Chronic jet lag reduces motivation and affects other mood-related behaviors in male mice.

Introduction: The circadian system regulates various physiological processes such as sleep-wake cycles, hormone secretion, metabolism, and the reaction to both natural and drug-based rewards. Chronic disruption of the circadian system caused by unsteady synchronization with light-dark (LD) schedules, such as advancing chronic jet lag (CJL), leads to adverse physiological effects and pathologies, and is linked with changes in mood and depressive behaviors in humans and rodent models. Methods: C57BL/6J male mice were subjected to circadian disruption through phase advances of 6 h every 2 days (CJL +6/2). Mice under 12:12-h LD cycle were used as controls. After 8 weeks under these conditions, a battery of behavioral tests was performed to assess if mood-related behaviors were affected. Results: Compared to controls under 24 h LD cycles, mice under CJL presented desynchronization of activity-rest rhythms that led to several behavioral impairments, including a decrease in motivation for food reward, and an increase in anxiety, anhedonia, and depressive-like behavior. Conclusion: Chronic circadian disruption, caused by an experimental CJL protocol, affects mood-related and reward-related behaviors in mice. Understanding the importance of the circadian system and its potential role for disruption due to CJL is important for maintaining good health and well-being.

Chronic circadian desynchronization of feeding-fasting rhythm generates alterations in daily glycemia, LDL cholesterolemia and microbiota composition in mice.

Introduction: The circadian system synchronizes behavior and physiology to the 24-h light- dark (LD) cycle. Timing of food intake and fasting periods provide strong signals for peripheral circadian clocks regulating nutrient assimilation, glucose, and lipid metabolism. Mice under 12 h light:12 h dark (LD) cycles exhibit behavioral activity and feeding during the dark period, while fasting occurs at rest during light. Disruption of energy metabolism, leading to an increase in body mass, was reported in experimental models of circadian desynchronization. In this work, the effects of chronic advances of the LD cycles (chronic jet-lag protocol, CJL) were studied on the daily homeostasis of energy metabolism and weight gain. Methods: Male C57 mice were subjected to a CJL or LD schedule, measuring IPGTT, insulinemia, microbiome composition and lipidemia. Results: Mice under CJL show behavioral desynchronization and feeding activity distributed similarly at the light and dark hours and, although feeding a similar daily amount of food as compared to controls, show an increase in weight gain. In addition, ad libitum glycemia rhythm was abolished in CJL-subjected mice, showing similar blood glucose values at light and dark. CJL also generated glucose intolerance at dark in an intraperitoneal glucose tolerance test (IPGTT), with increased insulin release at both light and dark periods. Low-density lipoprotein (LDL) cholesterolemia was increased under this condition, but no changes in HDL cholesterolemia were observed. Firmicutes/Bacteroidetes ratio was analyzed as a marker of circadian disruption of microbiota composition, showing opposite phases at the light and dark when comparing LD vs. CJL. Discussion: Chronic misalignment of feeding/fasting rhythm leads to metabolic disturbances generating nocturnal hyperglycemia, glucose intolerance and hyperinsulinemia in a IPGTT, increased LDL cholesterolemia, and increased weight gain, underscoring the importance of the timing of food consumption with respect to the circadian system for metabolic health.

Cysteine Oxidation Promotes Dimerization/Oligomerization of Circadian Protein Period 2.

The molecular circadian clock is based on a transcriptional/translational feedback loop in which the stability and half-life of circadian proteins is of importance. Cysteine residues of proteins are subject to several redox reactions leading to S-thiolation and disulfide bond formation, altering protein stability and function. In this work, the ability of the circadian protein period 2 (PER2) to undergo oxidation of cysteine thiols was investigated in HEK-293T cells. PER2 includes accessible cysteines susceptible to oxidation by nitroso cysteine (CysNO), altering its stability by decreasing its monomer form and subsequently increasing PER2 homodimers and multimers. These changes were reversed by treatment with 2-mercaptoethanol and partially mimicked by hydrogen peroxide. These results suggest that cysteine oxidation can prompt PER2 homodimer and multimer formation in vitro, likely by S-nitrosation and disulphide bond formation. These kinds of post-translational modifications of PER2 could be part of the redox regulation of the molecular circadian clock.

Redox and Antioxidant Modulation of Circadian Rhythms: Effects of Nitroxyl, N-Acetylcysteine and Glutathione.

