Recommendations for measuring and standardizing light for laboratory mammals to improve welfare and reproducibility in animal research.

Light enables vision and exerts widespread effects on physiology and behavior, including regulating circadian rhythms, sleep, hormone synthesis, affective state, and cognitive processes. Appropriate lighting in animal facilities may support welfare and ensure that animals enter experiments in an appropriate physiological and behavioral state. Furthermore, proper consideration of light during experimentation is important both when it is explicitly employed as an independent variable and as a general feature of the environment. This Consensus View discusses metrics to use for the quantification of light appropriate for nonhuman mammals and their application to improve animal welfare and the quality of animal research. It provides methods for measuring these metrics, practical guidance for their implementation in husbandry and experimentation, and quantitative guidance on appropriate light exposure for laboratory mammals. The guidance provided has the potential to improve data quality and contribute to reduction and refinement, helping to ensure more ethical animal use.

Machine learning estimation of human body time using metabolomic profiling.

Circadian rhythms influence physiology, metabolism, and molecular processes in the human body. Estimation of individual body time (circadian phase) is therefore highly relevant for individual optimization of behavior (sleep, meals, sports), diagnostic sampling, medical treatment, and for treatment of circadian rhythm disorders. Here, we provide a partial least squares regression (PLSR) machine learning approach that uses plasma-derived metabolomics data in one or more samples to estimate dim light melatonin onset (DLMO) as a proxy for circadian phase of the human body. For this purpose, our protocol was aimed to stay close to real-life conditions. We found that a metabolomics approach optimized for either women or men under entrained conditions performed equally well or better than existing approaches using more labor-intensive RNA sequencing-based methods. Although estimation of circadian body time using blood-targeted metabolomics requires further validation in shift work and other real-world conditions, it currently may offer a robust, feasible technique with relatively high accuracy to aid personalized optimization of behavior and clinical treatment after appropriate validation in patient populations.

Circadian rhythm entrainment of the jewel wasp, Nasonia vitripennis, by antagonistic interactions of multiple spectral inputs.

Circadian light entrainment in some insects is regulated by blue-light-sensitive cryptochrome (CRY) protein that is expressed in the clock neurons, but this is not the case in hymenopterans. The hymenopteran clock does contain CRY, but it appears to be light-insensitive. Therefore, we investigated the role of retinal photoreceptors in the photic entrainment of the jewel wasp Nasonia vitripennis. Application of monochromatic light stimuli at different light intensities caused phase shifts in the wasp's circadian activity from which an action spectrum with three distinct peaks was derived. Electrophysiological recordings from the compound eyes and ocelli revealed the presence of three photoreceptor classes, with peak sensitivities at 340 nm (ultraviolet), 450 nm (blue) and 530 nm (green). An additional photoreceptor class in the ocelli with sensitivity maximum at 560-580 nm (red) was found. Whereas a simple sum of photoreceptor spectral sensitivities could not explain the action spectrum of the circadian phase shifts, modelling of the action spectrum indicates antagonistic interactions between pairs of spectral photoreceptors, residing in the compound eyes and the ocelli. Our findings imply that the photic entrainment mechanism in N. vitripennis encompasses the neural pathways for measuring the absolute luminance as well as the circuits mediating colour opponency.

Circadian entrainment to red-light Zeitgebers and action spectrum for entrainment in the jewel wasp Nasonia vitripennis.

