Letteratura scientifica selezionata sul tema "Circadian cycle characterization"

This study reports for the first time the effects of retinoid-related orphan receptors [RORβ; receptor gene deletion RORβ(C3H)−/−] in C3H/HeN mice on behavioral and circadian phenotypes. Pineal melatonin levels showed a robust diurnal rhythm with high levels at night in wild-type (+/+), heterozygous (+/−), and knockout (−/−) mice. The RORβ(C3H)−/− mice displayed motor (“duck gait,” hind paw clasping reflex) and olfactory deficits, and reduced anxiety and learned helplessness-related behaviors. Circadian rhythms of wheel-running activity in all genotypes showed entrainment to the light-dark (LD) cycle, and free running in constant dark, with RORβ(C3H)−/− mice showing a significant increase in circadian period ( tau). Melatonin administration (90 μg/mouse sc for 3 days) at circadian time (CT) 10 induced phase advances, while exposure to a light pulse (300 lux) at CT 14 induced phase delays of circadian activity rhythms of the same magnitude in all genotypes. In RORβ(C3H)−/− mice a light pulse at CT 22 elicited a larger phase advance in activity rhythms and a slower rate of reentrainment after a 6-h advance in the LD cycle compared with (+/+) mice. Yet, the rate of reentrainment was significantly advanced by melatonin administration at the new dark onset in both (+/+) and (−/−) mice. We conclude that the RORβ nuclear receptor is not involved in either the rhythmic production of pineal melatonin or in mediating phase shifts of circadian rhythms by melatonin, but it may regulate clock responses to photic stimuli at certain time domains.

Key indicators of circadian regulation include the persistence of physiological rhythmicity in the absence of environmental time cues and entrainment of this rhythmicity by the ambient light cycle. In some teleosts, the inner segments of rod and cone photoreceptors contract and elongate according to changes in ambient lighting and the circadian cycle. Pigment granules in the retinal pigment epithelium (RPE) disperse and aggregate in a similar manner. Collectively, these movements are known as retinomotor movements. We report the histological characterization of diurnal and circadian retinomotor movements in zebrafish, Danio rerio. Adult fish subjected to a 14:10 light:dark (LD) cycle, constant darkness (DD), or constant light (LL) were sacrificed at 1–13 h intervals and processed for semithin sectioning of the retina. Using bright-field microscopy, 15 measurements of pigment granule position and the inner segment lengths of 30 rods and 30–45 cones were collected from the central third of the dorso-optic retina per time point. In LD, rods and cones followed a clear diurnal rhythm in their inner segment movements. Short-single, UV-sensitive cones were found to contract significantly 1 h before light onset in LD conditions. In DD conditions, the inner segments movements of short-single and double cones displayed statistically significant rhythms. RPE pigment granule movements are rhythmically regulated in both LD and DD although fluctuations are damped in the absence of photic cues. No significant retinomotor movements were observed in LL. These findings indicate retinomotor movements in zebrafish are differentially regulated by an endogenous oscillator and by light-dependent mechanisms.

The human circadian pacemaker entrains to the 24-h day, but interindividual differences in properties of the pacemaker, such as intrinsic period, affect chronotype and mediate responses to challenges to the circadian system, such as shift work and jet lag, and the efficacy of therapeutic interventions such as light therapy. Robust characterization of circadian properties requires desynchronization of the circadian system from the rest-activity cycle, and these forced desynchrony protocols are very time and resource intensive. However, circadian protocols designed to derive the relationship between light intensity and phase shift, which is inherently affected by intrinsic period, may be applied more broadly. To exploit this relationship, we applied a mathematical model of the human circadian pacemaker with a Markov-Chain Monte Carlo parameter estimation algorithm to estimate the representative group intrinsic period for a group of participants using their collective illuminance-response curve data. We first validated this methodology using simulated illuminance-response curve data in which the intrinsic period was known. Over a physiological range of intrinsic periods, this method accurately estimated the representative intrinsic period of the group. We also applied the method to previously published experimental data describing the illuminance-response curve for a group of healthy adult participants. We estimated the study participants’ representative group intrinsic period to be 24.26 and 24.27 h using uniform and normal priors, respectively, consistent with estimates of the average intrinsic period of healthy adults determined using forced desynchrony protocols. Our results establish an approach to estimate a population’s representative intrinsic period from illuminance-response curve data, thereby facilitating the characterization of intrinsic period across a broader range of participant populations than could be studied using forced desynchrony protocols. Future applications of this approach may improve the understanding of demographic differences in the intrinsic circadian period.

