Circadian expression of clock and putative clock-controlled genes in skeletal muscle of the zebrafish.

PubMed

Amaral, Ian P G; Johnston, Ian A

2012-01-01

To identify circadian patterns of gene expression in skeletal muscle, adult male zebrafish were acclimated for 2 wk to a 12:12-h light-dark photoperiod and then exposed to continuous darkness for 86 h with ad libitum feeding. The increase in gut food content associated with the subjective light period was much diminished by the third cycle, enabling feeding and circadian rhythms to be distinguished. Expression of zebrafish paralogs of mammalian transcriptional activators of the circadian mechanism (bmal1, clock1, and rora) followed a rhythmic pattern with a ∼24-h periodicity. Peak expression of rora paralogs occurred at the beginning of the subjective light period [Zeitgeber time (ZT)07 and ZT02 for roraa and rorab], whereas the highest expression of bmal1 and clock paralogs occurred 12 h later (ZT13-15 and ZT16 for bmal and clock paralogs). Expression of the transcriptional repressors cry1a, per1a/1b, per2, per3, nr1d2a/2b, and nr1d1 also followed a circadian pattern with peak expression at ZT0-02. Expression of the two paralogs of cry2 occurred in phase with clock1a/1b. Duplicated genes had a high correlation of expression except for paralogs of clock1, nr1d2, and per1, with cry1b showing no circadian pattern. The highest expression difference was 9.2-fold for the activator bmal1b and 51.7-fold for the repressor per1a. Out of 32 candidate clock-controlled genes, only myf6, igfbp3, igfbp5b, and hsf2 showed circadian expression patterns. Igfbp3, igfbp5b, and myf6 were expressed in phase with clock1a/1b and had an average of twofold change in expression from peak to trough, whereas hsf2 transcripts were expressed in phase with cry1a and had a 7.2-fold-change in expression. The changes in expression of clock and clock-controlled genes observed during continuous darkness were also observed at similar ZTs in fish exposed to a normal photoperiod in a separate control experiment. The role of circadian clocks in regulating muscle maintenance and growth are discussed.

Impaired light detection of the circadian clock in a zebrafish melanoma model.

PubMed

Hamilton, Noémie; Diaz-de-Cerio, Natalia; Whitmore, David

2015-01-01

The circadian clock controls the timing of the cell cycle in healthy tissues and clock disruption is known to increase tumourigenesis. Melanoma is one of the most rapidly increasing forms of cancer and the precise molecular circadian changes that occur in a melanoma tumor are unknown. Using a melanoma zebrafish model, we have explored the molecular changes that occur to the circadian clock within tumors. We have found disruptions in melanoma clock gene expression due to a major impairment to the light input pathway, with a parallel loss of light-dependent activation of DNA repair genes. Furthermore, the timing of mitosis in tumors is perturbed, as well as the regulation of certain key cell cycle regulators, such that cells divide arhythmically. The inability to co-ordinate DNA damage repair and cell division is likely to promote further tumourigenesis and accelerate melanoma development.

Impaired light detection of the circadian clock in a zebrafish melanoma model

PubMed Central

Hamilton, Noémie; Diaz-de-Cerio, Natalia; Whitmore, David

2015-01-01

The circadian clock controls the timing of the cell cycle in healthy tissues and clock disruption is known to increase tumourigenesis. Melanoma is one of the most rapidly increasing forms of cancer and the precise molecular circadian changes that occur in a melanoma tumor are unknown. Using a melanoma zebrafish model, we have explored the molecular changes that occur to the circadian clock within tumors. We have found disruptions in melanoma clock gene expression due to a major impairment to the light input pathway, with a parallel loss of light-dependent activation of DNA repair genes. Furthermore, the timing of mitosis in tumors is perturbed, as well as the regulation of certain key cell cycle regulators, such that cells divide arhythmically. The inability to co-ordinate DNA damage repair and cell division is likely to promote further tumourigenesis and accelerate melanoma development. PMID:25832911

The Importance of Stochastic Effects for Explaining Entrainment in the Zebrafish Circadian Clock.

PubMed

Heussen, Raphaela; Whitmore, David

2015-01-01

The circadian clock plays a pivotal role in modulating physiological processes and has been implicated, either directly or indirectly, in a range of pathological states including cancer. Here we investigate how the circadian clock is entrained by external cues such as light. Working with zebrafish cell lines and combining light pulse experiments with simulation efforts focused on the role of synchronization effects, we find that even very modest doses of light exposure are sufficient to trigger some entrainment, whereby a higher light intensity or duration correlates with strength of the circadian signal. Moreover, we observe in the simulations that stochastic effects may be considered an essential feature of the circadian clock in order to explain the circadian signal decay in prolonged darkness, as well as light initiated resynchronization as a strong component of entrainment.

Timing Embryo Segmentation: Dynamics and Regulatory Mechanisms of the Vertebrate Segmentation Clock

PubMed Central

Resende, Tatiana P.; Andrade, Raquel P.; Palmeirim, Isabel

2014-01-01

All vertebrate species present a segmented body, easily observed in the vertebrate column and its associated components, which provides a high degree of motility to the adult body and efficient protection of the internal organs. The sequential formation of the segmented precursors of the vertebral column during embryonic development, the somites, is governed by an oscillating genetic network, the somitogenesis molecular clock. Herein, we provide an overview of the molecular clock operating during somite formation and its underlying molecular regulatory mechanisms. Human congenital vertebral malformations have been associated with perturbations in these oscillatory mechanisms. Thus, a better comprehension of the molecular mechanisms regulating somite formation is required in order to fully understand the origin of human skeletal malformations. PMID:24895605

Analyzing notochord segmentation and intervertebral disc formation using the twhh:gfp transgenic zebrafish model.

PubMed

Haga, Yutaka; Dominique, Vincent J; Du, Shao Jun

2009-10-01

To characterize the process of vertebral segmentation and disc formation in living animals, we analyzed tiggy-winkle hedgehog (twhh):green fluorescent protein (gfp) and sonic hedgehog (shh):gfp transgenic zebrafish models that display notochord-specific GFP expression. We found that they showed distinct patterns of expression in the intervertebral discs of late stage fish larvae and adult zebrafish. A segmented pattern of GFP expression was detected in the intervertebral disc of twhh:gfp transgenic fish. In contrast, little GFP expression was found in the intervertebral disc of shh:gfp transgenic fish. Treating twhh:gfp transgenic zebrafish larvae with exogenous retinoic acid (RA), a teratogenic factor on normal development, resulted in disruption of notochord segmentation and formation of oversized vertebrae. Histological analysis revealed that the oversized vertebrae are likely due to vertebral fusion. These studies demonstrate that the twhh:gfp transgenic zebrafish is a useful model for studying vertebral segmentation and disc formation, and moreover, that RA signaling may play a role in this process.

Rhodopsin expression in the zebrafish pineal gland from larval to adult stage.

PubMed

Magnoli, Domenico; Zichichi, Rosalia; Laurà, Rosaria; Guerrera, Maria Cristina; Campo, Salvatore; de Carlos, Felix; Suárez, Alberto Álvarez; Abbate, Francesco; Ciriaco, Emilia; Vega, Jose Antonio; Germanà, Antonino

2012-03-09

The zebrafish pineal gland plays an important role in different physiological functions including the regulation of the circadian clock. In the fish pineal gland the pinealocytes are made up of different segments: outer segment, inner segment and basal pole. Particularly, in the outer segment the rhodopsin participates in the external environment light reception that represents the first biochemical step in the melatonin production. It is well known that the rhodopsin in the adult zebrafish is well expressed in the pineal gland but both the expression and the cellular localization of this protein during development remain still unclear. In this study using qRT-PCR, sequencing and immunohistochemistry the expression as well as the protein localization of the rhodopsin in the zebrafish from larval (10 dpf) to adult stage (90 dpf) were demonstrated. The rhodopsin mRNA expression presents a peak of expression at 10 dpf, a further reduction to 50 dpf before increasing again in the adult stage. Moreover, the cellular localization of the rhodopsin-like protein was always localized in the pinealocyte at all ages examined. Our results demonstrated the involvement of the rhodopsin in the zebrafish pineal gland physiology particularly in the light capture during the zebrafish lifespan. Copyright © 2012 Elsevier B.V. All rights reserved.

Chevron formation of the zebrafish muscle segments

PubMed Central

Rost, Fabian; Eugster, Christina; Schröter, Christian; Oates, Andrew C.; Brusch, Lutz

2014-01-01

The muscle segments of fish have a folded shape, termed a chevron, which is thought to be optimal for the undulating body movements of swimming. However, the mechanism shaping the chevron during embryogenesis is not understood. Here, we used time-lapse microscopy of developing zebrafish embryos spanning the entire somitogenesis period to quantify the dynamics of chevron shape development. By comparing such time courses with the start of movements in wildtype zebrafish and analysing immobile mutants, we show that the previously implicated body movements do not play a role in chevron formation. Further, the monotonic increase of chevron angle along the anteroposterior axis revealed by our data constrains or rules out possible contributions by previously proposed mechanisms. In particular, we found that muscle pioneers are not required for chevron formation. We put forward a tension-and-resistance mechanism involving interactions between intra-segmental tension and segment boundaries. To evaluate this mechanism, we derived and analysed a mechanical model of a chain of contractile and resisting elements. The predictions of this model were verified by comparison with experimental data. Altogether, our results support the notion that a simple physical mechanism suffices to self-organize the observed spatiotemporal pattern in chevron formation. PMID:25267843

Chevron formation of the zebrafish muscle segments.

PubMed

Rost, Fabian; Eugster, Christina; Schröter, Christian; Oates, Andrew C; Brusch, Lutz

2014-11-01

The muscle segments of fish have a folded shape, termed a chevron, which is thought to be optimal for the undulating body movements of swimming. However, the mechanism shaping the chevron during embryogenesis is not understood. Here, we used time-lapse microscopy of developing zebrafish embryos spanning the entire somitogenesis period to quantify the dynamics of chevron shape development. By comparing such time courses with the start of movements in wildtype zebrafish and analysing immobile mutants, we show that the previously implicated body movements do not play a role in chevron formation. Further, the monotonic increase of chevron angle along the anteroposterior axis revealed by our data constrains or rules out possible contributions by previously proposed mechanisms. In particular, we found that muscle pioneers are not required for chevron formation. We put forward a tension-and-resistance mechanism involving interactions between intra-segmental tension and segment boundaries. To evaluate this mechanism, we derived and analysed a mechanical model of a chain of contractile and resisting elements. The predictions of this model were verified by comparison with experimental data. Altogether, our results support the notion that a simple physical mechanism suffices to self-organize the observed spatiotemporal pattern in chevron formation. © 2014. Published by The Company of Biologists Ltd.

Faster embryonic segmentation through elevated Delta-Notch signalling

PubMed Central

Liao, Bo-Kai; Jörg, David J.; Oates, Andrew C.

2016-01-01

An important step in understanding biological rhythms is the control of period. A multicellular, rhythmic patterning system termed the segmentation clock is thought to govern the sequential production of the vertebrate embryo's body segments, the somites. Several genetic loss-of-function conditions, including the Delta-Notch intercellular signalling mutants, result in slower segmentation. Here, we generate DeltaD transgenic zebrafish lines with a range of copy numbers and correspondingly increased signalling levels, and observe faster segmentation. The highest-expressing line shows an altered oscillating gene expression wave pattern and shortened segmentation period, producing embryos with more, shorter body segments. Our results reveal surprising differences in how Notch signalling strength is quantitatively interpreted in different organ systems, and suggest a role for intercellular communication in regulating the output period of the segmentation clock by altering its spatial pattern. PMID:27302627

mir-125a-5p-mediated Regulation of Lfng is Essential for the Avian Segmentation Clock

PubMed Central

Riley, Maurisa F.; Bochter, Matthew S.; Wahi, Kanu; Nuovo, Gerard J.; Cole, Susan E.

2013-01-01

Summary Somites are embryonic precursors of the axial skeleton and skeletal muscles, and establish the segmental vertebrate body plan. Somitogenesis is controlled in part by a segmentation clock that requires oscillatory expression of genes including Lunatic fringe (Lfng). Oscillatory genes must be tightly regulated both at the transcriptional and post-transcriptional levels for proper clock function. Here we demonstrate that microRNA-mediated regulation of Lfng is essential for proper segmentation during chick somitogenesis. We find that mir-125a-5p targets evolutionarily conserved sequences in the Lfng 3′UTR, and that preventing interactions between mir-125a-5p and Lfng transcripts in vivo causes abnormal segmentation and perturbs clock activity. This provides strong evidence that miRNAs function in the post-transcriptional regulation of oscillatory genes in the segmentation clock. Further, this demonstrates that the relatively subtle effects of miRNAs on target genes can have broad effects in developmental situations that have critical requirements for tight post-transcriptional regulation. PMID:23484856

The circadian clock regulates autophagy directly through the nuclear hormone receptor Nr1d1/Rev-erbα and indirectly via Cebpb/(C/ebpβ) in zebrafish.

PubMed

Huang, Guodong; Zhang, Fanmiao; Ye, Qiang; Wang, Han

2016-08-02

Autophagy is a highly conserved intracellular degradation system, and recently was shown to display circadian rhythms in mice. The mechanisms underlying circadian regulation of autophagy, however, are still unclear. Here, we observed that numbers of autophagosomes and autolysosomes exhibit daily rhythms in the zebrafish liver, and cebpb/(c/ebpβ) and various autophagy genes are rhythmically expressed in zebrafish larvae but significantly upregulated in per1b and TALEN-generated nr1d1/rev-erbα mutant fish, indicating that both Per1b and Nr1d1 play critical roles in autophagy rhythms. Luciferase reporter and ChIP assays show that the circadian clock directly regulates autophagy genes through Nr1d1, and also regulates transcription of cebpb through Per1b. We also found that fasting leads to altered expression of both circadian clock genes and autophagy genes in zebrafish adult peripheral organs. Further, transcriptome analysis reveals multiple functions of Nr1d1 in zebrafish. Taken together, these findings provide evidence for how the circadian clock regulates autophagy, imply that nutritional signaling affects both circadian regulation and autophagy activities in peripheral organs, and shed light on how circadian gene mutations act through autophagy to contribute to common metabolic diseases such as obesity.

Abnormal photoreceptor outer segment development and early retinal degeneration in kif3a mutant zebrafish.

PubMed

Raghupathy, Rakesh K; Zhang, Xun; Alhasani, Reem H; Zhou, Xinzhi; Mullin, Margaret; Reilly, James; Li, Wenchang; Liu, Mugen; Shu, Xinhua

2016-08-01

Photoreceptors are highly specialized sensory neurons that possess a modified primary cilium called the outer segment. Photoreceptor outer segment formation and maintenance require highly active protein transport via a process known as intraflagellar transport. Anterograde transport in outer segments is powered by the heterotrimeric kinesin II and coordinated by intraflagellar transport proteins. Here, we describe a new zebrafish model carrying a nonsense mutation in the kinesin II family member 3A (kif3a) gene. Kif3a mutant zebrafish exhibited curved body axes and kidney cysts. Outer segments were not formed in most parts of the mutant retina, and rhodopsin was mislocalized, suggesting KIF3A has a role in rhodopsin trafficking. Both rod and cone photoreceptors degenerated rapidly between 4 and 9 days post fertilization, and electroretinography response was not detected in 7 days post fertilization mutant larvae. Loss of KIF3A in zebrafish also resulted in an intracellular transport defect affecting anterograde but not retrograde transport of organelles. Our results indicate KIF3A plays a conserved role in photoreceptor outer segment formation and intracellular transport. Copyright © 2016 John Wiley & Sons, Ltd.

Circadian Clock Synchronization of the Cell Cycle in Zebrafish Occurs through a Gating Mechanism Rather Than a Period-phase Locking Process.