Light is the most important environmental cue for the circadian system of most organisms to stay synchronized to daily environmental changes. Like many other insects, the wasp Nasonia vitripennis has trichromatic compound eye-based colour vision and is sensitive to the light spectrum ranging from UV to green. We recently described a red-sensitive, ocelli-based photoreceptor, but its contribution to circadian entrainment remains unclear. In this study, we investigated the possibility of Nasonia circadian light entrainment under long-wavelength red LED light-dark cycles and characterized the strength of red light as a potential Zeitgeber. Additionally, we measured the possibility of entrainment under various light intensities (from 5·1012 to 4·1015 photons·cm-2·s-1) and a broader range of wavelengths (455-656 nm) to construct corresponding action spectra for characterizing all circadian photoreceptors involved in photic entrainment. We also conducted electroretinogram (ERG) recordings for each wavelength in the compound eyes. Our findings demonstrate that Nasonia can entrain under red light dark cycles, and the sensory pathway underlying the red-light Zeitgeber response may reside in the ocelli. Combined with findings from previous research, we pose that blue- and green-sensitive rhodopsin photoreceptor cells function as the major circadian photoreceptors in both circadian entrainment by light-dark cycles and circadian phase shifts by light pulses, whereas the red-sensitive photoreceptor cell requires higher light intensity for its role in circadian entrainment by light-dark cycles.

Variation in photoperiod response corresponds to differences in circadian light sensitivity in northern and southern Nasonia vitripennis lines.

The circadian clock times physiological and behavioural processes and resets on a daily basis to synchronize with the environment. The involvement of the circadian clock in photoperiodic time measurement synchronising annual rhythms is still under debate and different models have been proposed explaining their integration. Insects overcome unfavourable conditions in diapause, a form of dormancy. A latitudinal cline in diapause induction in the parasitoid wasp Nasonia vitripennis as well as a difference in circadian light sensitivity between north and south provide us with additional evidence that the circadian system of Nasonia is involved in photoperiodic time measurement and that latitude-specific seasonality drives adaptive evolution in photoperiodism partly through adaptation responses in the circadian system. We tested diapause induction in a range of T-cycles and photoperiods and found diapause induction in short photoperiods in all T-cycles in the northern line but in the southern line, diapause only occurred in T-cycles close to 24 h. Due to a lower light sensitivity in the southern line, a wider distribution of phase angles of entrainment can be expected at a specific T-cycle duration, while the range of entrainment will decrease. Taking these oscillator properties into account, our data can be explained by an external coincidence model involving a single oscillator with a light-sensitive phase that drives annual timing of diapause in Nasonia vitripennis.

Adrenalectomy prevents the effects of social defeat stress on PER2 rhythms in some peripheral tissues in male mice.

While stress does not affect the phase or period of the central pacemaker in the suprachiasmatic nucleus, it can shift clocks in peripheral tissues. Our previous studies showed significant delays of the PER2 rhythms in lung and kidney following social defeat stress. The mechanism underlying these effects is not fully understood, but might involve glucocorticoids (GC) released during the stressor. In the present study, we performed social defeat stress in adrenalectomized (ADX) mice to see if the induction of endogenous GC is necessary for the stress-induced phase shifts of peripheral clocks. We used mice that carry a luciferase reporter gene fused to the circadian clock gene Period2 (PER2::LUC) to examine daily rhythms of PER2 expression in various peripheral tissues. Mice were exposed to 5 consecutive daily social defeat stress in the late dark phase (ZT21-22). Running wheel rotations were recorded during 7 baseline and 5 social defeat days, which showed that social defeat stress suppressed locomotor activity without affecting the phase of the rhythm. This suppression of activity was not prevented by ADX. One hour after the last stressor, tissue samples from the liver, kidney and lung were collected and cultured for ex vivo bioluminescence recordings. In the liver, PER2 rhythms were not affected by social defeat stress or ADX. In the kidney, social defeat stress caused a > 4 h phase delay of the PER2 rhythm, which was prevented by ADX, supporting the hypothesis of a crucial role of GC in this stress effect. In the lung, social defeat stress caused an 8 h phase delay, but, surprisingly, a similar phase delay was seen in ADX animals independent of defeat. The latter indicates complex effects of stress and stress hormones on the lung clock. In conclusion, the findings show that repeated social defeat stress in the dark phase can shift PER2 rhythms in some tissues (lung, kidney) and not others (liver). Moreover, the social defeat stress effect in some tissues appears to be mediated by glucocorticoids (kidney) whereas the mechanism in other tissues is more complex (lung).

