Circadian rhythms

Mammals have developed an endogenous circadian clock located in the brain suprachiasmatic nuclei (SCN) of the anterior hypothalamus that responds to the environmental light–dark cycle (Fig. 1). Light is absorbed through the retina and this information is transmitted to the SCN, which in turn relays the information via neuronal connections or circulating humoral factors to peripheral clocks, such as the liver, heart and lungs, regulating cellular and physiological functions⁽Reference Reppert and Weaver^1–Reference Schibler, Ripperger and Brown³⁾. The clock mechanism in both SCN neurons and peripheral tissues consists of CLOCK and BMAL1 (brain-muscle-Arnt-like 1) proteins that heterodimerise and bind to E-box sequences to mediate transcription of tissue-specific genes, including Periods (Per1, Per2, Per3) and Cryptochromes (Cry1, Cry2). PER and CRY constitute part of the negative feedback loop, which inhibits CLOCK:BMAL1-mediated transcription⁽Reference Reppert and Weaver^1, Reference Froy, Chang and Reppert⁴⁾.

Fig. 1. Effect of feeding diet regimens on circadian rhythms and health. SCN, suprachiasmatic nuclei.

Chronodisruption and ageing

Disruption of the coordination between the endogenous clock and the environment leads to symptoms of fatigue, disorientation and insomnia. Night-shift workers have disrupted circadian rhythms and they exhibit metabolic disorders, hormone imbalance⁽Reference Davis and Mirick⁵⁾, psychological and sleep disorders⁽Reference Qureshi and Mehler⁶⁾, and increased incidence of cancer and malignant growth⁽Reference Davis and Mirick⁵⁾. Longevity in hamsters is decreased with disruption of rhythmicity and is increased in older animals given fetal SCN implants that restore high-amplitude rhythms⁽Reference Hurd and Ralph⁷⁾. Even chronic reversal of the external light–dark cycle at weekly intervals results in a significant decrease in the survival time of cardiomyopathic hamsters⁽Reference Penev, Kolker and Zee⁸⁾.

It has been shown that circadian rhythms change with normal ageing, including a shift in the phase and decrease in amplitude⁽Reference Hofman and Swaab^9, Reference Froy and Miskin¹⁰⁾. Deficiency of the CLOCK protein significantly affects longevity, as the average lifespan of Clock^−/− mice was reduced by 15% compared with wild-type mice, while maximum life span was reduced by more than 20%. CLOCK deficiency also resulted in the development of cataracts and dermatitis, two age-specific pathologies⁽Reference Seyfarth, Schliemann and Antonov^11, Reference Iroku-Malize and Kirsch¹²⁾, at a much higher rate than in wild-type mice⁽Reference Dubrovsky, Samsa and Kondratov¹³⁾. In addition, Bmal1^−/− knockout mice have reduced life span and they display various symptoms of premature ageing, including cataracts and organ shrinkage⁽Reference Kondratov, Kondratova and Gorbacheva¹⁴⁾. Per1,2 ^−/− mice are morphologically indistinguishable from wild-type animals at birth, but as early as 12–14 months of age they start to develop features of premature ageing, such as a faster decline in fertility, loss of soft tissues and kyphosis⁽Reference Lee^15, Reference Froy¹⁶⁾.

It has been reported that old mice are approximately 20 times less sensitive to the synchronising effect of light compared with young animals⁽Reference Zhang, Brainard and Zee¹⁷⁾. When the SCN becomes less sensitive, the endogenous period (τ) becomes extremely important. A positive link between τ close to 24 h and survival has been previously suggested⁽Reference Hurd and Ralph^7, Reference Wyse, Coogan and Selman¹⁸⁾. According to this suggestion, τ longer or shorter than 24 h necessitates a daily synchronisation to external time cues (i.e. light–dark cycles) with a physiological cost proportional to the deviation. This cost might affect survival. We have recently shown that a long-lived transgenic mouse has a τ of 24 h at a young and old age compared with its short-lived genetic background whose τ is 23·5 h at young age and 25 h at old age⁽Reference Gutman, Genzer and Chapnik¹⁹⁾.

Circadian rhythms in metabolism

Obesity has become a serious and growing public health problem⁽Reference Wyatt, Winters and Dubbert²⁰⁾. Attempts to understand the causes of obesity and develop new therapeutic strategies have mostly focused on the imbalance between energy expenditure and energy intake. However, studies in the last decade link energy regulation to the circadian clock at the behavioural, physiological and molecular levels⁽Reference Oishi, Shirai and Ishida^21–Reference Froy²⁴⁾, emphasising that the timing of food intake itself may play a significant role in weight gain⁽Reference Arble, Bass and Laposky²⁵⁾. Obesity, which is characterised by the excess of fat accumulation in white adipose tissue, has been related to irregular sleep–wake schedules, high snacking frequency or social jet lag known to disrupt the circadian clock⁽Reference McHill and Wright²⁶⁾.