PubMed

Laranjeiro, Ricardo; Tamai, T Katherine; Letton, William; Hamilton, Noémie; Whitmore, David

2018-04-01

Studies from a number of model systems have shown that the circadian clock controls expression of key cell cycle checkpoints, thus providing permissive or inhibitory windows in which specific cell cycle events can occur. However, a major question remains: Is the clock actually regulating the cell cycle through such a gating mechanism or, alternatively, is there a coupling process that controls the speed of cell cycle progression? Using our light-responsive zebrafish cell lines, we address this issue directly by synchronizing the cell cycle in culture simply by changing the entraining light-dark (LD) cycle in the incubator without the need for pharmacological intervention. Our results show that the cell cycle rapidly reentrains to a shifted LD cycle within 36 h, with changes in p21 expression and subsequent S phase timing occurring within the first few hours of resetting. Reentrainment of mitosis appears to lag S phase resetting by 1 circadian cycle. The range of entrainment of the zebrafish clock to differing LD cycles is large, from 16 to 32 hour periods. We exploited this feature to explore cell cycle entrainment at both the population and single cell levels. At the population level, cell cycle length is shortened or lengthened under corresponding T-cycles, suggesting that a 1:1 coupling mechanism is capable of either speeding up or slowing down the cell cycle. However, analysis at the single cell level reveals that this, in fact, is not true and that a gating mechanism is the fundamental method of timed cell cycle regulation in zebrafish. Cell cycle length at the single cell level is virtually unaltered with varying T-cycles.

Circadian Clock Synchronization of the Cell Cycle in Zebrafish Occurs through a Gating Mechanism Rather Than a Period-phase Locking Process

PubMed Central

Tamai, T. Katherine; Letton, William; Hamilton, Noémie; Whitmore, David

2018-01-01

Studies from a number of model systems have shown that the circadian clock controls expression of key cell cycle checkpoints, thus providing permissive or inhibitory windows in which specific cell cycle events can occur. However, a major question remains: Is the clock actually regulating the cell cycle through such a gating mechanism or, alternatively, is there a coupling process that controls the speed of cell cycle progression? Using our light-responsive zebrafish cell lines, we address this issue directly by synchronizing the cell cycle in culture simply by changing the entraining light-dark (LD) cycle in the incubator without the need for pharmacological intervention. Our results show that the cell cycle rapidly reentrains to a shifted LD cycle within 36 h, with changes in p21 expression and subsequent S phase timing occurring within the first few hours of resetting. Reentrainment of mitosis appears to lag S phase resetting by 1 circadian cycle. The range of entrainment of the zebrafish clock to differing LD cycles is large, from 16 to 32 hour periods. We exploited this feature to explore cell cycle entrainment at both the population and single cell levels. At the population level, cell cycle length is shortened or lengthened under corresponding T-cycles, suggesting that a 1:1 coupling mechanism is capable of either speeding up or slowing down the cell cycle. However, analysis at the single cell level reveals that this, in fact, is not true and that a gating mechanism is the fundamental method of timed cell cycle regulation in zebrafish. Cell cycle length at the single cell level is virtually unaltered with varying T-cycles. PMID:29444612

A Blind Circadian Clock in Cavefish Reveals that Opsins Mediate Peripheral Clock Photoreception

PubMed Central

Cavallari, Nicola; Frigato, Elena; Vallone, Daniela; Fröhlich, Nadine; Lopez-Olmeda, Jose Fernando; Foà, Augusto; Berti, Roberto; Sánchez-Vázquez, Francisco Javier; Bertolucci, Cristiano; Foulkes, Nicholas S.

2011-01-01

The circadian clock is synchronized with the day-night cycle primarily by light. Fish represent fascinating models for deciphering the light input pathway to the vertebrate clock since fish cell clocks are regulated by direct light exposure. Here we have performed a comparative, functional analysis of the circadian clock involving the zebrafish that is normally exposed to the day-night cycle and a cavefish species that has evolved in perpetual darkness. Our results reveal that the cavefish retains a food-entrainable clock that oscillates with an infradian period. Importantly, however, this clock is not regulated by light. This comparative study pinpoints the two extra-retinal photoreceptors Melanopsin (Opn4m2) and TMT-opsin as essential upstream elements of the peripheral clock light input pathway. PMID:21909239

Light directs zebrafish period2 expression via conserved D and E boxes.

PubMed

Vatine, Gad; Vallone, Daniela; Appelbaum, Lior; Mracek, Philipp; Ben-Moshe, Zohar; Lahiri, Kajori; Gothilf, Yoav; Foulkes, Nicholas S

2009-10-01

For most species, light represents the principal environmental signal for entraining the endogenous circadian clock. The zebrafish is a fascinating vertebrate model for studying this process since unlike mammals, direct exposure of most of its tissues to light leads to local clock entrainment. Importantly, light induces the expression of a set of genes including certain clock genes in most zebrafish cell types in vivo and in vitro. However, the mechanism linking light to gene expression remains poorly understood. To elucidate this key mechanism, here we focus on how light regulates transcription of the zebrafish period2 (per2) gene. Using transgenic fish and stably transfected cell line-based assays, we define a Light Responsive Module (LRM) within the per2 promoter. The LRM lies proximal to the transcription start site and is both necessary and sufficient for light-driven gene expression and also for a light-dependent circadian clock regulation. Curiously, the LRM sequence is strongly conserved in other vertebrate per2 genes, even in species lacking directly light-sensitive peripheral clocks. Furthermore, we reveal that the human LRM can substitute for the zebrafish LRM to confer light-regulated transcription in zebrafish cells. The LRM contains E- and D-box elements that are critical for its function. While the E-box directs circadian clock regulation by mediating BMAL/CLOCK activity, the D-box confers light-driven expression. The zebrafish homolog of the thyrotroph embryonic factor binds efficiently to the LRM D-box and transactivates expression. We demonstrate that tef mRNA levels are light inducible and that knock-down of tef expression attenuates light-driven transcription from the per2 promoter in vivo. Together, our results support a model where a light-dependent crosstalk between E- and D-box binding factors is a central determinant of per2 expression. These findings extend the general understanding of the mechanism whereby the clock is entrained by light

Modelling Delta-Notch perturbations during zebrafish somitogenesis.

PubMed

Murray, Philip J; Maini, Philip K; Baker, Ruth E

2013-01-15

The discovery over the last 15 years of molecular clocks and gradients in the pre-somitic mesoderm of numerous vertebrate species has added significant weight to Cooke and Zeeman's 'clock and wavefront' model of somitogenesis, in which a travelling wavefront determines the spatial position of somite formation and the somitogenesis clock controls periodicity (Cooke and Zeeman, 1976). However, recent high-throughput measurements of spatiotemporal patterns of gene expression in different zebrafish mutant backgrounds allow further quantitative evaluation of the clock and wavefront hypothesis. In this study we describe how our recently proposed model, in which oscillator coupling drives the propagation of an emergent wavefront, can be used to provide mechanistic and testable explanations for the following observed phenomena in zebrafish embryos: (a) the variation in somite measurements across a number of zebrafish mutants; (b) the delayed formation of somites and the formation of 'salt and pepper' patterns of gene expression upon disruption of oscillator coupling; and (c) spatial correlations in the 'salt and pepper' patterns in Delta-Notch mutants. In light of our results, we propose a number of plausible experiments that could be used to further test the model. Copyright © 2012 Elsevier Inc. All rights reserved.

Circadian rhythmicity and light sensitivity of the zebrafish brain.

PubMed

Moore, Helen A; Whitmore, David

2014-01-01

Traditionally, circadian clocks have been thought of as a neurobiological phenomenon. This view changed somewhat over recent years with the discovery of peripheral tissue circadian oscillators. In mammals, however, the suprachiasmatic nucleus (SCN) in the hypothalamus still retains the critical role of a central synchronizer of biological timing. Zebrafish, in contrast, have always reflected a more highly decentralized level of clock organization, as individual cells and tissues contain directly light responsive circadian pacemakers. As a consequence, clock function in the zebrafish brain has remained largely unexplored, and the precise organization of rhythmic and light-sensitive neurons within the brain is unknown. To address this issue, we used the period3 (per3)-luciferase transgenic zebrafish to confirm that multiple brain regions contain endogenous circadian oscillators that are directly light responsive. In addition, in situ hybridization revealed localised neural expression of several rhythmic and light responsive clock genes, including per3, cryptochrome1a (cry1a) and per2. Adult brain nuclei showing significant clock gene expression include the teleost equivalent of the SCN, as well as numerous hypothalamic nuclei, the periventricular grey zone (PGZ) of the optic tectum, and granular cells of the rhombencephalon. To further investigate the light sensitive properties of neurons, expression of c-fos, a marker for neuronal activity, was examined. c-fos mRNA was upregulated in response to changing light conditions in different nuclei within the zebrafish brain. Furthermore, under constant dark (DD) conditions, c-fos shows a significant circadian oscillation. Taken together, these results show that there are numerous areas of the zebrafish central nervous system, which contain deep brain photoreceptors and directly light-entrainable circadian pacemakers. However, there are also multiple brain nuclei, which possess neither, demonstrating a degree of pacemaker

Circadian Rhythmicity and Light Sensitivity of the Zebrafish Brain

PubMed Central

Moore, Helen A.; Whitmore, David

2014-01-01

Traditionally, circadian clocks have been thought of as a neurobiological phenomenon. This view changed somewhat over recent years with the discovery of peripheral tissue circadian oscillators. In mammals, however, the suprachiasmatic nucleus (SCN) in the hypothalamus still retains the critical role of a central synchronizer of biological timing. Zebrafish, in contrast, have always reflected a more highly decentralized level of clock organization, as individual cells and tissues contain directly light responsive circadian pacemakers. As a consequence, clock function in the zebrafish brain has remained largely unexplored, and the precise organization of rhythmic and light-sensitive neurons within the brain is unknown. To address this issue, we used the period3 (per3)-luciferase transgenic zebrafish to confirm that multiple brain regions contain endogenous circadian oscillators that are directly light responsive. In addition, in situ hybridization revealed localised neural expression of several rhythmic and light responsive clock genes, including per3, cryptochrome1a (cry1a) and per2. Adult brain nuclei showing significant clock gene expression include the teleost equivalent of the SCN, as well as numerous hypothalamic nuclei, the periventricular grey zone (PGZ) of the optic tectum, and granular cells of the rhombencephalon. To further investigate the light sensitive properties of neurons, expression of c-fos, a marker for neuronal activity, was examined. c-fos mRNA was upregulated in response to changing light conditions in different nuclei within the zebrafish brain. Furthermore, under constant dark (DD) conditions, c-fos shows a significant circadian oscillation. Taken together, these results show that there are numerous areas of the zebrafish central nervous system, which contain deep brain photoreceptors and directly light-entrainable circadian pacemakers. However, there are also multiple brain nuclei, which possess neither, demonstrating a degree of pacemaker

Multiple PAR and E4BP4 bZIP transcription factors in zebrafish: diverse spatial and temporal expression patterns.

PubMed

Ben-Moshe, Zohar; Vatine, Gad; Alon, Shahar; Tovin, Adi; Mracek, Philipp; Foulkes, Nicholas S; Gothilf, Yoav

2010-09-01

Circadian rhythms of physiology and behavior are generated by an autonomous circadian oscillator that is synchronized daily with the environment, mainly by light input. The PAR subfamily of transcriptional activators and the related E4BP4 repressor belonging to the basic leucine zipper (bZIP) family are clock-controlled genes that are suggested to mediate downstream circadian clock processes and to feedback onto the core oscillator. Here, the authors report the characterization of these genes in the zebrafish, an increasingly important model in the field of chronobiology. Five novel PAR and six novel e4bp4 zebrafish homolog genes were identified using bioinformatic tools and their coding sequences were cloned. Based on their evolutionary relationships, these genes were annotated as ztef2, zhlf1 and zhlf2, zdbp1 and zdbp2, and ze4bp4-1 to -6. The spatial and temporal mRNA expression pattern of each of these factors was characterized in zebrafish embryos in the context of a functional circadian clock and regulation by light. Nine of the factors exhibited augmented and rhythmic expression in the pineal gland, a central clock organ in zebrafish. Moreover, these genes were found to be regulated, to variable extents, by the circadian clock and/or by light. Differential expression patterns of multiple paralogs in zebrafish suggest multiple roles for these factors within the vertebrate circadian clock. This study, in the genetically accessible zebrafish model, lays the foundation for further research regarding the involvement and specific roles of PAR and E4BP4 transcription factors in the vertebrate circadian clock mechanism.

The clock and wavefront model revisited.

PubMed

Murray, Philip J; Maini, Philip K; Baker, Ruth E

2011-08-21

The currently accepted interpretation of the clock and wavefront model of somitogenesis is that a posteriorly moving molecular gradient sequentially slows the rate of clock oscillations, resulting in a spatial readout of temporal oscillations. However, while molecular components of the clocks and wavefronts have now been identified in the pre-somitic mesoderm (PSM), there is not yet conclusive evidence demonstrating that the observed molecular wavefronts act to slow clock oscillations. Here we present an alternative formulation of the clock and wavefront model in which oscillator coupling, already known to play a key role in oscillator synchronisation, plays a fundamentally important role in the slowing of oscillations along the anterior-posterior (AP) axis. Our model has three parameters which can be determined, in any given species, by the measurement of three quantities: the clock period in the posterior PSM, somite length and the length of the PSM. A travelling wavefront, which slows oscillations along the AP axis, is an emergent feature of the model. Using the model we predict: (a) the distance between moving stripes of gene expression; (b) the number of moving stripes of gene expression and (c) the oscillator period profile along the AP axis. Predictions regarding the stripe data are verified using existing zebrafish data. We simulate a range of experimental perturbations and demonstrate how the model can be used to unambiguously define a reference frame along the AP axis. Comparing data from zebrafish, chick, mouse and snake, we demonstrate that: (a) variation in patterning profiles is accounted for by a single nondimensional parameter; the ratio of coupling strengths; and (b) the period profile along the AP axis is conserved across species. Thus the model is consistent with the idea that, although the genes involved in pattern propagation in the PSM vary, there is a conserved patterning mechanism across species. Copyright © 2011 Elsevier Ltd. All rights

Differential maturation of rhythmic clock gene expression during early development in medaka (Oryzias latipes).

PubMed

Cuesta, Ines H; Lahiri, Kajori; Lopez-Olmeda, Jose Fernando; Loosli, Felix; Foulkes, Nicholas S; Vallone, Daniela

2014-05-01

One key challenge for the field of chronobiology is to identify how circadian clock function emerges during early embryonic development. Teleosts such as the zebrafish are ideal models for studying circadian clock ontogeny since the entire process of development occurs ex utero in an optically transparent chorion. Medaka (Oryzias latipes) represents another powerful fish model for exploring early clock function with, like the zebrafish, many tools available for detailed genetic analysis. However, to date there have been no reports documenting circadian clock gene expression during medaka development. Here we have characterized the expression of key clock genes in various developmental stages and in adult tissues of medaka. As previously reported for other fish, light dark cycles are required for the emergence of clock gene expression rhythms in this species. While rhythmic expression of per and cry genes is detected very early during development and seems to be light driven, rhythmic clock and bmal expression appears much later around hatching time. Furthermore, the maturation of clock function seems to correlate with the appearance of rhythmic expression of these positive elements of the clock feedback loop. By accelerating development through elevated temperatures or by artificially removing the chorion, we show an earlier onset of rhythmicity in clock and bmal expression. Thus, differential maturation of key elements of the medaka clock mechanism depends on the developmental stage and the presence of the chorion.

Caudal migration and proliferation of renal progenitors regulates early nephron segment size in zebrafish.

PubMed

Naylor, Richard W; Dodd, Rachel C; Davidson, Alan J

2016-10-19

The nephron is the functional unit of the kidney and is divided into distinct proximal and distal segments. The factors determining nephron segment size are not fully understood. In zebrafish, the embryonic kidney has long been thought to differentiate in situ into two proximal tubule segments and two distal tubule segments (distal early; DE, and distal late; DL) with little involvement of cell movement. Here, we overturn this notion by performing lineage-labelling experiments that reveal extensive caudal movement of the proximal and DE segments and a concomitant compaction of the DL segment as it fuses with the cloaca. Laser-mediated severing of the tubule, such that the DE and DL are disconnected or that the DL and cloaca do not fuse, results in a reduction in tubule cell proliferation and significantly shortens the DE segment while the caudal movement of the DL is unaffected. These results suggest that the DL mechanically pulls the more proximal segments, thereby driving both their caudal extension and their proliferation. Together, these data provide new insights into early nephron morphogenesis and demonstrate the importance of cell movement and proliferation in determining initial nephron segment size.

spiel ohne grenzen/pou2 is required for zebrafish hindbrain segmentation.

