Chronobiology in mammalian health

Circadian rhythms are daily cycles of physiology and behavior that are driven by an endogenous oscillator with a period of approximately one day. In mammals, the hypothalamic suprachiasmatic nuclei are our principal circadian oscillators which influences peripheral tissue clocks via endocrine, autonomic and behavioral cues, and other brain regions and most peripheral tissues contain circadian clocks as well. The circadian molecular machinery comprises a group of circadian genes, namely Clock, Bmal1, Per1, Per2, Per3, Cry1 and Cry2. These circadian genes drive endogenous oscillations which promote rhythmically expression of downstream genes and thereby physiological and behavioral processes. Disruptions in circadian homeostasis have pronounced impact on physiological functioning, overall health and disease susceptibility. This review introduces the general profile of circadian gene expression and tissue-specific circadian regulation, highlights the connection between the circadian rhythms and physiological processes, and discusses the role of circadian rhythms in human disease.     

Altered feeding differentially regulates circadian rhythms and energy metabolism in liver and muscle of rats

Energy metabolism follows a diurnal pattern responding to the light/dark cycle and food availability. This study investigated the impact of restricting feeding to the daylight hours and feeding a high fat diet on circadian clock (bmal1, dbp, tef and e4bp4) and metabolic (pepck, fas, ucp3, pdk4) gene expression and markers of energy metabolism in muscle and liver of rats. The results show that in chow-fed rats switched to daylight feeding, the peak diurnal expression of genes in liver was shifted by 6–12 h while expression of these genes in muscle remained in a similar phase to rats feeding ad libitum. High fat feeding during the daylight hours had limited effect on clock gene expression in liver or muscle but shifted the peak expression of metabolic genes (pepck, fas) in liver by 6–12 h. The differential effects of daylight feeding on gene and protein expression in muscle and liver were accompanied by an 8% reduction in whole body energy expenditure, a 20–30% increased glycogen content during the light phase in muscle of day-fed rats and increased adipose tissue deposition per gram food consumed. These data demonstrate that a mismatch of feeding and light/dark cycle disrupts tissue metabolism in muscle with significant consequences for whole body energy homeostasis.     

Free Access to a Running-Wheel Advances the Phase of Behavioral and Physiological Circadian Rhythms and Peripheral Molecular Clocks in Mice

Behavioral and physiological circadian rhythms are controlled by endogenous oscillators in animals. Voluntary wheel-running in rodents is thought to be an appropriate model of aerobic exercise in humans. We evaluated the effects of chronic voluntary exercise on the circadian system by analyzing temporal profiles of feeding, core body temperature, plasma hormone concentrations and peripheral expression of clock and clock-controlled genes in mice housed under sedentary (SED) conditions or given free access to a running-wheel (RW) for four weeks. Voluntary wheel-running activity advanced the circadian phases of increases in body temperature, food intake and corticosterone secretion in the mice. The circadian expression of clock and clock-controlled genes was tissue- and gene-specifically affected in the RW mice. The temporal expression of E-box-dependent circadian clock genes such as Per1, Per2, Nr1d1 and Dbp were slightly, but significantly phase-advanced in the liver and white adipose tissue, but not in brown adipose tissue and skeletal muscle. Peak levels of Per1, Per2 and Nr1d1 expression were significantly increased in the skeletal muscle of RW mice. The circadian phase and levels of hepatic mRNA expression of the clock-controlled genes that are involved in cholesterol and fatty acid metabolism significantly differed between SED and RW mice. These findings indicated that endogenous clock-governed voluntary wheel-running activity provides feedback to the central circadian clock that systemically governs behavioral and physiological rhythms.     

Daily expression of genes coding for neurotransmitters in central and peripheral tissues of redheaded bunting: Implication for circadian regulation of physiology in songbirds

In birds, circadian control of tissue level communication is not well understood. The present study investigated this, by monitoring daily oscillation of genes coding for peptides (neuropeptide Y, NPY; vasoactive intestinal peptide, VIP; somatostatis, SST) and intermediary enzymes of amine and amino acid neurotransmitters (dopamine [tyrosine hydroxylase, TH]; glutamate [glutaminase, GLS; glutamate oxaloacetate transaminase 2, GOT2]; gamma amino butyric actid, GABA [glutamic acid decarboxylase 65, GAD65]) biosynthetic pathway, along with c-FOS as an activation marker, in different tissues of migratory redheaded buntings, Emberiza bruniceps. We cloned a partial sequence of these genes, and measured their mRNA expression in the ‘central’ clock (retina, hypothalamus) and peripheral (heart, stomach, gut, liver) tissues, collected at six times (ZT 2, 6, 11, 13, 18 and 23; ZT 0 = lights on) from birds (n = 4/ ZT) in the 12 h:12 h light-dark cycle. There were daily mRNA oscillations of all genes, although with a tissue-specific expression pattern as well as with the differential phase relationships in genes within and between tissues. These results support a conserved tissue level circadian regulation of genes coding for peptide, amine and amino acid neurotransmitters, and substantiate the expression and plausible role of neurotransmitters in the peripheral tissues. We suggest a tissue-specific contribution of neurotransmitters in the circadian regulation of physiology and behaviour in a seasonal migratory species, the redheaded bunting.     

