Publications by authors named "Michel Stephan"

59 Publications

Aging selectively dampens oscillation of lipid abundance in white and brown adipose tissue.

Sci Rep 2021 Mar 15;11(1):5932. Epub 2021 Mar 15.

Laboratory Genetic Metabolic Diseases, Amsterdam UMC-AMC, University of Amsterdam, Amsterdam Gastroenterology and Metabolism, Amsterdam Cardiovascular Sciences, Meibergdreef 9, 1105 AZ, Amsterdam, the Netherlands.

Lipid metabolism is under the control of the circadian system and circadian dysregulation has been linked to obesity and dyslipidemia. These factors and outcomes have also been associated to, or affected by, the process of aging. Here, we investigated whether murine white (WAT) and brown (BAT) adipose tissue lipids exhibit rhythmicity and if this is affected by aging. To this end, we have measured the 24 h lipid profiles of WAT and BAT using a global lipidomics analysis of > 1100 lipids. We observed rhythmicity in nearly all lipid classes including glycerolipids, glycerophospholipids, sterol lipids and sphingolipids. Overall, ~ 22% of the analyzed lipids were considered rhythmic in WAT and BAT. Despite a general accumulation of lipids upon aging the fraction of oscillating lipids decreased in both tissues to 14% and 18%, respectively. Diurnal profiles of lipids in BAT appeared to depend on the lipid acyl chain length and this specific regulation was lost in aged mice. Our study revealed how aging affects the rhythmicity of lipid metabolism and could contribute to the quest for targets that improve diurnal lipid homeostasis to maintain cardiometabolic health during aging.
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http://dx.doi.org/10.1038/s41598-021-85455-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7961067PMC
March 2021

A multi-level assessment of the bidirectional relationship between aging and the circadian clock.

J Neurochem 2021 04 23;157(1):73-94. Epub 2021 Jan 23.

Department of Cellular and Chemical Biology, Laboratory for Neurophysiology, Leiden University Medical Center, Leiden, the Netherlands.

The daily temporal order of physiological processes and behavior contribute to the wellbeing of many organisms including humans. The central circadian clock, which coordinates the timing within our body, is located in the suprachiasmatic nucleus (SCN) of the hypothalamus. Like in other parts of the brain, aging impairs the SCN function, which in turn promotes the development and progression of aging-related diseases. We here review the impact of aging on the different levels of the circadian clock machinery-from molecules to organs-with a focus on the role of the SCN. We find that the molecular clock is less effected by aging compared to other cellular components of the clock. Proper rhythmic regulation of intracellular signaling, ion channels and neuronal excitability of SCN neurons are greatly disturbed in aging. This suggests a disconnection between the molecular clock and the electrophysiology of these cells. The neuronal network of the SCN is able to compensate for some of these cellular deficits. However, it still results in a clear reduction in the amplitude of the SCN electrical rhythm, suggesting a weakening of the output timing signal. Consequently, other brain areas and organs not only show aging-related deficits in their own local clocks, but also receive a weaker systemic timing signal. The negative spiral completes with the weakening of positive feedback from the periphery to the SCN. Consequently, chronotherapeutic interventions should aim at strengthening overall synchrony in the circadian system using life-style and/or pharmacological approaches.
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http://dx.doi.org/10.1111/jnc.15286DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8048448PMC
April 2021

Electrophysiological Approaches to Studying the Suprachiasmatic Nucleus.

Methods Mol Biol 2021 ;2130:303-324

Department of Psychiatry and Biobehavioral Sciences, University of California, Los Angeles, Los Angeles, CA, USA.

In mammals, the part of the nervous system responsible for most circadian behavior can be localized to a bilaterally paired structure in the hypothalamus known as the suprachiasmatic nucleus (SCN). Understanding the mammalian circadian system will require a detailed multilevel analysis of neural SCN circuits ex vivo and in vivo. Many of the techniques and approaches that are used for the analysis of the circuitry driving circadian oscillations in the SCN are similar to those employed in other brain regions. There is, however, one fundamental difference that needs to be taken into consideration, that is, the physiological, cell, and molecular properties of SCN neurons vary with the time of day. In this chapter, we will consider the preparations and electrophysiological techniques that we have used to analyze the SCN circuit focusing on the acute brain slice and intact, freely moving animal.
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http://dx.doi.org/10.1007/978-1-0716-0381-9_23DOI Listing
March 2021

Brief light exposure at dawn and dusk can encode day-length in the neuronal network of the mammalian circadian pacemaker.

FASEB J 2020 10 31;34(10):13685-13695. Epub 2020 Aug 31.

Department of Cellular and Chemical Biology, Laboratory for Neurophysiology, Leiden University Medical Center, Leiden, the Netherlands.