PubMed

Hauptmann, Giselbert; Belting, Heinz-Georg; Wolke, Uta; Lunde, Karen; Söll, Iris; Abdelilah-Seyfried, Salim; Prince, Victoria; Driever, Wolfgang

2002-04-01

Segmentation of the vertebrate hindbrain leads to the formation of a series of rhombomeres with distinct identities. In mouse, Krox20 and kreisler play important roles in specifying distinct rhombomeres and in controlling segmental identity by directly regulating rhombomere-specific expression of Hox genes. We show that spiel ohne grenzen (spg) zebrafish mutants develop rhombomeric territories that are abnormal in both size and shape. Rhombomere boundaries are malpositioned or absent and the segmental pattern of neuronal differentiation is perturbed. Segment-specific expression of hoxa2, hoxb2 and hoxb3 is severely affected during initial stages of hindbrain development in spg mutants and the establishment of krx20 (Krox20 ortholog) and valentino (val; kreisler ortholog) expression is impaired. spg mutants carry loss-of-function mutations in the pou2 gene. pou2 is expressed at high levels in the hindbrain primordium of wild-type embryos prior to activation of krx20 and val. Widespread overexpression of Pou2 can rescue the segmental krx20 and val domains in spg mutants, but does not induce ectopic expression of these genes. This suggests that spg/pou2 acts in a permissive manner and is essential for normal expression of krx20 and val. We propose that spg/pou2 is an essential component of the regulatory cascade controlling hindbrain segmentation and acts before krx20 and val in the establishment of rhombomere precursor territories.

beamter/deltaC and the role of Notch ligands in the zebrafish somite segmentation, hindbrain neurogenesis and hypochord differentiation.

PubMed

Jülich, Dörthe; Hwee Lim, Chiaw; Round, Jennifer; Nicolaije, Claudia; Schroeder, Joshua; Davies, Alexander; Geisler, Robert; Lewis, Julian; Jiang, Yun-Jin; Holley, Scott A

2005-10-15

The Tübingen large-scale zebrafish genetic screen completed in 1996 identified a set of five genes required for orderly somite segmentation. Four of them have been molecularly identified and three were found to code for components of the Notch pathway, which are required for the coordinated oscillation of gene expression, known as the segmentation clock, in the presomitic mesoderm (PSM). Here, we show that the final member of the group, beamter (bea), codes for the Notch ligand DeltaC, and we present and characterize two new alleles, including one allele encoding for a protein truncated in the 7th EGF repeat and an allele deleting only the DSL domain which was previously shown to be necessary for ligand function. Interestingly however, when we over-express any of the mutant deltaC mRNAs, we observe antimorphic effects on both hindbrain neurogenesis and hypochord formation. Expression of bea/deltaC oscillates in the PSM, and a triple fluorescent in situ analysis of its oscillation in relation to that of other oscillating genes in the PSM reveals differences in subcellular localization of the oscillating mRNAs in individual cells in different oscillation phases. Mutations in aei/deltaD and bea/deltaC differ in the way they disrupt the oscillating expression of her1 and deltaC. Furthermore, we find that the double mutants have significantly stronger defects in hypochord formation but not in somitogenesis or hindbrain neurogenesis, indicating genetically that the two delta's may function either semi-redundantly or distinctly, depending upon context.

C2orf71a/pcare1 is important for photoreceptor outer segment morphogenesis and visual function in zebrafish.

PubMed

Corral-Serrano, Julio C; Messchaert, Muriël; Dona, Margo; Peters, Theo A; Kamminga, Leonie M; van Wijk, Erwin; Collin, Rob W J

2018-06-26

Mutations in C2orf71 are causative for autosomal recessive retinitis pigmentosa and occasionally cone-rod dystrophy. We have recently discovered that the protein encoded by this gene is important for modulation of the ciliary membrane through the recruitment of an actin assembly module, and have therefore renamed the gene to PCARE (photoreceptor cilium actin regulator). Here, we report on the identification of two copies of the c2orf71/pcare gene in zebrafish, pcare1 and pcare2. To study the role of the gene most similar to human PCARE, pcare1, we have generated a stable pcare1 mutant zebrafish model (designated pcare1 rmc100/rmc100 ) in which the coding sequence was disrupted using CRISPR/Cas9 technology. Retinas of both embryonic (5 dpf) and adult (6 mpf) pcare1 rmc100/rmc100 zebrafish display a clear disorganization of photoreceptor outer segments, resembling the phenotype observed in Pcare -/- mice. Optokinetic response and visual motor response measurements indicated visual impairment in pcare1 rmc100/rmc100 zebrafish larvae at 5 dpf. In addition, electroretinogram measurements showed decreased b-wave amplitudes in pcare1 rmc100/rmc100 zebrafish as compared to age- and strain-matched wild-type larvae, indicating a defect in the transretinal current. Altogether, our data show that lack of pcare1 causes a retinal phenotype in zebrafish and indicate that the function of the PCARE gene is conserved across species.

Development of the zebrafish mesonephros.

PubMed

Diep, Cuong Q; Peng, Zhenzhen; Ukah, Tobechukwu K; Kelly, Paul M; Daigle, Renee V; Davidson, Alan J

2015-01-01

The vertebrate kidney plays an essential role in removing metabolic waste and balancing water and salt. This is carried out by nephrons, which comprise a blood filter attached to an epithelial tubule with proximal and distal segments. In zebrafish, two nephrons are first formed as part of the embryonic kidney (pronephros) and hundreds are formed later to make up the adult kidney (mesonephros). Previous studies have focused on the development of the pronephros while considerably less is known about how the mesonephros is formed. Here, we characterize mesonephros development in zebrafish and examine the nephrons that form during larval metamorphosis. These nephrons, arising from proliferating progenitor cells that express the renal transcription factor genes wt1b, pax2a, and lhx1a, form on top of the pronephric tubules and develop a segmentation pattern similar to pronephric nephrons. We find that the pronephros acts as a scaffold for the mesonephros, where new nephrons fuse with the distal segments of the pronephric tubules to form the final branching network that characterizes the adult zebrafish kidney. © 2015 Wiley Periodicals, Inc.

Development of the zebrafish mesonephros

PubMed Central

Diep, Cuong Q.; Peng, Zhenzhen; Ukah, Tobechukwu K.; Kelly, Paul M.; Daigle, Renee V.; Davidson, Alan J.

2015-01-01

The vertebrate kidney plays an essential role in removing metabolic waste and balancing water and salt. This is carried out by nephrons, which comprise a blood filter attached to an epithelial tubule with proximal and distal segments. In zebrafish, two nephrons are first formed as part of the embryonic kidney (pronephros) and hundreds are formed later to make up the adult kidney (mesonephros). Previous studies have focused on the development of the pronephros while considerably less is known about how the mesonephros is formed. Here, we characterize mesonephros development in zebrafish and examine the nephrons that form during larval metamorphosis. These nephrons, arising from proliferating progenitor cells that express the renal transcription factor genes wt1b, pax2a, and lhx1a, form on top of the pronephric tubules and develop a segmentation pattern similar to pronephric nephrons. We find that the pronephros acts as a scaffold for the mesonephros, where new nephrons fuse with the distal segments of the pronephric tubules to form the final branching network that characterizes the adult zebrafish kidney. PMID:25677367

Coordination of Fictive Motor Activity in the Larval Zebrafish Is Generated by Non-Segmental Mechanisms

PubMed Central

Wiggin, Timothy D.; Peck, Jack H.; Masino, Mark A.

2014-01-01

The cellular and network basis for most vertebrate locomotor central pattern generators (CPGs) is incompletely characterized, but organizational models based on known CPG architectures have been proposed. Segmental models propose that each spinal segment contains a circuit that controls local coordination and sends longer projections to coordinate activity between segments. Unsegmented/continuous models propose that patterned motor output is driven by gradients of neurons and synapses that do not have segmental boundaries. We tested these ideas in the larval zebrafish, an animal that swims in discrete episodes, each of which is composed of coordinated motor bursts that progress rostrocaudally and alternate from side to side. We perturbed the spinal cord using spinal transections or strychnine application and measured the effect on fictive motor output. Spinal transections eliminated episode structure, and reduced both rostrocaudal and side-to-side coordination. Preparations with fewer intact segments were more severely affected, and preparations consisting of midbody and caudal segments were more severely affected than those consisting of rostral segments. In reduced preparations with the same number of intact spinal segments, side-to-side coordination was more severely disrupted than rostrocaudal coordination. Reducing glycine receptor signaling with strychnine reversibly disrupted both rostrocaudal and side-to-side coordination in spinalized larvae without disrupting episodic structure. Both spinal transection and strychnine decreased the stability of the motor rhythm, but this effect was not causal in reducing coordination. These results are inconsistent with a segmented model of the spinal cord and are better explained by a continuous model in which motor neuron coordination is controlled by segment-spanning microcircuits. PMID:25275377

Automatic multiple zebrafish larvae tracking in unconstrained microscopic video conditions.

PubMed

Wang, Xiaoying; Cheng, Eva; Burnett, Ian S; Huang, Yushi; Wlodkowic, Donald

2017-12-14

The accurate tracking of zebrafish larvae movement is fundamental to research in many biomedical, pharmaceutical, and behavioral science applications. However, the locomotive characteristics of zebrafish larvae are significantly different from adult zebrafish, where existing adult zebrafish tracking systems cannot reliably track zebrafish larvae. Further, the far smaller size differentiation between larvae and the container render the detection of water impurities inevitable, which further affects the tracking of zebrafish larvae or require very strict video imaging conditions that typically result in unreliable tracking results for realistic experimental conditions. This paper investigates the adaptation of advanced computer vision segmentation techniques and multiple object tracking algorithms to develop an accurate, efficient and reliable multiple zebrafish larvae tracking system. The proposed system has been tested on a set of single and multiple adult and larvae zebrafish videos in a wide variety of (complex) video conditions, including shadowing, labels, water bubbles and background artifacts. Compared with existing state-of-the-art and commercial multiple organism tracking systems, the proposed system improves the tracking accuracy by up to 31.57% in unconstrained video imaging conditions. To facilitate the evaluation on zebrafish segmentation and tracking research, a dataset with annotated ground truth is also presented. The software is also publicly accessible.

Transcription factors involved in retinogenesis are co-opted by the circadian clock following photoreceptor differentiation

PubMed Central

Laranjeiro, Ricardo; Whitmore, David

2014-01-01

The circadian clock is known to regulate a wide range of physiological and cellular processes, yet remarkably little is known about its role during embryo development. Zebrafish offer a unique opportunity to explore this issue, not only because a great deal is known about key developmental events in this species, but also because the clock starts on the very first day of development. In this study, we identified numerous rhythmic genes in zebrafish larvae, including the key transcriptional regulators neurod and cdx1b, which are involved in neuronal and intestinal differentiation, respectively. Rhythmic expression of neurod and several additional transcription factors was only observed in the developing retina. Surprisingly, these rhythms in expression commenced at a stage of development after these transcription factors are known to have played their essential role in photoreceptor differentiation. Furthermore, this circadian regulation was maintained in adult retina. Thus, once mature photoreceptors are formed, multiple retinal transcription factors fall under circadian clock control, at which point they appear to play a new and important role in regulating rhythmic elements in the phototransduction pathway. PMID:24924194

The orphan receptor Rev-erbα gene is a target of the circadian clock pacemaker

PubMed Central

Triqueneaux, Gérard; Thenot, Sandrine; Kakizawa, Tomoko; Antoch, Marina P; Safi, Rachid; Takahashi, Joseph S; Delaunay, Franck; Laudet, Vincent

2013-01-01

Rev-erbα is a ubiquitously expressed orphan nuclear receptor which functions as a constitutive transcriptional repressor and is expressed in vertebrates according to a robust circadian rhythm. We report here that two Rev-erbα mRNA isoforms, namely Rev-erbα1 and Rev-erbα2, are generated through alternative promoter usage and that both show a circadian expression pattern in an in vitro system using serum-shocked fibroblasts. Both promoter regions P1 (Rev-erbα1) and P2 (Rev-erbα2) contain several E-box DNA sequences, which function as response elements for the core circadian-clock components: CLOCK and BMAL1. The CLOCK–BMAL1 heterodimer stimulates the activity of both P1 and P2 promoters in transient transfection assay by 3–6-fold. This activation was inhibited by the overexpression of CRY1, a component of the negative limb of the circadian transcriptional loop. Critical E-box elements were mapped within both promoters. This regulation is conserved in vertebrates since we found that the CLOCK–BMAL1 heterodimer also regulates the zebrafish Rev-erbα gene. In line with these data Rev-erbα circadian expression was strongly impaired in the livers of Clock mutant mice and in the pineal glands of zebrafish embryos treated with Clock and Bmal1 antisense oligonucleotides. Together these data demonstrate that CLOCK is a critical regulator of Rev-erbα circadian gene expression in evolutionarily distant vertebrates and suggest a role for Rev-erbα in the circadian clock output. PMID:15591021

Transcription factors involved in retinogenesis are co-opted by the circadian clock following photoreceptor differentiation.

PubMed

Laranjeiro, Ricardo; Whitmore, David

2014-07-01

The circadian clock is known to regulate a wide range of physiological and cellular processes, yet remarkably little is known about its role during embryo development. Zebrafish offer a unique opportunity to explore this issue, not only because a great deal is known about key developmental events in this species, but also because the clock starts on the very first day of development. In this study, we identified numerous rhythmic genes in zebrafish larvae, including the key transcriptional regulators neurod and cdx1b, which are involved in neuronal and intestinal differentiation, respectively. Rhythmic expression of neurod and several additional transcription factors was only observed in the developing retina. Surprisingly, these rhythms in expression commenced at a stage of development after these transcription factors are known to have played their essential role in photoreceptor differentiation. Furthermore, this circadian regulation was maintained in adult retina. Thus, once mature photoreceptors are formed, multiple retinal transcription factors fall under circadian clock control, at which point they appear to play a new and important role in regulating rhythmic elements in the phototransduction pathway. © 2014. Published by The Company of Biologists Ltd.

The photoreceptive cells of the pineal gland in adult zebrafish (Danio rerio).

PubMed

Laurà, Rosaria; Magnoli, Domenico; Zichichi, Rosalia; Guerrera, Maria Cristina; De Carlos, Felix; Suárez, Alberto Álvarez; Abbate, Francesco; Ciriaco, Emilia; Vega, Jose Antonio; Germanà, Antonino

2012-03-01

The zebrafish pineal gland plays a fundamental role in the regulation of the circadian rhythm through the melatonin secretion. The pinealocytes, also called photoreceptive cells, are considered the morphofunctional unit of pineal gland. In literature, the anatomical features, the cellular characteristics, and the pinealocytes morphology of zebrafish pineal gland have not been previously described in detail. Therefore, this study was undertaken to analyze the structure and ultrastructure, as well as the immunohistochemical profile of the zebrafish pineal gland with particular reference to the pinealocytes. Here, we demonstrated, using RT-PCR, immunohistochemistry and transmission electron microscopy, the expression of the mRNA for rhodopsin in the pineal gland of zebrafish, as well as its cellular localization exclusively in the pinealocytes of adult zebrafish. Moreover, the ultrastructural observations demonstrated that the pinealocytes were constituted by an outer segment with numerous lamellar membranes, an inner segment with many mitochondria, and a basal pole with the synapses. Our results taken together demonstrated a central role of zebrafish pinealocytes in the control of pineal gland functions. Copyright © 2011 Wiley Periodicals, Inc.

