Spontaneous synchronization of coupled circadian oscillators

In mammals, the circadian pacemaker, which controls daily rhythms, is located in the suprachiasmatic nucleus (SCN). Circadian oscillations are generated in individual SCN neurons by a molecular regulatory network. Cells oscillate with periods ranging from 20 to 28 h, but at the tissue level, SCN neurons display significant synchrony, suggesting a robust intercellular coupling in which neurotransmitters are assumed to play a crucial role. We present a dynamical model for the coupling of a population of circadian oscillators in the SCN. The cellular oscillator, a three-variable model, describes the core negative feedback loop of the circadian clock. The coupling mechanism is incorporated through the global level of neurotransmitter concentration. Global coupling is efficient to synchronize a population of 10,000 cells. Synchronized cells can be entrained by a 24-h light-dark cycle. Simulations of the interaction between two populations representing two regions of the SCN show that the driven population can be phase-leading. Experimentally testable predictions are: 1), phases of individual cells are governed by their intrinsic periods; and 2), efficient synchronization is achieved when the average neurotransmitter concentration would dampen individual oscillators. However, due to the global neurotransmitter oscillation, cells are effectively synchronized.     

Noise Induces Oscillation and Synchronization of the Circadian Neurons

The principle clock of mammals, named suprachiasmatic nucleus (SCN), coordinates the circadian rhythms of behavioral and physiological activity to the external 24 h light-dark cycle. In the absence of the daily cycle, the SCN acts as an endogenous clock that regulates the ~24h rhythm of activity. Experimental and theoretical studies usually take the light-dark cycle as a main external influence, and often ignore light pollution as an external influence. However, in modern society, the light pollution such as induced by electrical lighting influences the circadian clock. In the present study, we examined the effect of external noise (light pollution) on the collective behavior of coupled circadian oscillators under constant darkness using a Goodwin model. We found that the external noise plays distinct roles in the network behavior of neurons for weak or strong coupling between the neurons. In the case of strong coupling, the noise reduces the synchronization and the period of the SCN network. Interestingly, in the case of weak coupling, the noise induces a circadian rhythm in the SCN network which is absent in noise-free condition. In addition, the noise increases the synchronization and decreases the period of the SCN network. Our findings may shed new light on the impact of the external noise on the collective behavior of SCN neurons.     

Synchronization-induced rhythmicity of circadian oscillators in the suprachiasmatic nucleus

The suprachiasmatic nuclei (SCN) host a robust, self-sustained circadian pacemaker that coordinates physiological rhythms with the daily changes in the environment. Neuronal clocks within the SCN form a heterogeneous network that must synchronize to maintain timekeeping activity. Coherent circadian output of the SCN tissue is established by intercellular signaling factors, such as vasointestinal polypeptide. It was recently shown that besides coordinating cells, the synchronization factors play a crucial role in the sustenance of intrinsic cellular rhythmicity. Disruption of intercellular signaling abolishes sustained rhythmicity in a majority of neurons and desynchronizes the remaining rhythmic neurons. Based on these observations, the authors propose a model for the synchronization of circadian oscillators that combines intracellular and intercellular dynamics at the single-cell level. The model is a heterogeneous network of circadian neuronal oscillators where individual oscillators are damped rather than self-sustained. The authors simulated different experimental conditions and found that: (1) in normal, constant conditions, coupled circadian oscillators quickly synchronize and produce a coherent output; (2) in large populations, such oscillators either synchronize or gradually lose rhythmicity, but do not run out of phase, demonstrating that rhythmicity and synchrony are codependent; (3) the number of oscillators and connectivity are important for these synchronization properties; (4) slow oscillators have a higher impact on the period in mixed populations; and (5) coupled circadian oscillators can be efficiently entrained by light–dark cycles. Based on these results, it is predicted that: (1) a majority of SCN neurons needs periodic synchronization signal to be rhythmic; (2) a small number of neurons or a low connectivity results in desynchrony; and (3) amplitudes and phases of neurons are negatively correlated. The authors conclude that to understand the orchestration of timekeeping in the SCN, intracellular circadian clocks cannot be isolated from their intercellular communication components.     