The central circadian pacemaker in mammals, the suprachiasmatic nucleus (SCN), is important for daily as well as seasonal rhythms. The SCN encodes seasonal changes in day length by adjusting phase distribution among oscillating neurons thereby shaping the output signal used for adaptation of physiology and behavior. It is well-established that brief light exposure at the beginning and end of the day, also referred to as "skeleton" light pulses, are sufficient to evoke the seasonal behavioral phenotype. However, the effect of skeleton light exposure on SCN network reorganization remains unknown. Therefore, we exposed mice to brief morning and evening light pulses that mark the time of dawn and dusk in a short winter- or a long summer day. Single-cell PER2::LUC recordings, electrophysiological recordings of SCN activity, and measurements of GABA response polarity revealed that skeleton light-regimes affected the SCN network to the same degree as full photoperiod. These results indicate the powerful, yet potentially harmful effects of even relatively short light exposures during the evening or night for nocturnal animals.
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http://dx.doi.org/10.1096/fj.202001133RRDOI Listing
October 2020

Aging Affects the Capacity of Photoperiodic Adaptation Downstream from the Central Molecular Clock.

J Biol Rhythms 2020 04 27;35(2):167-179. Epub 2020 Jan 27.

Department of Cellular and Chemical Biology, Laboratory for Neurophysiology, Leiden University Medical Center, Leiden, the Netherlands.

Aging impairs circadian clock function, leading to disrupted sleep-wake patterns and a reduced capability to adapt to changes in environmental light conditions. This makes shift work or the changing of time zones challenging for the elderly and, importantly, is associated with the development of age-related diseases. However, it is unclear what levels of the clock machinery are affected by aging, which is relevant for the development of targeted interventions. We found that naturally aged mice of >24 months had a reduced rhythm amplitude in behavior compared with young controls (3-6 months). Moreover, the old animals had a strongly reduced ability to adapt to short photoperiods. Recording PER2::LUC protein expression in the suprachiasmatic nucleus revealed no impairment of the rhythms in PER2 protein under the 3 different photoperiods tested (LD: 8:16, 12:12, and 16:8). Thus, we observed a discrepancy between the behavioral phenotype and the molecular clock, and we conclude that the aging-related deficits emerge downstream of the core molecular clock. Since it is known that aging affects several intracellular and membrane components of the central clock cells, it is likely that an impairment of the interaction between the molecular clock and these components is contributing to the deficits in photoperiod adaptation.
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http://dx.doi.org/10.1177/0748730419900867DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7134598PMC
April 2020

trans-Diastereoselective Ru(II)-Catalyzed Asymmetric Transfer Hydrogenation of α-Acetamido Benzocyclic Ketones via Dynamic Kinetic Resolution.

Org Lett 2019 05 6;21(10):3644-3648. Epub 2019 May 6.

National Institute of Chemistry , Hajdrihova 19 , SI-1000 Ljubljana , Slovenia.

A highly efficient enantio- and diastereoselective catalyzed asymmetric transfer hydrogenation via dynamic kinetic resolution (DKR-ATH) of α,β-dehydro-α-acetamido and α-acetamido benzocyclic ketones to ent- trans-β-amido alcohols is disclosed employing a new ansa-Ru(II) complex of an enantiomerically pure syn- N, N-ligand, i.e. ent- syn-ULTAM-(CH)Ph. DFT calculations of the transition state structures revealed an atypical two-pronged substrate attractive stabilization engaging the commonly encountered CH/π electrostatic interaction and a new additional O═S═O···HNAc H-bond hence favoring the trans-configured products.
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http://dx.doi.org/10.1021/acs.orglett.9b01069DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6750876PMC
May 2019

Uncovering functional signature in neural systems via random matrix theory.

PLoS Comput Biol 2019 05 1;15(5):e1006934. Epub 2019 May 1.

Instituut-Lorentz for Theoretical Physics, Leiden Institute of Physics, University of Leiden, Leiden, The Netherlands.

Neural systems are organized in a modular way, serving multiple functionalities. This multiplicity requires that both positive (e.g. excitatory, phase-coherent) and negative (e.g. inhibitory, phase-opposing) interactions take place across brain modules. Unfortunately, most methods to detect modules from time series either neglect or convert to positive, any measured negative correlation. This may leave a significant part of the sign-dependent functional structure undetected. Here we present a novel method, based on random matrix theory, for the identification of sign-dependent modules in the brain. Our method filters out both local (unit-specific) noise and global (system-wide) dependencies that typically obfuscate the presence of such structure. The method is guaranteed to identify an optimally contrasted functional 'signature', i.e. a partition into modules that are positively correlated internally and negatively correlated across. The method is purely data-driven, does not use any arbitrary threshold or network projection, and outputs only statistically significant structure. In measurements of neuronal gene expression in the biological clock of mice, the method systematically uncovers two otherwise undetectable, negatively correlated modules whose relative size and mutual interaction strength are found to depend on photoperiod.
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http://dx.doi.org/10.1371/journal.pcbi.1006934DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6513117PMC
May 2019

The influence of neuronal electrical activity on the mammalian central clock metabolome.

Metabolomics 2018 09 17;14(10):122. Epub 2018 Sep 17.

Department of Cellular and Chemical Biology, Leiden University Medical Center, Einthovenweg 20, 2333 ZC, Leiden, The Netherlands.

Introduction: Most organisms display circadian rhythms in physiology and behaviour. In mammals, these rhythms are orchestrated by the suprachiasmatic nucleus (SCN). Recently, several metabolites have emerged as important regulators of circadian timekeeping. Metabolomics approaches have aided in identifying some key metabolites in circadian processes in peripheral tissue, but methods to routinely measure metabolites in small brain areas are currently lacking.