The circadian clock at the hypothalamic suprachiasmatic nucleus (SCN) entrains output rhythms to 24-h light cycles. To entrain by phase-advances, light signaling at the end of subjective night (circadian time 18, CT18) requires free radical nitric oxide (NO•) binding to soluble guanylate cyclase (sGC) heme group, activating the cyclic guanosine monophosphate (cGMP)-dependent protein kinase (PKG). Phase-delays at CT14 seem to be independent of NO•, whose redox-related species were yet to be investigated. Here, the one-electron reduction of NO• nitroxyl was pharmacologically delivered by Angeli's salt (AS) donor to assess its modulation on phase-resetting of locomotor rhythms in hamsters. Intracerebroventricular AS generated nitroxyl at the SCN, promoting phase-delays at CT14, but potentiated light-induced phase-advances at CT18. Glutathione/glutathione disulfide (GSH/GSSG) couple measured in SCN homogenates showed higher values at CT14 (i.e., more reduced) than at CT18 (oxidized). In addition, administration of antioxidants N-acetylcysteine (NAC) and GSH induced delays per se at CT14 but did not affect light-induced advances at CT18. Thus, the relative of NO• nitroxyl generates phase-delays in a reductive SCN environment, while an oxidative favors photic-advances. These data suggest that circadian phase-locking mechanisms should include redox SCN environment, generating relatives of NO•, as well as coupling with the molecular oscillator.

Role of G-Substrate in the NO/cGMP/PKG Signal Transduction Pathway for Photic Entrainment of the Hamster Circadian Clock.

The mammalian circadian clock at the hypothalamic suprachiasmatic nuclei (SCN) entrains biological rhythms to the 24-h cyclic environment, by encoding light-dark transitions in SCN neurons. Light pulses induce phase shifts in the clock and in circadian rhythms; photic signaling for circadian phase advances involves a nitric oxide (NO)/cyclic guanosine monophosphate (cGMP)/cGMP-dependent protein kinase (PKG) pathway, increasing the expression of Period (Per) genes. Effectors downstream of PKG remain unknown. Here we investigate the role of G-substrate (GS), a PKG substrate, in the hamster SCN. GS and phosphorylated G-substrate (p-GS) were present in a subset of SCN cells. Moreover, GS phosphorylation (p-GS/GS ratio) increased in SCN homogenates after light pulses delivered at circadian time (CT) 18 and intraperitoneal treatment with sildenafil, an inhibitor of phosphodiesterase 5 (a cGMP-specific phosphodiesterase). On the other hand, intracerebroventricular treatment with the PKG inhibitor KT5823, reduced photic phosphorylation of GS to basal levels. Since p-GS could act as a protein phosphatase 2 A (PP2A) inhibitor, we demonstrated physical interaction between p-GS and PP2A in SCN homogenates, and also a light-pulse dependent decrease of PP2A activity. Intracerebroventricular treatment with okadaic acid, a PP2A inhibitor, increased the magnitude of light-induced phase advances of locomotor rhythms. We provide evidence on the physiological phosphorylation of GS as a new downstream effector in the NO/cGMP/PKG photic pathway in the hamster SCN, including its role as a PP2A inhibitor.

Coping with Antarctic demands: Psychological implications of isolation and confinement.

Working in extreme environments requires a wide range of cognitive, psychological and social competences. Antarctica represents one of the most challenging habitats to work in due to its aridity, extremely cold weather, and isolated conditions. This study aimed to assess mood variations and coping strategies, as well as their possible modulation by group dynamics in a crew at the Belgrano II Argentine Antarctic Station throughout 1 year of confinement. Thirteen members of the Argentine Army completed emotional, coping and social dynamics questionnaires bimonthly in March, May, July, September and November. Results showed a significant decline in social dynamics scales, evidenced by decreases in perceived peer and hierarchical support. Additionally, coping strategies displayed a drop in mature defence throughout the expedition. A positive correlation was found between social support and recovery from stress. Our results highlight the importance of interpersonal relationships in psychological adjustment to isolation and extreme environments.

Access to electric light is associated with delays of the dim-light melatonin onset in a traditionally hunter-gatherer Toba/Qom community.