Culturable Gut Microbiota Diversity in Zebrafish

PubMed Central

Sørby, Jan Roger Torp; Aleström, Peter; Sørum, Henning

2012-01-01

Abstract The zebrafish (Danio rerio) is an increasingly used laboratory animal model in basic biology and biomedicine, novel drug development, and toxicology. The wide use has increased the demand for optimized husbandry protocols to ensure animal health care and welfare. The knowledge about the correlation between culturable zebrafish intestinal microbiota and health in relation to environmental factors and management procedures is very limited. A semi-quantitative level of growth of individual types of bacteria was determined and associated with sampling points. A total of 72 TAB line zebrafish from four laboratories (Labs A–D) in the Zebrafish Network Norway were used. Diagnostic was based on traditional bacterial culture methods and biochemical characterization using commercial kits, followed by 16S rDNA gene sequencing from pure subcultures. Also selected Gram-negative isolates were analyzed for antibiotic susceptibility to 8 different antibiotics. A total of 13 morphologically different bacterial species were the most prevalent: Aeromonas hydrophila, Aeromonas sobria, Vibrio parahaemolyticus, Photobacterium damselae, Pseudomonas aeruginosa, Pseudomonas fluorescens, Pseudomonas luteola, Comamonas testosteroni, Ochrobactrum anthropi, Staphylococcus cohnii, Staphylococcus epidermidis, Staphylococcus capitis, and Staphylococcus warneri. Only Lab B had significantly higher levels of total bacterial growth (OR=2.03), whereas numbers from Lab C (OR=1.01) and Lab D (OR=1.12) were found to be similar to the baseline Lab A. Sexually immature individuals had a significantly higher level of harvested total bacterial growth than mature fish (OR=0.82), no statistically significant differences were found between male and female fish (OR=1.01), and the posterior intestinal segment demonstrated a higher degree of culturable bacteria than the anterior segment (OR=4.1). Multiple antibiotic (>3) resistance was observed in 17% of the strains. We propose that a rapid

Culturable gut microbiota diversity in zebrafish.

PubMed

Cantas, Leon; Sørby, Jan Roger Torp; Aleström, Peter; Sørum, Henning

2012-03-01

The zebrafish (Danio rerio) is an increasingly used laboratory animal model in basic biology and biomedicine, novel drug development, and toxicology. The wide use has increased the demand for optimized husbandry protocols to ensure animal health care and welfare. The knowledge about the correlation between culturable zebrafish intestinal microbiota and health in relation to environmental factors and management procedures is very limited. A semi-quantitative level of growth of individual types of bacteria was determined and associated with sampling points. A total of 72 TAB line zebrafish from four laboratories (Labs A-D) in the Zebrafish Network Norway were used. Diagnostic was based on traditional bacterial culture methods and biochemical characterization using commercial kits, followed by 16S rDNA gene sequencing from pure subcultures. Also selected Gram-negative isolates were analyzed for antibiotic susceptibility to 8 different antibiotics. A total of 13 morphologically different bacterial species were the most prevalent: Aeromonas hydrophila, Aeromonas sobria, Vibrio parahaemolyticus, Photobacterium damselae, Pseudomonas aeruginosa, Pseudomonas fluorescens, Pseudomonas luteola, Comamonas testosteroni, Ochrobactrum anthropi, Staphylococcus cohnii, Staphylococcus epidermidis, Staphylococcus capitis, and Staphylococcus warneri. Only Lab B had significantly higher levels of total bacterial growth (OR=2.03), whereas numbers from Lab C (OR=1.01) and Lab D (OR=1.12) were found to be similar to the baseline Lab A. Sexually immature individuals had a significantly higher level of harvested total bacterial growth than mature fish (OR=0.82), no statistically significant differences were found between male and female fish (OR=1.01), and the posterior intestinal segment demonstrated a higher degree of culturable bacteria than the anterior segment (OR=4.1). Multiple antibiotic (>3) resistance was observed in 17% of the strains. We propose that a rapid conventional

The light-induced transcriptome of the zebrafish pineal gland reveals complex regulation of the circadian clockwork by light

PubMed Central

Ben-Moshe, Zohar; Alon, Shahar; Mracek, Philipp; Faigenbloom, Lior; Tovin, Adi; Vatine, Gad D.; Eisenberg, Eli; Foulkes, Nicholas S.; Gothilf, Yoav

2014-01-01

Light constitutes a primary signal whereby endogenous circadian clocks are synchronized (‘entrained’) with the day/night cycle. The molecular mechanisms underlying this vital process are known to require gene activation, yet are incompletely understood. Here, the light-induced transcriptome in the zebrafish central clock organ, the pineal gland, was characterized by messenger RNA (mRNA) sequencing (mRNA-seq) and microarray analyses, resulting in the identification of multiple light-induced mRNAs. Interestingly, a considerable portion of the molecular clock (14 genes) is light-induced in the pineal gland. Four of these genes, encoding the transcription factors dec1, reverbb1, e4bp4-5 and e4bp4-6, differentially affected clock- and light-regulated promoter activation, suggesting that light-input is conveyed to the core clock machinery via diverse mechanisms. Moreover, we show that dec1, as well as the core clock gene per2, is essential for light-entrainment of rhythmic locomotor activity in zebrafish larvae. Additionally, we used microRNA (miRNA) sequencing (miR-seq) and identified pineal-enhanced and light-induced miRNAs. One such miRNA, miR-183, is shown to downregulate e4bp4-6 mRNA through a 3′UTR target site, and importantly, to regulate the rhythmic mRNA levels of aanat2, the key enzyme in melatonin synthesis. Together, this genome-wide approach and functional characterization of light-induced factors indicate a multi-level regulation of the circadian clockwork by light. PMID:24423866

Estimation of network path segment delays

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nichols, Kathleen Marie

A method for estimation of a network path segment delay includes determining a scaled time stamp for each packet of a plurality of packets by scaling a time stamp for each respective packet to minimize a difference of at least one of a frequency and a frequency drift between a transport protocol clock of a host and a monitoring point. The time stamp for each packet is provided by the transport protocol clock of the host. A corrected time stamp for each packet is determined by removing from the scaled time stamp for each respective packet, a temporal offset betweenmore » the transport protocol clock and the monitoring clock by minimizing a temporal delay variation of the plurality of packets traversing a segment between the host and the monitoring point.« less

Recent Advances in Elucidating the Genetic Mechanisms of Nephrogenesis Using Zebrafish

PubMed Central

Cheng, Christina N.; Verdun, Valerie A.; Wingert, Rebecca A.

2015-01-01

The kidney is comprised of working units known as nephrons, which are epithelial tubules that contain a series of specialized cell types organized into a precise pattern of functionally distinct segment domains. There is a limited understanding of the genetic mechanisms that establish these discrete nephron cell types during renal development. The zebrafish embryonic kidney serves as a simplified yet conserved vertebrate model to delineate how nephron segments are patterned from renal progenitors. Here, we provide a concise review of recent advances in this emerging field, and discuss how continued research using zebrafish genetics can be applied to gain insightsabout nephrogenesis. PMID:26024215

Analysis of a Gene Regulatory Cascade Mediating Circadian Rhythm in Zebrafish

PubMed Central

Wang, Haifang; Du, Jiulin; Yan, Jun

2013-01-01

In the study of circadian rhythms, it has been a puzzle how a limited number of circadian clock genes can control diverse aspects of physiology. Here we investigate circadian gene expression genome-wide using larval zebrafish as a model system. We made use of a spatial gene expression atlas to investigate the expression of circadian genes in various tissues and cell types. Comparison of genome-wide circadian gene expression data between zebrafish and mouse revealed a nearly anti-phase relationship and allowed us to detect novel evolutionarily conserved circadian genes in vertebrates. We identified three groups of zebrafish genes with distinct responses to light entrainment: fast light-induced genes, slow light-induced genes, and dark-induced genes. Our computational analysis of the circadian gene regulatory network revealed several transcription factors (TFs) involved in diverse aspects of circadian physiology through transcriptional cascade. Of these, microphthalmia-associated transcription factor a (mitfa), a dark-induced TF, mediates a circadian rhythm of melanin synthesis, which may be involved in zebrafish's adaptation to daily light cycling. Our study describes a systematic method to discover previously unidentified TFs involved in circadian physiology in complex organisms. PMID:23468616

Automated feature extraction for retinal vascular biometry in zebrafish using OCT angiography

NASA Astrophysics Data System (ADS)

Bozic, Ivan; Rao, Gopikrishna M.; Desai, Vineet; Tao, Yuankai K.

2017-02-01

Zebrafish have been identified as an ideal model for angiogenesis because of anatomical and functional similarities with other vertebrates. The scale and complexity of zebrafish assays are limited by the need to manually treat and serially screen animals, and recent technological advances have focused on automation and improving throughput. Here, we use optical coherence tomography (OCT) and OCT angiography (OCT-A) to perform noninvasive, in vivo imaging of retinal vasculature in zebrafish. OCT-A summed voxel projections were low pass filtered and skeletonized to create an en face vascular map prior to connectivity analysis. Vascular segmentation was referenced to the optic nerve head (ONH), which was identified by automatically segmenting the retinal pigment epithelium boundary on the OCT structural volume. The first vessel branch generation was identified as skeleton segments with branch points closest to the ONH, and subsequent generations were found iteratively by expanding the search space outwards from the ONH. Biometric parameters, including length, curvature, and branch angle of each vessel segment were calculated and grouped by branch generation. Despite manual handling and alignment of each animal over multiple time points, we observe distinct qualitative patterns that enable unique identification of each eye from individual animals. We believe this OCT-based retinal biometry method can be applied for automated animal identification and handling in high-throughput organism-level pharmacological assays and genetic screens. In addition, these extracted features may enable high-resolution quantification of longitudinal vascular changes as a method for studying zebrafish models of retinal neovascularization and vascular remodeling.

On estimating the effects of clock instability with flicker noise characteristics

NASA Technical Reports Server (NTRS)

Wu, S. C.

1981-01-01

A scheme for flicker noise generation is given. The second approach is that of successive segmentation: A clock fluctuation is represented by 2N piecewise linear segments and then converted into a summation of N+1 triangular pulse train functions. The statistics of the clock instability are then formulated in terms of two sample variances at N+1 specified averaging times. The summation converges very rapidly that a value of N 6 is seldom necessary. An application to radio interferometric geodesy shows excellent agreement between the two approaches. Limitations to and the relative merits of the two approaches are discussed.

A Spatio-Temporal Model of Notch Signalling in the Zebrafish Segmentation Clock: Conditions for Synchronised Oscillatory Dynamics

PubMed Central

Terry, Alan J.; Sturrock, Marc; Dale, J. Kim; Maroto, Miguel; Chaplain, Mark A. J.

2011-01-01

In the vertebrate embryo, tissue blocks called somites are laid down in head-to-tail succession, a process known as somitogenesis. Research into somitogenesis has been both experimental and mathematical. For zebrafish, there is experimental evidence for oscillatory gene expression in cells in the presomitic mesoderm (PSM) as well as evidence that Notch signalling synchronises the oscillations in neighbouring PSM cells. A biological mechanism has previously been proposed to explain these phenomena. Here we have converted this mechanism into a mathematical model of partial differential equations in which the nuclear and cytoplasmic diffusion of protein and mRNA molecules is explictly considered. By performing simulations, we have found ranges of values for the model parameters (such as diffusion and degradation rates) that yield oscillatory dynamics within PSM cells and that enable Notch signalling to synchronise the oscillations in two touching cells. Our model contains a Hill coefficient that measures the co-operativity between two proteins (Her1, Her7) and three genes (her1, her7, deltaC) which they inhibit. This coefficient appears to be bounded below by the requirement for oscillations in individual cells and bounded above by the requirement for synchronisation. Consistent with experimental data and a previous spatially non-explicit mathematical model, we have found that signalling can increase the average level of Her1 protein. Biological pattern formation would be impossible without a certain robustness to variety in cell shape and size; our results possess such robustness. Our spatially-explicit modelling approach, together with new imaging technologies that can measure intracellular protein diffusion rates, is likely to yield significant new insight into somitogenesis and other biological processes. PMID:21386903

A spatio-temporal model of Notch signalling in the zebrafish segmentation clock: conditions for synchronised oscillatory dynamics.

PubMed

Terry, Alan J; Sturrock, Marc; Dale, J Kim; Maroto, Miguel; Chaplain, Mark A J

2011-02-28

In the vertebrate embryo, tissue blocks called somites are laid down in head-to-tail succession, a process known as somitogenesis. Research into somitogenesis has been both experimental and mathematical. For zebrafish, there is experimental evidence for oscillatory gene expression in cells in the presomitic mesoderm (PSM) as well as evidence that Notch signalling synchronises the oscillations in neighbouring PSM cells. A biological mechanism has previously been proposed to explain these phenomena. Here we have converted this mechanism into a mathematical model of partial differential equations in which the nuclear and cytoplasmic diffusion of protein and mRNA molecules is explicitly considered. By performing simulations, we have found ranges of values for the model parameters (such as diffusion and degradation rates) that yield oscillatory dynamics within PSM cells and that enable Notch signalling to synchronise the oscillations in two touching cells. Our model contains a Hill coefficient that measures the co-operativity between two proteins (Her1, Her7) and three genes (her1, her7, deltaC) which they inhibit. This coefficient appears to be bounded below by the requirement for oscillations in individual cells and bounded above by the requirement for synchronisation. Consistent with experimental data and a previous spatially non-explicit mathematical model, we have found that signalling can increase the average level of Her1 protein. Biological pattern formation would be impossible without a certain robustness to variety in cell shape and size; our results possess such robustness. Our spatially-explicit modelling approach, together with new imaging technologies that can measure intracellular protein diffusion rates, is likely to yield significant new insight into somitogenesis and other biological processes.

Ion transport in the zebrafish kidney from a human disease angle: possibilities, considerations, and future perspectives.

PubMed

Kersten, Simone; Arjona, Francisco J

2017-01-01

Unique experimental advantages, such as its embryonic/larval transparency, high-throughput nature, and ease of genetic modification, underpin the rapid emergence of the zebrafish (Danio rerio) as a preeminent model in biomedical research. Particularly in the field of nephrology, the zebrafish provides a promising model for studying the physiological implications of human solute transport processes along consecutive nephron segments. However, although the zebrafish might be considered a valuable model for numerous renal ion transport diseases and functional studies of many channels and transporters, not all human renal electrolyte transport mechanisms and human diseases can be modeled in the zebrafish. With this review, we explore the ontogeny of zebrafish renal ion transport, its nephron structure and function, and thereby demonstrate the clinical translational value of this model. By critical assessment of genomic and amino acid conservation of human proteins involved in renal ion handling (channels, transporters, and claudins), kidney and nephron segment conservation, and renal electrolyte transport physiology in the zebrafish, we provide researchers and nephrologists with an indication of the possibilities and considerations of the zebrafish as a model for human renal ion transport. Combined with advanced techniques envisioned for the future, implementation of the zebrafish might expand beyond unraveling pathophysiological mechanisms that underlie distinct genetic or environmentally, i.e., pharmacological and lifestyle, induced renal transport deficits. Specifically, the ease of drug administration and the exploitation of improved genetic approaches might argue for the adoption of the zebrafish as a model for preclinical personalized medicine for distinct renal diseases and renal electrolyte transport proteins. Copyright © 2017 the American Physiological Society.

Spine Patterning Is Guided by Segmentation of the Notochord Sheath.

PubMed

Wopat, Susan; Bagwell, Jennifer; Sumigray, Kaelyn D; Dickson, Amy L; Huitema, Leonie F A; Poss, Kenneth D; Schulte-Merker, Stefan; Bagnat, Michel

2018-02-20

The spine is a segmented axial structure made of alternating vertebral bodies (centra) and intervertebral discs (IVDs) assembled around the notochord. Here, we show that, prior to centra formation, the outer epithelial cell layer of the zebrafish notochord, the sheath, segments into alternating domains corresponding to the prospective centra and IVD areas. This process occurs sequentially in an anteroposterior direction via the activation of Notch signaling in alternating segments of the sheath, which transition from cartilaginous to mineralizing domains. Subsequently, osteoblasts are recruited to the mineralized domains of the notochord sheath to form mature centra. Tissue-specific manipulation of Notch signaling in sheath cells produces notochord segmentation defects that are mirrored in the spine. Together, our findings demonstrate that notochord sheath segmentation provides a template for vertebral patterning in the zebrafish spine. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

Zebrafish Meis functions to stabilize Pbx proteins and regulate hindbrain patterning.

