Sleep and Thermoregulation in Birds: Cold Exposure Reduces Brain Temperature but Has Little Influence on Sleep Time and Sleep Architecture in Jackdaws (Coloeus monedula).

Birds have an electrophysiological sleep state that resembles mammalian rapid-eye-movement (REM) sleep. However, whether its regulation and function are similar is unclear. In the current experiment, we studied REM sleep regulation in jackdaws (Coloeus monedula) by exposing the birds to low ambient temperature, a procedure that selectively suppresses REM sleep in mammals. Eight jackdaws were equipped with electrodes to record brain activity and neck muscle activity and a thermistor to record cortical brain temperature. Recordings covered a three-day period starting with a 24 h baseline day at an ambient temperature of 21 °C, followed by a 12 h cold night at 4 °C, after which the ambient temperature was restored to 21 °C for the remaining recovery period. Cold exposure at night caused a significant drop in brain temperature of 1.4 °C compared to the baseline night. However, throughout the cold night, jackdaws expressed NREM sleep and REM sleep levels that were not significantly different from the baseline. Also, EEG spectral power during NREM sleep was unaffected by cold exposure. Thus, while cold exposure had a clear effect on brain temperature in jackdaws, it did not have the same REM sleep suppressing effect reported for mammals. These findings suggest that the REM-sleep-like state in birds, unlike REM sleep in mammals, is protected against the influence of low temperature.

Semelparous marsupials reduce sleep for sex.

Sleep is a prominent, seemingly universal animal behavior. Although sleep maintains optimal waking performance, the biological drive to sleep may be incompatible with the life history of some species. In a multi-year study on semelparous marsupials in Australia, we provide the first direct evidence of ecological sleep restriction in a terrestrial mammal. Dusky (Antechinus swainsonii) and agile (A. agilis) antechinus have an unusual reproductive strategy characterized by the synchronous death of all males at the end of their only breeding season. Using accelerometry, electrophysiology, and metabolomics, we show that males, but not females, increase their activity during the breeding season by reducing sleep. In a trade-off between the neurophysiological requirements for sleep and evolutionary necessity for reproduction, strong sexual selection might drive males to sacrifice sleep to increase access to fertile females and ultimately maximize their fitness.

Melatonin Does Not Affect the Stress-Induced Phase Shifts of Peripheral Clocks in Male Mice.

Repeated or chronic stress can change the phase of peripheral circadian rhythms. Melatonin (Mel) is thought to be a circadian clock-controlled signal that might play a role in synchronizing peripheral rhythms, in addition to its direct suppressing effects on the stress axis. In this study we test whether Mel can reduce the social-defeat stress-induced phase shifts in peripheral rhythms, either by modulating circadian phase or by modulating the stress axis. Two experiments were performed with male Mel-deficient C57BL/6J mice carrying the circadian reporter gene construct (PER2::LUC). In the first experiment, mice received night-restricted (ZT11-21) Mel in their drinking water, resulting in physiological levels of plasma Mel peaking in the early dark phase. This treatment facilitated re-entrainment of the activity rhythm to a shifted light-dark cycle, but did not prevent the stress-induced (ZT21-22) reduction of activity during stress days. Also, this treatment did not attenuate the phase-delaying effects of stress in peripheral clocks in the pituitary, lung, and kidney. In a second experiment, pituitary, lung, and kidney collected from naive mice (ZT22-23), were treated with Mel, dexamethasone (Dex), or a combination of the two. Dex application affected PER2 rhythms in the pituitary, kidney, and lung by changing period, phase, or both. Administering Mel did not influence PER2 rhythms nor did it alleviate Dex-induced delays in PER2 rhythms in those tissues. We conclude that exogenous Mel is insufficient to affect peripheral PER2 rhythms and reduce stress effects on locomotor activity and phase changes in peripheral tissues.

Seasonal variation in sleep time: jackdaws sleep when it is dark, but do they really need it?

