting that it may act as an endocrine entraining factor. The clock gene rhythms in the peripheral tissues were different in timing compared with the ovine suprachiasmatic nucleus (SCN, central pacemaker) and pars tuberalis (melatonin target tissue), and the hepatic rhythms were of lower amplitude compared with photoperiodic rodents. Thus, there are likely to be important species differences in the way the central and peripheral clockwork encodes external photoperiod.

035. Circadian rhythms and the early life programming of adult physiological systems

We are all familiar with the idea that the external environment influences many diverse physiological systems. For example, the level of nutrition can not only influence adult health directly, but also fetal development and subsequently many adult functions in the offspring. Maternal stress can affect fetal outcomes as can the administration of drugs during pregnancy. Until recently, however, the daily changes in environmental light have been considered to really only influence the time that we sleep and in many other species the optimal time to mate. The impact of circadian rhythms on life trajectory has had little attention. In the last 5 years it has become clear that circadian rhythmicity is entrenched in virtually every cell of our bodies. A suite of clock gene transcription factors that include Clock, Bmal1 and the period and cryptochrome genes, generate a robust daily cycle of transcription and translation of hundreds of proteins. This cellular clock system is synchronised with the external photoperiod through retinal light perception, the hypothalamic suprachiasmatic nucleus (SCN) and neural and hormonal pathways. Most importantly when the clock system in peripheral tissues is disrupted, a growing list of detrimental consequences are being uncovered. As an example, mice with mutations in either Clock or Bmal1 have non-rhythmic peripheral tissues and exhibit mild to severe reproductive failure and metabolic dysfunction. Null per2 mice have a higher incidence of salivary gland hyperplasia, teratomas and increased susceptibility to radiation induced lymphomas. It is also apparent that intrauterine insults (e.g. cocaine administration, poor nutrition and stress) can have long term effects on the central circadian timing system in the SCN. Whether this involves alterations in neural development or gene function is not known. Nevertheless it is time we paid more attention to the temporal nature of our environment as a possible contributor to lifetime disorders and diseases.