The circadian clock regulates metabolism and energy homeostasis in peripheral tissues⁽Reference Froy^24, Reference Garaulet and Madrid^27, Reference Kuehn²⁸⁾. This is achieved by mediating the expression and/or activity of certain metabolic enzymes and transport systems⁽Reference Hirota and Fukada^29, Reference Kohsaka and Bass³⁰⁾ involved in cholesterol metabolism, amino acid regulation, drug and toxin metabolism, the citric acid cycle, and glycogen and glucose metabolism⁽Reference Froy^24, Reference Garaulet and Madrid^27, Reference La Fleur, Kalsbeek and Wortel^31–Reference Ramsey, Marcheva and Kohsaka³⁴⁾. Moreover, lesions of rat central clock in the SCN abolishes diurnal variations in whole body glucose homeostasis⁽Reference Cailotto, La Fleur and Van Heijningen³⁵⁾, altering not only rhythms in glucose utilisation rates but also endogenous hepatic glucose production. Indeed, the SCN projects to the pre-autonomic paraventricular nucleus neurons to control hepatic glucose production⁽Reference Kalsbeek, Ruiter and La Fleur³⁶⁾. Similarly, glucose uptake and the concentration of the primary cellular metabolic currency ATP in the brain and peripheral tissues have been found to fluctuate around the circadian cycle⁽Reference La Fleur^32, Reference Kalsbeek, Ruiter and La Fleur^36, Reference Yamazaki, Ishida and Inouye³⁷⁾. In addition, many hormones involved in metabolism, such as insulin ⁽Reference La Fleur, Kalsbeek and Wortel³¹⁾, glucagon⁽Reference Ruiter, La Fleur and van Heijningen³⁸⁾, adiponectin⁽Reference Ando, Yanagihara and Hayashi³⁹⁾, corticosterone⁽Reference De Boer and Van der Gugten⁴⁰⁾, leptin and ghrelin⁽Reference Ahima, Prabakaran and Flier^41, Reference Bodosi, Gardi and Hajdu⁴²⁾, have been shown to exhibit circadian oscillation.

However, the most compelling connection between the circadian clock and metabolism is achieved by genetic knockout or mutated clock genes. Homozygous Clock mutant mice have a greatly attenuated diurnal feeding rhythm, are hyperphagic and obese, and develop a metabolic syndrome of hyperleptinaemia, hyperlipidaemias, hepatic steatosis and hyperglycaemia⁽Reference Turek, Joshu and Kohsaka²²⁾. Combination of this mutation with the leptin knockout (ob/ob) resulted in significantly heavier mice than the ob/ob phenotype⁽Reference Oishi, Ohkura and Wakabayashi⁴³⁾, emphasising the inter-relations between leptin and the circadian clock⁽Reference Froy^24, Reference Garaulet and Madrid^27, Reference Green, Takahashi and Bass⁴⁴⁾. In addition, Bmal1^−/− knockout mice, similarly to Clock mutant mice, exhibit suppressed diurnal variations in glucose and TAG as well as abolished gluconeogenesis⁽Reference Rudic, McNamara and Curtis⁴⁵⁾.