Sleep is an important behavioural and physiological state that is ubiquitous throughout the animal kingdom. Birds are an interesting group to study sleep since they share similar sleep features with mammals. Interestingly, sleep time in birds has been shown to vary greatly amongst seasons. To understand the mechanisms behind these variations in sleep time, we did an electro-encephalogram (EEG) study in eight European jackdaws (Coloeus monedula) in winter and summer under outdoor seminatural conditions. To assess whether the amount and pattern of sleep is determined by the outdoor seasonal state of the animals or directly determined by the indoor light-dark cycle, we individually housed them indoors where we manipulated the light-dark (LD) cycles to mimic long winter nights (8:16 LD) and short summer nights (16:8 LD) within both seasons. Jackdaws showed under seminatural outdoor conditions 5 h less sleep in summer compared to winter. During the indoor conditions, the birds rapidly adjusted their sleep time to the new LD cycle. Although they swiftly increased or decreased their sleep time, sleep intensity did not vary. The results indicate that the strong seasonal differences in sleep time are largely and directly driven by the available dark time, rather than an endogenous annual clock. Importantly, these findings confirm that sleep in birds is not a rigid phenomenon but highly sensitive to environmental factors.

Cofilin overactivation improves hippocampus-dependent short-term memory.

Many living organisms of the animal kingdom have the fundamental ability to form and retrieve memories. Most information is initially stored as short-term memory, which is then converted to a more stable long-term memory through a process called memory consolidation. At the neuronal level, synaptic plasticity is crucial for memory storage. It includes the formation of new spines, as well as the modification of existing spines, thereby tuning and shaping synaptic efficacy. Cofilin critically contributes to memory processes as upon activation, it regulates the shape of dendritic spines by targeting actin filaments. We previously found that prolonged activation of cofilin in hippocampal neurons attenuated the formation of long-term object-location memories. Because the modification of spine shape and structure is also essential for short-term memory formation, we determined whether overactivation of hippocampal cofilin also influences the formation of short-term memories. To this end, mice were either injected with an adeno-associated virus expressing catalytically active cofilin, or an eGFP control, in the hippocampus. We show for the first time that cofilin overactivation improves short-term memory formation in the object-location memory task, without affecting anxiety-like behavior. Surprisingly, we found no effect of cofilin overactivation on AMPA receptor expression levels. Altogether, while cofilin overactivation might negatively impact the formation of long-lasting memories, it may benefit short-term plasticity.

A Study on REM Sleep Homeostasis in the Day-Active Tree Shrew (Tupaia belangeri): Cold-Induced Suppression of REM Sleep Is Not Followed by a Rebound.

The function and regulation of rapid-eye-movement (REM) sleep is a topic of ongoing debate. It is often assumed that REM sleep is a homeostatically regulated process and that a need for REM sleep builds up, either during prior wakefulness or during preceding slow wave sleep. In the current study, we tested this hypothesis in six diurnal tree shrews (Tupaia belangeri), small mammals closely related to primates. All animals were individually housed and kept under a 12:12 light-dark cycle with an ambient temperature of 24 °C. We recorded sleep and temperature in the tree shrews for 3 consecutive 24 h days. During the second night, we exposed the animals to a low ambient temperature of 4 °C, a procedure that is known to suppress REM sleep. Cold exposure caused a significant drop in brain temperature and body temperature and also resulted in a strong and selective suppression of REM sleep by 64.9%. However, contrary to our expectation, the loss of REM sleep was not recovered during the subsequent day and night. These findings in a diurnal mammal confirm that the expression of REM sleep is highly sensitive to environmental temperature but do not support the view that REM sleep is homeostatically regulated in this species.

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).

Recovering object-location memories after sleep deprivation-induced amnesia.

It is well established that sleep deprivation after learning impairs hippocampal memory processes and can cause amnesia. It is unknown, however, whether sleep deprivation leads to the loss of information or merely the suboptimal storage of information that is difficult to retrieve. Here, we show that hippocampal object-location memories formed under sleep deprivation conditions can be successfully retrieved multiple days following training, using optogenetic dentate gyrus (DG) memory engram activation or treatment with the clinically approved phosphodiesterase 4 (PDE4) inhibitor roflumilast. Moreover, the combination of optogenetic DG memory engram activation and roflumilast treatment, 2 days following training and sleep deprivation, made the memory more persistently accessible for retrieval even several days later (i.e., without further optogenetic or pharmacological manipulation). Altogether, our studies in mice demonstrate that sleep deprivation does not necessarily cause memory loss but instead leads to the suboptimal storage of information that cannot be retrieved without drug treatment or optogenetic stimulation. Furthermore, our findings suggest that object-location memories, consolidated under sleep deprivation conditions and thought to be lost, can be made accessible again several days after the learning and sleep deprivation episode, using the clinically approved PDE4 inhibitor roflumilast.