Come together, right...now: synchronization of rhythms in a mammalian circadian clock

In mammals, the suprachiasmatic nuclei (SCN) of the hypothalamus act as a dominant circadian pacemaker, coordinating rhythms throughout the body and regulating daily and seasonal changes in physiology and behavior. This review focuses on the mechanisms that mediate synchronization of circadian rhythms between SCN neurons. Understanding how these neurons communicate as a network of circadian oscillators has begun to shed light on the adaptability and dysfunction of the brain's master clock.     

Multiple oscillators in the suprachiasmatic nucleus

The suprachiasmatic nucleus (SCN) of the hypothalamus is the site of the pacemaker that controls circadian rhythms of a variety of physiological functions. Data strongly indicate the majority of the SCN neurons express self-sustaining oscillations that can be detected as rhythms in the spontaneous firing of individual neurons. The period of single SCN neurons in a dissociated cell culture is dispersed in a wide range (from 20h to 28h in rats), but that of the locomotor rhythm is close to 24h, suggesting individual oscillators are coupled to generate an averaged circadian period in the nucleus. Electrical coupling via gap junctions, glial regulation, calcium spikes, ephaptic interactions, extracellular ion flux, and diffusible substances have been discussed as possible mechanisms that mediate the interneuronal rhythm synchrony. Recently, GABA (γ-aminobutyric acid), a major neurotransmitter in the SCN, was reported to regulate cellular communication and to synchronize rhythms through GABA_A receptors. At present, subsequent intracellular processes that are able to reset the genetic loop of oscillations are unknown. There may be diverse mechanisms for integrating the multiple circadian oscillators in the SCN. This article reviews the knowledge about the various circadian oscillations intrinsic to the SCN, with particular focus on the intercellular signaling of coupled oscillators. (Chronobiology International, 18(3), 371-387, 2001)     

Cellular communication and coupling within the suprachiasmatic nucleus

In mammals, the part of the nervous system responsible for most circadian behavior can be localized to a pair of structures in the hypothalamus known as the suprachiasmatic nucleus (SCN). Importantly, when SCN neurons are removed from the organism and maintained in a brain slice preparation, they continue to generate 24h rhythms in electrical activity, secretion, and gene expression. Previous studies suggest that the basic mechanism responsible for the generation of these rhythms is intrinsic to individual cells in the SCN. If we assume that individual cells in the SCN are competent circadian oscillators, it is obviously important to understand how these cells communicate and remain synchronized with each other. Cell-to-cell communication is clearly necessary for conveying inputs to and outputs from the SCN and may be involved in ensuring the high precision of the observed rhythm. In addition, there is a growing body of evidence that a number of systems-level phenomena could be dependent on the cellular communication between circadian pacemaker neurons. It is not yet known how this cellular synchronization occurs, but it is likely that more than one of the already proposed mechanisms is utilized. The purpose of this review is to summarize briefly the possible mechanisms by which the oscillatory cells in the SCN communicate with each other. (Chronobiology International, 18(4)579-600, 2001)     

MULTIPLE OSCILLATORS IN THE SUPRACHIASMATIC NUCLEUS

The suprachiasmatic nucleus (SCN) of the hypothalamus is the site of the pacemaker that controls circadian rhythms of a variety of physiological functions. Data strongly indicate the majority of the SCN neurons express self-sustaining oscillations that can be detected as rhythms in the spontaneous firing of individual neurons. The period of single SCN neurons in a dissociated cell culture is dispersed in a wide range (from 20h to 28h in rats), but that of the locomotor rhythm is close to 24h, suggesting individual oscillators are coupled to generate an averaged circadian period in the nucleus. Electrical coupling via gap junctions, glial regulation, calcium spikes, ephaptic interactions, extracellular ion flux, and diffusible substances have been discussed as possible mechanisms that mediate the interneuronal rhythm synchrony. Recently, GABA (γ-aminobutyric acid), a major neurotransmitter in the SCN, was reported to regulate cellular communication and to synchronize rhythms through GABA_A receptors. At present, subsequent intracellular processes that are able to reset the genetic loop of oscillations are unknown. There may be diverse mechanisms for integrating the multiple circadian oscillators in the SCN. This article reviews the knowledge about the various circadian oscillations intrinsic to the SCN, with particular focus on the intercellular signaling of coupled oscillators. (Chronobiology International, 18(3), 371–387, 2001)     