Abstract Introduction Misalignment between circadian phase (i.e., timing) and the desired sleep window is associated with sleep disturbances including insomnia and circadian rhythm sleep-wake disorders. Identifying circadian-driven sleep disruption requires assessment of circadian phase, the gold-standard of which is dim light melatonin onset (DLMO). While DLMO is traditionally measured in the lab using saliva or blood sampled over multiple hours, novel methods of estimating DLMO using activity and light data obtained from wearable sensors (i.e., digital DLMO) have been recently validated in samples with both healthy and disordered sleep. Such methods could potentially provide pragmatic, low-burden ways of assessing circadian phase, which in turn could be used to aid diagnostic decisions and to adjust circadian-targeted interventions in real time. However, digital DLMO has yet to be characterized within the broader adult population, which is needed to differentiate between normative and clinically salient values. Therefore, this study will examine digital DLMO in a large, population-based sample, as well as explore the potential clinical utility of digital biomarkers related to circadian phase. Methods This study will be a secondary analysis of data obtained during the ancillary sleep study of the Multi-Ethnic Study of Atherosclerosis (i.e., MESA Sleep). Seven days of activity and light data, measured via wrist-worn actigraphy, were obtained from 2,237 participants. Digital DLMO will be estimated using the extended Kronauer limit-cycle model of the human circadian pacemaker. Phase angles, or the differences in clock time, between digital DLMO and the following will be calculated: self-reported in-bed time; and sleep onset, midpoint, and offset measured by actigraphy. The associations between these phase angles and sleep outcomes (i.e., sleep onset latency, sleep efficiency, and sleep quality) will be explored using linear regression. Covariates will include the age, gender, and race/ethnicity of the participant. Weekend and weekday averages will be evaluated separately. Results Results will include the distribution of digital DLMO in a population-based sample, as well as the associations between digital DLMO phase angles and sleep outcomes. Conclusion This study will inform future research into the clinical potential of digital circadian biomarkers. Support (if any)

Women with primary vasospastic syndrome (VS), but otherwise healthy, exhibit a functional disorder of vascular regulation (main symptom: cold extremities) and often suffer from difficulties initiating sleep (DIS). Diverse studies have shown a close association between distal vasodilatation before lights off and a rapid onset of sleep. Therefore, we hypothesized that DIS in women with VS could be due to a reduced heat loss capacity in the evening, i.e., subjects are physiologically not ready for sleep. The aim of the study was to elucidate whether women having both VS and DIS (WVD) or not (controls) show different circadian characteristics (e.g., phase delay of the circadian thermoregulatory system with respect to the sleep-wake cycle). Healthy young women ( n = 9 WVD and n = 9 control) completed a 40-h constant routine protocol (adjusted to habitual bedtime) before and after an 8-h sleep episode. Skin temperatures [off-line calculated as distal-proximal skin temperature gradient (DPG)] and core body temperature (CBT; rectal) were continuously recorded. Half-hourly saliva samples were collected for melatonin assay and subjective sleepiness was assessed on the Karolinska Sleepiness Scale (KSS). Compared with control, WVD showed no differences in habitual bed times, but a 1-h circadian phase delay of dim light-melatonin onset (hours after lights on: WVD 14.6 ± 0.3 h; control 13.5 ± 0.2 h; P = 0.01). Similar phase shifts were observed in CBT, DPG, and KSS ratings. In conclusion, WVD exhibit a phase delay of the endogenous circadian system with respect to their habitual sleep-wake cycle, which could be a cause of DIS.

In humans, the connection between sleep and mood has long been recognized, although direct molecular evidence is lacking. We identified two rare variants in the circadian clock gene PERIOD3 (PER3-P415A/H417R) in humans with familial advanced sleep phase accompanied by higher Beck Depression Inventory and seasonality scores. hPER3-P415A/H417R transgenic mice showed an altered circadian period under constant light and exhibited phase shifts of the sleep-wake cycle in a short light period (photoperiod) paradigm. Molecular characterization revealed that the rare variants destabilized PER3 and failed to stabilize PERIOD1/2 proteins, which play critical roles in circadian timing. Although hPER3-P415A/H417R-Tg mice showed a mild depression-like phenotype, Per3 knockout mice demonstrated consistent depression-like behavior, particularly when studied under a short photoperiod, supporting a possible role for PER3 in mood regulation. These findings suggest that PER3 may be a nexus for sleep and mood regulation while fine-tuning these processes to adapt to seasonal changes.