Chronic Ethanol Consumption Disrupts the Core Molecular Clock and Diurnal Rhythms of Metabolic Genes in the Liver without Affecting the Suprachiasmatic Nucleus

Chronic ethanol consumption disrupts several metabolic pathways including β-oxidation and lipid biosynthesis, facilitating the development of alcoholic fatty liver disease. Many of these same metabolic pathways are directly regulated by cell autonomous circadian clocks, and recent studies suggest that disruption of daily rhythms in metabolism contributes to multiple common cardiometabolic diseases (including non-alcoholic fatty liver disease). However, it is not known whether ethanol disrupts the core molecular clock in the liver, nor whether this, in turn, alters rhythms in lipid metabolism. Herein, we tested the hypothesis that chronic ethanol consumption disrupts the molecular circadian clock in the liver and potentially changes the diurnal expression patterns of lipid metabolism genes. Consistent with previous studies, male C57BL/6J mice fed an ethanol-containing diet exhibited higher levels of liver triglycerides compared to control mice, indicating hepatic steatosis. Further, the diurnal oscillations of core clock genes (Bmal1, Clock, Cry1, Cry2, Per1, and Per2) and clock-controlled genes (Dbp, Hlf, Nocturnin, Npas2, Rev-erbα, and Tef) were altered in livers from ethanol-fed mice. In contrast, ethanol had only minor effects on the expression of core clock genes in the suprachiasmatic nucleus (SCN). These results were confirmed in Per2^Luciferase knock-in mice, in which ethanol induced a phase advance in PER2::LUC bioluminescence oscillations in liver, but not SCN. Further, there was greater variability in the phase of PER2::LUC oscillations in livers from ethanol-fed mice. Ethanol consumption also affected the diurnal oscillations of metabolic genes, including Adh1, Cpt1a, Cyp2e1, Pck1, Pdk4, Ppargc1a, Ppargc1b and Srebp1c, in the livers of C57BL/6J mice. In summary, chronic ethanol consumption alters the function of the circadian clock in liver. Importantly, these results suggest that chronic ethanol consumption, at levels sufficient to cause steatosis, disrupts the core hepatic clock as well as the diurnal rhythms of key lipid metabolism genes.     

DNA Methylation Landscapes of Human Fetal Development

Remodelling the methylome is a hallmark of mammalian development and cell differentiation. However, current knowledge of DNA methylation dynamics in human tissue specification and organ development largely stems from the extrapolation of studies in vitro and animal models. Here, we report on the DNA methylation landscape using the 450k array of four human tissues (amnion, muscle, adrenal and pancreas) during the first and second trimester of gestation (9,18 and 22 weeks). We show that a tissue-specific signature, constituted by tissue-specific hypomethylated CpG sites, was already present at 9 weeks of gestation (W9). Furthermore, we report large-scale remodelling of DNA methylation from W9 to W22. Gain of DNA methylation preferentially occurred near genes involved in general developmental processes, whereas loss of DNA methylation mapped to genes with tissue-specific functions. Dynamic DNA methylation was associated with enhancers, but not promoters. Comparison of our data with external fetal adrenal, brain and liver revealed striking similarities in the trajectory of DNA methylation during fetal development. The analysis of gene expression data indicated that dynamic DNA methylation was associated with the progressive repression of developmental programs and the activation of genes involved in tissue-specific processes. The DNA methylation landscape of human fetal development provides insight into regulatory elements that guide tissue specification and lead to organ functionality.     

Circadian rhythm of serum and tissue lipids in fed and fasted rats

The oscillations of the free fatty acid concentration in the serum and white (epididymal) adipose tissue, of triglycerides in the serum and liver, of total serum, liver and adrenal cholesterol and of serum phospholipids were studied at 3-hour intervals for a period of 24 hours in fed male Wistar rats and in animals fasted for 24 hours (both adapted to an illumination regimen of 12 hours' light and 12 hours' darkness. The rhythm--studied by means of the cosinor analysis--was present in most of the given parameters; it was not recorded in the liver triglycerides and serum phospholipids of fasted rats and in the adrenal cholesterol of fed animals. Apart from the circadian rhythm, many parameters distinctly displayed an ultradian rhythm, mainly an approximately 12-hour period. In general, one day's starvation did not significantly affect the course of the circadian oscillations of the given indicators of rat lipid metabolism.     

Signalling entrains the peripheral circadian clock

In mammals, 24-h rhythms of behaviour and physiology are regulated by the circadian clock. The circadian clock is controlled by a central clock in the brain's suprachiasmatic nucleus (SCN) that synchronizes peripheral clocks in peripheral tissues. Clock genes in the SCN are primarily entrained by light. Increasing evidence has shown that peripheral clocks are also regulated by light and hormones independent of the SCN. How the peripheral clocks deal with internal signals is dependent on the relevance of a specific cue to a specific tissue. In different tissues, most genes that are under circadian control are not overlapping, revealing the tissue-specific control of peripheral clocks. We will discuss how different signals control the peripheral clocks in different peripheral tissues, such as the liver, gastrointestinal tract, and pancreas, and discuss the organ-to-organ communication between the peripheral clocks at the molecular level.     

Digital signal processing reveals circadian baseline oscillation in majority of mammalian genes

In mammals, circadian periodicity has been described for gene expression in the hypothalamus and multiple peripheral tissues. It is accepted that 10%–15% of all genes oscillate in a daily rhythm, regulated by an intrinsic molecular clock. Statistical analyses of periodicity are limited by the small size of datasets and high levels of stochastic noise. Here, we propose a new approach applying digital signal processing algorithms separately to each group of genes oscillating in the same phase. Combined with the statistical tests for periodicity, this method identifies circadian baseline oscillation in almost 100% of all expressed genes. Consequently, circadian oscillation in gene expression should be evaluated in any study related to biological pathways. Changes in gene expression caused by mutations or regulation of environmental factors (such as photic stimuli or feeding) should be considered in the context of changes in the amplitude and phase of genetic oscillations.