scholarly journals Gpr19 is a circadian clock-controlled orphan GPCR with a role in modulating free-running period and light resetting capacity of the circadian clock

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yoshiaki Yamaguchi ◽  
Iori Murai ◽  
Kaoru Goto ◽  
Shotaro Doi ◽  
Huihua Zhou ◽  
...  
Keyword(s):  
Circadian Clock ◽  
Light Exposure ◽  
Physiological Role ◽  
Delayed Initiation ◽  
Free Running ◽  
Responsive Element ◽  
Free Running Period ◽  
Ventral Area ◽  
Orphan Gpcr

AbstractGpr19 encodes an evolutionarily conserved orphan G-protein-coupled receptor (GPCR) with currently no established physiological role in vivo. We characterized Gpr19 expression in the suprachiasmatic nucleus (SCN), the locus of the master circadian clock in the brain, and determined its role in the context of the circadian rhythm regulation. We found that Gpr19 is mainly expressed in the dorsal part of the SCN, with its expression fluctuating in a circadian fashion. A conserved cAMP-responsive element in the Gpr19 promoter was able to produce circadian transcription in the SCN. Gpr19−/− mice exhibited a prolonged circadian period and a delayed initiation of daily locomotor activity. Gpr19 deficiency caused the downregulation of several genes that normally peak during the night, including Bmal1 and Gpr176. In response to light exposure at night, Gpr19−/− mice had a reduced capacity for light-induced phase-delays, but not for phase-advances. This defect was accompanied by reduced response of c-Fos expression in the dorsal region of the SCN, while apparently normal in the ventral area of the SCN, in Gpr19−/− mice. Thus, our data demonstrate that Gpr19 is an SCN-enriched orphan GPCR with a distinct role in circadian regulation and may provide a potential target option for modulating the circadian clock.

scholarly journals Gpr19 is a circadian clock-controlled orphan GPCR with a role in modulating free-running period and light resetting capacity of the circadian clock

2021 ◽  
Author(s):  
Yoshiaki Yamaguchi ◽  
Iori Murai ◽  
Kaoru Goto ◽  
Shotaro Doi ◽  
Huihua Zhou ◽  
...  
Keyword(s):  
Circadian Clock ◽  
Clock Genes ◽  
Dorsal Region ◽  
Circadian Expression ◽  
Delayed Initiation ◽  
Free Running ◽  
Responsive Element ◽  
Free Running Period ◽  
Orphan Gpcr

Background and Purpose: Gpr19 encodes an evolutionarily conserved orphan G-protein-coupled receptor (GPCR) with no established physiological function in vivo. The purpose of this study was to determine the role of Gpr19 in the circadian clock system. Experimental Approach: We examined whether and how the master circadian clock neurons in the suprachiasmatic nucleus (SCN) express Gpr19. By analysing Gpr19-deficient (Gpr19−/−) mice, we asked whether Gpr19 has a role in modulating free-running period and light resetting capacity of the circadian clock. Key Results: Compared with the known common core clock genes, Gpr19 was identified to show several distinct yet limited features related to the circadian clock. Gpr19 mRNA was mainly expressed in the middle-to-dorsal region of the SCN. A conserved cAMP-responsive element within the Gpr19 promoter drove the circadian expression of Gpr19. Gpr19−/− mice exhibited a prolonged circadian period and a delayed initiation of daily locomotor activity in a 12-h light/12-h dark cycle. Gpr19 deficiency caused the downregulation of several genes that normally peak during the night, including Bmal1 and Gpr176. Gpr19−/− mice had a reduced capacity for phase shift to early subjective night light. The defect was only observed for phase-delay, but not phase-advance, and accompanied by reduced response of c-Fos expression in the dorsal region of the SCN, while apparently normal in the ventral part of the SCN, in Gpr19−/− mice. Conclusion and Implications: Gpr19 is an SCN-enriched orphan GPCR with a distinct role in circadian regulation and thus may be a potential target for alleviating circadian clock disorders.