ABSTRACT Genes showing differential expression related to the early G1 phase of the cell cycle during synchronized circadian growth of the toxic dinoflagellate Alexandrium fundyensewere identified and characterized by differential display (DD). The determination in our previous work that toxin production inAlexandrium is relegated to a narrow time frame in early G1 led to the hypothesis that transcriptionally up- or downregulated genes during this subphase of the cell cycle might be related to toxin biosynthesis. Three genes, encodingS-adenosylhomocysteine hydrolase (Sahh), methionine aminopeptidase (Map), and a histone-like protein (HAf), were isolated. Sahh was downregulated, while Map and HAf were upregulated, during the early G1 phase of the cell cycle. Sahh andMap encoded amino acid sequences with about 90 and 70% similarity to those encoded by several eukaryotic and prokaryoticSahh and Map genes, respectively. The partialMap sequence also contained three cobalt binding motifs characteristic of all Map genes. HAf encoded an amino acid sequence with 60% similarity to those of two histone-like proteins from the dinoflagellate Crypthecodinium cohniiBiecheler. This study documents the potential of applying DD to the identification of genes that are related to physiological processes or cell cycle events in phytoplankton under conditions where small sample volumes represent an experimental constraint. The identification of an additional 21 genes with various cell cycle-related DD patterns also provides evidence for the importance of pretranslational or transcriptional regulation in dinoflagellates, contrary to previous reports suggesting the possibility that translational mechanisms are the primary means of circadian regulation in this group of organisms.

Dynamic studies in time course experimental designs and clinical approaches have been widely used by the biomedical community. These applications are particularly relevant in stimuli-response models under environmental conditions, characterization of gradient biological processes in developmental biology, identification of therapeutic effects in clinical trials, disease progressive models, cell-cycle, and circadian periodicity. Despite their feasibility and popularity, sophisticated dynamic methods that are well validated in large-scale comparative studies, in terms of statistical and computational rigor, are less benchmarked, comparing to their static counterparts. To date, a number of novel methods in bulk RNA-Seq data have been developed for the various time-dependent stimuli, circadian rhythms, cell-lineage in differentiation, and disease progression. Here, we comprehensively review a key set of representative dynamic strategies and discuss current issues associated with the detection of dynamically changing genes. We also provide recommendations for future directions for studying non-periodical, periodical time course data, and meta-dynamic datasets.

Complex brain functions, including learning and memory, arise in part from the modulatory role of astrocytes on neuronal circuits. Functionally, the dentate gyrus (DG) exhibits differences in the acquisition of long-term potentiation (LTP) between day and night. We hypothesize that the dynamic nature of astrocyte morphology plays an important role in the functional circuitry of hippocampal learning and memory, specifically in the DG. Standard microscopy techniques, such as differential interference contrast (DIC), present insufficient contrast for detecting changes in astrocyte structure and function and are unable to inform on the intrinsic structure of the sample in a quantitative manner. Recently, gradient light interference microscopy (GLIM) has been developed to upgrade a DIC microscope with quantitative capabilities such as single-cell dry mass and volume characterization. Here, we present a methodology for combining GLIM and electrophysiology to quantify the astrocyte morphological behavior over the day-night cycle. Colocalized measurements of GLIM and fluorescence allowed us to quantify the dry masses and volumes of hundreds of astrocytes. Our results indicate that, on average, there is a 25% cell volume reduction during the nocturnal cycle. Remarkably, this cell volume change takes place at constant dry mass, which suggests that the volume regulation occurs primarily through aqueous medium exchange with the environment.

The identification and characterization of rhythmically expressed mRNAs have been an active area of research over the past 20 years, as these mRNAs are believed to produce the daily rhythms in a wide range of biological processes. Circadian transcriptome studies have used mature mRNA as a primary readout and focused largely on rhythmic RNA synthesis as a regulatory mechanism underlying rhythmic mRNA expression. However, RNA synthesis, RNA degradation, or a combination of both must be rhythmic to drive rhythmic RNA profiles, and it is still unclear to what extent rhythmic synthesis leads to rhythmic RNA profiles. In addition, circadian RNA expression is also often tissue specific. Although a handful of genes cycle in all or most tissues, others are rhythmic only in certain tissues, even though the same core clock mechanism is believed to control the rhythmic RNA profiles in all tissues. This review focuses on the dynamics of rhythmic RNA synthesis and degradation and discusses how these steps collectively determine the rhythmicity, phase, and amplitude of RNA accumulation. In particular, we highlight a possible role of RNA degradation in driving tissue-specific RNA rhythms. By unifying findings from experimental and theoretical studies, we will provide a comprehensive overview of how rhythmic gene expression can be achieved and how each regulatory step contributes to tissue-specific circadian transcriptome output in mammals.

