Laboratory of Biological Rhythms

Laboratory of Biological Rhythms - bg 3

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Laboratory of Biological Rhythms - 24 uvodni

About the Laboratory

Please, consider filing out anonymous questionnaire regarding your chronotype and social jet lag.

In the center of our interest is the endogenous time-keeping system, circadian clock of mammals, including humans. The system temporally regulates rhythmic processes in our body so that they take place at the proper time of day and are optimally synchronized relative to each other.

A failure of the temporal regulation has a negative impact on human health. Using in vivo and in vitro models, we study the molecular mechanisms underlying the circadian clock as well as the resulting physiological and behavioral rhythms.

We focus on the following questions:

  • What are the mechanisms for entraining (synchronizing) the circadian system and what are the consequences of its failure?
  • How does the circadian system develop and how is it being entrained during ontogeny?
  • How is the circadian system of humans entrained in real life conditions, both in health and in disease?
Laboratory of Biological Rhythms - scn e17 and p1 ms

Fig. PER2 protein in the prenatal and early postnatal SCN, ex vivo luminescence microscopy.

Achievements

Martin Sládek, Pavel Houdek, Jihwan Myung, Kateryna Semenovykh, Tereza Dočkal, Alena Sumová. Fluids Barriers CNS. 2024 May 27;21(1):46. doi: 10.1186/s12987-024-00547-3.

Choroid plexus (ChP), the brain structure primarily responsible for cerebrospinal fluid production, contains a robust circadian clock with unknown function. We used genetic mouse models (WT, mPer2Luc, ChP-specific Bmal1 knockout) combined with surgical lesion of the SCN (SCNx), time-resolved transcriptomics, and single cell luminescence microscopy. We found that the ChP clock regulates cerebrospinal fluid circadian secretome, precisely times endoplasmic reticulum stress response, and controls genes involved in neurodegenerative diseases. In ChP of SCNx mice, the rhythms were severely dampened to a comparable extent as in mice with ChP-specific Bmal1 knockout; they were restored by daily injections of dexamethasone. Our data demonstrate that the ChP clock controls tissue-specific gene expression and is strongly dependent on the presence of a functional connection with the SCN. The results may contribute to the search for a novel link between ChP clock disruption and impaired brain health.


Laboratory of Biological Rhythms - fig

Fig: A. Workflow. B. Period of locomotor activity in Control and SCNx mice. C. SCN after sham surgery and after lesion. D. Rhythms in SCNx ChP are dampened to similar extent as rhythms in ChP from tissues-specific Bmal1-KO. E. Rhythmic genes in Control (left) or SCNx (right) ChP samples. F. Phase histogram of genes Control (left) or SCNx (right) ChP samples. G. PER2::LUCIFERASE rhythms in single-cell sized ROIs across an explanted ChP from either Control or SCNx mice.

Milica Drapšin, Tereza Dočkal, Pavel Houdek, Martin Sládek, Kateryna Semenovykh, Alena Sumová. Brain Behav Immun. 2024 Jan 25:S0889-1591(24)00229-0. doi: 10.1016/j.bbi.2024.01.217.

Choroid plexus (ChP) in brain ventricles plays significant role in brain homeostasis. ChP cells contain robust circadian oscillators that control its function. Chronodisruption in the form of exposure to constant light (cLL) and chronic advance of the light-dark cycle (cLD-shifts) impaired the ChP clock due to disrupted signals originating from the SCN involving glucocorticoid signaling. Sleep patterns and immune markers in the brain were also affected. Interestingly, LPS systemic inflammation did not influence the ChP clock, but disrupted the liver clock. The data highlight the role of ChP in protecting the brain from neuroinflammation. Our results provide a novel link between human lifestyle-induced chronodisruption and the impairment of ChP-dependent brain homeostasis.

Laboratory of Biological Rhythms - bbi figv2

Fig. Choroid plexus (ChP) chronodisruption. (A) Total locomotor activity, (B) relative amplitude, and (C) sleep duration of mice reared on

normal light-dark regime (LD12:12), on a chronic phase-shifted light-dark (cLD-shifts) cycle regime, or on a constant light (cLL) regime.