Key to the transition of humans from nomadic hunting-gathering groups to industrialized and highly urbanized societies was the creation of protected and artificially lit environments that extended the natural daylight hours and consolidated sleep away from nocturnal threats. These conditions isolated humans from the natural regulators of sleep and exposed them to higher levels of light during the evening, which are associated with a later sleep onset. Here, we investigated the extent to which this delayed timing of sleep is due to a delayed circadian system. We studied two communities of Toba/Qom in the northern region of Argentina, one with and the other without access to electricity. These communities have recently transitioned from a hunting-gathering subsistence to mixed subsistence systems and represent a unique model in which to study the potential effects of the access to artificial light on sleep physiology. We have previously shown that participants in the community with access to electricity had, compared to participants in the community without electricity, later sleep onsets, and shorter sleep bouts. Here, we show they also have a delayed dim-light melatonin onset (DLMO). This difference is present during the winter but not during the spring when the influence of evening artificial light is likely less relevant. Our results support the notion that the human transition into artificially lit environments had a major impact on physiological systems that regulate sleep timing, including the phase of the master circadian clock.

Subjective time estimation in Antarctica: The impact of extreme environments and isolation on a time production task.

Interval timing measures time estimation in the seconds-to-minutes range. Antarctica provides a real-world context to study the effect of extreme photoperiods and isolation on time perception. The aim of this study was to explore interval timing as a cognitive measure in the crew of Belgrano II Argentine Antarctic Station. A total of 13 subjects were assessed for interval timing in short (3 s), intermediate (6 s) and long (12 s) duration stimuli. Measures were taken during the morning and evening, five times along the year. Significant variations were found for 3 s and 6 s during the morning and 6 s during the evening. Results suggest an impact of isolation on morning performances and an effect of the polar night on evening measures. These findings shed some light on the use of interval timing as a cognitive test to assess performance in extreme environments.

Sleep misalignment and circadian rhythm impairment in long-haul bus drivers under a two-up operations system.

OBJECTIVES: The objective of the study was to describe working and sleep conditions and to assess how sleep opportunities are associated with obtained sleep and alertness, in a sample of long-haul bus drivers working with a two-up operations system. METHODS: Measures of subjective sleep and sleepiness, actigraphy, circadian temperature rhythm, and psychomotor vigilance tasks were obtained from a sample of 122 drivers from Argentina. Variables were compared between high and low fatigue risk groups, which were formed using a median split of a fatigue risk score. The score was calculated based on drivers' total working hours, maximum shift duration, minimum short break duration, maximum night work per seven days, and long break frequencies. RESULTS: Considering a standardized one-day period, sleep in the bus accounted for 1.9±0.1 h of total sleep (57±1% efficiency), sleep at destination for 1.6±0.2 h of total sleep (90±1% efficiency), and sleep at home for 3.8±0.2 h of total sleep (89±1% nap efficiency and 90±1% anchor sleep efficiency). In drivers exposed to high-risk working schedules, the circadian temperature rhythm was weaker (lower % of variance explained by the model) (22.0±1.7% vs. 27.6±2.0%, p <0.05) and without a significant acrophase. CONCLUSIONS: Drivers obtained a total amount of weekly sleep similar to the recommended levels for adults, but distributed at different locations and at different times during the day. High-risk working schedules were associated with disruption of circadian temperature rhythms. These results point out to the need of the implementation of shift-work scheduling strategies to minimize sleep misalignment and circadian desynchronization in long-haul bus drivers.

Sleep, napping and alertness during an overwintering mission at Belgrano II Argentine Antarctic station.

During Antarctic isolation personnel are exposed to extreme photoperiods. A frequent observation is a sleep onset phase delay during winter. It is not known if, as a result, daytime sleeping in the form of naps increases. We sought to assess sleep patterns - with focus on daytime sleeping - and alertness in a Latin American crew overwintering in Argentine Antarctic station Belgrano II. Measurements were collected in 13 males during March, May, July, September and November, and included actigraphy and psychomotor vigilance tasks. Sleep duration significantly decreased during winter. A total of eight participants took at least one weekly nap across all measurement points. During winter, the nap onset was delayed, its duration increased and its efficiency improved. We observed a significant effect of seasonality in the association of evening alertness with sleep onset. Our results replicate previous findings regarding sleep during overwintering in Antarctica, adding the description of the role of napping and the report of a possible modulatory effect of seasonality in the relation between sleep and alertness. Napping should be considered as an important factor in the scheduling of activities of multicultural crews that participate in Antarctica.