CELLULAR COMMUNICATION AND COUPLING WITHIN THE SUPRACHIASMATIC NUCLEUS

In mammals, the part of the nervous system responsible for most circadian behavior can be localized to a pair of structures in the hypothalamus known as the suprachiasmatic nucleus (SCN). Importantly, when SCN neurons are removed from the organism and maintained in a brain slice preparation, they continue to generate 24h rhythms in electrical activity, secretion, and gene expression. Previous studies suggest that the basic mechanism responsible for the generation of these rhythms is intrinsic to individual cells in the SCN. If we assume that individual cells in the SCN are competent circadian oscillators, it is obviously important to understand how these cells communicate and remain synchronized with each other. Cell-to-cell communication is clearly necessary for conveying inputs to and outputs from the SCN and may be involved in ensuring the high precision of the observed rhythm. In addition, there is a growing body of evidence that a number of systems-level phenomena could be dependent on the cellular communication between circadian pacemaker neurons. It is not yet known how this cellular synchronization occurs, but it is likely that more than one of the already proposed mechanisms is utilized. The purpose of this review is to summarize briefly the possible mechanisms by which the oscillatory cells in the SCN communicate with each other. (Chronobiology International, 18(4)579–600, 2001)     

Simulation of circadian rhythm generation in the suprachiasmatic nucleus with locally coupled self-sustained oscillators

In mammals, circadian rhythms are driven by a pacemaker located in the suprachiasmatic nuclei (SCN) of the anterior hypothalamus. The firing rate of neurons within the SCN exhibits a circadian rhythm. There is evidence that individual neurons within the SCN act as circadian oscillators. Rhythm generation in the SCN was therefore modeled by a system of self-sustained oscillators. The model is composed of up to 10000 oscillatory elements arranged in a square array. Each oscillator has its own (randomly determined) intrinsic period reflecting the widely dispersed periods observed in the SCN. The model behavior was investigated mainly in the absence of synchronizing zeitgebers. Due to local coupling the oscillators synchronized and an overall rhythm emerged. This indicates that a locally coupled system is capable of integrating the output of individual clock cells with widely dispersed periods. The period of the global output (average of all oscillators) corresponded to the average of the intrinsic periods and was stable even for small amplitudes and during transients. Noise, reflecting biological fluctuations at the cellular level, distorted the global rhythm in small arrays. The period of the rhythm could be stabilized by increasing the array size, which thus increased the robustness against noise. Since different regions of the SCN have separate output pathways, the array of oscillators was subdivided into four quadrants. Sudden deviations of periodicity sometimes appeared in one quadrant, while the periods of the other quadrants were largely unaffected. This result could represent a model for splitting, which has been observed in animal experiments. In summary, the multi-oscillator model of the SCN showed a broad repertoire of dynamic patterns, revealed a stable period (even during transients) with robustness against noise, and was able to account for such a complex physiological behavior as splitting.     

A GABAergic mechanism is necessary for coupling dissociable ventral and dorsal regional oscillators within the circadian clock

BACKGROUND: Circadian rhythms in mammalian behavior, physiology, and biochemistry are controlled by the central clock of the suprachiasmatic nucleus (SCN). The clock is synchronized to environmental light-dark cycles via the retino-hypothalamic tract, which terminates predominantly in the ventral SCN of the rat. In order to understand synchronization of the clock to the external light-dark cycle, we performed ex vivo recordings of spontaneous impulse activity in SCN slices of the rat. RESULTS: We observed bimodal patterns of spontaneous impulse activity in the dorsal and ventral SCN after a 6 hr delay of the light schedule. Bisection of the SCN slice revealed a separate fast-resetting oscillator in the ventral SCN and a distinct slow-resetting oscillator in the dorsal SCN. Continuous application of the GABA(A) antagonist bicuculline yielded similar results as cut slices. Short application of bicuculline at different phases of the circadian cycle increased the electrical discharge rate in the ventral SCN but, unexpectedly, decreased activity in the dorsal SCN. CONCLUSIONS: GABA transmits phase information between the ventral and dorsal SCN oscillators. GABA can act excitatory in the dorsal SCN and inhibits neurons in the ventral SCN. We hypothesize that this difference results in asymmetrical interregional coupling within the SCN, with a stronger phase-shifting effect of the ventral on the dorsal SCN than vice versa. A model is proposed that focuses on this asymmetry and on the role of GABA in phase regulation.     