PubMed

Waskiewicz, A J; Rikhof, H A; Hernandez, R E; Moens, C B

2001-11-01

Homeodomain-containing Hox proteins regulate segmental identity in Drosophila in concert with two partners known as Extradenticle (Exd) and Homothorax (Hth). These partners are themselves DNA-binding, homeodomain proteins, and probably function by revealing the intrinsic specificity of Hox proteins. Vertebrate orthologs of Exd and Hth, known as Pbx and Meis (named for a myeloid ecotropic leukemia virus integration site), respectively, are encoded by multigene families and are present in multimeric complexes together with vertebrate Hox proteins. Previous results have demonstrated that the zygotically encoded Pbx4/Lazarus (Lzr) protein is required for segmentation of the zebrafish hindbrain and proper expression and function of Hox genes. We demonstrate that Meis functions in the same pathway as Pbx in zebrafish hindbrain development, as expression of a dominant-negative mutant Meis results in phenotypes that are remarkably similar to that of lzr mutants. Surprisingly, expression of Meis protein partially rescues the lzr(-) phenotype. Lzr protein levels are increased in embryos overexpressing Meis and are reduced for lzr mutants that cannot bind to Meis. This implies a mechanism whereby Meis rescues lzr mutants by stabilizing maternally encoded Lzr. Our results define two functions of Meis during zebrafish hindbrain segmentation: that of a DNA-binding partner of Pbx proteins, and that of a post-transcriptional regulator of Pbx protein levels.

Zebrafish Health Conditions in the China Zebrafish Resource Center and 20 Major Chinese Zebrafish Laboratories.

PubMed

Liu, Liyue; Pan, Luyuan; Li, Kuoyu; Zhang, Yun; Zhu, Zuoyan; Sun, Yonghua

2016-07-01

In China, the use of zebrafish as an experimental animal in the past 15 years has widely expanded. The China Zebrafish Resource Center (CZRC), which was established in 2012, is becoming one of the major resource centers in the global zebrafish community. Large-scale use and regular exchange of zebrafish resources have put forward higher requirements on zebrafish health issues in China. This article reports the current aquatic infrastructure design, animal husbandry, and health-monitoring programs in the CZRC. Meanwhile, through a survey of 20 Chinese zebrafish laboratories, we also describe the current health status of major zebrafish facilities in China. We conclude that it is of great importance to establish a widely accepted health standard and health-monitoring strategy in the Chinese zebrafish research community.

Automated analysis of brain activity for seizure detection in zebrafish models of epilepsy.

PubMed

Hunyadi, Borbála; Siekierska, Aleksandra; Sourbron, Jo; Copmans, Daniëlle; de Witte, Peter A M

2017-08-01

Epilepsy is a chronic neurological condition, with over 30% of cases unresponsive to treatment. Zebrafish larvae show great potential to serve as an animal model of epilepsy in drug discovery. Thanks to their high fecundity and relatively low cost, they are amenable to high-throughput screening. However, the assessment of seizure occurrences in zebrafish larvae remains a bottleneck, as visual analysis is subjective and time-consuming. For the first time, we present an automated algorithm to detect epileptic discharges in single-channel local field potential (LFP) recordings in zebrafish. First, candidate seizure segments are selected based on their energy and length. Afterwards, discriminative features are extracted from each segment. Using a labeled dataset, a support vector machine (SVM) classifier is trained to learn an optimal feature mapping. Finally, this SVM classifier is used to detect seizure segments in new signals. We tested the proposed algorithm both in a chemically-induced seizure model and a genetic epilepsy model. In both cases, the algorithm delivered similar results to visual analysis and found a significant difference in number of seizures between the epileptic and control group. Direct comparison with multichannel techniques or methods developed for different animal models is not feasible. Nevertheless, a literature review shows that our algorithm outperforms state-of-the-art techniques in terms of accuracy, precision and specificity, while maintaining a reasonable sensitivity. Our seizure detection system is a generic, time-saving and objective method to analyze zebrafish LPF, which can replace visual analysis and facilitate true high-throughput studies. Copyright © 2017 Elsevier B.V. All rights reserved.

Differences in neural crest sensitivity to ethanol account for the infrequency of anterior segment defects in the eye compared with craniofacial anomalies in a zebrafish model of fetal alcohol syndrome.

PubMed

Eason, Jessica; Williams, Antionette L; Chawla, Bahaar; Apsey, Christian; Bohnsack, Brenda L

2017-09-01

Ethanol (ETOH) exposure during pregnancy is associated with craniofacial and neurologic abnormalities, but infrequently disrupts the anterior segment of the eye. In these studies, we used zebrafish to investigate differences in the teratogenic effect of ETOH on craniofacial, periocular, and ocular neural crest. Zebrafish eye and neural crest development was analyzed by means of live imaging, TUNEL (terminal deoxynucleotidyl transferase dUTP nick end labeling) assay, immunostaining, detection of reactive oxygen species, and in situ hybridization. Our studies demonstrated that foxd3-positive neural crest cells in the periocular mesenchyme and developing eye were less sensitive to ETOH than sox10-positive craniofacial neural crest cells that form the pharyngeal arches and jaw. ETOH increased apoptosis in the retina, but did not affect survival of periocular and ocular neural crest cells. ETOH also did not increase reactive oxygen species within the eye. In contrast, ETOH increased ventral neural crest apoptosis and reactive oxygen species production in the facial mesenchyme. In the eye and craniofacial region, sod2 showed high levels of expression in the anterior segment and in the setting of Sod2 knockdown, low levels of ETOH decreased migration of foxd3-positive neural crest cells into the developing eye. However, ETOH had minimal effect on the periocular and ocular expression of transcription factors (pitx2 and foxc1) that regulate anterior segment development. Neural crest cells contributing to the anterior segment of the eye exhibit increased ability to withstand ETOH-induced oxidative stress and apoptosis. These studies explain the rarity of anterior segment dysgenesis despite the frequent craniofacial abnormalities in fetal alcohol syndrome. Birth Defects Research 109:1212-1227, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Development of the Astyanax mexicanus circadian clock and non-visual light responses.

PubMed

Frøland Steindal, Inga A; Beale, Andrew D; Yamamoto, Yoshiyuki; Whitmore, David

2018-06-23

addition, the light induction of clock genes is developmentally delayed in cave populations. Zebrafish embryonic light sensitivity has been shown to be critical not only for clock entrainment, but also for transcriptional activation of DNA repair processes. Similar downstream transcriptional responses to light also occur in Astyanax. Interestingly, the establishment of the adult timing profile of clock gene expression takes several days to become apparent. This fact may provide mechanistic insight into the key differences between the cave and surface fish clock mechanisms. Copyright © 2018. Published by Elsevier Inc.

Characterization of Na+ and Ca2+ Channels in Zebrafish Dorsal Root Ganglion Neurons

PubMed Central

Won, Yu-Jin; Ono, Fumihito; Ikeda, Stephen R.

2012-01-01

Background Dorsal root ganglia (DRG) somata from rodents have provided an excellent model system to study ion channel properties and modulation using electrophysiological investigation. As in other vertebrates, zebrafish (Danio rerio) DRG are organized segmentally and possess peripheral axons that bifurcate into each body segment. However, the electrical properties of zebrafish DRG sensory neurons, as compared with their mammalian counterparts, are relatively unexplored because a preparation suitable for electrophysiological studies has not been available. Methodology/Principal Findings We show enzymatically dissociated DRG neurons from juvenile zebrafish expressing Isl2b-promoter driven EGFP were easily identified with fluorescence microscopy and amenable to conventional whole-cell patch-clamp studies. Two kinetically distinct TTX-sensitive Na+ currents (rapidly- and slowly-inactivating) were discovered. Rapidly-inactivating INa were preferentially expressed in relatively large neurons, while slowly-inactivating INa was more prevalent in smaller DRG neurons. RT-PCR analysis suggests zscn1aa/ab, zscn8aa/ab, zscn4ab and zscn5Laa are possible candidates for these INa components. Voltage-gated Ca2+ currents (ICa) were primarily (87%) comprised of a high-voltage activated component arising from ω-conotoxin GVIA-sensitive CaV2.2 (N-type) Ca2+ channels. A few DRG neurons (8%) displayed a miniscule low-voltage-activated component. ICa in zebrafish DRG neurons were modulated by neurotransmitters via either voltage-dependent or -independent G-protein signaling pathway with large cell-to-cell response variability. Conclusions/Significance Our present results indicate that, as in higher vertebrates, zebrafish DRG neurons are heterogeneous being composed of functionally distinct subpopulations that may correlate with different sensory modalities. These findings provide the first comparison of zebrafish and rodent DRG neuron electrical properties and thus provide a basis for

Grouper tshβ Promoter-Driven Transgenic Zebrafish Marks Proximal Kidney Tubule Development

PubMed Central

Wang, Yang; Sun, Zhi-Hui; Zhou, Li; Li, Zhi; Gui, Jian-Fang

2014-01-01

Kidney tubule plays a critical role in recovering or secreting solutes, but the detailed morphogenesis remains unclear. Our previous studies have found that grouper tshβ (gtshβ) is also expressed in kidney, however, the distribution significance is still unknown. To understand the gtshβ role and kidney tubule morphogenesis, here, we have generated a transgenic zebrafish line Tg(gtshβ:GFP) with green fluorescent protein driven by the gtshβ promoter. Similar to the endogenous tshβ in zebrafish or in grouper, the gtshβ promoter-driven GFP is expressed in pituitary and kidney, and the developing details of proximal kidney tubule are marked in the transgenic zebrafish line. The gfp initially transcribes at 16 hours post fertilization (hpf) above the dorsal mesentery, and partially co-localizes with pronephric tubular markers slc20a1a and cdh17. Significantly, the GFP specifically localizes in proximal pronephric segments during embryogenesis and resides at kidney duct epithelium in adult fish. To test whether the gtshβ promoter-driven GFP may serve as a readout signal of the tubular development, we have treated the embryos with retinoic acid signaing (RA) reagents, in which exogenous RA addition results in a distal extension of the proximal segments, while RA inhibition induces a weakness and shortness of the proximal segments. Therefore, this transgenic line provides a useful tool for genetic or chemical analysis of kidney tubule. PMID:24905828

Masking of a circadian behavior in larval zebrafish involves the thalamo-habenula pathway.

PubMed

Lin, Qian; Jesuthasan, Suresh

2017-06-22

Changes in illumination can rapidly influence behavior that is normally controlled by the circadian clock. This effect is termed masking. In mice, masking requires melanopsin-expressing retinal ganglion cells that detect blue light and project to the thalamus. It is not known whether masking is wavelength-dependent in other vertebrates, nor is it known whether the thalamus is also involved or how it influences masking. Here, we address these questions in zebrafish. We find that diel vertical migration, a circadian behavior in larval zebrafish, is effectively triggered by blue, but not by red light. Two-photon calcium imaging reveals that a thalamic nucleus and a downstream structure, the habenula, have a sustained response to blue but not to red light. Lesioning the habenula reduces light-evoked climbing. These data suggest that the thalamo-habenula pathway is involved in the ability of blue light to influence a circadian behavior.

Continuum theory of gene expression waves during vertebrate segmentation.

PubMed

Jörg, David J; Morelli, Luis G; Soroldoni, Daniele; Oates, Andrew C; Jülicher, Frank

2015-09-01

The segmentation of the vertebrate body plan during embryonic development is a rhythmic and sequential process governed by genetic oscillations. These genetic oscillations give rise to traveling waves of gene expression in the segmenting tissue. Here we present a minimal continuum theory of vertebrate segmentation that captures the key principles governing the dynamic patterns of gene expression including the effects of shortening of the oscillating tissue. We show that our theory can quantitatively account for the key features of segmentation observed in zebrafish, in particular the shape of the wave patterns, the period of segmentation and the segment length as a function of time.

Continuum theory of gene expression waves during vertebrate segmentation

PubMed Central

Jörg, David J; Morelli, Luis G; Soroldoni, Daniele; Oates, Andrew C; Jülicher, Frank

2015-01-01

Abstract The segmentation of the vertebrate body plan during embryonic development is a rhythmic and sequential process governed by genetic oscillations. These genetic oscillations give rise to traveling waves of gene expression in the segmenting tissue. Here we present a minimal continuum theory of vertebrate segmentation that captures the key principles governing the dynamic patterns of gene expression including the effects of shortening of the oscillating tissue. We show that our theory can quantitatively account for the key features of segmentation observed in zebrafish, in particular the shape of the wave patterns, the period of segmentation and the segment length as a function of time. PMID:28725158

Lego clocks: building a clock from parts.

PubMed

Brunner, Michael; Simons, Mirre J P; Merrow, Martha

2008-06-01

A new finding opens up speculation that the molecular mechanism of circadian clocks in Synechococcus elongatus is composed of multiple oscillator systems (Kitayama and colleagues, this issue, pp. 1513-1521), as has been described in many eukaryotic clock model systems. However, an alternative intepretation is that the pacemaker mechanism-as previously suggested-lies primarily in the rate of ATP hydrolysis by the clock protein KaiC.

Regulation of circadian clock transcriptional output by CLOCK:BMAL1

PubMed Central

Trott, Alexandra J.

2018-01-01

The mammalian circadian clock relies on the transcription factor CLOCK:BMAL1 to coordinate the rhythmic expression of 15% of the transcriptome and control the daily regulation of biological functions. The recent characterization of CLOCK:BMAL1 cistrome revealed that although CLOCK:BMAL1 binds synchronously to all of its target genes, its transcriptional output is highly heterogeneous. By performing a meta-analysis of several independent genome-wide datasets, we found that the binding of other transcription factors at CLOCK:BMAL1 enhancers likely contribute to the heterogeneity of CLOCK:BMAL1 transcriptional output. While CLOCK:BMAL1 rhythmic DNA binding promotes rhythmic nucleosome removal, it is not sufficient to generate transcriptionally active enhancers as assessed by H3K27ac signal, RNA Polymerase II recruitment, and eRNA expression. Instead, the transcriptional activity of CLOCK:BMAL1 enhancers appears to rely on the activity of ubiquitously expressed transcription factors, and not tissue-specific transcription factors, recruited at nearby binding sites. The contribution of other transcription factors is exemplified by how fasting, which effects several transcription factors but not CLOCK:BMAL1, either decreases or increases the amplitude of many rhythmically expressed CLOCK:BMAL1 target genes. Together, our analysis suggests that CLOCK:BMAL1 promotes a transcriptionally permissive chromatin landscape that primes its target genes for transcription activation rather than directly activating transcription, and provides a new framework to explain how environmental or pathological conditions can reprogram the rhythmic expression of clock-controlled genes. PMID:29300726

Zebrafish Pronephros Development.

PubMed

Naylor, Richard W; Qubisi, Sarah S; Davidson, Alan J

The pronephros is the first kidney type to form in vertebrate embryos. The first step of pronephrogenesis in the zebrafish is the formation of the intermediate mesoderm during gastrulation, which occurs in response to secreted morphogens such as BMPs and Nodals. Patterning of the intermediate mesoderm into proximal and distal cell fates is induced by retinoic acid signaling with downstream transcription factors including wt1a, pax2a, pax8, hnf1b, sim1a, mecom, and irx3b. In the anterior intermediate mesoderm, progenitors of the glomerular blood filter migrate and fuse at the midline and recruit a blood supply. More posteriorly localized tubule progenitors undergo epithelialization and fuse with the cloaca. The Notch signaling pathway regulates the formation of multi-ciliated cells in the tubules and these cells help propel the filtrate to the cloaca. The lumenal sheer stress caused by flow down the tubule activates anterior collective migration of the proximal tubules and induces stretching and proliferation of the more distal segments. Ultimately these processes create a simple two-nephron kidney that is capable of reabsorbing and secreting solutes and expelling excess water-processes that are critical to the homeostasis of the body fluids. The zebrafish pronephric kidney provides a simple, yet powerful, model system to better understand the conserved molecular and cellular progresses that drive nephron formation, structure, and function.