Regularly scheduled voluntary exercise synchronizes the mouse circadian clock

1991 ◽  
Vol 261 (4) ◽  
pp. R928-R933 ◽  
Author(s):  
D. M. Edgar ◽  
W. C. Dement
Keyword(s):  
Circadian Rhythms ◽  
Phase Shift ◽  
Circadian Clock ◽  
Wheel Running ◽  
External Environment ◽  
Voluntary Exercise ◽  
Wheel Rotation ◽  
Exercise Activity ◽  
Free Running ◽  
Free Running Period

Circadian rhythm entrainment has long been thought to depend exclusively on periodic cues in the external environment. However, evidence now suggests that appropriately timed vigorous activity may also phase shift the circadian clock. Previously it was not known whether levels of exercise/activity associated with spontaneous behavior provided sufficient feedback to phase shift or synchronize circadian rhythms. The present study investigated this issue by monitoring the sleep-wake, drinking, and wheel-running circadian rhythms of mice (Mus musculus) during unrestricted access to running wheels and when free wheel rotation was limited to either 12- or 6-h intervals with a fixed period of 24 h. Wheel rotation was controlled remotely. Mice spontaneously ran in wheels during scheduled access, and free-running sleep-wake and drinking circadian rhythms became entrained to scheduled exercise in 11 of 15 animals. However, steady-state entrainment was achieved only when exercise commenced several hours into the subjective night. The temporal placement of running during entrainment was related (r = 0.7003, P less than 0.02) to free-running period before entrainment. Mice with a free-running period less than 23.0 h did not entrain but exhibited relative coordination between free-running variables and the wheel availability schedule. Thus the circadian timekeeping system responds to temporal feedback arising from the timing of volitional exercise/activity, suggesting that the biological clock not only is responsive to periodic geophysical events in the external environment but also derives physiological feedback from the spontaneous activity behaviors of the organism.

scholarly journals Intracellular Ca2+ Regulates Free-Running Circadian Clock Oscillation In Vivo

2007 ◽  
Vol 27 (46) ◽  
pp. 12489-12499 ◽  
Author(s):  
M. C. Harrisingh ◽  
Y. Wu ◽  
G. A. Lnenicka ◽  
M. N. Nitabach
Keyword(s):  
Circadian Clock ◽  
Free Running

scholarly journals Single adeno-associated virus-based multiplexed CRISPR-Cas9 system to nullify core components of the mammalian molecular clock

2020 ◽  
Author(s):  
Boil Kim ◽  
Jihoon Kim ◽  
Minjeong Chun ◽  
Inah Park ◽  
Mijung Choi ◽  
...  
Keyword(s):  
Circadian Clock ◽  
Molecular Clock ◽  
Viral Vectors ◽  
Clock Genes ◽  
Ex Vivo ◽  
Physiological Role ◽  
Guide Rnas ◽  
Multiple Genes

ABSTRACTThe mammalian molecular clock is based on a transcription-translation feedback loop (TTFL) containing Period1, 2 (Per1, 2), Cryptochrome1, 2 (Cry1, 2), and Brain and Muscle ARNT-Like 1 (Bmal1). TTFL robustness is endowed by genetic complementation between these components; therefore, multiple genes must be knocked out to physiologically investigate the molecular clock, which requires extensive research resources. To facilitate molecular clock disruption, we developed a CRISPR-Cas9-based single adeno-associated viral (AAV) system targeting the circadian clock (CSAC) for Pers, Crys, or Bmal1. First, we designed single guide RNAs (sgRNAs) targeting individual clock genes using an in silico approach and validated their efficiency in Neuro2a cells. To target multiple genes, multiplex sgRNA plasmids were constructed using Golden Gate assembly and expressed in viral vectors. CSAC efficiency was demonstrated by decreased protein expression in vitro and ablated molecular oscillation ex vivo. We also measured locomotor activity and body temperature in Cas9-expressing mice injected with CSAC at the suprachiasmatic nucleus. Circadian rhythm disruption was observed under free-running conditions, indicating that CSAC can efficiently and robustly disrupt molecular circadian clock. Thus, CSAC is a simple and powerful tool for investigating the physiological role of the molecular clock in vivo.

scholarly journals The Phosphorylation of CREB at Serine 133 Is a Key Event for Circadian Clock Timing and Entrainment in the Suprachiasmatic Nucleus

2018 ◽  
Vol 33 (5) ◽  
pp. 497-514 ◽  
Author(s):  
Kelin L. Wheaton ◽  
Katelin F. Hansen ◽  
Sydney Aten ◽  
Kyle A. Sullivan ◽  
Hyojung Yoon ◽  
...  
Keyword(s):  
Circadian Clock ◽  
Suprachiasmatic Nucleus ◽  
Wheel Running ◽  
Slice Culture ◽  
Creb Phosphorylation ◽  
Running Activity ◽  
Wheel Running Activity ◽  
Free Running