Circadian gene expression in individual fibroblasts: cell-autonomous and self-sustained oscillators pass time to daughter cells

The mammalian circadian timing system is composed of a central pacemaker in the suprachiasmatic nucleus (SCN) of the brain and subsidiary oscillators in most peripheral cell types. While oscillators in SCN neurons are known to function in a self-sustained fashion, peripheral oscillators have been thought to damp rapidly when disconnected from the control exerted by the SCN. Using two reporter systems, we monitored circadian gene expression in NIH3T3 mouse fibroblasts in real time and in individual cells. In conjunction with mathematical modeling and cell co-culture experiments, these data demonstrated that in vitro cultured fibroblasts harbor self-sustained and cell-autonomous circadian clocks similar to those operative in SCN neurons. Circadian gene expression in fibroblasts continues during cell division, and our experiments unveiled unexpected interactions between the circadian clock and the cell division clock. Specifically, the circadian oscillator gates cytokinesis to defined time windows, and mitosis elicits phase shifts in circadian cycles.     

Gates and oscillators: a network model of the brain clock

The suprachiasmatic nuclei (SCN) control circadian oscillations of physiology and behavior. Measurements of electrical activity and of gene expression indicate that these heterogeneous structures are composed of both rhythmic and nonrhythmic cells. A fundamental question with regard to the organization of the circadian system is how the SCN achieve a coherent output while their constituent independent cellular oscillators express a wide range of periods. Previously, the consensus output of individual oscillators had been attributed to coupling among cells. The authors propose a model that incorporates nonrhythmic "gate" cells and rhythmic oscillator cells with a wide range of periods, that neither requires nor excludes a role for interoscillator coupling. The gate provides daily input to oscillator cells and is in turn regulated (directly or indirectly) by the oscillator cells. In the authors' model, individual oscillators with initial random phases are able to self-assemble so as to maintain cohesive rhythmic output. In this view, SCN circuits are important for self-sustained oscillation, and their network properties distinguish these nuclei from other tissues that rhythmically express clock genes. The model explains how individual SCN cells oscillate independently and yet work together to produce a coherent rhythm.     

Forced desynchronization of dual circadian oscillators within the rat suprachiasmatic nucleus

The circadian clock in the suprachiasmatic nucleus of the hypothalamus (SCN) contains multiple autonomous single-cell circadian oscillators and their basic intracellular oscillatory mechanism is beginning to be identified. Less well understood is how individual SCN cells create an integrated tissue pacemaker that produces a coherent read-out to the rest of the organism. Intercellular coupling mechanisms must coordinate individual cellular periods to generate the averaged, genotype-specific circadian period of whole animals. To noninvasively dissociate this circadian oscillatory network in vivo, we (T.C. and A.D.-N.) have developed an experimental paradigm that exposes animals to exotic light-dark (LD) cycles with periods close to the limits of circadian entrainment. If individual oscillators with different periods are loosely coupled within the network, perhaps some of them would be synchronized to the external cycle while others remain unentrained. In fact, rats exposed to an artificially short 22 hr LD cycle express two stable circadian motor activity rhythms with different period lengths in individual animals. Our analysis of SCN gene expression under such conditions suggests that these two motor activity rhythms reflect the separate activities of two oscillators in the anatomically defined ventrolateral and dorsomedial SCN subdivisions. Our "forced desychronization" protocol has allowed the first stable separation of these two regional oscillators in vivo, correlating their activities to distinct behavioral outputs, and providing a powerful approach for understanding SCN tissue organization and signaling mechanisms in behaving animals.     

Clock genes, oscillators, and cellular networks in the suprachiasmatic nuclei

The mammalian SCN contains a biological clock that drives remarkably precise circadian rhythms in vivo and in vitro. Recent advances have revealed molecular and cellular mechanisms required for the generation of these daily rhythms and their synchronization between SCN neurons and to the environmental light cycle. This review of the evidence for a cell-autonomous circadian pacemaker within specialized neurons of the SCN focuses on 6 genes implicated within the pace making mechanism, an additional 4 genes implicated in pathways from the pacemaker, and the intercellular and intracellular mechanisms that synchronize SCN neurons to each other and to solar time.     