Seasonal variation in rest-activity patterns in barnacle geese: are measurements of activity a good indicator of sleep-wake patterns?

Sleep is a widely spread phenomenon in the animal kingdom and is thought to serve important functions. Yet, the function of sleep remains an enigma. Studies in non-model animal species in their natural habitat might provide more insight into the evolution and function of sleep. However, polysomnography in the wild may not always be an option or first choice and some studies may need to rely on rest-activity recordings as a proxy for sleep and wakefulness. In the current paper, we analyzed how accelerometry-based activity data correlate with electroencephalogram (EEG)-based sleep-wake patterns in barnacle geese under seminatural conditions across different seasons. In winter, the geese had pronounced daily rhythms in rest and activity, with most activity occurring during the daytime. In summer, activity was more spread out over the 24 h cycle. Hourly activity scores strongly correlated with EEG-determined time awake, but the strength of the correlation varied with phase of the day and season. In winter, the correlations between activity and waking time were weaker for daytime than for night-time. Furthermore, the correlations between activity and waking during daytime were weaker in winter than in summer. During daytime in winter, there were many instances where the birds were awake but not moving. Experimental sleep deprivation had no effect on the strength of the correlation between activity scores and EEG-based wake time. Overall, hourly activity scores also showed significant inverse correlation with the time spent in non-rapid eye movement (NREM) sleep. However, correlation between activity scores and time spent in REM sleep was weak. In conclusion, accelerometry-based activity scores can serve as a good estimate for time awake or even the specific time spent in NREM sleep. However, activity scores cannot reliably predict REM sleep and sleep architecture.

Chronic Social Defeat Stress Shifts Peripheral Circadian Clocks in Male Mice in a Tissue-Specific and Time-of-Day Dependent Fashion.

Uncontrollable stress is linked to the development of many diseases, some of which are associated with disrupted daily rhythms in physiology and behavior. While available data indicate that the master circadian pacemaker in the suprachiasmatic nucleus (SCN) is unaffected by stress, accumulating evidence suggest that circadian oscillators in peripheral tissues and organs can be shifted by a variety of stressors and stress hormones. In the present study, we examined effects of acute and chronic social defeat stress in mice and addressed the question of whether effects of uncontrollable stress on peripheral clocks are tissue specific and depend on time of day of stress exposure. 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 social defeat stress in the early (ZT13-14) or late (ZT21-22) dark phase, either once (acute stress) or repeatedly on 10 consecutive days (chronic stress). One hour after the last stressor, tissue samples from liver, lung, kidney, and white adipose tissue (WAT) were collected. Social defeat stress caused a phase delay of several hours in the rhythm of PER2 expression in lung and kidney, but this delay was stronger after chronic than after acute stress. Moreover, shifts only occurred after stress in the late dark phase, not in the early dark phase. PER2 rhythms in liver and WAT were not significantly shifted by social defeat, suggesting a different response of various peripheral clocks to stress. This study indicates that uncontrollable social defeat stress is capable of shifting peripheral clocks in a time of day dependent and tissue specific manner. These shifts in peripheral clocks were smaller or absent after a single stress exposure and may therefore be the consequence of a cumulative chronic stress effect.

Sleep deprivation reduces the density of individual spine subtypes in a branch-specific fashion in CA1 neurons.

Sleep deprivation has a negative impact on hippocampus-dependent memory, which is thought to depend on cellular plasticity. We previously found that 5 h of sleep deprivation robustly decreases dendritic spine density in the CA1 area of the hippocampus in adult male mice. However, recent work by others suggests that sleep deprivation increases the density of certain spine types on specific dendritic branches. Based on these recent findings and our previous work, we conducted a more in-depth analysis of different spine types on branches 1, 2 and 5 of both apical and basal dendrites to assess whether 5 h of sleep deprivation may have previously unrecognized spine-type and branch-specific effects. This analysis shows no spine-type specific changes on branch 1 and 2 of apical dendrites after sleep deprivation. In contrast, sleep deprivation decreases the number of mushroom and branched spines on branch 5. Likewise, sleep deprivation reduces thin, mushroom and filopodia spine density on branch 5 of the basal dendrites, without affecting spines on branch 1 and 2. Our findings indicate that sleep deprivation leads to local branch-specific reduction in the density of individual spine types, and that local effects might not reflect the overall impact of sleep deprivation on CA1 structural plasticity. Moreover, our analysis underscores that focusing on a subset of dendritic branches may lead to potential misinterpretation of the overall impact of, in this case, sleep deprivation on structural plasticity.

