Entrainment of the circadian clock in the liver by feeding

Circadian rhythms of behavior are driven by oscillators in the brain that are coupled to the environmental light cycle. Circadian rhythms of gene expression occur widely in peripheral organs. It is unclear how these multiple rhythms are coupled together to form a coherent system. To study such coupling, we investigated the effects of cycles of food availability (which exert powerful entraining effects on behavior) on the rhythms of gene expression in the liver, lung, and suprachiasmatic nucleus (SCN). We used a transgenic rat model whose tissues express luciferase in vitro. Although rhythmicity in the SCN remained phase-locked to the light-dark cycle, restricted feeding rapidly entrained the liver, shifting its rhythm by 10 hours within 2 days. Our results demonstrate that feeding cycles can entrain the liver independently of the SCN and the light cycle, and they suggest the need to reexamine the mammalian circadian hierarchy. They also raise the possibility that peripheral circadian oscillators like those in the liver may be coupled to the SCN primarily through rhythmic behavior, such as feeding.     

Resetting central and peripheral circadian oscillators in transgenic rats

In multicellular organisms, circadian oscillators are organized into multitissue systems which function as biological clocks that regulate the activities of the organism in relation to environmental cycles and provide an internal temporal framework. To investigate the organization of a mammalian circadian system, we constructed a transgenic rat line in which luciferase is rhythmically expressed under the control of the mouse Per1 promoter. Light emission from cultured suprachiasmatic nuclei (SCN) of these rats was invariably and robustly rhythmic and persisted for up to 32 days in vitro. Liver, lung, and skeletal muscle also expressed circadian rhythms, which damped after two to seven cycles in vitro. In response to advances and delays of the environmental light cycle, the circadian rhythm of light emission from the SCN shifted more rapidly than did the rhythm of locomotor behavior or the rhythms in peripheral tissues. We hypothesize that a self-sustained circadian pacemaker in the SCN entrains circadian oscillators in the periphery to maintain adaptive phase control, which is temporarily lost following large, abrupt shifts in the environmental light cycle.     

Multiple circadian oscillators regulate the timing of behavioral and endocrine rhythms in female golden hamsters

A single daily "surge" in pituitary luteinizing hormone release was observed in ovariectomized-estrogen-treated hamsters expressing an intact circadian rhythm of locomotor activity. In contrast, two luteinizing hormone surges occurred within a single 24-hour period in hamsters whose activity rhythm had dissociated or "split" into two distinct components. These observations indicate that both behavioral and endocrine circadian rhythms are regulated by the same multioscillator system, which seems to be composed of at least two distinct circadian oscillators.     

Differential rescue of light- and food-entrainable circadian rhythms

When food is plentiful, circadian rhythms of animals are powerfully entrained by the light-dark cycle. However, if animals have access to food only during their normal sleep cycle, they will shift most of their circadian rhythms to match the food availability. We studied the basis for entrainment of circadian rhythms by food and light in mice with targeted disruption of the clock gene Bmal1, which lack circadian rhythmicity. Injection of a viral vector containing the Bmal1 gene into the suprachiasmatic nuclei of the hypothalamus restored light-entrainable, but not food-entrainable, circadian rhythms. In contrast, restoration of the Bmal1 gene only in the dorsomedial hypothalamic nucleus restored the ability of animals to entrain to food but not to light. These results demonstrate that the dorsomedial hypothalamus contains a Bmal1-based oscillator that can drive food entrainment of circadian rhythms.     

Temperature as a universal resetting cue for mammalian circadian oscillators

Environmental temperature cycles are a universal entraining cue for all circadian systems at the organismal level with the exception of homeothermic vertebrates. We report here that resistance to temperature entrainment is a property of the suprachiasmatic nucleus (SCN) network and is not a cell-autonomous property of mammalian clocks. This differential sensitivity to temperature allows the SCN to drive circadian rhythms in body temperature, which can then act as a universal cue for the entrainment of cell-autonomous oscillators throughout the body. Pharmacological experiments show that network interactions in the SCN are required for temperature resistance and that the heat shock pathway is integral to temperature resetting and temperature compensation in mammalian cells. These results suggest that the evolutionarily ancient temperature resetting response can be used in homeothermic animals to enhance internal circadian synchronization.     

Antiphase oscillation of the left and right suprachiasmatic nuclei

An unusual property of the circadian timekeeping systems of animals is rhythm "splitting," in which a single daily period of physical activity (usually measured as wheel running) dissociates into two stably coupled components about 12 hours apart; this behavior has been ascribed to a clock composed of two circadian oscillators cycling in antiphase. We analyzed gene expression in the hypothalamic circadian clock, the suprachiasmatic nucleus (SCN), of behaviorally "split" hamsters housed in constant light. The results show that the two oscillators underlying the split condition correspond to the left and right sides of the bilaterally paired SCN.     