Direct activation of chordoblasts by retinoic acid is required for segmented centra mineralization during zebrafish spine development.

PubMed

Pogoda, Hans-Martin; Riedl-Quinkertz, Iris; Löhr, Heiko; Waxman, Joshua S; Dale, Rodney M; Topczewski, Jacek; Schulte-Merker, Stefan; Hammerschmidt, Matthias

2018-05-08

Zebrafish mutants with increased retinoic acid (RA) signaling due to the loss of the RA-inactivating enzyme Cyp26b1 develop a hyper-mineralized spine with gradually fusing vertebral body precursors (centra). However, the underlying cellular mechanisms remain incompletely understood. Here, we show that cells of the notochord epithelium named chordoblasts are sensitive to RA signaling. Chordoblasts are uniformly distributed along the anteroposterior axis and initially generate the continuous collagenous notochord sheath. However, subsequently and iteratively, subsets of these cells undergo further RA-dependent differentiation steps, acquire a stellate-like shape, downregulate expression of the collagen gene col2a1a , switch on cyp26b1 expression and trigger metameric sheath mineralization. This mineralization fails to appear upon chordoblast-specific cell ablation or RA signal transduction blockade. Together, our data reveal that, despite their different developmental origins, the activities and regulation of chordoblasts are very similar to those of osteoblasts, including their RA-induced transition from osteoid-producing cells to osteoid-mineralizing ones. Furthermore, our data point to a requirement for locally controlled RA activity within the chordoblast layer in order to generate the segmented vertebral column. © 2018. Published by The Company of Biologists Ltd.

Smoc2 modulates embryonic myelopoiesis during zebrafish development.

PubMed

Mommaerts, Hendrik; Esguerra, Camila V; Hartmann, Ursula; Luyten, Frank P; Tylzanowski, Przemko

2014-11-01

SMOC2 is a member of the BM-40 (SPARC) family of matricellular proteins, reported to influence signaling in the extracellular compartment. In mice, Smoc2 is expressed in many different tissues and was shown to enhance the response to angiogenic growth factors, mediate cell adhesion, keratinocyte migration, and metastasis. Additionally, SMOC2 is associated with vitiligo and craniofacial and dental defects. The function of Smoc2 during early zebrafish development has not been determined to date. In pregastrula zebrafish embryos, smoc2 is expressed ubiquitously. As development progresses, the expression pattern becomes more anteriorly restricted. At the onset of blood cell circulation, smoc2 morphants presented a mild ventralization of posterior structures. Molecular analysis of the smoc2 morphants indicated myelopoietic defects in the rostral blood islands during segmentation stages. Hemangioblast development and further specification of the myeloid progenitor cells were shown to be impaired. Additional experiments indicated that Bmp target genes were down-regulated in smoc2 morphants. Our findings reveal that Smoc2 is an essential player in the development of myeloid cells of the anterior lateral plate mesoderm during embryonic zebrafish development. Furthermore, our data show that Smoc2 affects the transcription of Bmp target genes without affecting initial dorsoventral patterning or mesoderm development. Copyright © 2014 Wiley Periodicals, Inc.

Simulating Future GPS Clock Scenarios with Two Composite Clock Algorithms

NASA Technical Reports Server (NTRS)

Suess, Matthias; Matsakis, Demetrios; Greenhall, Charles A.

2010-01-01

Using the GPS Toolkit, the GPS constellation is simulated using 31 satellites (SV) and a ground network of 17 monitor stations (MS). At every 15-minutes measurement epoch, the monitor stations measure the time signals of all satellites above a parameterized elevation angle. Once a day, the satellite clock estimates the station and satellite clocks. The first composite clock (B) is based on the Brown algorithm, and is now used by GPS. The second one (G) is based on the Greenhall algorithm. The composite clock of G and B performance are investigated using three ground-clock models. Model C simulates the current GPS configuration, in which all stations are equipped with cesium clocks, except for masers at USNO and Alternate Master Clock (AMC) sites. Model M is an improved situation in which every station is equipped with active hydrogen masers. Finally, Models F and O are future scenarios in which the USNO and AMC stations are equipped with fountain clocks instead of masers. Model F is a rubidium fountain, while Model O is more precise but futuristic Optical Fountain. Each model is evaluated using three performance metrics. The timing-related user range error having all satellites available is the first performance index (PI1). The second performance index (PI2) relates to the stability of the broadcast GPS system time itself. The third performance index (PI3) evaluates the stability of the time scales computed by the two composite clocks. A distinction is made between the "Signal-in-Space" accuracy and that available through a GNSS receiver.

Clock Scan Protocol for Image Analysis: ImageJ Plugins.

PubMed

Dobretsov, Maxim; Petkau, Georg; Hayar, Abdallah; Petkau, Eugen

2017-06-19

The clock scan protocol for image analysis is an efficient tool to quantify the average pixel intensity within, at the border, and outside (background) a closed or segmented convex-shaped region of interest, leading to the generation of an averaged integral radial pixel-intensity profile. This protocol was originally developed in 2006, as a visual basic 6 script, but as such, it had limited distribution. To address this problem and to join similar recent efforts by others, we converted the original clock scan protocol code into two Java-based plugins compatible with NIH-sponsored and freely available image analysis programs like ImageJ or Fiji ImageJ. Furthermore, these plugins have several new functions, further expanding the range of capabilities of the original protocol, such as analysis of multiple regions of interest and image stacks. The latter feature of the program is especially useful in applications in which it is important to determine changes related to time and location. Thus, the clock scan analysis of stacks of biological images may potentially be applied to spreading of Na + or Ca ++ within a single cell, as well as to the analysis of spreading activity (e.g., Ca ++ waves) in populations of synaptically-connected or gap junction-coupled cells. Here, we describe these new clock scan plugins and show some examples of their applications in image analysis.

The circadian clock of teleost fish: a comparative analysis reveals distinct fates for duplicated genes.

PubMed

Toloza-Villalobos, Jessica; Arroyo, José Ignacio; Opazo, Juan C

2015-01-01

The circadian clock is a central oscillator that coordinates endogenous rhythms. Members of six gene families underlie the metabolic machinery of this system. Although this machinery appears to correspond to a highly conserved genetic system in metazoans, it has been recognized that vertebrates possess a more diverse gene inventory than that of non-vertebrates. This difference could have originated in the two successive rounds of whole-genome duplications that took place in the common ancestor of the group. Teleost fish underwent an extra event of whole-genome duplication, which is thought to have provided an abundance of raw genetic material for the biological innovations that facilitated the radiation of the group. In this study, we assessed the relative contributions of whole-genome duplication and small-scale gene duplication to generate the repertoire of genes associated with the circadian clock of teleost fish. To achieve this goal, we annotated genes from six gene families associated with the circadian clock in eight teleost fish species, and we reconstructed their evolutionary history by inferring phylogenetic relationships. Our comparative analysis indicated that teleost species possess a variable repertoire of genes related to the circadian clock gene families and that the actual diversity of these genes has been shaped by a variety of phenomena, such as the complete deletion of ohnologs, the differential retention of genes, and lineage-specific gene duplications. From a functional perspective, the subfunctionalization of two ohnolog genes (PER1a and PER1b) in zebrafish highlights the power of whole-genome duplications to generate biological diversity.

Episodic swimming in the larval zebrafish is generated by a spatially distributed spinal network with modular functional organization

PubMed Central

Wiggin, Timothy D.; Anderson, Tatiana M.; Eian, John; Peck, Jack H.

2012-01-01

Despite the diverse methods vertebrates use for locomotion, there is evidence that components of the locomotor central pattern generator (CPG) are conserved across species. When zebrafish begin swimming early in development, they perform short episodes of activity separated by periods of inactivity. Within these episodes, the trunk flexes with side-to-side alternation and the traveling body wave progresses rostrocaudally. To characterize the distribution of the swimming CPG along the rostrocaudal axis, we performed transections of the larval zebrafish spinal cord and induced fictive swimming using N-methyl-d-aspartate (NMDA). In both intact and spinalized larvae, bursting is found throughout the rostrocaudal extent of the spinal cord, and the properties of fictive swimming observed were dependent on the concentration of NMDA. We isolated series of contiguous spinal segments by performing multiple spinal transections on the same larvae. Although series from all regions of the spinal cord have the capacity to produce bursts, the capacity to produce organized episodes of fictive swimming has a rostral bias: in the rostral spinal cord, only 12 contiguous body segments are necessary, whereas 23 contiguous body segments are necessary in the caudal spinal cord. Shorter series of segments were often active but produced either continuous rhythmic bursting or sporadic, nonrhythmic bursting. Both episodic and continuous bursting alternated between the left and right sides of the body and showed rostrocaudal progression, demonstrating the functional dissociation of the circuits responsible for episodic structure and fine burst timing. These findings parallel results in mammalian locomotion, and we propose a hierarchical model of the larval zebrafish swimming CPG. PMID:22572943

Effect of rearing temperatures during embryonic development on the phenotypic sex in zebrafish (Danio rerio).

PubMed

Abozaid, H; Wessels, S; Hörstgen-Schwark, G

2011-01-01

In zebrafish, Danio rerio, a polygenic pattern of sex determination or a female heterogamety with possible influences of environmental factors is assumed. The present study focuses on the effects of an elevated water temperature (35° C) during the embryonic development on sex determination in zebrafish. Eggs derived from 3 golden females were fertilized by the same mitotic gynogenetic male and exposed to a water temperature of 35° C, applied from 5 to 10 h post fertilization (hpf), from 5 to 24 hpf, and from 5 to 48 hpf, which correspond to the following developmental stages: gastrula, gastrula to segmentation, and gastrula to pharyngula stage, respectively. Hatching and survival rates decreased with increasing exposure to high water temperatures. Reductions in the hatching and survival rates were not responsible for differences in sex ratios. Accordingly, exposition of the fertilized eggs to a high temperature (35° C) leads to an increase of the male proportion from 22.0% in the controls to a balanced sex ratio (48.3, 47.5, and 52.6%) in the gastrula, segmentation, and pharyngula groups, respectively. These results prove the possibility to change the pathway of sexual determination during early embryonic stages in zebrafish by exposure to a high water temperature. Copyright © 2011 S. Karger AG, Basel.

RH1020 Single Event Clock Upset Summary Report

NASA Technical Reports Server (NTRS)

Katz, Richard B.; Wang, J. J.

1998-01-01

This report summarizes the testing and analysis of "single event clock upset' in the RH1020. Also included are SEU-rate predictions and design recommendations for risk analysis and reduction. The subject of "upsets" in the RH1020 is best understood by using a model consisting of a global clock buffer and a D-type flip-flop as the basic memory unit. The RH1020 is built on the ACT 1 family architecture. As such, it has one low-skew global clock buffer with a TTL-level input threshold that is accessed via a single dedicated pin. The clock signal is driven to full CMOS levels, buffered, and sent to individual row buffers with one buffer per channel. For low-skew performance, the outputs of all of the RH1020 row buffers are shorted together via metal lines, as is done in the A1020B. All storage in the RH1020 consists of routed flip-flops, constructed with multiplexors and feedback through the routing segments. A simple latch can be constructed from a single (combinatorial or C) module; an edge-triggered flip-flop is constructed using two concatenated latches. There is no storage in the I/O modules. The front end of the clock buffering circuitry, at a common point relative to the row buffer, is a sub-circuit that was determined to be the most susceptible to heavy ions. This is due, in part, to its smaller transistors compared to the rest of the circuitry. This conclusion is also supported by SPICE simulations and an analysis of the heavy ion data, described in this report. The edge triggered D flip-flop has two single-event-upset modes. Mode one, called C-module upset, is caused by a heavy ion striking the C-module's sensitive area on the silicon and produces a soft single bit error at the output of the flip-flop. Mode two, called clock upset, is caused by a heavy ion strike on the clock buffer, generating a runt pulse interpreted as a false clock signal and consequently producing errors at the flip-flop outputs. C-module upset sensitivity in the RH1020 is essentially

Sprouting Buds of Zebrafish Research in Malaysia: First Malaysia Zebrafish Disease Model Workshop.

PubMed

Okuda, Kazuhide Shaun; Tan, Pei Jean; Patel, Vyomesh

2016-04-01

Zebrafish is gaining prominence as an important vertebrate model for investigating various human diseases. Zebrafish provides unique advantages such as optical clarity of embryos, high fecundity rate, and low cost of maintenance, making it a perfect complement to the murine model equivalent in biomedical research. Due to these advantages, researchers in Malaysia are starting to take notice and incorporate the zebrafish model into their research activities. However, zebrafish research in Malaysia is still in its infancy stage and many researchers still remain unaware of the full potential of the zebrafish model or have limited access to related tools and techniques that are widely utilized in many zebrafish laboratories worldwide. To overcome this, we organized the First Malaysia Zebrafish Disease Model Workshop in Malaysia that took place on 11th and 12th of November 2015. In this workshop, we showcased how the zebrafish model is being utilized in the biomedical field in international settings as well as in Malaysia. For this, notable international speakers and those from local universities known to be carrying out impactful research using zebrafish were invited to share some of the cutting edge techniques that are used in their laboratories that may one day be incorporated in the Malaysian scientific community.

Habenular commissure formation in zebrafish is regulated by the pineal gland-specific gene unc119c.

PubMed

Toyama, Reiko; Kim, Mi Ha; Rebbert, Martha L; Gonzales, John; Burgess, Harold; Dawid, Igor B

2013-09-01

The zebrafish pineal gland (epiphysis) is a site of melatonin production, contains photoreceptor cells, and functions as a circadian clock pacemaker. Since it is located on the surface of the forebrain, it is accessible for manipulation and, therefore, is a useful model system to analyze pineal gland function and development. We previously analyzed the pineal transcriptome during development and showed that many genes exhibit a highly dynamic expression pattern in the pineal gland. Among genes preferentially expressed in the zebrafish pineal gland, we identified a tissue-specific form of the unc119 gene family, unc119c, which is highly preferentially expressed in the pineal gland during day and night at all stages examined from embryo to adult. When expression of unc119c was inhibited, the formation of the habenular commissure (HC) was specifically compromised. The Unc119c interacting factors Arl3l1 and Arl3l2 as well as Wnt4a also proved indispensible for HC formation. We suggest that Unc119c, together with Arl3l1/2, plays an important role in modulating Wnt4a production and secretion during HC formation in the forebrain of the zebrafish embryo. Copyright © 2013 Wiley Periodicals, Inc.

Habenular commissure formation in zebrafish is regulated by the pineal gland specific gene unc119c

PubMed Central

Toyama, Reiko; Kim, Mi Ha; Rebbert, Martha L.; Gonzales, John; Burgess, Harold; Dawid, Igor B.

2013-01-01

Background The zebrafish pineal gland (epiphysis) is a site of melatonin production, contains photoreceptor cells, and functions as a circadian clock pacemaker. Since it is located on the surface of the forebrain, it is accessible for manipulation and therefore is a useful model system to analyze pineal gland function and development. We previously analyzed the pineal transcriptome during development and showed that many genes exhibit a highly dynamic expression pattern in the pineal gland. Results Among genes preferentially expressed in the zebrafish pineal gland, we identified a tissue-specific form of the unc119 gene family, unc119c, which is highly preferentially expressed in the pineal gland during day and night at all stages examined from embryo to adult. When expression of unc119c was inhibited, the formation of the habenular commissure (HC) was specifically compromised. The Unc119c interacting factors Arl3l1 and Arl3l2 as well as Wnt4a also proved indispensible for HC formation. Conclusions We suggest that Unc119c, together with Arl3l1/2, plays an important role in modulating Wnt4a production and secretion during HC formation in the forebrain of the zebrafish embryo. PMID:23749482

Real-time simulation clock

NASA Technical Reports Server (NTRS)

Bennington, Donald R. (Inventor); Crawford, Daniel J. (Inventor)

1990-01-01

The invention is a clock for synchronizing operations within a high-speed, distributed data processing network. The clock is actually a distributed system comprising a central clock and multiple site clock interface units (SCIUs) which are connected by means of a fiber optic star network and which operate under control of separate clock software. The presently preferred embodiment is a part of the flight simulation system now in current use at the NASA Langley Research Center.