A comparison of continuous and intermittent EEG recordings in geese: How much data are needed to reliably estimate sleep-wake patterns?

Recent technological advancements allow researchers to measure electrophysiological parameters of animals, such as sleep, in remote locations by using miniature dataloggers. Yet, continuous recording of sleep might be constrained by the memory and battery capacity of the recording devices. These limitations can be alleviated by recording intermittently instead of continuously, distributing the limited recording capacity over a longer period. We assessed how reduced sampling of sleep recordings affected measurement precision of NREM sleep, REM sleep, and Wake. We analysed a dataset on sleep in barnacle geese that we resampled following 12 different recording schemes, with data collected for 1 min per 5 min up to 1 min per 60 min in steps of 5 min. Recording 1 min in 5 min still yielded precise estimates of hourly sleep-wake values (correlations of 0.9) while potentially extending the total recording period by a factor of 5. The correlation strength gradually decreased to 0.5 when recording 1 min per 60 min. For hourly values of Wake and NREM sleep, the correlation strength in winter was higher compared with summer, reflecting more fragmented sleep in summer. Interestingly for hourly values of REM sleep, correlations were unaffected by season. Estimates of total 24 h sleep-wake values were similar for all intermittent recording schedules compared to the continuous recording. These data indicate that there is a large safe range in which researchers can periodically record sleep. Increasing the sample size while maintaining precision can substantially increase the statistical power, and is therefore recommended whenever the total recording time is limited.

The continued need for animals to advance brain research.

Policymakers aim to move toward animal-free alternatives for scientific research and have introduced very strict regulations for animal research. We argue that, for neuroscience research, until viable and translational alternatives become available and the value of these alternatives has been proven, the use of animals should not be compromised.

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.

The role of clock genes in sleep, stress and memory.

Circadian clock genes serve as the molecular basis for animals' ~24-h internal timekeeping. Clock gene expression inside and outside of the mammalian brain's circadian pacemaker (i.e. the SCN) integrates temporal information into a wealth of physiological processes. Ample data suggests that in addition to canonical cellular timekeeping functions, clock proteins also interact with proteins involved in cellular processes not related to timekeeping, including protein regulation and the interaction with other signaling mechanisms not directly linked to the regulation of circadian rhythms. Indeed, recent data suggests that clock genes outside the SCN are involved in fundamental brain processes such as sleep/wakefulness, stress and memory. The role of clock genes in these brain processes are complex and divers, influencing many molecular pathways and phenotypes. In this review, we will discuss recent work on the involvement of clock genes in sleep, stress, and memory. Moreover, we raise the controversial possibility that these functions may be under certain circumstances independent of their circadian timekeeping function.

Cloud cover amplifies the sleep-suppressing effect of artificial light at night in geese.

In modern society the night sky is lit up not only by the moon but also by artificial light devices. Both of these light sources can have a major impact on wildlife physiology and behaviour. For example, a number of bird species were found to sleep several hours less under full moon compared to new moon and a similar sleep-suppressing effect has been reported for artificial light at night (ALAN). Cloud cover at night can modulate the light levels perceived by wildlife, yet, in opposite directions for ALAN and moon. While clouds will block moon light, it may reflect and amplify ALAN levels and increases the night glow in urbanized areas. As a consequence, cloud cover may also modulate the sleep-suppressing effects of moon and ALAN in different directions. In this study we therefore measured sleep in barnacle geese (Branta leucopsis) under semi-natural conditions in relation to moon phase, ALAN and cloud cover. Our analysis shows that, during new moon nights stronger cloud cover was indeed associated with increased ALAN levels at our study site. In contrast, light levels during full moon nights were fairly constant, presumably because of moonlight on clear nights or because of reflected artificial light on cloudy nights. Importantly, cloud cover caused an estimated 24.8% reduction in the amount of night-time NREM sleep from nights with medium to full cloud cover, particularly during new moon when sleep was unaffected by moon light. In conclusion, our findings suggest that cloud cover can, in a rather dramatic way, amplify the immediate effects of ALAN on wildlife. Sleep appears to be highly sensitive to ALAN and may therefore be a good indicator of its biological effects.