The impact of stress and stress hormones on endogenous clocks and circadian rhythms.

In mammals, daily rhythms in physiology and behavior are under control of a circadian pacemaker situated in the suprachiasmatic nucleus (SCN). This master clock receives photic input from the retina and coordinates peripheral oscillators present in other tissues, maintaining all rhythms in the body synchronized to the environmental light-dark cycle. In line with its function as a master clock, the SCN appears to be well protected against unpredictable stressful stimuli. However, available data indicate that stress and stress hormones at certain times of day are capable of shifting peripheral oscillators in, e.g., liver, kidney and heart, which are normally under control of the SCN. Such shifts of peripheral oscillators may represent a temporary change in circadian organization that facilitates adaptation to repeated stress. Alternatively, these shifts of internal rhythms may represent an imbalance between precisely orchestrated physiological and behavioral processes that may have severe consequences for health and well-being.

Differential temperature effects on photoperiodism in female voles: A possible explanation for declines in vole populations.

Many mammalian species use photoperiod as a predictive cue to time seasonal reproduction. In addition, metabolic effects on the reproductive axis may also influence seasonal timing, especially in female small, short-lived mammals. To get a better understanding of how annual cycling environmental cues impact reproductive function and plasticity in small, short-lived herbivores with different geographic origins, we investigated the mechanisms underlying integration of temperature in the photoperiodic-axis regulating female reproduction in a Northern vole species (tundra vole, Microtus oeconomus) and in a Southern vole species (common vole, Microtus arvalis). We show that photoperiod and temperature interact to determine appropriate physiological responses; there is species-dependent annual variation in the sensitivity to temperature for reproductive organ development. In common voles, temperature can overrule photoperiodical spring-programmed responses, with reproductive organ mass being higher at 10°C than at 21°C, whereas in autumn they are less sensitive to temperature. These findings are in line with our census data, showing an earlier onset of spring reproduction in cold springs, while reproductive offset in autumn is synchronized to photoperiod. The reproductive organs of tundra voles were relatively insensitive to temperature, whereas hypothalamic gene expression was generally upregulated at 10°C. Thus, both vole species use photoperiod, whereas only common voles use temperature as a cue to control spring reproduction, which indicates species-specific reproductive strategies. Due to global warming, spring reproduction in common voles will be delayed, perhaps resulting in shorter breeding seasons and thus declining populations, as observed throughout Europe.

Transcriptional regulation underlying the temperature response of embryonic development rate in the winter moth.

Climate change will strongly affect the developmental timing of insects, as their development rate depends largely on ambient temperature. However, we know little about the genetic mechanisms underlying the temperature sensitivity of embryonic development in insects. We investigated embryonic development rate in the winter moth (Operophtera brumata), a species with egg dormancy which has been under selection due to climate change. We used RNA sequencing to investigate which genes are involved in the regulation of winter moth embryonic development rate in response to temperature. Over the course of development, we sampled eggs before and after an experimental change in ambient temperature, including two early development weeks when the temperature sensitivity of eggs is low and two late development weeks when temperature sensitivity is high. We found temperature-responsive genes that responded in a similar way across development, as well as genes with a temperature response specific to a particular development week. Moreover, we identified genes whose temperature effect size changed around the switch in temperature sensitivity of development rate. Interesting candidate genes for regulating the temperature sensitivity of egg development rate included genes involved in histone modification, hormonal signalling, nervous system development and circadian clock genes. The diverse sets of temperature-responsive genes we found here indicate that there are many potential targets of selection to change the temperature sensitivity of embryonic development rate. Identifying for which of these genes there is genetic variation in wild insect populations will give insight into their adaptive potential in the face of climate change.

Mechanisms of temperature modulation in mammalian seasonal timing.