FishFace: interactive atlas of zebrafish craniofacial development at cellular resolution

PubMed Central

2013-01-01

Background The vertebrate craniofacial skeleton may exhibit anatomical complexity and diversity, but its genesis and evolution can be understood through careful dissection of developmental programs at cellular resolution. Resources are lacking that include introductory overviews of skeletal anatomy coupled with descriptions of craniofacial development at cellular resolution. In addition to providing analytical guidelines for other studies, such an atlas would suggest cellular mechanisms underlying development. Description We present the Fish Face Atlas, an online, 3D-interactive atlas of craniofacial development in the zebrafish Danio rerio. Alizarin red-stained skulls scanned by fluorescent optical projection tomography and segmented into individual elements provide a resource for understanding the 3D structure of the zebrafish craniofacial skeleton. These data provide the user an anatomical entry point to confocal images of Alizarin red-stained zebrafish with transgenically-labelled pharyngeal arch ectomesenchyme, chondrocytes, and osteoblasts, which illustrate the appearance, morphogenesis, and growth of the mandibular and hyoid cartilages and bones, as viewed in live, anesthetized zebrafish during embryonic and larval development. Confocal image stacks at high magnification during the same stages provide cellular detail and suggest developmental and evolutionary hypotheses. Conclusion The FishFace Atlas is a novel learning tool for understanding craniofacial skeletal development, and can serve as a reference for a variety of studies, including comparative and mutational analyses. PMID:23714426

Automatic zebrafish heartbeat detection and analysis for zebrafish embryos.

PubMed

Pylatiuk, Christian; Sanchez, Daniela; Mikut, Ralf; Alshut, Rüdiger; Reischl, Markus; Hirth, Sofia; Rottbauer, Wolfgang; Just, Steffen

2014-08-01

A fully automatic detection and analysis method of heartbeats in videos of nonfixed and nonanesthetized zebrafish embryos is presented. This method reduces the manual workload and time needed for preparation and imaging of the zebrafish embryos, as well as for evaluating heartbeat parameters such as frequency, beat-to-beat intervals, and arrhythmicity. The method is validated by a comparison of the results from automatic and manual detection of the heart rates of wild-type zebrafish embryos 36-120 h postfertilization and of embryonic hearts with bradycardia and pauses in the cardiac contraction.

A clock-aided positioning algorithm based on Kalman model of GNSS receiver clock bias

NASA Astrophysics Data System (ADS)

Zhu, Lingyao; Li, Zishen; Yuan, Hong

2017-10-01

The modeling and forecasting of the receiver clock bias is of practical significance, including the improvement of positioning accuracy, etc. When the clock frequency of the receiver is stable, the model can be established according to the historical clock bias data and the clock bias of the following time can be predicted. For this, we adopted the Kalman model to predict the receiver clock bias based on the calculated clock bias data obtained from the laboratory via sliding mode. Meanwhile, the relevant clock-aided positioning algorithm was presented. The results show that: the Kalman model can be used in practical work; and that under the condition that only 3 satellite signal can be received, this clock-aided positioning results can meet the needs of civilian users, which improves the continuity of positioning in harsh conditions.

Variable frequency microprocessor clock generator

DOE Office of Scientific and Technical Information (OSTI.GOV)

Branson, C.N.

A microprocessor-based system is described comprising: a digital central microprocessor provided with a clock input and having a rate of operation determined by the frequency of a clock signal input thereto; memory means operably coupled to the central microprocessor for storing programs respectively including a plurality of instructions and addressable by the central microprocessor; peripheral device operably connected to the central microprocessor, the first peripheral device being addressable by the central microprocessor for control thereby; a system clock generator for generating a digital reference clock signal having a reference frequency rate; and frequency rate reduction circuit means connected between themore » clock generator and the clock input of the central microprocessor for selectively dividing the reference clock signal to generate a microprocessor clock signal as an input to the central microprocessor for clocking the central microprocessor.« less

Clock Drawing in Spatial Neglect: A Comprehensive Analysis of Clock Perimeter, Placement, and Accuracy

PubMed Central

Chen, Peii; Goedert, Kelly M.

2012-01-01

Clock drawings produced by right-brain-damaged (RBD) individuals with spatial neglect often contain an abundance of empty space on the left while numbers and hands are placed on the right. However, the clock perimeter is rarely compromised in neglect patients’ drawings. By analyzing clock drawings produced by 71 RBD and 40 healthy adults, this study investigated whether the geometric characteristics of the clock perimeter reveal novel insights to understanding spatial neglect. Neglect participants drew smaller clocks than either healthy or non-neglect RBD participants. While healthy participants’ clock perimeter was close to circular, RBD participants drew radially extended ellipses. The mechanisms for these phenomena were investigated by examining the relation between clock-drawing characteristics and performance on six subtests of the Behavioral Inattention Test (BIT). The findings indicated that the clock shape was independent of any BIT subtest or the drawing placement on the test sheet and that the clock size was significantly predicted by one BIT subtest: the poorer the figure and shape copying, the smaller the clock perimeter. Further analyses revealed that in all participants, clocks decreased in size as they were placed farther from the center of the paper. However, even when neglect participants placed their clocks towards the center of the page, they were smaller than those produced by healthy or non-neglect RBD participants. These results suggest a neglect-specific reduction in the subjectively available workspace for graphic production from memory, consistent with the hypothesis that neglect patients are impaired in the ability to enlarge the attentional aperture. PMID:22390278

Automatic segmentation of time-lapse microscopy images depicting a live Dharma embryo.

PubMed

Zacharia, Eleni; Bondesson, Maria; Riu, Anne; Ducharme, Nicole A; Gustafsson, Jan-Åke; Kakadiaris, Ioannis A

2011-01-01

Biological inferences about the toxicity of chemicals reached during experiments on the zebrafish Dharma embryo can be greatly affected by the analysis of the time-lapse microscopy images depicting the embryo. Among the stages of image analysis, automatic and accurate segmentation of the Dharma embryo is the most crucial and challenging. In this paper, an accurate and automatic segmentation approach for the segmentation of the Dharma embryo data obtained by fluorescent time-lapse microscopy is proposed. Experiments performed in four stacks of 3D images over time have shown promising results.

Room 103, transom woodwork and original clock. All clocks are ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

Room 103, transom woodwork and original clock. All clocks are driven by a common signal. - San Bernardino Valley College, Life Science Building, 701 South Mount Vernon Avenue, San Bernardino, San Bernardino County, CA

Oxytocin reversed MK-801-induced social interaction and aggression deficits in zebrafish.

PubMed

Zimmermann, Fernanda Francine; Gaspary, Karina Vidarte; Siebel, Anna Maria; Bonan, Carla Denise

2016-09-15

Changes in social behavior occur in several neuropsychiatric disorders such as schizophrenia and autism. The interaction between individuals is an essential aspect and an adaptive response of several species, among them the zebrafish. Oxytocin is a neuroendocrine hormone associated with social behavior. The aim of the present study was to investigate the effects of MK-801, a non-competitive antagonist of glutamate NMDA receptors, on social interaction and aggression in zebrafish. We also examined the modulation of those effects by oxytocin, the oxytocin receptor agonist carbetocin and the oxytocin receptor antagonist L-368,899. Our results showed that MK-801 induced a decrease in the time spent in the segment closest to the conspecific school and in the time spent in the segment nearest to the mirror image, suggesting an effect on social behavior. The treatment with oxytocin after the exposure to MK-801 was able to reestablish the time spent in the segment closest to the conspecific school, as well as the time spent in the segment nearest to the mirror image. In addition, in support of the role of the oxytocin pathway in modulating those responses, we showed that the oxytocin receptor agonist carbetocin reestablished the social and aggressive behavioral deficits induced by MK-801. However, the oxytocin receptor antagonist L-368,899 was not able to reverse the behavioral changes induced by MK-801. This study supports the critical role for NMDA receptors and the oxytocinergic system in the regulation of social behavior and aggression which may be relevant for the mechanisms associated to autism and schizophrenia. Copyright © 2016 Elsevier B.V. All rights reserved.

Common features in diverse insect clocks.

PubMed

Numata, Hideharu; Miyazaki, Yosuke; Ikeno, Tomoko

2015-01-01

This review describes common features among diverse biological clocks in insects, including circadian, circatidal, circalunar/circasemilunar, and circannual clocks. These clocks control various behaviors, physiological functions, and developmental events, enabling adaptation to periodic environmental changes. Circadian clocks also function in time-compensation for celestial navigation and in the measurement of day or night length for photoperiodism. Phase response curves for such clocks reported thus far exhibit close similarities; specifically, the circannual clock in Anthrenus verbasci shows striking similarity to circadian clocks in its phase response. It is suggested that diverse biological clocks share physiological properties in their phase responses irrespective of period length. Molecular and physiological mechanisms are best understood for the optic-lobe and mid-brain circadian clocks, although there is no direct evidence that these clocks are involved in rhythmic phenomena other than circadian rhythms in daily events. Circadian clocks have also been localized in peripheral tissues, and research on their role in various rhythmic phenomena has been started. Although clock genes have been identified as controllers of circadian rhythms in daily events, some of these genes have also been shown to be involved in photoperiodism and possibly in time-compensated celestial navigation. In contrast, there is no experimental evidence indicating that any known clock gene is involved in biological clocks other than circadian clocks.

Afferent Connectivity of the Zebrafish Habenulae

PubMed Central

Turner, Katherine J.; Hawkins, Thomas A.; Yáñez, Julián; Anadón, Ramón; Wilson, Stephen W.; Folgueira, Mónica

2016-01-01

The habenulae are bilateral nuclei located in the dorsal diencephalon that are conserved across vertebrates. Here we describe the main afferents to the habenulae in larval and adult zebrafish. We observe afferents from the subpallium, nucleus rostrolateralis, posterior tuberculum, posterior hypothalamic lobe, median raphe; we also see asymmetric afferents from olfactory bulb to the right habenula, and from the parapineal to the left habenula. In addition, we find afferents from a ventrolateral telencephalic nucleus that neurochemical and hodological data identify as the ventral entopeduncular nucleus (vENT), confirming and extending observations of Amo et al. (2014). Fate map and marker studies suggest that vENT originates from the diencephalic prethalamic eminence and extends into the lateral telencephalon from 48 to 120 hour post-fertilization (hpf). No afferents to the habenula were observed from the dorsal entopeduncular nucleus (dENT). Consequently, we confirm that the vENT (and not the dENT) should be considered as the entopeduncular nucleus “proper” in zebrafish. Furthermore, comparison with data in other vertebrates suggests that the vENT is a conserved basal ganglia nucleus, being homologous to the entopeduncular nucleus of mammals (internal segment of the globus pallidus of primates) by both embryonic origin and projections, as previously suggested by Amo et al. (2014). PMID:27199671

USNO Master Clock - Naval Oceanography Portal

Science.gov Websites

section Advanced Search... Sections Home Time Earth Orientation Astronomy Meteorology Oceanography Ice You are here: Home › USNO › Precise Time › Master Clock USNO Logo USNO Navigation Master Clock GPS Display Clocks TWSTT Telephone Time NTP Info USNO Master Clock clock vault The USNO Master Clock is the

Cloning of zebrafish Mustn1 orthologs and their expression during early development.

PubMed

Camarata, Troy; Vasilyev, Aleksandr; Hadjiargyrou, Michael

2016-11-15

Mustn1 is a small nuclear protein that is involved in the development and regeneration of the musculoskeletal system. Previous work established a role for Mustn1 in myogenic and chondrogenic differentiation. In addition, recent evidence suggests a potential role for Mustn1 in cilia function in zebrafish. A detailed study of Mustn1 expression has yet to be conducted in zebrafish. As such, we report herein the cloning of the zebrafish Mustn1 orthologs, mustn1a and mustn1b, and their expression during zebrafish embryonic and larval development. Results indicate a 44% nucleotide identity between the two paralogs. Phylogenetic analysis further confirmed that the Mustn1a and 1b predicted proteins were highly related to other vertebrate members of the Mustn1 protein family. Whole mount in situ hybridization revealed expression of both mustn1a and 1b at the 7-somite stage through 72hpf in structures such as Kupffer's vesicle, segmental mesoderm, head structures, and otic vesicle. Additionally, in 5day old larva, mustn1a and 1b expression is detected in the neurocranium, otic capsule, and the gut. Although both were expressed in the neurocranium, mustn1a was localized in the hypophyseal fenestra whereas mustn1b was found near the posterior basicapsular commissure. mustn1b also displayed expression in the ceratohyal and ceratobranchial elements of the pharyngeal skeleton. These expression patterns were verified temporally by q-PCR analysis. Taken together, we conclude that Mustn1 expression is conserved in vertebrates and that the variations in expression of the two zebrafish paralogs suggest different modes of molecular regulation. Copyright © 2016 Elsevier B.V. All rights reserved.

Neurochemical measurements in the zebrafish brain

PubMed Central

Jones, Lauren J.; McCutcheon, James E.; Young, Andrew M. J.; Norton, William H. J.

2015-01-01

The zebrafish is an ideal model organism for behavioral genetics and neuroscience. The high conservation of genes and neurotransmitter pathways between zebrafish and other vertebrates permits the translation of research between species. Zebrafish behavior can be studied at both larval and adult stages and recent research has begun to establish zebrafish models for human disease. Fast scan cyclic voltammetry (FSCV) is an electrochemical technique that permits the detection of neurotransmitter release and reuptake. In this study we have used in vitro FSCV to measure the release of analytes in the adult zebrafish telencephalon. We compare different stimulation methods and present a characterization of neurochemical changes in the wild-type zebrafish brain. This study represents the first FSCV recordings in zebrafish, thus paving the way for neurochemical analysis of the fish brain. PMID:26441575

Zebrafish as tools for drug discovery.

PubMed

MacRae, Calum A; Peterson, Randall T

2015-10-01

The zebrafish has become a prominent vertebrate model for disease and has already contributed to several examples of successful phenotype-based drug discovery. For the zebrafish to become useful in drug development more broadly, key hurdles must be overcome, including a more comprehensive elucidation of the similarities and differences between human and zebrafish biology. Recent studies have begun to establish the capabilities and limitations of zebrafish for disease modelling, drug screening, target identification, pharmacology, and toxicology. As our understanding increases and as the technologies for manipulating zebrafish improve, it is hoped that the zebrafish will have a key role in accelerating the emergence of precision medicine.

Countdown Clock Ribbon Cutting

NASA Image and Video Library

2016-03-01

Confetti is launched as the spaceport's historic countdown clock is dedicated as the newest display at the Kennedy Space Center Visitor Complex. Now located at the entrance to the visitor complex, the spaceport's historic countdown clock was used starting with the launch of Apollo 12 on Nov. 14, 1969. Originally set up at the space center's Press Site, the clock operated through the final space shuttle mission, STS-135, launched on July 8, 2011. The old countdown clock was replaced in 2014 with a modern light emitting diode, or LED, display.

Physiological links of circadian clock and biological clock of aging.

PubMed

Liu, Fang; Chang, Hung-Chun

2017-07-01

Circadian rhythms orchestrate biochemical and physiological processes in living organisms to respond the day/night cycle. In mammals, nearly all cells hold self-sustained circadian clocks meanwhile couple the intrinsic rhythms to systemic changes in a hierarchical manner. The suprachiasmatic nucleus (SCN) of the hypothalamus functions as the master pacemaker to initiate daily synchronization according to the photoperiod, in turn determines the phase of peripheral cellular clocks through a variety of signaling relays, including endocrine rhythms and metabolic cycles. With aging, circadian desynchrony occurs at the expense of peripheral metabolic pathologies and central neurodegenerative disorders with sleep symptoms, and genetic ablation of circadian genes in model organisms resembled the aging-related features. Notably, a number of studies have linked longevity nutrient sensing pathways in modulating circadian clocks. Therapeutic strategies that bridge the nutrient sensing pathways and circadian clock might be rational designs to defy aging.