Within the suprachiasmatic nucleus (SCN)—the locus of the master circadian clock— transcriptional regulation via the CREB/CRE pathway is implicated in the functioning of the molecular clock timing process, and is a key conduit through which photic input entrains the oscillator. One event driving CRE-mediated transcription is the phosphorylation of CREB at serine 133 (Ser133). Indeed, numerous reporter gene assays have shown that an alanine point mutation in Ser133 reduces CREB-mediated transcription. Here, we sought to examine the contribution of Ser133 phosphorylation to the functional role of CREB in SCN clock physiology in vivo. To this end, we used a CREB knock-in mouse strain, in which Ser133 was mutated to alanine (S/A CREB). Under a standard 12 h light-dark cycle, S/A CREB mice exhibited a marked alteration in clock-regulated wheel running activity. Relative to WT mice, S/A CREB mice had highly fragmented bouts of locomotor activity during the night phase, elevated daytime activity, and a delayed phase angle of entrainment. Further, under free-running conditions, S/A CREB mice had a significantly longer tau than WT mice and reduced activity amplitude. In S/A CREB mice, light-evoked clock entrainment, using both Aschoff type 1 and 6 h “jet lag” paradigms, was markedly reduced relative to WT mice. S/A CREB mice exhibited attenuated transcriptional drive, as assessed by examining both clock-gated and light-evoked gene expression. Finally, SCN slice culture imaging detected a marked disruption in cellular clock phase synchrony following a phase-resetting stimulus in S/A CREB mice. Together, these data indicate that signaling through CREB phosphorylation at Ser133 is critical for the functional fidelity of both SCN timing and entrainment.

scholarly journals Cyclin-dependent kinase 5 (CDK5) regulates the circadian clock

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Andrea Brenna ◽  
Iwona Olejniczak ◽  
Rohit Chavan ◽  
Jürgen A Ripperger ◽  
Sonja Langmesser ◽  
...  
Keyword(s):  
Circadian Clock ◽  
Cyclin Dependent Kinase ◽  
Suprachiasmatic Nuclei ◽  
Nuclear Entry ◽  
Period 2 ◽  
Clock Component ◽  
Free Running ◽  
Free Running Period ◽  
Cryptochrome 1 ◽  
Cyclin Dependent Kinase 5

Circadian oscillations emerge from transcriptional and post-translational feedback loops. An important step in generating rhythmicity is the translocation of clock components into the nucleus, which is regulated in many cases by kinases. In mammals, the kinase promoting the nuclear import of the key clock component Period 2 (PER2) is unknown. Here, we show that the cyclin-dependent kinase 5 (CDK5) regulates the mammalian circadian clock involving phosphorylation of PER2. Knock-down of Cdk5 in the suprachiasmatic nuclei (SCN), the main coordinator site of the mammalian circadian system, shortened the free-running period in mice. CDK5 phosphorylated PER2 at serine residue 394 (S394) in a diurnal fashion. This phosphorylation facilitated interaction with Cryptochrome 1 (CRY1) and nuclear entry of the PER2-CRY1 complex. Taken together, we found that CDK5 drives nuclear entry of PER2, which is critical for establishing an adequate circadian period of the molecular circadian cycle. Of note is that CDK5 may not exclusively phosphorylate PER2, but in addition may regulate other proteins that are involved in the clock mechanism. Taken together, it appears that CDK5 is critically involved in the regulation of the circadian clock and may represent a link to various diseases affected by a derailed circadian clock.

scholarly journals Optogenetic stimulation of VIPergic SCN neurons induces photoperiodic changes in the mammalian circadian clock

2021 ◽  
Author(s):  
Michael C. Tackenberg ◽  
Jacob J. Hughey ◽  
Douglas G. McMahon
Keyword(s):  
Ex Vivo ◽  
Day Length ◽  
Optogenetic Stimulation ◽  
Period 2 ◽  
Free Running ◽  
Free Running Period ◽  
The Brain ◽  
Long Photoperiod ◽  
Stimulation Of

SummaryCircadian clocks play key roles in how organisms respond to and even anticipate seasonal change in day length, or photoperiod. In mammals, photoperiod is encoded by the central circadian pacemaker in the brain, the suprachiasmatic nucleus (SCN). The subpopulation of SCN neurons that secrete the neuropeptide VIP mediate the transmission of light information within the SCN neural network, suggesting a role for these neurons in circadian plasticity in response to light information that has yet to be directly tested. Here, we used in vivo optogenetic stimulation of VIPergic SCN neurons followed by ex vivo PERIOD 2::LUCIFERASE (PER2::LUC) bioluminescent imaging to test whether activation of this SCN neuron sub-population can induce SCN network changes that are hallmarks of photoperiodic encoding. We found that optogenetic stimulation designed to mimic a long photoperiod indeed altered subsequent SCN entrained phase, increased the phase dispersal of PER2 rhythms within the SCN network, and shortened SCN free-running period – similar to the effects of a true extension of photoperiod. Optogenetic stimulation also induced analogous changes on related aspects of locomotor behavior in vivo. Thus, selective activation of VIPergic SCN neurons induces photoperiodic network plasticity in the SCN which underpins photoperiodic entrainment of behavior.