Moreover, several key metabolic factors have been shown to participate in the core clock mechanism. REV-ERBα, the negative regulator of Bmal1 ⁽Reference Preitner, Damiola and Lopez-Molina⁴⁶⁾, is induced during normal adipogenesis⁽Reference Chawla and Lazar⁴⁷⁾. The positive regulators of Bmal1 expression, retinoid-related orphan receptor α and PPARα, regulate lipid metabolism⁽Reference Sato, Panda and Miraglia^48, Reference Canaple, Rambaud and Dkhissi-Benyahya⁴⁹⁾. In turn, CLOCK:BMAL1 heterodimer regulates the expression of Rev-erbα, Pparα and Rora (retinoid-related orphan receptor α)⁽Reference Oishi, Shirai and Ishida^21, Reference Preitner, Damiola and Lopez-Molina^46, Reference Sato, Panda and Miraglia^48–Reference Inoue, Shinoda and Ikeda⁵¹⁾. PPARγ co-activator-1α, a PPARγ transcriptional co-activator that regulates energy metabolism, stimulates the expression of the clock genes, Bmal1 and Rev-erbα, through co-activation of the retinoid-related orphan receptors; mice lacking PPARγ co-activator-1α show abnormal diurnal rhythms of activity, body temperature and metabolic rate⁽Reference Liu, Li and Liu⁵²⁾. AMP-activated protein kinase, a sensitive sensor of low energy and nutrient state in the cell, leads to the degradation of PER and CRY proteins⁽Reference Eide, Woolf and Kang^53, Reference Lamia, Sachdeva and DiTacchio⁵⁴⁾. Degradation of the negative feedback loop leads to a phase advance in the circadian expression pattern of clock genes in mice⁽Reference Um, Yang and Yamazaki^55, Reference Barnea, Madar and Froy⁵⁶⁾. Mammalian target of rapamycin, which functions as a sensor of cellular nutrient and energy levels, is regulated by light in the SCN⁽Reference Cao, Lee and Cho⁵⁷⁾. One of the key factors in the mammalian target of rapamycin pathway, protein 70 S6 kinase 1, rhythmically phosphorylates BMAL1 allowing it to both associate with the translational machinery and stimulate circadian oscillations of protein synthesis⁽Reference Lipton, Yuan and Boyle⁵⁸⁾. SIRT1, a key factor involved in metabolism and life span, interacts directly with CLOCK and deacetylates BMAL1 and PER2⁽Reference Asher, Gatfield and Stratmann^59–Reference Nakahata, Kaluzova and Grimaldi⁶¹⁾.

Effect of restricted feeding on circadian rhythms

Limiting the time and duration of food availability with no energy reduction is termed restricted feeding (RF)⁽Reference Schibler, Ripperger and Brown^3, Reference Hirota and Fukada^29, Reference Stephan^62, Reference Cassone and Stephan⁶³⁾. Animals which receive food ad libitum every day at the same time for only a few hours, adjust to the feeding period and consume their daily food intake during that limited time⁽Reference Honma, Honma and Hiroshige^64–Reference Froy, Chapnik and Miskin⁶⁶⁾. Restricting food to a particular time of day has profound effects on the behaviour and physiology of animals. Two to four hours before the meal, the animals display food anticipatory behaviour, which is demonstrated by an increase in locomotor activity, body temperature, corticosterone secretion, gastrointestinal motility and activity of digestive enzymes⁽Reference Stephan^62, Reference Honma, Honma and Hiroshige^64, Reference Saito, Murakami and Suda^67, Reference Comperatore and Stephan⁶⁸⁾, all are known output systems of the circadian clock. RF is dominant over the SCN and drives rhythms in arrhythmic and clock mutant mice and animals with lesioned SCN, regardless of the lighting conditions⁽Reference Stephan^62, Reference Stephan, Swann and Sisk^69–Reference Horikawa, Minami and Iijima⁷³⁾. In most incidents, RF affects circadian oscillators in peripheral tissues, with no effect on the central pacemaker in the SCN⁽Reference Schibler, Ripperger and Brown^3, Reference Hirota and Fukada^29, Reference Cassone and Stephan^63, Reference Hara, Wan and Wakamatsu^71, Reference Oishi, Miyazaki and Ishida^72, Reference Damiola, Le Minh and Preitner^74, Reference Stokkan, Yamazaki and Tei⁷⁵⁾. Thus, RF uncouples the SCN from the periphery⁽Reference Lin, Liu and Li⁷⁶⁾. We have shown that long-term daytime RF can increase the amplitude of clock gene expression, increase expression of catabolic factors and reduce the levels of disease markers leading to better health⁽Reference Sherman, Frumin and Gutman⁷⁷⁾ (Fig. 1). RF diet regimen resembles the month of Ramadan, as Muslims abstain from eating and drinking during the activity period. The average low levels of cholesterol and TAG found during RF are in agreement with those found during Ramadan⁽Reference Ibrahim, Habib and Jarrar^78, Reference Salehi and Neghab⁷⁹⁾. Aksungar et al. ⁽Reference Aksungar, Topkaya and Akyildiz⁸⁰⁾ demonstrated that Ramadan fasting has some positive effects on the inflammatory state and on risk factors for CVD, such as C reactive protein and homocysteine.