Objective: The aim of the study was to establish a reliable method for metabolite quantifications in the central circadian clock and relate them to different states of neuronal excitability.

Methods: We developed a method to collect and process small brain tissue samples (0.2 mm), suitable for liquid chromatography-mass spectrometry. Metabolites were analysed in the SCN and one of its main hypothalamic targets, the paraventricular nucleus (PVN). Tissue samples were taken at peak (midday) and trough (midnight) of the endogenous rhythm in SCN electrical activity. Additionally, neuronal activity was altered pharmacologically.

Results: We found a minor effect of day/night fluctuations in electrical activity or silencing activity during the day. In contrast, increasing electrical activity during the night significantly upregulated many metabolites in SCN and PVN.

Conclusion: Our method has shown to produce reliable and physiologically relevant metabolite data from small brain samples. Inducing electrical activity at night mimics the effect of a light pulses in the SCN, producing phase shifts of the circadian rhythm. The upregulation of metabolites could have a functional role in this process, since they are not solely products of physiological states, they are significant parts of cellular signalling pathways.
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http://dx.doi.org/10.1007/s11306-018-1423-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6153692PMC
September 2018

From clock to functional pacemaker.

Eur J Neurosci 2020 01 2;51(1):482-493. Epub 2019 May 2.

Group Neurophysiology, Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, The Netherlands.

In mammals, the central pacemaker that coordinates 24-hr rhythms is located in the suprachiasmatic nucleus (SCN). Individual neurons of the SCN have a molecular basis for rhythm generation and hence, they function as cell autonomous oscillators. Communication and synchronization among these neurons are crucial for obtaining a coherent rhythm at the population level, that can serve as a pace making signal for brain and body. Hence, the ability of single SCN neurons to produce circadian rhythms is equally important as the ability of these neurons to synchronize one another, to obtain a bona fide pacemaker at the SCN tissue level. In this chapter we will discuss the mechanisms underlying synchronization, and plasticity herein, which allows adaptation to changes in day length. Furthermore, we will discuss deterioration in synchronization among SCN neurons in aging, and gain in synchronization by voluntary physical activity or exercise.
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http://dx.doi.org/10.1111/ejn.14388DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7027845PMC
January 2020

Effects of an intensive long-term prevention programme after myocardial infarction - a randomized trial.

Eur J Prev Cardiol 2019 03 18;26(5):522-530. Epub 2018 Jun 18.

1 Bremen Institute for Heart and Circulation Research (BIHKF) at the Klinikum Links der Weser, Germany.

Background: Long-term risk factor control after myocardial infarction (MI) is currently inadequate and there is an unmet need for effective secondary prevention programmes.

Design And Methods: It was the aim of the study to compare a 12-month intensive prevention programme (IPP), coordinated by prevention assistants and including education sessions, telephone visits and telemetric risk factor control, with usual care after MI. Three hundred and ten patients were randomized to IPP vs. usual care one month after hospital discharge for MI in two German heart centres. Primary study endpoint was the IPP Prevention Score (0-15 points) quantifying global risk factor control.

Results: Global risk factor control was strongly improved directly after MI before the beginning of the randomized study (30% increase IPP Prevention Score). During the 12-month course of the randomized trial the IPP Prevention Score was improved by a further 14.3% in the IPP group ( p < 0.001), while it decreased by 11.8% in the usual care group ( p < 0.001). IPP significantly reduced smoking, low-density lipoprotein cholesterol, systolic blood pressure and physical inactivity compared with usual care ( p < 0.05). Step counters with online documentation were used by the majority of patients (80%). Quality of life was significantly improved by IPP ( p < 0.05). The composite endpoint of adverse clinical events was slightly lower in the IPP group during 12 months (13.8% vs. 18.9%, p = 0.25).

Conclusions: A novel intensive prevention programme after MI, coordinated by prevention assistants and using personal teachings and telemetric strategies for 12 months, was significantly superior to usual care in providing sustainable risk factor control and better quality of life.
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http://dx.doi.org/10.1177/2047487318781109DOI Listing
March 2019

Differential Phase Arrangement of Cellular Clocks along the Tonotopic Axis of the Mouse Cochlea Ex Vivo.

Curr Biol 2017 Sep 17;27(17):2623-2629.e2. Epub 2017 Aug 17.

Department of Physiology and Pharmacology, Karolinska Institutet, 17177 Stockholm, Sweden. Electronic address:

Topological distributions of individual cellular clocks have not been demonstrated in peripheral organs. The cochlea displays circadian patterns of core clock gene expression [1, 2]. PER2 protein is expressed in the hair cells and spiral ganglion neurons of the cochlea in the spiral ganglion neurons [1]. To investigate the topological organization of cellular oscillators in the cochlea, we recorded circadian rhythms from mouse cochlear explants using highly sensitive real-time tracking of PER2::LUC bioluminescence. Here, we show cell-autonomous and self-sustained oscillations originating from hair cells and spiral ganglion neurons. Multi-phased cellular clocks were arranged along the length of the cochlea with oscillations initiating at the apex (low-frequency region) and traveling toward the base (high-frequency region). Phase differences of 3 hr were found between cellular oscillators in the apical and middle regions and from isolated individual cochlear regions, indicating that cellular networks organize the rhythms along the tonotopic axis. This is the first demonstration of a spatiotemporal arrangement of circadian clocks at the cellular level in a peripheral organ. Cochlear rhythms were disrupted in the presence of either voltage-gated potassium channel blocker (TEA) or extracellular calcium chelator (BAPTA), demonstrating that multiple types of ion channels contribute to the maintenance of coherent rhythms. In contrast, preventing action potentials with tetrodotoxin (TTX) or interfering with cell-to-cell communication the broad-spectrum gap junction blocker (CBX [carbenoxolone]) had no influence on cochlear rhythms. These findings highlight a dynamic regulation and longitudinal distribution of cellular clocks in the cochlea.
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http://dx.doi.org/10.1016/j.cub.2017.07.019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6899219PMC
September 2017