scholarly journals Cyclin Dependent Kinase 5 (CDK5) Regulates the Circadian Clock

2019 ◽  
Author(s):  
Andrea Brenna ◽  
Iwona Olejniczak ◽  
Rohit Chavan ◽  
Jürgen A. Ripperger ◽  
Sonja Langmesser ◽  
...  
Keyword(s):  
Circadian Clock ◽  
Cyclin Dependent Kinase ◽  
Suprachiasmatic Nuclei ◽  
Nuclear Entry ◽  
Period 2 ◽  
Clock Component ◽  
Free Running ◽  
Free Running Period ◽  
Cryptochrome 1 ◽  
Cyclin Dependent Kinase 5

AbstractCircadian oscillations emerge from transcriptional and post-translational feedback loops. An important step in generating rhythmicity is the translocation of clock components into the nucleus, which is regulated in many cases by kinases. In mammals, the kinase promoting the nuclear import of the key clock component Period 2 (PER2) is unknown. Here we show that the cyclin-dependent kinase 5 (CDK5) regulates the mammalian circadian clock involving phosphorylation of PER2. Knock-down of Cdk5 in the suprachiasmatic nuclei (SCN), the main coordinator site of the mammalian circadian system, shortened the free-running period in mice. CDK5 phosphorylated PER2 at serine residue 394 (S394) in a diurnal fashion. This phosphorylation facilitated interaction with Cryptochrome 1 (CRY1) and nuclear entry of the PER2-CRY1 complex. Taken together, we found that CDK5 drives nuclear entry of PER2, which is critical for establishing an adequate circadian period of the molecular circadian cycle. Therefore, CDK5 is critically involved in the regulation of the circadian clock and may represent a link to various diseases affected by the circadian clock.

A New Model for Quantitative In Vivo Microscopic Analysis of Thrombus Formation and Vascular Recanalisation: The Ear of the Hairless (hr/hr) Mouse

1997 ◽  
Vol 78 (05) ◽  
pp. 1408-1414 ◽  
Author(s):  
Frank Roesken ◽  
Martin Ruecker ◽  
Brigitte Vollmar ◽  
Nicole Boeckel ◽  
Eberhard Morgenstern ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Light Exposure ◽  
Thrombus Formation ◽  
Microscopic Analysis ◽  
Cell Interaction ◽  
Vascular Perfusion ◽  
Vessel Occlusion ◽  
Endothelial Cell Interaction ◽  
Platelet Deposition

SummaryThe alteration of rheological blood properties as well as deterioration of vascular perfusion conditions and cell-cell interactions are major determinants of thrombus formation. Herein, we present an experimental model which allows for quantitative in vivo microscopic analysis of these determinants during both thrombus formation and vascular recanalisation. The model does not require surgical preparation procedures, and enables for repeated analysis of identical microvessels over time periods of days or months, respectively. After i.v. administration of FITC-dextran thrombus formation was induced photochemically by light exposure to individual arterioles and venules of the ear of ten anaesthetised hairless mice. In venules, epiillumination induced rapid thrombus formation with first platelet deposition after 0.59 ± 0.04 min and complete vessel occlusion within 7.48 ±1.31 min. After a 24-h time period, 75% of the thrombosed venules were found recanalised. Marked leukocyte-endothelial cell interaction in those venules indicated persistent endothelial cell activation and/or injury, even after an observation period of 7 days. In arterioles, epi-illumination provoked vasomotion, while thrombus formation was significantly (p <0.05) delayed with first platelet deposition after 2.32 ± 0.22 min and complete vessel occlusion within 20.07 ±3.84 min. Strikingly, only one of the investigated arterioles was found recanalised after 24 h, which, however, did not show leukocyte-endothelial cell interaction. Heparin (300 U/kg, i.v.) effectively counteracted the process of thrombus formation in this model, including both first platelet deposition and vessel occlusion. We conclude that the model of the ear of the hairless mouse allows for distinct in vivo analysis of arteriolar and venular thrombus formation/ recanalisation, and, thus, represents an interesting tool for the study of novel antithrombotic and thrombolytic strategies, respectively.

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