Effect of energy restriction on circadian rhythms

Calorie restriction (CR) refers to a dietary regimen low in energy without malnutrition. CR restricts the amount of energy to 60–75% of ad libitum-fed animals⁽Reference Masoro, Shimokawa and Higami⁸¹⁾. It has been documented that CR significantly extends the life span of rodents by up to 50%⁽Reference Koubova and Guarente^82, Reference Masoro⁸³⁾. In addition to the increase in life span, CR also delays the occurrence of age-related diseases, such as cancer, diabetes and cataracts⁽Reference Masoro^83–Reference Roth, Mattison and Ottinger⁸⁶⁾. Theories on how CR modulates ageing and longevity abound, but the exact mechanism is still unknown⁽Reference Masoro⁸³⁾. The reduction of energy intake, and, as a result, in oxidative stress, is considered the critical beneficial factor in the CR diet regimen⁽Reference Masoro⁸³⁾. It has been argued that in mice, the oxidative stress theory can account for age-related diseases, such as cancer, but not for longevity per se ⁽Reference Muller, Lustgarten and Jang⁸⁷⁾.

As opposed to RF, CR entrains the clock in the SCN⁽Reference Challet, Caldelas and Graff^88–Reference Resuehr and Olcese⁹¹⁾, indicating that energy reduction could affect the central oscillator. CR during the daytime affects the temporal organisation of the SCN clockwork and circadian outputs in mice under light–dark cycle. In addition, CR affects photic responses of the circadian system, indicating that energy metabolism modulates gating of photic inputs in mammals⁽Reference Mendoza, Drevet and Pevet⁹²⁾. These findings suggest that synchronisation of peripheral oscillators during CR could be achieved directly due to the temporal eating, as has been reported for RF⁽Reference Hara, Wan and Wakamatsu^71, Reference Damiola, Le Minh and Preitner^74, Reference Stokkan, Yamazaki and Tei⁷⁵⁾, or by synchronising the SCN⁽Reference Challet, Caldelas and Graff^88–Reference Mendoza, Graff and Dardente⁹⁰⁾, which entrains the peripheral tissues⁽Reference Froy, Chapnik and Miskin^93, Reference Froy and Miskin⁹⁴⁾ (Fig. 1).

Effect of intermittent fasting on circadian rhythms

Intermittent fasting (IF) allows food to be available ad libitum every other day. Similarly to energetically restricted animals, IF-fed animals exhibit increased life span as well as improved cardio- and neuro-protection and increased resistance to cancer⁽Reference Mattson⁹⁵⁾. One suggested mechanism for its beneficial effects is the stimulation of cellular stress pathways induced by the IF diet regimen⁽Reference Mattson^95, Reference Anson, Guo and de Cabo⁹⁶⁾. IF alters circadian rhythms depending on the time of food introduction (Fig. 1). When food was introduced during the light period, mice exhibited almost arrhythmicity in clock gene expression in the liver. Unlike daytime feeding, night-time feeding yielded rhythms similar to those generated during ad libitum feeding⁽Reference Froy, Chapnik and Miskin⁹⁷⁾.

Effect of high-fat diet on circadian rhythms

Several studies have shown that a high-fat diet leads to disruptions in locomotor activity in total darkness and to elevated food intake during the rest phase under light–dark conditions⁽Reference Kohsaka, Laposky and Ramsey⁹⁸⁾. These changes were also manifested by disrupted clock gene expression in the hypothalamus, liver and adipose tissue as well as altered cycling of hormones in mice, rats and human subjects⁽Reference Barnea, Madar and Froy^56, Reference Kohsaka, Laposky and Ramsey^98–Reference Barnea, Madar and Froy¹⁰²⁾. In addition, a high-fat diet induced a phase delay in clock and clock-controlled genes⁽Reference Barnea, Madar and Froy^56, Reference Barnea, Madar and Froy¹⁰²⁾ (Fig. 1). Combining high-fat diet with RF led to a leaner phenotype although the energy intake was the same as mice fed a low-fat diet⁽Reference Sherman, Genzer and Cohen¹⁰³⁾. Altogether, these studies demonstrate the importance of timing of feeding over its content.

Effect of breakfast on circadian metabolism

Breakfast has previously been demonstrated to be of major importance for the 24-h regulation of glucose⁽Reference Mekary, Giovannucci and Willett¹⁰⁴⁾. Indeed, skipping breakfast has been shown to be associated with weight gain and other adverse health outcomes, including insulin resistance and increased risk for developing type 2 diabetes. In contrast, consumption of a high-energy breakfast and a low-energy dinner resulted in a significant reduction of all-day postprandial glycaemia and body weight⁽Reference Jakubowicz, Wainstein and Ahren^105–Reference Jakubowicz, Barnea and Wainstein¹⁰⁷⁾. The importance of breakfast has recently been demonstrated in type 2 diabetic patient who skipped breakfast and had increased postprandial hyperglycaemia after both lunch and dinner in association with impaired insulin response⁽Reference Jakubowicz, Wainstein and Ahren¹⁰⁸⁾.