(D-E) 6-day luminescence microscopy of the circadian expression of PER2 protein imaged in ChP explants of analyzed mice,

quantitative analysis of amplitude (F) and period (G) of individual ChP cells. (I) Multiplex analysis of cytokines in plasma of analyzed mice.

Martin Sládek, Jan Klusáček, Dana Hamplová and Alena Sumová. Sleep 2023 Feb 24, 10.1093/sleep/zsad037. Social jetlag, or chronic misalignment between biological and social time, is the most common disturbance of circadian rhythms in modern society.

It remains unclear, however, whether social jetlag has any negative health effects. By investigating a unique population-representative dataset, including 1957 blood samples analyzed for nine biomarkers, we report significant associations between sleep phase preference, social jetlag and cardio-metabolic biomarkers. We identify novel factors affecting social jetlag and demonstrate that flexible working schedule efficiently alleviates social jetlag.

Laboratory of Biological Rhythms - 1 graphical abstract sleep

Karolína Liška , Tereza Dočkal , Pavel Houdek , Martin Sládek, Vendula Lužná , Kateryna Semenovykh , Milica Drapšin and Alena Sumová.

Lithium affects the circadian clock in the choroid plexus – A new role for an old mechanism. Biomedicine & Pharmacotherapy 159 (2023), 114292

Lithium is an effective mood stabilizer used in the treatment of Bipolar Disorder (BD), but the mechanism of its therapeutic action is not well understood. We investigated the effect of lithium on the circadian clock located in the ventricle barrier complex containing the choroid plexus (CP), a part of the glymphatic system that influences gross brain function via the production of cerebrospinal fluid. Using in vivo and ex vivo methods, analyzing clock gene expression and probing cell-signaling pathways of the CP, we demonstrated that lithium strongly affected the circadian clock in the CP. The effect of lithium was not mediated by GSK3b-exclusive mechanism, but rather by modulation of PKC activity. The effects on the CP clock may be involved in the therapeutic effects of lithium and improve brain function in BD patients by aligning the function of the CP clock-related glymphatic system with the sleep-wake cycle. Importantly, our data argue for personalized timing of lithium treatment in BD patients.

Laboratory of Biological Rhythms - lithium

Fig: Lithium affects the circadian clock in the choroid plexus independently of GSK3, resulting in time-dependent phase shifts that may affect CSF production, brain function and its therapeutic outcome.

Laboratory of Biological Rhythms - pditc1r27853eyy05pci
Fig. Maternal signals drive rhythmicity in the fetal suprachiasmatic nuclei before their molecular clock develops.

doi:10.1371/journal.pbio.3001637.g005. Our results indicate the importance of a well-functioning maternal biological clock in providing rhythmic environment during the fetal brain development.

 

During development in of the fetal suprachiasmatic nuclei, maternal stimuli may substitute for an absent inter-cellular web of synapses and drive cell-population rhythms before the SCN clock fully matures. Distinct maternal signals rhythmically control a variety of neuronal processes in the fetal rat suprachiasmatic nuclei before they begin to operate as the central circadian clock.

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Fig: Feeding aligned with activity status (as occurs in rats under ad libitum conditions) is associated with coupling between the peaks in insulin and corticosterone levels and the phases of the clocks in the endocrine and exocrine parts of the pancreas.
doi:10.1007/s00018-022-04354-7. Misaligned meals become a frequent lifestyle transgression related to current societal pressure imposed on behavior.
 
Here we provide the first evidence in rats and mice that whereas the clock in the endocrine pancreas shifts according to mealtime, the clock in the exocrine pancreas can be severely impaired by incorrect meal timing. Our findings pave a new avenue for future investigation of the relationship between long-term circadian misalignment and a higher risk of development of metabolic disorders.

doi:10.1016/j.neuropharm.2021.108455. The paper provides new insights into the modulatory role of endocannabinoid signaling during the light entrainment of the SCN.