La actigrafía como herramienta diagnóstica / ROLE OF ACTIGRAPHY IN THE STUDY OF SLEEP AND CIRCADIAN RHYTHMS : ACCURACY, SENSITIVITY AND SPECIFICITY OF ACTIGRAPHY FOR THE ASSESSMENT OF SLEEP DISORDERS

El diagnóstico y tratamiento de los trastornos de sueño, especialmente los asociados al Ritmo Circadiano, utilizan métodos costosos, invasivos e incómodos tanto para los pacientes como para los médicos, quienes deben realizar un seguimiento de los hábitos de sueño. La actigrafía ha sido aceptada como una herramienta válida para el estudio y diagnóstico de trastornos circadianos. Más de 300 dispositivos se comercializan actualmente para el uso personal, pero pocos de estos han sido probados para un uso diagnóstico. En este estudio comparativo compuesto por 21 sujetos, se informa acerca de los patrones de sueño y actividad registrados por algunos dispositivos, como Micro-Mini Motionlogger Watch, Condor Act Trust, MisFit Flash y Fitbit Flex. No se observan diferencias significativas en el análisis del patrón de actividad de descanso entre dispositivos. Tampoco se observan para el sueño Onset (inicio), el Tiempo Total de Sueño y la Eficiencia del Sueño. Según el tipo de estudio y análisis deseado, éstos dispositivos pueden resultar alternativos para los registros de actividad y sueño.

This is a comparative analysis of actigraphy performance in comparison with different sleep Parameters. Actigraphy is a non-invasive and valid method of monitoring human rest activity cycles. The report describes the role of actigraphy to assess the study of sleep-wake patterns and circadian rhythms, evaluating its development as a diagnostic tool, with a comparative analysis of actigraphy performance in comparison with different sleep parameters. The diagnosis and treatment of sleep disorders, especially those associated with the cicardian rhythm, employ very expensive costs, invasives or unconfortable for the patients the same as for physicians, who must perform a demand of the sleeping habits. The International Classification of Sleep Disorders has identified more than 80 sleep disorders, all of them have associated treatments. Actinography has been accepted as a valid tool for the study and diagnosis of circadian disorders. All these aspects are discussed in the article

Circadian and Metabolic Effects of Light: Implications in Weight Homeostasis and Health.

Daily interactions between the hypothalamic circadian clock at the suprachiasmatic nucleus (SCN) and peripheral circadian oscillators regulate physiology and metabolism to set temporal variations in homeostatic regulation. Phase coherence of these circadian oscillators is achieved by the entrainment of the SCN to the environmental 24-h light:dark (LD) cycle, coupled through downstream neural, neuroendocrine, and autonomic outputs. The SCN coordinate activity and feeding rhythms, thus setting the timing of food intake, energy expenditure, thermogenesis, and active and basal metabolism. In this work, we will discuss evidences exploring the impact of different photic entrainment conditions on energy metabolism. The steady-state interaction between the LD cycle and the SCN is essential for health and wellbeing, as its chronic misalignment disrupts the circadian organization at different levels. For instance, in nocturnal rodents, non-24 h protocols (i.e., LD cycles of different durations, or chronic jet-lag simulations) might generate forced desynchronization of oscillators from the behavioral to the metabolic level. Even seemingly subtle photic manipulations, as the exposure to a "dim light" scotophase, might lead to similar alterations. The daily amount of light integrated by the clock (i.e., the photophase duration) strongly regulates energy metabolism in photoperiodic species. Removing LD cycles under either constant light or darkness, which are routine protocols in chronobiology, can also affect metabolism, and the same happens with disrupted LD cycles (like shiftwork of jetlag) and artificial light at night in humans. A profound knowledge of the photic and metabolic inputs to the clock, as well as its endocrine and autonomic outputs to peripheral oscillators driving energy metabolism, will help us to understand and alleviate circadian health alterations including cardiometabolic diseases, diabetes, and obesity.

Alterations in Metabolism and Diurnal Rhythms following Bilateral Surgical Removal of the Superior Cervical Ganglia in Rats.