Regulation of mammalian circadian behavior by non-rod, non-cone, ocular photoreceptors

Circadian rhythms of mammals are entrained by light to follow the daily solar cycle (photoentrainment). To determine whether retinal rods and cones are required for this response, the effects of light on the regulation of circadian wheel-running behavior were examined in mice lacking these photoreceptors. Mice without cones (cl) or without both rods and cones (rdta/cl) showed unattenuated phase-shifting responses to light. Removal of the eyes abolishes this behavior. Thus, neither rods nor cones are required for photoentrainment, and the murine eye contains additional photoreceptors that regulate the circadian clock.     

Bright light resets the human circadian pacemaker independent of the timing of the sleep-wake cycle

Human circadian rhythms were once thought to be insensitive to light, with synchronization to the 24-hour day accomplished either through social contacts or the sleep-wake schedule. Yet the demonstration of an intensity-dependent neuroendocrine response to bright light has led to renewed consideration of light as a possible synchronizer of the human circadian pacemaker. In a laboratory study, the output of the circadian pacemaker of an elderly woman was monitored before and after exposure to 4 hours of bright light for seven consecutive evenings, and before and after a control study in ordinary room light while her sleep-wake schedule and social contacts remained unchanged. The exposure to bright light in the evening induced a 6-hour delay shift of her circadian pacemaker, as indicated by recordings of body temperature and cortisol secretion. The unexpected magnitude, rapidity, and stability of the shift challenge existing concepts regarding circadian phase-resetting capacity in man and suggest that exposure to bright light can indeed reset the human circadian pacemaker, which controls daily variations in physiologic, behavioral, and cognitive function.     

Molecular mechanisms of the biological clock in cultured fibroblasts

In mammals, the central circadian pacemaker resides in the hypothalamic suprachiasmatic nucleus (SCN), but circadian oscillators also exist in peripheral tissues. Here, using wild-type and cryptochrome (mCry)-deficient cell lines derived from mCry mutant mice, we show that the peripheral oscillator in cultured fibroblasts is identical to the oscillator in the SCN in (i) temporal expression profiles of all known clock genes, (ii) the phase of the various mRNA rhythms (i.e., antiphase oscillation of Bmal1 and mPer genes), (iii) the delay between maximum mRNA levels and appearance of nuclear mPER1 and mPER2 protein, (iv) the inability to produce oscillations in the absence of functional mCry genes, and (v) the control of period length by mCRY proteins.     

Melanopsin (Opn4) requirement for normal light-induced circadian phase shifting

The master circadian oscillator in the hypothalamic suprachiasmatic nucleus is entrained to the day/night cycle by retinal photoreceptors. Melanopsin (Opn4), an opsin-based photopigment, is a primary candidate for photoreceptor-mediated entrainment. To investigate the functional role of melanopsin in light resetting of the oscillator, we generated melanopsin-null mice (Opn4-/-). These mice entrain to a light/dark cycle and do not exhibit any overt defect in circadian activity rhythms under constant darkness. However, they display severely attenuated phase resetting in response to brief pulses of monochromatic light, highlighting the critical role of melanopsin in circadian photoentrainment in mammals.     

Light pulses that shift rhythms induce gene expression in the suprachiasmatic nucleus

Lighting cycles synchronize (entrain) mammalian circadian rhythms by altering activity of cells in the suprachiasmatic nucleus (SCN) of the hypothalamus, a circadian pacemaker. Exposure of hamsters and rats to light pulses at those phases of the circadian rhythm during which light can shift the rhythm caused increased immunoreactivity for the product of the immediate-early gene c-fos in cells in the region of the SCN that receives retinal fibers. Light pulses also increased messenger RNA for the Fos protein and for the immediate-early protein NGFI-A in the rat SCN. Similar increases in mRNA for NGFI-A were seen in the SCN of hamsters. Thus cells in this portion of the SCN undergo alterations in gene expression in response to retinal illumination, but only at times in the circadian cycle when light is capable of influencing entrainment.     

Illuminating the circadian clock in monarch butterfly migration

Migratory monarch butterflies use a time-compensated Sun compass to navigate to their overwintering grounds in Mexico. Here, we report that constant light, which disrupts circadian clock function at both the behavioral and molecular levels in monarchs, also disrupts the time-compensated component of flight navigation. We further show that ultraviolet light is important for flight navigation but is not required for photic entrainment of circadian rhythms. Tracing these distinct light-input pathways into the brain should aid our understanding of the clock-compass mechanisms necessary for successful migration.     