La función circadiana es esencial para el crecimiento y adaptación de las plantas a su entorno. La maquinaria molecular responsable de la generación de ritmos circadianos está basada en la expresión rítmica de genes cuyo pico de expresión oscila en diferentes fases durante el día y la noche. Los ritmos de expresión génica se traducen en oscilaciones de procesos fisiológicos y de desarrollo. El crecimiento de las plantas está regulado por una plétora de procesos que en última instancia operan a través del control de la proliferación y diferenciación celular. La proliferación celular depende de la progresión del ciclo mitótico, el cual está dividido en 4 fases: S (Síntesis del ADN), M (Mitosis) y de las interfases G1 y G2 (en inglés Gap 1 y 2) que ocurren antes de las fases S y M respectivamente. El proceso de diferenciación celular coincide con el cambio al endociclo, una variante del ciclo mitótico en la que el ADN genómico se duplica pero sin posterior división, es decir en ausencia de fase M. Aunque la regulación circadiana y el ciclo celular han sido individualmente estudiados en plantas, no se ha demostrado hasta la fecha la posible conexión de ambos ciclos en plantas. El trabajo realizado durante esta Tesis Doctoral se ha centrado en el estudio del papel del reloj circadiano en el control del ciclo celular durante la regulación del crecimiento de la planta. Los resultados obtenidos muestran que plantas con un reloj circadiano de ritmo lento desaceleran la progresión del ciclo celular, mientras que un reloj de ritmo rápido lo acelera. El componente esencial del reloj denominado en inglés TIMING OF CAB EXPRESSION 1 (TOC1) controla la transición de la fase G1 a la fase S, regulando así el ritmo del ciclo mitótico durante los estadios tempranos del desarrollo foliar. Asimismo, TOC1 también controla la ploidía somática característica del endociclo durante estadios tardíos del desarrollo foliar y en las células del hipocotilo. Utilizando técnicas de citometría de flujo y parámetros de cinéticas de crecimiento foliar se pudo determinar que en plantas que sobre-expresan TOC1 la fase S es más corta, lo que se correlaciona con la represión diurna del gen CELL DIVISION CONTROL 6 (CDC6). Este gen codifica un factor esencial en la formación de los complejos de pre-replicación que determinan los orígenes de replicación del ADN. Mediante técnicas de inmunoprecipitación de cromatina encontramos que la represión de CDC6 ocurre a través de la unión directa de TOC1 al promotor de CDC6. Los análisis de interacción genética demostraron que los fenotipos de crecimiento reducido y de ploidía somática alterada observados en plantas que sobre-expresan TOC1, quedaban revertidos al sobre-expresarse también CDC6. Estos resultados confirman que la función de TOC1 en el ciclo celular ocurre en gran medida a través de la represión de CDC6. La desaceleración de la progresión del ciclo celular en plantas que sobre-expresan TOC1 afecta no solo el desarrollo de los órganos de la planta, sino también el desarrollo tumoral en los tallos de las inflorescencias. Por lo tanto, nuestros estudios demuestran que la función de TOC1 es importante en la regulación rítmica de la maquinaria pre-replicativa del ADN para controlar el crecimiento de las plantas en resonancia con el medio ambiente.
The circadian function is essential for plant growth and its adaptation to the environment. The molecular machinery responsible for the establishment of the circadian rhythmicity relies on the rhythmic oscillation of differentially expressed genes with different peaks of expression along the day and night. The rhythms in gene expression are translated into oscillations of physiological and developmental processes. Plant growth is controlled by a plethora of different processes that ultimately work through the control of cell proliferation and differentiation. Cell proliferation relies on the proper progression of the mitotic cycle, which is divided in 4 phases: S (DNA synthesis), M (Mitosis) and two gap phases G1 and G2, that take place before S and M phases, respectively. Cell differentiation coincides with the entry into the endocycle, a variant of the mitotic cycle in which genomic DNA duplicates without further division or mitosis. Even though the circadian clock and cell cycle as separate pathways have been well documented in plants, the possible direct interplay between these two cyclic processes has not been previously addressed. The work performed during this Thesis has focused on the characterization of the role of the circadian clock in the control of the cell cycle during plant growth. We found that plants with slower than Wild-Type circadian clocks slow down the progression of the cell cycle, while plants with faster clocks speed it up. The core clock component TIMING OF CAB EXPRESSION 1 (TOC1) controls the G1 to S-phase transition, thereby regulating the rhythm of the mitotic cycle during the early stages of leaf development. Likewise, TOC1 controls somatic ploidy during later stages of leaf development and of hypocotyl cell elongation. The use of flow cytometry analyses and of leaf growth kinetics showed that in plants over-expressing TOC1, the S-phase is shorter, which correlates with the diurnal repression of the CELL DIVISION CONTROL 6 (CDC6) gene. This gene encodes an essential component of the pre-replication complex, which is responsible for the specification of DNA origins of replication. Chromatin immunoprecipitation assays showed that the diurnal repression of CDC6 most likely relies on the direct binding of TOC1 to the CDC6 promoter. Genetic interaction analyses showeed that the reduced growth and altered somatic ploidy phenotypes observed in plants over-expressing TOC1 were reverted when CDC6 was over-expressed. Thus, our results confirm that TOC1 regulation of the cell cycle occurs through CDC6 repression. The slow cell cycle progression in plants over-expressing TOC1 has an impact not only in organ development but also on tumor growth in stems and inflorescences. Thus, TOC1 sets the time of the DNA pre-replicative machinery to control plant growth in resonance with the environment.