Mammalian circadian rhythms are controlled by a master pacemaker located in the suprachiasmatic nuclei (SCN), which is synchronized to the environment by photic and nonphotic stimuli. One of the main functions of the SCN is to regulate peripheral oscillators to set temporal variations in the homeostatic control of physiology and metabolism. In this sense, the SCN coordinate the activity/rest and feeding/fasting rhythms setting the timing of food intake, energy expenditure, thermogenesis, and active and basal metabolism. One of the major time cues to the periphery is the nocturnal melatonin, which is synthesized and secreted by the pineal gland. Under SCN control, arylalkylamine N-acetyltransferase (AA-NAT)-the main enzyme regulating melatonin synthesis in vertebrates-is activated at night by sympathetic innervation that includes the superior cervical ganglia (SCG). Bilateral surgical removal of the superior cervical ganglia (SCGx) is considered a reliable procedure to completely prevent the nocturnal AA-NAT activation, irreversibly suppressing melatonin rhythmicity. In the present work, we studied the effects of SCGx on rat metabolic parameters and diurnal rhythms of feeding and locomotor activity. We found a significant difference between SCGx and sham-operated rats in metabolic variables such as an increased body weight/food intake ratio, increased adipose tissue, and decreased glycemia with a normal glucose tolerance. An analysis of locomotor activity and feeding rhythms showed an increased daytime (lights on) activity (including food consumption) in the SCGx group. These alterations suggest that superior cervical ganglia-related feedback mechanisms play a role in SCN-periphery phase coordination and that SCGx is a valid model without brain-invasive surgery to explore how sympathetic innervation affects daily (24 h) patterns of activity, food consumption and, ultimately, its role in metabolism homeostasis.

Comparative analysis of actigraphy performance in healthy young subjects.

Sleep-related health disorders are increasing worldwide; diagnosis and treatment of such sleep diseases are commonly invasive and sometimes unpractical or expensive. Actigraphy has been recently introduced as a tool for the study of sleep and circadian disorders; however, there are several devices that claim to be useful for research and have not been thoroughly tested. This comparative study provides activity, sleep and temperature information regarding several of the most commonly used actigraphers: Micro-Mini Motion Logger; Act Trust; Misfit Flash; Fitbit Flex & Thermochron. Twenty-two healthy young subjects were assessed with five different commercial actigraphs (Micro-Mini Motionlogger Watch, Condor Act Trust, MisFit Flash and Fitbit Flex) and a temperature recorder (Thermochron), and also completed a sleep diary for a week. There were not significant differences in the analysis of rest-activity pattern between devices. Temperature rhythm comparison between the Act Trust and the Thermochron showed significant differences in rhythm percentage (p<0.05) and mesor (p<0.0563) but not in amplitude or acrophase. Although data accessibility and ease of use was very different for the diverse devices, there were no significant differences for sleep onset, total sleep time and sleep efficiency recordings, where applicable. In conclusion, depending on the type of study and analysis desired (as well as cost and compliance of use), we propose some relative advantages for the different actigraphy/temperature recording devices.

N-nitrosomelatonin enhances photic synchronization of mammalian circadian rhythms.

Most physiological processes in mammals are synchronized to the daily light:dark cycle by a circadian clock located in the hypothalamic suprachiasmatic nucleus. Signal transduction of light-induced phase advances of the clock is mediated through a neuronal nitric oxide synthase-guanilyl cyclase pathway. We have employed a novel nitric oxide-donor, N-nitrosomelatonin, to enhance the photic synchronization of circadian rhythms in hamsters. The intraperitoneal administration of this drug before a sub-saturating light pulse at circadian time 18 generated a twofold increase of locomotor rhythm phase-advances, having no effect over saturating light pulses. This potentiation was also obtained even when inhibiting suprachiasmatic nitric oxide synthase activity. However, N-nitrosomelatonin had no effect on light-induced phase delays at circadian time 14. The photic-enhancing effects were correlated with an increased suprachiasmatic immunoreactivity of FBJ murine osteosarcoma viral oncogene and period1. Moreover, in vivo nitric oxide release by N-nitrosomelatonin was verified by measuring nitrate and nitrite levels in suprachiasmatic nuclei homogenates. The compound also accelerated resynchronization to an abrupt 6-h advance in the light:dark cycle (but not resynchronization to a 6-h delay). Here, we demonstrate the chronobiotic properties of N-nitrosomelatonin, emphasizing the importance of nitric oxide-mediated transduction for circadian phase advances.

The times they're a-changing: effects of circadian desynchronization on physiology and disease.