Stereopure Functionalized Benzosultams via Ruthenium(II)-Catalyzed Dynamic Kinetic Resolution-Asymmetric Transfer Hydrogenation.

Org Lett 2017 04 13;19(8):2042-2045. Epub 2017 Apr 13.

National Institute of Chemistry, Hajdrihova 19, 1001 Ljubljana, Slovenia.

A highly diastereo- and enantioselective Ru(II)-catalyzed dynamic kinetic resolution-asymmetric transfer hydrogenation (DKR-ATH) of α-(N-sulfonylimino) and α-(N-sulfonylamino) aryl ketones to 4-hydroxy-benzo-δ- and 3-(α-hydroxy-arylmethyl)-benzo-γ-sultams is presented. By employing enantiopure ansa-Ru[PipSODPEN(CH)Ph] cat. II with S/C = 10 000 in a HCOH/EtN binary mix, up to >99.9% ee and dr >99:1 are obtained with 100% conversion under mild conditions. Application to access the stereopure "structurally simplified TsDPEN" N,N-ligand syn-3-(α-aminobenzyl)-benzo-γ-sultam ("syn-ULTAM") and its structural isomer trans-4-amino-3-phenyl-benzo-δ-sultam (trans-4) is demonstrated.
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http://dx.doi.org/10.1021/acs.orglett.7b00670DOI Listing
April 2017

Evidence for Weakened Intercellular Coupling in the Mammalian Circadian Clock under Long Photoperiod.

PLoS One 2016 22;11(12):e0168954. Epub 2016 Dec 22.

Department of Molecular Cell Biology, Laboratory for Neurophysiology, Leiden University Medical Center, Leiden, The Netherlands.

For animals living in temperate latitudes, seasonal changes in day length are an important cue for adaptations of their physiology and behavior to the altered environmental conditions. The suprachiasmatic nucleus (SCN) is known as the central circadian clock in mammals, but may also play an important role in adaptations to different photoperiods. The SCN receives direct light input from the retina and is able to encode day-length by approximating the waveform of the electrical activity rhythm to the duration of daylight. Changing the overall waveform requires a reorganization of the neuronal network within the SCN with a change in the degree of synchrony between the neurons; however, the underlying mechanisms are yet unknown. In the present study we used PER2::LUC bioluminescence imaging in cultured SCN slices to characterize network dynamics on the single-cell level and we aimed to provide evidence for a role of modulations in coupling strength in the photoperiodic-induced phase dispersal. Exposure to long photoperiod (LP) induced a larger distribution of peak times of the single-cell PER2::LUC rhythms in the anterior SCN, compared to short photoperiod. Interestingly, the cycle-to-cycle variability in single-cell period of PER2::LUC rhythms is also higher in the anterior SCN in LP, and is positively correlated with peak time dispersal. Applying a new, impartial community detection method on the time series data of the PER2::LUC rhythm revealed two clusters of cells with a specific spatial distribution, which we define as dorsolateral and ventromedial SCN. Post hoc analysis of rhythm characteristics of these clusters showed larger cycle-to-cycle single-cell period variability in the dorsolateral compared to the ventromedial cluster in the anterior SCN. We conclude that a change in coupling strength within the SCN network is a plausible explanation to the observed changes in single-cell period variability, which can contribute to the photoperiod-induced phase distribution.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0168954PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5179103PMC
July 2017

Usefulness of Iron Deficiency Correction in Management of Patients With Heart Failure [from the Registry Analysis of Iron Deficiency-Heart Failure (RAID-HF) Registry].

Am J Cardiol 2016 Dec 15;118(12):1875-1880. Epub 2016 Sep 15.

Bremer Institut für Herz- und Kreislaufforschung am Klinikum Links der Weser, Bremen, Germany.

Iron deficiency (ID) has been identified as an important co-morbidity in patients with heart failure (HF). Intravenous iron therapy reduced symptoms and rehospitalizations of iron-deficient patients with HF in randomized trials. The present multicenter study investigated the "real-world" management of iron status in patients with HF. Consecutive patients with HF and ejection fraction ≤40% were recruited and analyzed from December 2010 to October 2015 by 11 centers in Germany and Switzerland. Of 1,484 patients with HF, iron status was determined in only 923 patients (62.2%), despite participation of the centers in a registry focusing on ID and despite guideline recommendation to determine iron status. In patients with determined iron status, a prevalence of 54.7% (505 patients) for ID was observed. Iron therapy was performed in only 8.5% of the iron-deficient patients with HF; 2.6% were treated with intravenous iron therapy. The patients with iron therapy were characterized by a high rate of symptomatic HF and anemia. In conclusion, despite strong evidence of beneficial effects of iron therapy on symptoms and rehospitalizations, diagnostic and therapeutic efforts on ID in HF are low in the actual clinical practice, and the awareness to diagnose and treat ID in HF should be strongly enforced.
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http://dx.doi.org/10.1016/j.amjcard.2016.08.081DOI Listing
December 2016

Photoperiod Modulates Fast Delayed Rectifier Potassium Currents in the Mammalian Circadian Clock.