Global warming is predicted to have major effects on the annual time windows during which species may successfully reproduce. At the organismal level, climatic shifts engage with the control mechanism for reproductive seasonality. In mammals, laboratory studies on neuroendocrine mechanism emphasize photoperiod as a predictive cue, but this is based on a restricted group of species. In contrast, field-oriented comparative analyses demonstrate that proximate bioenergetic effects on the reproductive axis are a major determinant of seasonal reproductive timing. The interaction between proximate energetic and predictive photoperiodic cues is neglected. Here, we focused on photoperiodic modulation of postnatal reproductive development in common voles (Microtus arvalis), a herbivorous species in which a plastic timing of breeding is well documented. We demonstrate that temperature-dependent modulation of photoperiodic responses manifest in the thyrotrophin-sensitive tanycytes of the mediobasal hypothalamus. Here, the photoperiod-dependent expression of type 2 deiodinase expression, associated with the summer phenotype was enhanced by 21°C, whereas the photoperiod-dependent expression of type 3 deiodinase expression, associated with the winter phenotype, was enhanced by 10°C in spring voles. Increased levels of testosterone were found at 21°C, whereas somatic and gonadal growth were oppositely affected by temperature. The magnitude of these temperature effects was similar in voles photoperiodical programmed for accelerated maturation (ie, born early in the breeding season) and in voles photoperiodical programmed for delayed maturation (ie, born late in the breeding season). The melatonin-sensitive pars tuberalis was relatively insensitive to temperature. These data define a mechanistic hierarchy for the integration of predictive temporal cues and proximate thermo-energetic effects in mammalian reproduction.

Food and temperature change photoperiodic responses in two vole species.

Seasonal timing of reproduction in voles is driven by photoperiod. We hypothesized that a negative energy balance can modify spring-programmed photoperiodic responses in the hypothalamus, controlling reproductive organ development. We manipulated energy balance by the 'work-for-food' protocol, in which voles were exposed to increasing levels of food scarcity at different ambient temperatures under long photoperiod. We found that in common voles (Microtus arvalis) and tundra voles (Microtus oeconomus), photoperiod-induced pars tuberalis thyroid-stimulating hormone ß-subunit (Tshß) expression is reduced to potentially inhibit gonadal development when food is scarce. Reduction in gonadal size is more pronounced in tundra voles, in which anterior hypothalamic Kiss1 is additionally downregulated, especially in males. Low temperature additionally leads to decreased hypothalamic Rfrp expression, which potentially may facilitate further suppression of gonadal growth. Shutting off the photoperiodic axis when food is scarce in spring may be an adaptive response to save energy, leading to delayed reproductive organ development until food resources are sufficient for reproduction, lactation and offspring survival. Defining the mechanisms through which metabolic cues modify photoperiodic responses will be important for a better understanding of how environmental cues impact reproduction.

Timing of increased temperature sensitivity coincides with nervous system development in winter moth embryos.

Climate change is rapidly altering the environment and many species will need to genetically adapt their seasonal timing to keep up with these changes. Insect development rate is largely influenced by temperature, but we know little about the mechanisms underlying the temperature sensitivity of development. Here, we investigate seasonal timing of egg hatching in the winter moth, one of the few species which has been found to genetically adapt to climate change, likely through selection on temperature sensitivity of egg development rate. To study when during development winter moth embryos are most sensitive to changes in ambient temperature, we gave eggs an increase or decrease in temperature at different moments during their development. We measured their developmental progression and time of egg hatching, and used fluorescence microscopy to construct a timeline of embryonic development for the winter moth. We found that egg development rate responded more strongly to temperature once embryos were in the fully extended germband stage. This is the phylotypic stage at which all insect embryos have developed a rudimentary nervous system. Furthermore, at this stage, timing of ecdysone signaling determines developmental progression, which could act as an environment dependent gateway. Intriguingly, this may suggest that, from the phylotypic stage onward, insect embryos can start to integrate internal and environmental stimuli to actively regulate important developmental processes. As we found evidence that there is genetic variation for temperature sensitivity of egg development rate in our study population, such regulation could be a target of selection imposed by climate change.