Phototransduction by retinal ganglion cells that set the circadian clock

Light synchronizes mammalian circadian rhythms with environmental time by modulating retinal input to the circadian pacemaker-the suprachiasmatic nucleus (SCN) of the hypothalamus. Such photic entrainment requires neither rods nor cones, the only known retinal photoreceptors. Here, we show that retinal ganglion cells innervating the SCN are intrinsically photosensitive. Unlike other ganglion cells, they depolarized in response to light even when all synaptic input from rods and cones was blocked. The sensitivity, spectral tuning, and slow kinetics of this light response matched those of the photic entrainment mechanism, suggesting that these ganglion cells may be the primary photoreceptors for this system.     

Circadian rhythm of brain self-stimulation behavior

Under constant conditions of light, sound, temperature, and humidity, rats exhibited circadian rhythmicity in rate of bar-pressing with hypothalamic and septal reinforcing brain stimulation. Variations in reinforcer magnitude aflected absolute levels of operant response emission but not the frequency of the circadian rhythm. In long sessions, the time of peak responding deviated systematically from a strict 24-hour period. Such data show marked similarity to free-running rhythms of motor activity.     

A mutation of the circadian system in golden hamsters

A mutation has been found that dramatically shortens the period of the circadian locomotor rhythm of golden hamsters. The pattern of inheritance of this mutation suggests that it occurred at a single, autosomal locus (tau). Wild-type animals have rhythms with free-running periods averaging about 24 hours; animals heterozygous for the mutation have periods of about 22 hours, whereas homozygous animals have rhythms with periods close to 20 hours. Animals that carry the mutant alleles exhibit abnormal entrainment to 24-hour light:dark cycles or are unable to entrain.     

Potassium Channels in Samanea saman Protoplasts Controlled by Phytochrome and the Biological Clock

Leaflet movement in legumes depends on rhythmic, light-regulated ion fluxes in opposing regions of the leaf-moving organ. In flexor and extensor protoplasts from Samanea saman Merrill, opening and closing of K(+) channels were rhythmic in constant darkness. When channels were open in flexor protoplasts they were closed in extensor protoplasts, and vice versa. The rhythms were shifted by a delay in the onset of constant darkness, a response typical of endogenous circadian rhythms. During the light period, the channels in flexor protoplasts were sensitive to red light that was followed by premature darkness; phytochrome was implicated as the photoreceptor.     

Testosterone induces "splitting" of circadian locomotor activity rhythms in birds

Under the influence of testosterone, the free-running circadian rhythm of locomotor activity of the starling, Sturnus vulgaris, tends to "split" into two components which temporarily run with different circadian frequencies: "splitting" occurred in intact birds whose testes grew, and in castrated birds that were injected with testosterone. Since "splitting" most probably reflects the temporal separation of two (or two groups of) circadian oscillators, these results suggest that testosterone affects the mutual coupling of circadian oscillators controlling locomotor activity.     

光照周期对尼罗罗非鱼幼鱼昼夜节律调节的作用

为了研究尼罗罗非鱼幼鱼游泳行为在不喂食时是否存在昼夜节律和光照周期的调节作用,设计了光周期为光照(L)∶黑暗(D)=12 h∶12 h,持续的黑暗(DD),持续的光照(LL),光周期为L∶D=6 h∶6 h和光周期为L∶D=2 h∶2 h。结果表明:(1)光周期为L∶D=12 h∶12 h时,尼罗罗非鱼具有明显的昼夜节律,昼夜节律周期为(24. 3±0. 2) h;(2)尼罗罗非鱼的昼夜节律在持续的黑暗和光照下仍然存在,分别为(25. 1±1. 1) h和(25. 6±1. 0) h;(3)光周期为L∶D=6 h∶6 h时,尼罗罗非鱼仍具有明显的昼夜节律(12. 6±0. 5) h;(4)在光周期为L∶D=2 h∶2 h时,尼罗罗非鱼的昼夜游泳行为仍具有明显的昼夜节律,节律周期为(4. 0±2. 0) h。这些结果表明,尼罗罗非鱼具有以24 h为周期的内源性生物钟,但相比与外源性光照调控,内源性的生物钟对罗非鱼的调控较弱,外源性的光照周期才是调节尼罗罗非鱼幼鱼昼夜行为节律的主要因素。     

Time measurement in insect photoperiodism: reversal of a photoperiod effect by chilling

Spectacular reversals of the photoperiodic control of diapause are obtained if females of Nasonia vitripennis are chilled for 4 hours in certain lightdark cycles. Experiments in which chilling is combined with short light breaks (night interruptions) show that the first peak of diapause inhibition moves in response to the chilling. This result provides an explanation for the photoperiodic reversal; it is also circumstantial evidence for the participation of circadian rhythms in photoperiodism.