ASN Neuro 2016 10 3;8(5). Epub 2016 Oct 3.

Leiden University Medical Center, Leiden, The Netherlands.

One feature of the mammalian circadian clock, situated in the suprachiasmatic nucleus (SCN), is its ability to measure day length and thereby contribute to the seasonal adaptation of physiology and behavior. The timing signal from the SCN, namely the 24 hr pattern of electrical activity, is adjusted according to the photoperiod being broader in long days and narrower in short days. Vasoactive intestinal peptide and gamma-aminobutyric acid play a crucial role in intercellular communication within the SCN and contribute to the seasonal changes in phase distribution. However, little is known about the underlying ionic mechanisms of synchronization. The present study was aimed to identify cellular mechanisms involved in seasonal encoding by the SCN. Mice were adapted to long-day (light-dark 16:8) and short-day (light-dark 8:16) photoperiods and membrane properties as well as K currents activity of SCN neurons were measured using patch-clamp recordings in acute slices. Remarkably, we found evidence for a photoperiodic effect on the fast delayed rectifier K current, that is, the circadian modulation of this ion channel's activation reversed in long days resulting in 50% higher peak values during the night compared with the unaltered day values. Consistent with fast delayed rectifier enhancement, duration of action potentials during the night was shortened and afterhyperpolarization potentials increased in amplitude and duration. The slow delayed rectifier, transient K currents, and membrane excitability were not affected by photoperiod. We conclude that photoperiod can change intrinsic ion channel properties of the SCN neurons, which may influence cellular communication and contribute to photoperiodic phase adjustment.
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http://dx.doi.org/10.1177/1759091416670778DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5051630PMC
October 2016

Ryanodine-sensitive intracellular Ca channels are involved in the output from the SCN circadian clock.

Eur J Neurosci 2016 10 2;44(7):2504-2514. Epub 2016 Sep 2.

Neuroscience, Karolinska Institutet, Stockholm, Sweden.

The suprachiasmatic nuclei (SCN) contain the major circadian clock responsible for generation of circadian rhythms in mammals. The time measured by the molecular circadian clock must eventually be translated into a neuronal firing rate pattern to transmit a meaningful signal to other tissues and organs in the animal. Previous observations suggest that circadian modulation of ryanodine receptors (RyR) is a key element of the output pathway from the molecular circadian clock. To directly test this hypothesis, we studied the effects of RyR activation and inhibition on real time expression of PERIOD2::LUCIFERASE, intracellular calcium levels and spontaneous firing frequency in mouse SCN neurons. Furthermore, we determined whether the RyR-2 mRNA is expressed with a daily variation in SCN neurons. We provide evidence that pharmacological manipulation of RyR in mice SCN neurons alters the free [Ca ] in the cytoplasm and the spontaneous firing without affecting the molecular clock mechanism. Our data also show a daily variation in RyR-2 mRNA from single mouse SCN neurons with highest levels during the day. Together, these results confirm the hypothesis that RyR-2 is a key element of the circadian clock output from SCN neurons.
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http://dx.doi.org/10.1111/ejn.13368DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5053303PMC
October 2016

γ-Sultam-cored N,N-ligands in the ruthenium(ii)-catalyzed asymmetric transfer hydrogenation of aryl ketones.

Org Biomol Chem 2016 Feb;14(6):2112-20

National Institute of Chemistry, Hajdrihova 19, 1001 Ljubljana, Slovenia.

The synthesis of new enantiopure syn- and anti-3-(α-aminobenzyl)-benzo-γ-sultam ligands 6 and their application in the ruthenium(ii)-catalyzed asymmetric transfer hydrogenation (ATH) of ketones using formic acid/triethylamine is described. In particular, benzo-fused cyclic ketones afforded excellent enantioselectivities in reasonable time employing a low loading of the syn ligand-containing catalyst. A never-before-seen dynamic kinetic resolution (DKR) during reduction of a γ-keto carboxylic ester (S7) derivative of 1-indanone is realized leading as well to excellent induction.
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http://dx.doi.org/10.1039/c5ob02352aDOI Listing
February 2016

Role of vasoactive intestinal peptide in the light input to the circadian system.

Eur J Neurosci 2015 Jul 25;42(2):1839-48. Epub 2015 May 25.

Department of Psychiatry & Biobehavioral Sciences, University of California - Los Angeles, Los Angeles, CA, 90024, USA.