Melanopsin- and L-cone-induced pupil constriction is inhibited by S- and M-cones in humans.

The human retina contains five photoreceptor types: rods; short (S)-, mid (M)-, and long (L)-wavelength-sensitive cones; and melanopsin-expressing ganglion cells. Recently, it has been shown that selective increments in M-cone activation are paradoxically perceived as brightness decrements, as opposed to L-cone increments. Here we show that similar effects are also observed in the pupillary light response, whereby M-cone or S-cone increments lead to pupil dilation whereas L-cone or melanopic illuminance increments resulted in pupil constriction. Additionally, intermittent photoreceptor activation increased pupil constriction over a 30-min interval. Modulation of L-cone or melanopic illuminance within the 0.25-4-Hz frequency range resulted in more sustained pupillary constriction than light of constant intensity. Opposite results were found for S-cone and M-cone modulations (2 Hz), mirroring the dichotomy observed in the transient responses. The transient and sustained pupillary light responses therefore suggest that S- and M-cones provide inhibitory input to the pupillary control system when selectively activated, whereas L-cones and melanopsin response fulfill an excitatory role. These findings provide insight into functional networks in the human retina and the effect of color-coding in nonvisual responses to light, and imply that nonvisual and visual brightness discrimination may share a common pathway that starts in the retina.

Gonads or body? Differences in gonadal and somatic photoperiodic growth response in two vole species.

To optimally time reproduction, seasonal mammals use a photoperiodic neuroendocrine system (PNES) that measures photoperiod and subsequently drives reproduction. To adapt to late spring arrival at northern latitudes, a lower photoperiodic sensitivity and therefore a higher critical photoperiod for reproductive onset is necessary in northern species to arrest reproductive development until spring onset. Temperature-photoperiod relationships, and hence food availability-photoperiod relationships, are highly latitude dependent. Therefore, we predict PNES sensitivity characteristics to be latitude dependent. Here, we investigated photoperiodic responses at different times during development in northern (tundra or root vole, Microtus oeconomus) and southern vole species (common vole, Microtus arvalis) exposed to constant short (SP) or long photoperiod (LP). Although the tundra vole grows faster under LP, no photoperiodic effect on somatic growth is observed in the common vole. In contrast, gonadal growth is more sensitive to photoperiod in the common vole, suggesting that photoperiodic responses in somatic and gonadal growth can be plastic, and might be regulated through different mechanisms. In both species, thyroid-stimulating hormone ß-subunit (Tshß) and iodothyronine deiodinase 2 (Dio2) expression is highly increased under LP, whereas Tshr and Dio3 decrease under LP. High Tshr levels in voles raised under SP may lead to increased sensitivity to increasing photoperiods later in life. The higher photoperiodic-induced Tshr response in tundra voles suggests that the northern vole species might be more sensitive to thyroid-stimulating hormone when raised under SP. In conclusion, species differences in developmental programming of the PNES, which is dependent on photoperiod early in development, may form different breeding strategies as part of latitudinal adaptation.

Maximising survival by shifting the daily timing of activity.

Maximising survival requires animals to balance the competing demands of maintaining energy balance and avoiding predation. Here, quantitative modelling shows that optimising the daily timing of activity and rest based on the encountered environmental conditions enables small mammals to maximise survival. Our model shows that nocturnality is typically beneficial when predation risk is higher during the day than during the night, but this is reversed by the energetic benefit of diurnality when food becomes scarce. Empirical testing under semi-natural conditions revealed that the daily timing of activity and rest in mice exposed to manipulations in energy availability and perceived predation risk is in line with the model's predictions. Low food availability and decreased perceived daytime predation risk promote diurnal activity patterns. Overall, our results identify temporal niche switching in small mammals as a strategy to maximise survival in response to environmental changes in food availability and perceived predation risk.