Suprachiasmatic nucleus (SCN) of the hypothalamus is the master clock that drives circadian rhythms in physiology and behavior and adjusts their timing to external cues. Neurotransmitter glutamate and glutamatergic receptors sensitive to N-methyl-D-aspartate (NMDA) play a dual role in the SCN by coupling astrocytic and neuronal single cell oscillators and by resetting their phase in response to light. Recent reports suggested that signaling by endogenous cannabinoids (ECs) participates in both of these functions. ECs, such as 2-arachidonoylglycerol (2-AG), act via presynaptic CB1 receptors to modulate synaptic activity and affect the SCN response to light-mimicking NMDA stimulus in a time-dependent manner. We demonstrate that circadian clock in the rat SCN regulates expression of 2-AG transport, synthesis and degradation enzymes as well as its receptors. Inhibition of the major 2-AG synthesis enzyme, diacylglycerol lipase, enhances the phase delay and lowers the amplitude of explanted SCN rhythm in response to NMDAR activation. Using PER2 bioluminescence imaging, we show how single cell oscillators reflect the phase response of the whole SCN. Additionally, we present strong evidence that the zero amplitude behavior of the SCN in response to single NMDA stimulus in the middle of subjective night is the result of a loss of rhythm in individual SCN cells. The paper provides new insights into the modulatory role of endocannabinoid signaling during the light entrainment of the SCN.

Laboratory of Biological Rhythms - rhythmic Laboratory of Biological Rhythms - e8 scn1

Fig 1. Explanted SCN from mPer2Luc mouse oscillating in vitro was treated by diacylglycerol lipase inhibitor and NMDA during daytime (left) or in the middle of subjective nighttime (right).

Laboratory of Biological Rhythms - single cell

Fig 2. PER2 levels in a single representative neuron from the explants shown in Fig. 2. Red line shows the signal before treatment.

 

 

Polidarová, L., Sládek, M., Nováková, M., Parkanová, D. & Sumová, A. Increased Sensitivity of the Circadian System to Temporal Changes in the Feeding Regime of Spontaneously Hypertensive Rats – A Potential Role for Bmal2 in the Liver. PLoS One 8, e75690 (2013).

The spontaneously hypertensive rat (SHR) is an inbred laboratory model with a number of metabolic disorders. In the previous work, we showed that SHR circadian system is disrupted, resulting in phase advancing clock and desynchrony of peripheral organs. In the current paper, we describe the increased sensitivity of the circadian system of the SHR to the time-restricted feeding regime and its association with Bmal2 gene activity in the liver.

Laboratory of Biological Rhythms - polidarova1

Daily rhythm of BMAL2 protein in the liver is phase advanced in SHR compared to Wistar (A, B). Locomotor activity, just before the food is accessible (C), is significantly higher in SHR.

Nováková M, Sládek M, Sumová A. (2013): Human chronotype is determined in bodily cells under real-life conditions. Chronobiol Int. 30(4):607-17.

http://informahealthcare.com/doi/abs/10.3109/07420528.2012.754455

The chronotype is influenced both genetically and by the environment, mainly by the light. It is dependent on the central clock located in the hypothalamus. The chronotype affects e.g. the individual tolerance to shift work. Therefore, it is useful to know the individual chronotype to assign the best working schedule. The chronotype can be determined from the timing of sleep and the secretion of melatonin which are controlled by the central clock in the brain. In our study, we found that the individual chronotype also determines the timing of clock gene expression in peripheral clocks in cells acquired by scrubbing the mucosa of the inner cheek. Our study showed that the analysis of clock gene expression in peripheral cells can be used as a novel marker of the phase of the inner clock. This method can be used also in real-life field conditions.

Laboratory of Biological Rhythms - novakova2

Phase advanced melatonin secretion in early chronotypes (“larks“) as compared to late chronotypes (“owls“) (Nováková et al., Chronobiology International, 2013)

Laboratory of Biological Rhythms - novakova2a

Although individuals are exposed to the same lighting conditions, they differ in preferred sleep/wake schedule.

Sládek, M. & Sumová, A. Entrainment of Spontaneously Hypertensive Rat Fibroblasts by Temperature Cycles. PLoS One 8, e77010 (2013).

Regular daily changes in body temperature are one of the signals that entrain the circadian clock in peripheral organs. In this paper, we used isolated cell cultures from Wistar rats and SHR transfected with circadian luminescent reporter to investigate the sensitivity of the peripheral clock to the phase setting through an external 2.5 ° C temperature cycle. We have shown that a mere two-day temperature cycle is sufficient to restore dampened circadian rhythms and we constructed a curve of phase response to the temperature cycle.