The neuropeptide vasoactive intestinal peptide (VIP) is expressed at high levels in a subset of neurons in the ventral region of the suprachiasmatic nucleus (SCN). While VIP is known to be important for the synchronization of the SCN network, the role of VIP in photic regulation of the circadian system has received less attention. In the present study, we found that the light-evoked increase in electrical activity in vivo was unaltered by the loss of VIP. In the absence of VIP, the ventral SCN still exhibited N-methyl-d-aspartate-evoked responses in a brain slice preparation, although the absolute levels of neural activity before and after treatment were significantly reduced. Next, we used calcium imaging techniques to determine if the loss of VIP altered the calcium influx due to retinohypothalamic tract stimulation. The magnitude of the evoked calcium influx was not reduced in the ventral SCN, but did decline in the dorsal SCN regions. We examined the time course of the photic induction of Period1 in the SCN using in situ hybridization in VIP-mutant mice. We found that the initial induction of Period1 was not reduced by the loss of this signaling peptide. However, the sustained increase in Period1 expression (after 30 min) was significantly reduced. Similar results were found by measuring the light induction of cFOS in the SCN. These findings suggest that VIP is critical for longer-term changes within the SCN circuit, but does not play a role in the acute light response.
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http://dx.doi.org/10.1111/ejn.12919DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4509820PMC
July 2015

Age-related changes in large-conductance calcium-activated potassium channels in mammalian circadian clock neurons.

Neurobiol Aging 2015 Jun 31;36(6):2176-83. Epub 2015 Jan 31.

Laboratory of Neurophysiology, Department of Molecular Cell Biology, Leiden University Medical Center, Leiden, The Netherlands. Electronic address:

Aging impairs the function of the suprachiasmatic nucleus (SCN, the central mammalian clock), leading to a decline in the circadian rhythm of many physiological processes, including sleep-wake rhythms. Recent studies have found evidence of age-related changes in the circadian regulation of potassium currents; these changes presumably lead to a decrease in the SCN's electrical rhythm amplitude. Current through large-conductance Ca(2+)-activated K(+) (BK) channels promote rhythmicity in both SCN neuronal activity and behavior. In many neuron types, changes in BK activity are correlated with changes in intracellular Ca(2+) concentration ([Ca(2+)]i). We performed patch-clamp recordings of SCN neurons in aged mice and observed that the circadian modulation of BK channel activity was lost because of a reduction in BK currents during the night. This reduced current diminished the afterhyperpolarization, depolarized the resting membrane potential, widened the action potential, and increased [Ca(2+)]i. These data suggest that reduced BK current increases [Ca(2+)]i by altering the action potential waveform, possibly contributing to the observed age-related phenotype.
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http://dx.doi.org/10.1016/j.neurobiolaging.2014.12.040DOI Listing
June 2015

Neurophysiological analysis of the suprachiasmatic nucleus: a challenge at multiple levels.

Methods Enzymol 2015 26;552:75-102. Epub 2014 Dec 26.

Department of Molecular Cell Biology, Leiden University Medical Center, Leiden, The Netherlands.

Understanding the neurophysiology of the circadian timing system requires investigation at multiple levels of organization. Neurons of the suprachiasmatic nucleus (SCN) function as autonomous single-cell oscillators, which warrant studies at the single-cell level. Combining patch-clamp recordings of ion channels with imaging techniques to measure clock gene expression and intracellular calcium has proven extremely valuable to study cellular properties. To achieve and maintain rhythmic activity, SCN neurons require sufficient stimulation (i.e., input) from surrounding cells. At the network level, SCN rhythms are robust and can be measured in vitro, for example, in brain slices that contain the SCN. These recordings revealed that the collective behavior of the SCN neuronal network is strongly determined by the phase dispersal of the neurons. This phase dispersal is plastic, with high synchronization in short photoperiod, desynchronization in long photoperiod, and antiphase oscillations in aging and/or continuous light. In vivo recordings are needed in order to study the SCN as part of a larger network (i.e., interacting with other brain centers) and to study the SCN's response to light. Interestingly, superimposed on the circadian waveform are higher frequency fluctuations that are present in vivo but not in vitro. These fluctuations are attributed to input from other brain centers and computational analyses suggest that these fluctuations are beneficial to the system. Hence, the SCN's properties arise from several organizational levels, and a combination of approaches is needed in order to fully understand the circadian system.
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http://dx.doi.org/10.1016/bs.mie.2014.11.001DOI Listing
November 2015

Seasonal induction of GABAergic excitation in the central mammalian clock.

Proc Natl Acad Sci U S A 2014 Jul 16;111(26):9627-32. Epub 2014 Jun 16.

Department of Molecular Cell Biology, Laboratory of Neurophysiology, Leiden University Medical Center, 2300 RC Leiden, The Netherlands

The balance between excitation and inhibition is essential for the proper function of neuronal networks in the brain. The inhibitory neurotransmitter γ-aminobutyric acid (GABA) contributes to the network dynamics within the suprachiasmatic nucleus (SCN), which is involved in seasonal encoding. We investigated GABAergic activity and observed mainly inhibitory action in SCN neurons of mice exposed to a short-day photoperiod. Remarkably, the GABAergic activity in a long-day photoperiod shifts from inhibition toward excitation. The mechanistic basis for this appears to be a change in the equilibrium potential of GABA-evoked current. These results emphasize that environmental conditions can have substantial effects on the function of a key neurotransmitter in the central nervous system.
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http://dx.doi.org/10.1073/pnas.1319820111DOI Listing
July 2014

Efficient asymmetric syntheses of 1-phenyl-phosphindane, derivatives, and 2- or 3-oxa analogues: mission accomplished.