Dynamic expressions of hypothalamic genes regulate seasonal breeding in a natural rodent population.

Seasonal breeding is a universal reproductive strategy in many animals. Hypothalamic genes, especially type 2 and 3 iodothyronine deiodinases (Dio2/3), RFamide-related peptide 3 (Rfrp-3), kisspeptin (Kiss-1) and gonadotropin-releasing hormone (GnRH), are involved in a photoperiodic pathway that encodes seasonal signals from day length in many vertebrate species. However, the seasonal expression patterns of these genes in wild mammals are less studied. Here, we present a four-year field investigation to reveal seasonal rhythm and age-dependent reproductive activity in male Brandt's voles (Lasiopodomys brandtii) and to detect relationships among seasonal expression profiles of hypothalamic genes, testicular activity, age and annual day length. From breeding season (April) to nonbreeding season (October), adult male voles displayed a synchronous peak in gonadal activity with annual day length around summer solstice, which was jointly caused by age structure shifts and age-dependent gonadal development patterns. Overwintered males maintained reproductive activity until late in the breeding season, whereas most newborn males terminated gonadal development completely, except for a minority of males born early in spring. Consistently, the synchronous and opposite expression profiles of Dio2/3 suggest their central function to decode photoperiodic signals and to predict the onset of the nonbreeding season. Moreover, changes in Dio2/3 signals may guide the actions of Kiss-1 and Rfrp-3 to regulate the age-dependent divergence of reproductive strategy in wild Brandt's vole. Our results provide evidence on how hypothalamic photoperiod genes regulate seasonal breeding in a natural rodent population.

Daytime melatonin and light independently affect human alertness and body temperature.

Light significantly improves alertness during the night (Cajochen, Sleep Med Rev, 11, 2007 and 453; Ruger et al., AJP Regul Integr Comp Physiol, 290, 2005 and R1413), but results are less conclusive at daytime (Lok et al., J Biol Rhythms, 33, 2018 and 589). Melatonin and core body temperature levels at those times of day may contribute to differences in alerting effects of light. In this experiment, the combined effect of daytime exogenous melatonin administration and light intensity on alertness, body temperature, and skin temperature was studied. The goal was to assess whether (a) alerting effects of light are melatonin dependent, (b) soporific effects of melatonin are mediated via the thermoregulatory system, and (c) light can improve alertness after melatonin-induced sleepiness during daytime. 10 subjects (5 females, 5 males) received melatonin (5 mg) in dim (10 lux) and, on a separate occasion, in bright polychromatic white light (2000 lux). In addition, they received placebo both under dim and bright light conditions. Subjects participated in all four conditions in a balanced order, yielding a balanced within-subject design, lasting from noon to 04:00 pm. Alertness and performance were assessed half hourly, while body temperature and skin temperature were measured continuously. Saliva samples to detect melatonin concentrations were collected half hourly. Melatonin administration increased melatonin concentrations in all subjects. Subjective sleepiness and distal skin temperature increased after melatonin ingestion. Bright light exposure after melatonin administration did not change subjective alertness scores, but body temperature and proximal skin temperature increased, while distal skin temperature decreased. Light exposure did not significantly affect these parameters in the placebo condition. These results indicate that (a) exogenous melatonin administration during daytime increases subjective sleepiness, confirming a role for melatonin in sleepiness regulation, (b) bright light exposure after melatonin ingestion significantly affected thermoregulatory parameters without altering subjective sleepiness, therefore temperature changes seem nonessential for melatonin-induced sleepiness, (c) subjective sleepiness was increased by melatonin ingestion, but bright light administration was not able to improve melatonin-induced sleepiness feelings nor performance. Other (physiological) factors may therefore contribute to differences in alerting effects of light during daytime and nighttime.

Investigation of metabolites for estimating blood deposition time.