Circadian rhythms are endogenous and need to be continuously entrained (synchronized) with the environment. Entrainment includes both coupling internal oscillators to external periodic changes as well as synchrony between the central clock and peripheral oscillators, which have been shown to exhibit different phases and resynchronization speed. Temporal desynchronization induces diverse physiological alterations that ultimately decrease quality of life and induces pathological situations. Indeed, there is a considerable amount of evidence regarding the deleterious effect of circadian dysfunction on overall health or on disease onset and progression, both in human studies and in animal models. In this review we discuss the general features of circadian entrainment and introduce diverse experimental models of desynchronization. In addition, we focus on metabolic, immune and cognitive alterations under situations of acute or chronic circadian desynchronization, as exemplified by jet-lag and shiftwork schedules. Moreover, such situations might lead to an enhanced susceptibility to diverse cancer types. Possible interventions (including light exposure, scheduled timing for meals and use of chronobiotics) are also discussed.

cGMP-phosphodiesterase inhibition enhances photic responses and synchronization of the biological circadian clock in rodents.

The master circadian clock in mammals is located in the hypothalamic suprachiasmatic nuclei (SCN) and is synchronized by several environmental stimuli, mainly the light-dark (LD) cycle. Light pulses in the late subjective night induce phase advances in locomotor circadian rhythms and the expression of clock genes (such as Per1-2). The mechanism responsible for light-induced phase advances involves the activation of guanylyl cyclase (GC), cGMP and its related protein kinase (PKG). Pharmacological manipulation of cGMP by phosphodiesterase (PDE) inhibition (e.g., sildenafil) increases low-intensity light-induced circadian responses, which could reflect the ability of the cGMP-dependent pathway to directly affect the photic sensitivity of the master circadian clock within the SCN. Indeed, sildenafil is also able to increase the phase-shifting effect of saturating (1200 lux) light pulses leading to phase advances of about 9 hours, as well as in C57 a mouse strain that shows reduced phase advances. In addition, sildenafil was effective in both male and female hamsters, as well as after oral administration. Other PDE inhibitors (such as vardenafil and tadalafil) also increased light-induced phase advances of locomotor activity rhythms and accelerated reentrainment after a phase advance in the LD cycle. Pharmacological inhibition of the main downstream target of cGMP, PKG, blocked light-induced expression of Per1. Our results indicate that the cGMP-dependent pathway can directly modulate the light-induced expression of clock-genes within the SCN and the magnitude of light-induced phase advances of overt rhythms, and provide promising tools to design treatments for human circadian disruptions.

Effect of experimental glaucoma on the non-image forming visual system.

Glaucoma is a leading cause of blindness worldwide, characterized by retinal ganglion cell degeneration and damage to the optic nerve. We investigated the non-image forming visual system in an experimental model of glaucoma in rats induced by weekly injections of chondroitin sulphate (CS) in the eye anterior chamber. Animals were unilaterally or bilaterally injected with CS or vehicle for 6 or 10 weeks. In the retinas from eyes injected with CS, a similar decrease in melanopsin and Thy-1 levels was observed. CS injections induced a similar decrease in the number of melanopsin-containing cells and superior collicular retinal ganglion cells. Experimental glaucoma induced a significant decrease in the afferent pupil light reflex. White light significantly decreased nocturnal pineal melatonin content in control and glaucomatous animals, whereas blue light decreased this parameter in vehicle- but not in CS-injected animals. A significant decrease in light-induced c-Fos expression in the suprachiasmatic nuclei was observed in glaucomatous animals. General rhythmicity and gross entrainment appear to be conserved, but glaucomatous animals exhibited a delayed phase angle with respect to lights off and a significant increase in the percentage of diurnal activity. These results indicate the glaucoma induced significant alterations in the non-image forming visual system.

Paying the circadian toll: the circadian response to LPS injection is dependent on the Toll-like receptor 4.

Systemic low doses of the endotoxin lipopolysaccharide (LPS) administered at CT15 (circadian time 12 corresponds to locomotor activity onset) induce phase delays of locomotor activity rhythms in mice. To evaluate if this effect was mediated by the Toll-like receptor 4 (TLR4), our present aim was to characterize the circadian behavior and LPS-induced circadian response of TLR4 (LPS receptor)-deficient mice (in C57bl/10 and C3H backgrounds). In mutants, we observed a free-running period and a light-induced phase delay similar to the one observed in their corresponding wild-type (WT) littermates. The LPS-induced phase delay, wheel running inhibition and c-Fos/Per-1 immunoreactivity in the paraventricular nuclei observed in WT mice was absent or significantly decreased in the TLR4-deficient mice. In conclusion, we show that LPS-induced circadian responses are mediated by TLR4.