Org Lett 2014 May 28;16(10):2688-91. Epub 2014 Apr 28.

National Institute of Chemistry , Hajdrihova 19, 1000 Ljubljana, Slovenia.

A highly enantioselective synthesis of unsubstituted 1-phenyl-phosphindane and its P-borane and P-oxide derivatives was effectively established via a new fluoride-triggered desilylative carbocyclization strategy. Preparation of the "oxygen atom-doped" 1-phenyl-3-oxa-1-phosphindane-P-borane analogue was otherwise achieved via a tandem P-α-iodination-intra-O-alkylation.
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http://dx.doi.org/10.1021/ol500970xDOI Listing
May 2014

Enhanced phase resetting in the synchronized suprachiasmatic nucleus network.

J Biol Rhythms 2014 Feb;29(1):4-15

Laboratory for Neurophysiology, Department of Molecular Cell Biology, Leiden University Medical Center, Leiden, the Netherlands.

The suprachiasmatic nucleus (SCN) adapts to both the external light-dark (LD) cycle and seasonal changes in day length. In short photoperiods, single-cell activity patterns are tightly synchronized (i.e., in phase); in long photoperiods, these patterns are relatively dispersed, causing lower amplitude rhythms. The limit cycle oscillator has been used to describe the SCN's circadian rhythmicity and predicts that following a given perturbation, high-amplitude SCN rhythms will shift less than low-amplitude rhythms. Some studies reported, however, that phase delays are larger when animals are entrained to a short photoperiod. Because phase advances and delays are mediated by partially distinct (i.e., nonoverlapping) biochemical pathways, we investigated the effect of a 4-h phase advance of the LD cycle in mice housed in either short (LD 8:16) or long (LD 16:8) photoperiods. In vitro recordings revealed a significantly larger phase advance in the SCN of mice entrained to short as compared to long photoperiods (4.2 ± 0.3 h v. 1.4 ± 0.9 h, respectively). Surprisingly, in mice with long photoperiods, the behavioral phase shift was larger than the phase shift of the SCN (3.7 ± 0.4 h v. 1.4 ± 0.9 h, respectively). To exclude a confounding influence of running-wheel activity on the magnitude of the shifts of the SCN, we repeated the experiments in the absence of running wheels and found similar shifts in the SCN in vitro in short and long days (3.0 ± 0.5 h v. 0.4 ± 0.9 h, respectively). Interestingly, removal of the running wheel reduced the phase-shifting capacity of mice in long days, leading to similar behavioral shifts in short and long photoperiods (1.0 ± 0.1 h v. 1.0 ± 0.4 h). As the behavioral shifts in the presence of wheels were larger than the shift of the SCN, it is suggested that additional, non-SCN neuronal networks in the brain are involved in regulating the timing of behavioral activity. On the basis of the phase shifts observed in vitro, we conclude that highly synchronized SCN networks with high-amplitude rhythms show a larger phase-shifting capacity than desynchronized networks of low amplitude.
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http://dx.doi.org/10.1177/0748730413516750DOI Listing
February 2014

Aging of the suprachiasmatic clock.

Neuroscientist 2014 Feb 7;20(1):44-55. Epub 2013 Aug 7.

1Department of Molecular Cell Biology, Leiden University Medical Center, Leiden, The Netherlands.

More than half of the elderly in today's society suffer from sleep disorders with detrimental effects on brain function, behavior, and social life. A major contribution to the regulation of sleep stems from the circadian system. The central circadian clock located in the suprachiasmatic nucleus of the hypothalamus is like other brain regions subject to age-associated changes. Age affects different levels of the clock machinery from molecular rhythms, intracellular messenger, and membrane properties to neuronal network synchronization. While some of the age-sensitive components of the circadian clock, like ion channels and neurotransmitters, have been described, little is known about the underlying mechanisms. In any case, the result is a reduction in the amplitude of the circadian timing signal produced by the suprachiasmatic nucleus, a weakening in the control of peripheral oscillators and a decrease in amplitude and precision of daily rhythms in physiology and behavior. The distortion in temporal organization is thought to be related to a number of serious health problems and promote neurodegeneration. Understanding the mechanisms underlying age-related deficits in circadian clock function will therefore not only benefit rhythm disorders but also alleviate age-associated diseases aggravated by clock dysfunction.
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http://dx.doi.org/10.1177/1073858413498936DOI Listing
February 2014

P-stereogenic phospholanes or phosphorinanes from o-biarylylphosphines: two bridges not too far.

J Org Chem 2013 May 6;78(10):4665-73. Epub 2013 May 6.

National Institute of Chemistry, Hajdrihova 19, SI-1000 Ljubljana, Slovenia.

The discovery of a concise regiodivergent asymmetric route to nonclassical P-stereogenic 5- or 6-membered benzophosphacycles, under conditions-dependent radical (oxidative addition) versus anionic (S(N)Ar) benzannulation, is reported.
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http://dx.doi.org/10.1021/jo400565bDOI Listing
May 2013

Asymmetric transfer hydrogenation of 1-naphthyl ketones by an ansa-Ru(II) complex of a DPEN-SO2N(Me)-(CH2)2(η(6)-p-Tol) combined ligand.