Les horloges circadiennes, présentes dans les cellules de pratiquement tous les êtres vivants, sont essentielles dans la régulation rythmique de nombreux processus biologiques. Le bon fonctionnement des organismes dépend de la cohérence de phase de ces oscillateurs génétiques. Pourtant, chez les mammifères, les mécanismes sous-jacents à la synchronisation des horloges périphériques demeurent méconnus. Cette thèse se concentre sur l'étude de la synchronisation des horloges circadiennes périphériques des mammifères et sur l'analyse de la dynamique du cycle circadien. Dans un premier temps, nous supposons que les horloges périphériques sont capables d'assurer leur synchronisation grâce à des mécanismes de couplage, comparables à ceux observés entre les cellules de l'horloge centrale. Nous étudions cette hypothèse numériquement, à l'aide d'un modèle d'un réseau d'horloges périphériques couplées, construit avec des équations différentielles ordinaires. Nos simulations nous permettent d'identifier des facteurs favorisant la synchronisation des oscillateurs circadiens. Dans un second temps, nous nous focalisons sur la dynamique d'un unique cycle circadien, que nous caractérisons théoriquement via la construction d'un modèle affine par morceaux approximant un modèle continu construit avec des termes d'action de masse. Notre approche repose sur l'identification d'une séquence de transitions périodique entre les régions de l'espace de phases discrétisé du modèle continu et sur le développement d'un algorithme calculant des valeurs réelles de seuils qui garantissent une trajectoire périodique aux oscillateurs du modèle affine par morceaux et la reproduction des principales propriétés qualitatives des cycles circadiens. Nous proposons ensuite une méthode générale et automatisée pour caractériser la dynamique de n'importe quel cycle circadien dont les séries temporelles des (complexes de) protéines CLOCK:BMAL1, REV-ERB et PER:CRY sont connues. Notre méthode sert d'outil pour tester et comparer les dynamiques de différents cycles circadiens, tout en mettant en évidence des propriétés qu'ils partagent. Ces méthodes nous permettent finalement de mieux comprendre l'influence du couplage sur la dynamique des cycles d'un réseau d'horloges périphériques
Circadian clocks, present in the cells of virtually all living beings, are essential for the rhythmic regulation of many biological processes. The healthy functioning of organisms depends on the phase coherence of these genetic oscillators. However, in mammals, the mechanisms underlying the synchronization of peripheral clocks remain poorly understood. This thesis focuses on the study of the synchronization of mammalian peripheral circadian clocks and on the analysis of circadian cycle dynamics.First, we hypothesize that peripheral clocks can achieve synchronization through coupling mechanisms, comparable to those observed between central clock cells. We investigate this hypothesis numerically, using a model of a network of coupled peripheral clocks, constructed with ordinary differential equations. Our simulations lead to the identification of factors promoting the synchronization of circadian oscillators. Secondly, we focus on the dynamics of a single circadian cycle, which we characterize theoretically through the construction of a piecewise affine model approximating a continuous model including mass action terms. Our approach is based on the identification of a sequence of periodic transitions between regions of the discretized phase space of the continuous model, and on the development of an algorithm generating real threshold values that guarantee a periodic trajectory for the oscillators of the piecewise affine model and the reproduction of the main qualitative properties of circadian cycles. We then propose a general and automated method for characterizing the behaviour of any circadian cycle whose time series of CLOCK:BMAL1, REV-ERB and PER:CRY protein (complexes) are known. Our method provides a benchmark for testing and comparing the dynamics of different circadian cycles, while highlighting properties they share. Finally, these methods allow us to better understand the influence of coupling on the cycle dynamics of a network of peripheral clocks