Trace deposition timing reflects a novel concept in forensic molecular biology involving the use of rhythmic biomarkers for estimating the time within a 24-h day/night cycle a human biological sample was left at the crime scene, which in principle allows verifying a sample donor's alibi. Previously, we introduced two circadian hormones for trace deposition timing and recently demonstrated that messenger RNA (mRNA) biomarkers significantly improve time prediction accuracy. Here, we investigate the suitability of metabolites measured using a targeted metabolomics approach, for trace deposition timing. Analysis of 171 plasma metabolites collected around the clock at 2-h intervals for 36 h from 12 male participants under controlled laboratory conditions identified 56 metabolites showing statistically significant oscillations, with peak times falling into three day/night time categories: morning/noon, afternoon/evening and night/early morning. Time prediction modelling identified 10 independently contributing metabolite biomarkers, which together achieved prediction accuracies expressed as AUC of 0.81, 0.86 and 0.90 for these three time categories respectively. Combining metabolites with previously established hormone and mRNA biomarkers in time prediction modelling resulted in an improved prediction accuracy reaching AUCs of 0.85, 0.89 and 0.96 respectively. The additional impact of metabolite biomarkers, however, was rather minor as the previously established model with melatonin, cortisol and three mRNA biomarkers achieved AUC values of 0.88, 0.88 and 0.95 for the same three time categories respectively. Nevertheless, the selected metabolites could become practically useful in scenarios where RNA marker information is unavailable such as due to RNA degradation. This is the first metabolomics study investigating circulating metabolites for trace deposition timing, and more work is needed to fully establish their usefulness for this forensic purpose.

The flexible clock: predictive and reactive homeostasis, energy balance and the circadian regulation of sleep-wake timing.

The Darwinian fitness of mammals living in a rhythmic environment depends on endogenous daily (circadian) rhythms in behavior and physiology. Here, we discuss the mechanisms underlying the circadian regulation of physiology and behavior in mammals. We also review recent efforts to understand circadian flexibility, such as how the phase of activity and rest is altered depending on the encountered environment. We explain why shifting activity to the day is an adaptive strategy to cope with energetic challenges and show how this can reduce thermoregulatory costs. A framework is provided to make predictions about the optimal timing of activity and rest of non-model species for a wide range of habitats. This Review illustrates how the timing of daily rhythms is reciprocally linked to energy homeostasis, and it highlights the importance of this link in understanding daily rhythms in physiology and behavior.

Cold and hunger induce diurnality in a nocturnal mammal.

The mammalian circadian system synchronizes daily timing of activity and rest with the environmental light-dark cycle. Although the underlying molecular oscillatory mechanism is well studied, factors that influence phenotypic plasticity in daily activity patterns (temporal niche switching, chronotype) are presently unknown. Molecular evidence suggests that metabolism may influence the circadian molecular clock, but evidence at the level of the organism is lacking. Here we show that a metabolic challenge by cold and hunger induces diurnality in otherwise nocturnal mice. Lowering ambient temperature changes the phase of circadian light-dark entrainment in mice by increasing daytime and decreasing nighttime activity. This effect is further enhanced by simulated food shortage, which identifies metabolic balance as the underlying common factor influencing circadian organization. Clock gene expression analysis shows that the underlying neuronal mechanism is downstream from or parallel to the main circadian pacemaker (the hypothalamic suprachiasmatic nucleus) and that the behavioral phenotype is accompanied by phase adjustment of peripheral tissues. These findings indicate that nocturnal mammals can display considerable plasticity in circadian organization and may adopt a diurnal phenotype when energetically challenged. Our previously defined circadian thermoenergetics hypothesis proposes that such circadian plasticity, which naturally occurs in nocturnal mammals, reflects adaptive maintenance of energy balance. Quantification of energy expenditure shows that diurnality under natural conditions reduces thermoregulatory costs in small burrowing mammals like mice. Metabolic feedback on circadian organization thus provides functional benefits by reducing energy expenditure. Our findings may help to clarify relationships between sleep-wake patterns and metabolic phenotypes in humans.