Org Lett 2013 Apr 19;15(7):1614-7. Epub 2013 Mar 19.

National Institute of Chemistry, Hajdrihova 19, SI-1000 Ljubljana, Slovenia.

The first second-generation designer Ru(II) catalyst 1b featuring an enantiopure N,C-(N-ethylene-N-methyl-sulfamoyl)-tethered (DPEN-κ(2)N,N')/η(6)-toluene hybrid ligand is introduced. Using an S/C = 1000 in HCO2H-Et3N 5:2 transfer hydrogenation medium, secondary 1-naphthyl alcohols are obtained in up to >99.9% ee under mild conditions. Mechanistic factors are discussed.
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http://dx.doi.org/10.1021/ol400393jDOI Listing
April 2013

Mechanism of bilateral communication in the suprachiasmatic nucleus.

Eur J Neurosci 2013 Mar 14;37(6):964-71. Epub 2013 Jan 14.

Laboratory for Neurophysiology, Department of Molecular Cell Biology, Leiden University Medical Center, LUMC PZ S5-P, PO 9600, 2300 RC, Leiden, The Netherlands.

The central circadian pacemaker of the suprachiasmatic nuclei (SCN) is a bilaterally symmetrical structure. Little is known about the physiological mechanisms underlying communication between the left and right SCN and yet the degree of synchronization between SCN neurons can have a critical impact on the properties of the circadian system. In this study, we used electrophysiological tools and calcium (Ca(2+) ) imaging to examine the mechanisms underlying bilateral signaling in mouse SCN. Electrical stimulation of one SCN produced responses in the contralateral SCN with a short delay (approximately 5 ms) and Ca(2+) -dependence that are consistent with action potential-mediated chemical synaptic transmission. Patch-clamp recordings of stimulated cells revealed excitatory postsynaptic inward-currents (EPSCs), which were sufficient in magnitude to elicit action potentials. Electrical stimulation evoked tetrodotoxin-dependent Ca(2+) transients in about 30% of all contralateral SCN neurons recorded. The responding neurons were widely distributed within the SCN with a highest density in the posterior SCN. EPSCs and Ca(2+) responses were significantly reduced after application of a glutamate receptor antagonist. Application of antagonists for receptors of other candidate transmitters inhibited the Ca(2+) responses in some of the cells but overall the impact of these antagonists was variable. In a functional assay, electrical stimulation of the SCN produced phase shifts in the circadian rhythm in the frequency of multiunit activity rhythm in the contralateral SCN. These phase shifts were blocked by a glutamate receptor antagonist. Taken together, these results implicate glutamate as a transmitter required for communication between the left and right SCN.
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http://dx.doi.org/10.1111/ejn.12109DOI Listing
March 2013

Dynamic neuronal network organization of the circadian clock and possible deterioration in disease.

Prog Brain Res 2012 ;199:143-162

Laboratory for Neurophysiology, Department of Molecular Cell Biology, Leiden University Medical Center, Leiden, The Netherlands.

In mammals, the suprachiasmatic nuclei (SCNs) function as a circadian pacemaker that drives 24-h rhythms in physiology and behavior. The SCN is a multicellular clock in which the constituent oscillators show dynamics in their functional organization and phase coherence. Evidence has emerged that plasticity in phase synchrony among SCN neurons determines (i) the amplitude of the rhythm, (ii) the response to continuous light, (iii) the capacity to respond to seasonal changes, and (iv) the phase-resetting capacity. A decrease in circadian amplitude and phase-resetting capacity is characteristic during aging and can be a result of disease processes. Whether the decrease in amplitude is caused by a loss of synchronization or by a loss of single-cell rhythmicity remains to be determined and is important for the development of strategies to ameliorate circadian disorders.
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http://dx.doi.org/10.1016/B978-0-444-59427-3.00009-5DOI Listing
January 2013

Evidence for neuronal desynchrony in the aged suprachiasmatic nucleus clock.

J Neurosci 2012 Apr;32(17):5891-9

Department of Neurophysiology, Leiden University Medical Center, 2300 RC Leiden, The Netherlands.

Aging is associated with a deterioration of daily (circadian) rhythms in physiology and behavior. Deficits in the function of the central circadian pacemaker in the suprachiasmatic nucleus (SCN) have been implicated, but the responsible mechanisms have not been clearly delineated. In this report, we characterize the progression of rhythm deterioration in mice to 900 d of age. Longitudinal behavioral and sleep-wake recordings in up to 30-month-old mice showed strong fragmentation of rhythms, starting at the age of 700 d. Patch-clamp recordings in this age group revealed deficits in membrane properties and GABAergic postsynaptic current amplitude. A selective loss of circadian modulation of fast delayed-rectifier and A-type K+ currents was observed. At the tissue level, phase synchrony of SCN neurons was grossly disturbed, with some subpopulations peaking in anti-phase and a reduction in amplitude of the overall multiunit activity rhythm. We propose that aberrant SCN rhythmicity in old animals--with electrophysiological arrhythmia at the single-cell level and phase desynchronization at the network level--can account for defective circadian function with aging.
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http://dx.doi.org/10.1523/JNEUROSCI.0469-12.2012DOI Listing
April 2012
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