Elucidating the role of protein synthesis in hippocampus-dependent memory consolidation across the day and night.

It is widely acknowledged that de novo protein synthesis is crucial for the formation and consolidation of long-term memories. While the basal activity of many signaling cascades that modulate protein synthesis fluctuates in a circadian fashion, it is unclear whether the temporal dynamics of protein synthesis-dependent memory consolidation vary depending on the time of day. More specifically, it is unclear whether protein synthesis inhibition affects hippocampus-dependent memory consolidation in rodents differentially across the day (i.e., the inactive phase with an abundance of sleep) and night (i.e., the active phase with little sleep). To address this question, male and female C57Bl6/J mice were trained in a contextual fear conditioning task at the beginning or the end of the light phase. Animals received a single systemic injection with the protein synthesis inhibitor anisomycin or vehicle directly, 4, 8 hr, or 11.5 hr following training, and memory was assessed after 24 hr. Here, we show that protein synthesis inhibition impaired the consolidation of context-fear memories selectively when the protein synthesis inhibitor was administered at the first three time points, irrespective of timing of training. Even though the basal activity of signaling pathways regulating de novo protein synthesis may fluctuate across the 24-hr cycle, these results suggest that the temporal dynamics of protein synthesis-dependent memory consolidation are similar for day-time and night-time learning.

Seasonal variation in sleep homeostasis in migratory geese: a rebound of NREM sleep following sleep deprivation in summer but not in winter.

Sleep is a behavioral and physiological state that is thought to serve important functions. Many animals go through phases in the annual cycle where sleep time might be limited, for example, during the migration and breeding phases. This leads to the question whether there are seasonal changes in sleep homeostasis. Using electroencephalogram (EEG) data loggers, we measured sleep in summer and winter in 13 barnacle geese (Branta leucopsis) under semi-natural conditions. During both seasons, we examined the homeostatic regulation of sleep by depriving the birds of sleep for 4 and 8 h after sunset. In winter, barnacle geese showed a clear diurnal rhythm in sleep and wakefulness. In summer, this rhythm was less pronounced, with sleep being spread out over the 24-h cycle. On average, the geese slept 1.5 h less per day in summer compared with winter. In both seasons, the amount of NREM sleep was additionally affected by the lunar cycle, with 2 h NREM sleep less during full moon compared to new moon. During summer, the geese responded to 4 and 8 h of sleep deprivation with a compensatory increase in NREM sleep time. In winter, this homeostatic response was absent. Overall, sleep deprivation only resulted in minor changes in the spectral composition of the sleep EEG. In conclusion, barnacle geese display season-dependent homeostatic regulation of sleep. These results demonstrate that sleep homeostasis is not a rigid phenomenon and suggest that some species may tolerate sleep loss under certain conditions or during certain periods of the year.

A brief period of sleep deprivation negatively impacts the acquisition, consolidation, and retrieval of object-location memories.

Memory is a cognitive concept and refers to the storage of information over a longer time period. It exists of a series of complementary processes; acquisition, consolidation, and retrieval. Each of these processes has its own partly unique neurobiological signature. Sleep deprivation is known to impair hippocampus-dependent long-term memories. Many studies have used extended periods of wakefulness, affecting all three memory processes, thereby making it unable to determine how each of the processes is affected by sleep loss, separately. Others have extensively examined the effects on memory consolidation, showing the detrimental effect of sleep deprivation during the consolidation process on memory formation. Few studies have investigated how memory acquisition and its retrieval are affected by sleep loss. In the present study, we therefore assessed in mice how sleep deprivation negatively impacts memory acquisition, consolidation, and retrieval, in the Object Location Memory task. Mice were sleep deprived for six hours at the beginning of the light phase using the gentle handling method, 1) directly preceding the learning trial (acquisition), 2) immediately after the learning trial (consolidation), or 3) directly preceding the test trial (retrieval). Memory was assessed at either a 24-h or 1-h interval. Using this approach, we show for the first time that six hours of sleep deprivation attenuates the acquisition, consolidation, and retrieval of object-location memories in mice.