Effects of vasoactive intestinal peptide genotype on circadian gene expression in the suprachiasmatic nucleus and peripheral organs

The neuropeptide vasoactive intestinal polypeptide (VIP) has emerged as a key candidate molecule mediating the synchronization of rhythms in clock gene expression within the suprachiasmatic nucleus (SCN). In addition, neurons expressing VIP are anatomically well positioned to mediate communication between the SCN and peripheral oscillators. In this study, we examined the temporal expression profile of 3 key circadian genes: Per1, Per2 , and Bmal1 in the SCN, the adrenal glands and the liver of mice deficient for the Vip gene (VIP KO), and their wild-type counterparts. We performed these measurements in mice held in a light/dark cycle as well as in constant darkness and found that rhythms in gene expression were greatly attenuated in the VIP-deficient SCN. In the periphery, the impact of the loss of VIP varied with the tissue and gene measured. In the adrenals, rhythms in Per1 were lost in VIP-deficient mice, while in the liver, the most dramatic impact was on the phase of the diurnal expression rhythms. Finally, we examined the effects of the loss of VIP on ex vivo explants of the same central and peripheral oscillators using the PER2::LUC reporter system. The VIP-deficient mice exhibited low amplitude rhythms in the SCN as well as altered phase relationships between the SCN and the peripheral oscillators. Together, these data suggest that VIP is critical for robust rhythms in clock gene expression in the SCN and some peripheral organs and that the absence of this peptide alters both the amplitude of circadian rhythms as well as the phase relationships between the rhythms in the SCN and periphery.     

Photic desynchronization of two subgroups of circadian oscillators in a network model of the suprachiasmatic nucleus with dispersed coupling strengths

The suprachiasmatic nucleus (SCN) is the master circadian clock in mammals and is composed of thousands of neuronal oscillators expressing different intrinsic periods. These oscillators form a coupled network with a free-running period around 24 h in constant darkness and entrainable to the external light-dark cycle (T cycle). Coupling plays an important role in setting the period of the network and its range of entrainment. Experiments in rats have shown that two subgroups of oscillators within the SCN, a ventrolateral (VL) subgroup that receives photic input and a dorsomedial (DM) subgroup that is coupled to VL, can be desynchronized under a short (22-h) T cycle, with VL entrained to the cycle and DM free-running. We use a modified Goodwin model to understand how entrainment of the subgroups to short (22-h) and long (26-h) T cycles is influenced by light intensity, the proportion of neurons that receives photic input, and coupling heterogeneity. We find that the model's critical value for the proportion of photically-sensitive neurons is in accord with actual experimental estimates, while the model's inclusion of dispersed coupling can account for the experimental observation that VL and DM desynchronize more readily under the 22-h than under the 26-h T cycle. Heterogeneous intercellular coupling within the SCN is likely central to the generation of complex behavioral patterns.     

Intercellular coupling confers robustness against mutations in the SCN circadian clock network

Molecular mechanisms of the mammalian circadian clock have been studied primarily by genetic perturbation and behavioral analysis. Here, we used bioluminescence imaging to monitor Per2 gene expression in tissues and cells from clock mutant mice. We discovered that Per1 and Cry1 are required for sustained rhythms in peripheral tissues and cells, and in neurons dissociated from the suprachiasmatic nuclei (SCN). Per2 is also required for sustained rhythms, whereas Cry2 and Per3 deficiencies cause only period length defects. However, oscillator network interactions in the SCN can compensate for Per1 or Cry1 deficiency, preserving sustained rhythmicity in mutant SCN slices and behavior. Thus, behavior does not necessarily reflect cell-autonomous clock phenotypes. Our studies reveal previously unappreciated requirements for Per1, Per2, and Cry1 in sustaining cellular circadian rhythmicity and demonstrate that SCN intercellular coupling is essential not only to synchronize component cellular oscillators but also for robustness against genetic perturbations.