Laboratory of Biological Rhythms - sladek1

The phase change after the temperature cycle depends on the phase of the clock at the beginning of the temperature cycle.

Nováková MNevšímalová SPříhodová ISládek MSumová A. (2012): Alteration of the circadian clock in children with Smith-Magenis syndrome. J Clin Endocrinol Metab. 97(2):E312-8. Smith-Magenis Syndrome (SMS) is a rare genetic disorder. Patients with SMS are mentally retarded, not able to control their emotions, they often have hearing and speech impairment and lowered sensitivity to pain. Moreover, they suffer from severe sleep disturbances and have disrupted pattern in melatonin secretion. In healthy individuals, melatonin secretion from the pineal gland is strictly driven by the central clock in the hypothalamus. Therefore, melatonin levels are low during the day and high during the night. However, in SMS patients, melatonin levels could be high during the day and low during the night. We ascertained that this disruption does probably not originate directly in the pineal gland but rather arises due to disrupted timing cues, which are being send to the pineal gland from the central clock. This finding could be used in the future as the basis for the chronobiological treatment of some of the accompanying SMS symptoms. Laboratory of Biological Rhythms - novakova1 While healthy children have melatonin secretion high during the night (A), melatonin secretion patterns in children with SMS are severely disrupted (B). (Nováková et al., J Clin Endocrinol Metab., 2012)

Publications

Drapšin; Milica - Dočkal; Tereza - Houdek; Pavel - Sládek; Martin - Semenovykh; Kateryna - Sumová; Alena Circadian clock in choroid plexus is resistant to immune challenge but dampens in response to chronodisruption. Brain Behavior and Immunity. 2024; 117(March); 255-269.

IF = 15.1

Marhefková; N. - Sládek; Martin - Sumová; Alena - Dubský; M. Circadian dysfunction and cardio-metabolic disorders in humans. Frontiers in Endocrinology. 2024; 15(29 April); 1328139.

IF = 3.9

Sládek; Martin - Houdek; Pavel - Myung; J. - Semenovykh; Kateryna - Dočkal; Tereza - Sumová; Alena The circadian clock in the choroid plexus drives rhythms in multiple cellular processes under the control of the suprachiasmatic nucleus. Fluids and Barriers of the CNS. 2024; 21(27 May); 46.

IF = 5.9

Honzlová; Petra - Semenovykh; Kateryna - Sumová; Alena The Circadian Clock of Polarized Microglia and Its Interaction with Mouse Brain Oscillators. Cellular and Molecular Neurobiology. 2023; 43(3); 1319-1333.

IF = 4.0

Sládek; Martin - Klusáček; Jan - Hamplová; Dana - Sumová; Alena Population-representative study reveals cardiovascular and metabolic disease biomarkers associated with misaligned sleep schedules. Sleep. 2023; 46(6)); zsad037.

IF = 5.6

Honzlová; Petra - Sumová; Alena Metabolic regulation of the circadian clock in classically and alternatively activated macrophages. Immunology and Cell Biology. 2023; 101(5); 428-443.

IF = 4.0

People

Head of Laboratory

Vedoucí vědecký pracovník
Tel: 2528
Email: alena.sumova@fgu.cas.cz

Deputy Head of Laboratory

Vědecký pracovník
Tel: 2609
Email: martin.sladek@fgu.cas.cz

Laboratory staff

Tel: 2407
Email: tereza.dockal@fgu.cas.cz
Tel: 2407
Email: milica.drapsin@fgu.cas.cz
Odborný pracovník VaV
Tel: 2545
Email: pavel.houdek@fgu.cas.cz
Tel: 3733
Tel: 2407
Email: pedro.cadenato@fgu.cas.cz
Tel: 2775
Email: dmytro.semenovykh
Tel: 2545
Email: kateryna.semenovykh@fgu.cas.cz
Tel: 2775
Email: dmytro.semenovykh@fgu.cas.cz
Laborantka
Tel: 2609
Email: eva.tlusta@fgu.cas.cz