Non-invasive imaging of zebrafish with spinal deformities using optical coherence tomography: a preliminary study

NASA Astrophysics Data System (ADS)

Bernstein, Liane; Beaudette, Kathy; Patten, Kessen; Beaulieu-Ouellet, Émilie; Strupler, Mathias; Moldovan, Florina; Boudoux, Caroline

2013-03-01

A zebrafish model has recently been introduced to study various genetic mutations that could lead to spinal deformities such as scoliosis. However, current imaging techniques make it difficult to perform longitudinal studies of this condition in zebrafish, especially in the early stages of development. The goal of this project is to determine whether optical coherence tomography (OCT) is a viable non-invasive method to image zebrafish exhibiting spinal deformities. Images of both live and fixed malformed zebrafish (5 to 21 days postfertilization) as well as wild-type fish (5 to 29 days postfertilization) were acquired non-invasively using a commercial SD-OCT system, with a laser source centered at 930nm (λ=100nm), permitting axial and lateral resolutions of 7 and 8μm respectively. Using two-dimensional images and three-dimensional reconstructions, it was possible to identify the malformed notochord as well as deformities in other major organs at different stages of formation. Visualization of the notochord was facilitated with the development of a segmentation algorithm. OCT images were compared to HE histological sections and images obtained by calcein staining. Because of the possibility of performing longitudinal studies on a same fish and reducing image processing time as compared with staining techniques and histology, the use of OCT could facilitate phenotypic characterization in studying genetic factors leading to spinal deformities in zebrafish and could eventually contribute to the identification of the genetic causes of spinal deformities such as scoliosis.

Entanglement of quantum clocks through gravity

NASA Astrophysics Data System (ADS)

Castro Ruiz, Esteban; Giacomini, Flaminia; Brukner, Časlav

2017-03-01

In general relativity, the picture of space-time assigns an ideal clock to each world line. Being ideal, gravitational effects due to these clocks are ignored and the flow of time according to one clock is not affected by the presence of clocks along nearby world lines. However, if time is defined operationally, as a pointer position of a physical clock that obeys the principles of general relativity and quantum mechanics, such a picture is, at most, a convenient fiction. Specifically, we show that the general relativistic mass-energy equivalence implies gravitational interaction between the clocks, whereas the quantum mechanical superposition of energy eigenstates leads to a nonfixed metric background. Based only on the assumption that both principles hold in this situation, we show that the clocks necessarily get entangled through time dilation effect, which eventually leads to a loss of coherence of a single clock. Hence, the time as measured by a single clock is not well defined. However, the general relativistic notion of time is recovered in the classical limit of clocks.

Entanglement of quantum clocks through gravity.

PubMed

Castro Ruiz, Esteban; Giacomini, Flaminia; Brukner, Časlav

2017-03-21

In general relativity, the picture of space-time assigns an ideal clock to each world line. Being ideal, gravitational effects due to these clocks are ignored and the flow of time according to one clock is not affected by the presence of clocks along nearby world lines. However, if time is defined operationally, as a pointer position of a physical clock that obeys the principles of general relativity and quantum mechanics, such a picture is, at most, a convenient fiction. Specifically, we show that the general relativistic mass-energy equivalence implies gravitational interaction between the clocks, whereas the quantum mechanical superposition of energy eigenstates leads to a nonfixed metric background. Based only on the assumption that both principles hold in this situation, we show that the clocks necessarily get entangled through time dilation effect, which eventually leads to a loss of coherence of a single clock. Hence, the time as measured by a single clock is not well defined. However, the general relativistic notion of time is recovered in the classical limit of clocks.

Entanglement of quantum clocks through gravity

PubMed Central

Castro Ruiz, Esteban; Giacomini, Flaminia; Brukner, Časlav

2017-01-01

In general relativity, the picture of space–time assigns an ideal clock to each world line. Being ideal, gravitational effects due to these clocks are ignored and the flow of time according to one clock is not affected by the presence of clocks along nearby world lines. However, if time is defined operationally, as a pointer position of a physical clock that obeys the principles of general relativity and quantum mechanics, such a picture is, at most, a convenient fiction. Specifically, we show that the general relativistic mass–energy equivalence implies gravitational interaction between the clocks, whereas the quantum mechanical superposition of energy eigenstates leads to a nonfixed metric background. Based only on the assumption that both principles hold in this situation, we show that the clocks necessarily get entangled through time dilation effect, which eventually leads to a loss of coherence of a single clock. Hence, the time as measured by a single clock is not well defined. However, the general relativistic notion of time is recovered in the classical limit of clocks. PMID:28270623

Countdown Clock Ribbon Cutting

NASA Image and Video Library

2016-03-01

Therrin Protze, chief operating officer of the Kennedy Space Center Visitor Complex, speaks at the dedication of the center's historic countdown clock. To the right is space center director Bob Cabana. Now located at the entrance to the visitor complex, the spaceport's historic countdown clock was used starting with the launch of Apollo 12 on Nov. 14, 1969. Originally set up at the space center's Press Site, the clock was used through the final space shuttle mission, STS-135, launched on July 8, 2011. The old countdown clock was replaced in 2014 with a modern light emitting diode, or LED, display.

Countdown Clock Ribbon Cutting

NASA Image and Video Library

2016-03-01

Therrin Protze, chief operating officer of the Kennedy Space Center Visitor Complex, left, and center director Bob Cabana watch as confetti was launched as the spaceport's historic countdown clock is dedicated as the newest display at the entrance to Kennedy's visitor complex. The spaceport's historic countdown clock was used beginning with the launch of Apollo 12 on Nov. 14, 1969. Originally set up at the space center's Press Site, the clock was used through the final space shuttle mission, STS-135, launched on July 8, 2011. The old countdown clock was replaced in 2014 with a modern light emitting diode, or LED, display.

Master/slave clock arrangement for providing reliable clock signal

NASA Technical Reports Server (NTRS)

Abbey, Duane L. (Inventor)

1977-01-01

The outputs of two like frequency oscillators are combined to form a single reliable clock signal, with one oscillator functioning as a slave under the control of the other to achieve phase coincidence when the master is operative and in a free-running mode when the master is inoperative so that failure of either oscillator produces no effect on the clock signal.

Single-transistor-clocked flip-flop

DOEpatents

Zhao, Peiyi; Darwish, Tarek; Bayoumi, Magdy

2005-08-30

The invention provides a low power, high performance flip-flop. The flip-flop uses only one clocked transistor. The single clocked transistor is shared by the first and second branches of the device. A pulse generator produces a clock pulse to trigger the flip-flop. In one preferred embodiment the device can be made as a static explicit pulsed flip-flop which employs only two clocked transistors.

A Light Clock Satisfying the Clock Hypothesis of Special Relativity

ERIC Educational Resources Information Center

West, Joseph

2007-01-01

The design of the FMEL, a floor-mirrored Einstein-Langevin "light clock", is introduced. The clock provides a physically intuitive manner to calculate and visualize the time dilation effects for a spatially extended set of observers (an accelerated "frame") undergoing unidirectional acceleration or observers on a rotating cylinder of constant…

The Clock Mission Optis

NASA Astrophysics Data System (ADS)

Dittus, Hansjörg; Lämmerzahl, Claus

Clocks are an almost universal tool for exploring the fundamental structure of theories related to relativity. For future clock experiments, it is important for them to be performed in space. One mission which has the capability to perform and improve all relativity tests based on clocks by several orders of magnitude is OPTIS. These tests consist of (i) tests of the isotropy of light propagation (from which information about the matter sector which the optical resonators are made of can also be drawn), (ii) tests of the constancy of the speed of light, (iii) tests of the universality of the gravitational redshift by comparing clocks based on light propagation, like light clocks and various atomic clocks, (iv) time dilation based on the Doppler effect, (v) measuring the absolute gravitational redshift, (vi) measuring the perihelion advance of the satellite's orbit by using very precise tracking techniques, (vii) measuring the Lense-Thirring effect, and (viii) testing Newton's gravitational potential law on the scale of Earth-bound satellites. The corresponding tests are not only important for fundamental physics but also indispensable for practical purposes like navigation, Earth sciences, metrology, etc.

Countdown Clock Ribbon Cutting

NASA Image and Video Library

2016-03-01

Kennedy Space Center Director Bob Cabana, left, and Therrin Protze, chief operating officer of Kennedy's Visitor Complex, celebrate the dedication of the spaceport's historic countdown clock as the newest display at the center's visitor complex. Now located at the entrance to the visitor complex, the spaceport's historic countdown clock was used starting with the launch of Apollo 12 on Nov. 14, 1969. Originally set up at the space center's Press Site, the clock operated through the final space shuttle mission, STS-135, launched on July 8, 2011. The old countdown clock was replaced in 2014 with a modern light emitting diode, or LED, display.

The space optical clocks project

NASA Astrophysics Data System (ADS)

Schiller, S.; Tino, G. M.; Lemonde, P.; Sterr, U.; Lisdat, Ch.; Görlitz, A.; Poli, N.; Nevsky, A.; Salomon, C.

2017-11-01

The Space Optical Clocks project aims at operating lattice clocks on the ISS for tests of fundamental physics and for providing high-accuracy comparisons of future terrestrial optical clocks. A pre-phase-A study (2007- 10), funded partially by ESA and DLR, included the implementation of several optical lattice clock systems using Strontium and Ytterbium as atomic species and their characterization. Subcomponents of clock demonstrators with the added specification of transportability and using techniques suitable for later space use, such as all-solid-state lasers, low power consumption, and compact dimensions, have been developed and have been validated. This included demonstration of laser-cooling and magneto-optical trapping of Sr atoms in a compact breadboard apparatus and demonstration of a transportable clock laser with 1 Hz linewidth. With two laboratory Sr lattice clock systems a number of fundamental results were obtained, such as observing atomic resonances with linewidths as low as 3 Hz, non-destructive detection of atom excitation, determination of decoherence effects and reaching a frequency instability of 1×10-16.

Experimental validation of clock synchronization algorithms

NASA Technical Reports Server (NTRS)

Palumbo, Daniel L.; Graham, R. Lynn

1992-01-01

The objective of this work is to validate mathematically derived clock synchronization theories and their associated algorithms through experiment. Two theories are considered, the Interactive Convergence Clock Synchronization Algorithm and the Midpoint Algorithm. Special clock circuitry was designed and built so that several operating conditions and failure modes (including malicious failures) could be tested. Both theories are shown to predict conservative upper bounds (i.e., measured values of clock skew were always less than the theory prediction). Insight gained during experimentation led to alternative derivations of the theories. These new theories accurately predict the behavior of the clock system. It is found that a 100 percent penalty is paid to tolerate worst-case failures. It is also shown that under optimal conditions (with minimum error and no failures) the clock skew can be as much as three clock ticks. Clock skew grows to six clock ticks when failures are present. Finally, it is concluded that one cannot rely solely on test procedures or theoretical analysis to predict worst-case conditions.

Zebrafish and Streptococcal Infections.

PubMed

Saralahti, A; Rämet, M

2015-09-01

Streptococcal bacteria are a versatile group of gram-positive bacteria capable of infecting several host organisms, including humans and fish. Streptococcal species are common colonizers of the human respiratory and gastrointestinal tract, but they also cause some of the most common life-threatening, invasive infections in humans and aquaculture. With its unique characteristics and efficient tools for genetic and imaging applications, the zebrafish (Danio rerio) has emerged as a powerful vertebrate model for infectious diseases. Several zebrafish models introduced so far have shown that zebrafish are suitable models for both zoonotic and human-specific infections. Recently, several zebrafish models mimicking human streptococcal infections have also been developed. These models show great potential in providing novel information about the pathogenic mechanisms and host responses associated with human streptococcal infections. Here, we review the zebrafish infection models for the most relevant streptococcal species: the human-specific Streptococcus pneumoniae and Streptococcus pyogenes, and the zoonotic Streptococcus iniae and Streptococcus agalactiae. The recent success and the future potential of these models for the study of host-pathogen interactions in streptococcal infections are also discussed. © 2015 The Foundation for the Scandinavian Journal of Immunology.

Measuring zebrafish turning rate.

PubMed

Mwaffo, Violet; Butail, Sachit; di Bernardo, Mario; Porfiri, Maurizio

2015-06-01

Zebrafish is becoming a popular animal model in preclinical research, and zebrafish turning rate has been proposed for the analysis of activity in several domains. The turning rate is often estimated from the trajectory of the fish centroid that is output by commercial or custom-made target tracking software run on overhead videos of fish swimming. However, the accuracy of such indirect methods with respect to the turning rate associated with changes in heading during zebrafish locomotion is largely untested. Here, we compare two indirect methods for the turning rate estimation using the centroid velocity or position data, with full shape tracking for three different video sampling rates. We use tracking data from the overhead video recorded at 60, 30, and 15 frames per second of zebrafish swimming in a shallow water tank. Statistical comparisons of absolute turning rate across methods and sampling rates indicate that, while indirect methods are indistinguishable from full shape tracking, the video sampling rate significantly influences the turning rate measurement. The results of this study can aid in the selection of the video capture frame rate, an experimental design parameter in zebrafish behavioral experiments where activity is an important measure.

INDUCED AND SPONTANEOUS NEOPLASIA IN ZEBRAFISH.

EPA Science Inventory

To address the potential of zebrafish as a cancer model, it is important to determine the susceptibility of zebrafish to tumors, and to compare zebrafish tumors with human tumors. To determine whether the commonly-used germ line mutagen, ethylnitrosourea (ENU) induces tumors, we ...

Automatic control of clock duty cycle

NASA Technical Reports Server (NTRS)

Feng, Xiaoxin (Inventor); Roper, Weston (Inventor); Seefeldt, James D. (Inventor)

2010-01-01

In general, this disclosure is directed to a duty cycle correction (DCC) circuit that adjusts a falling edge of a clock signal to achieve a desired duty cycle. In some examples, the DCC circuit may generate a pulse in response to a falling edge of an input clock signal, delay the pulse based on a control voltage, adjust the falling edge of the input clock signal based on the delayed pulse to produce an output clock signal, and adjust the control voltage based on the difference between a duty cycle of the output clock signal and a desired duty cycle. Since the DCC circuit adjusts the falling edge of the clock cycle to achieve a desired duty cycle, the DCC may be incorporated into existing PLL control loops that adjust the rising edge of a clock signal without interfering with the operation of such PLL control loops.

Circadian molecular clock in lung pathophysiology

PubMed Central

Sundar, Isaac K.; Yao, Hongwei; Sellix, Michael T.

2015-01-01

Disrupted daily or circadian rhythms of lung function and inflammatory responses are common features of chronic airway diseases. At the molecular level these circadian rhythms depend on the activity of an autoregulatory feedback loop oscillator of clock gene transcription factors, including the BMAL1:CLOCK activator complex and the repressors PERIOD and CRYPTOCHROME. The key nuclear receptors and transcription factors REV-ERBα and RORα regulate Bmal1 expression and provide stability to the oscillator. Circadian clock dysfunction is implicated in both immune and inflammatory responses to environmental, inflammatory, and infectious agents. Molecular clock function is altered by exposomes, tobacco smoke, lipopolysaccharide, hyperoxia, allergens, bleomycin, as well as bacterial and viral infections. The deacetylase Sirtuin 1 (SIRT1) regulates the timing of the clock through acetylation of BMAL1 and PER2 and controls the clock-dependent functions, which can also be affected by environmental stressors. Environmental agents and redox modulation may alter the levels of REV-ERBα and RORα in lung tissue in association with a heightened DNA damage response, cellular senescence, and inflammation. A reciprocal relationship exists between the molecular clock and immune/inflammatory responses in the lungs. Molecular clock function in lung cells may be used as a biomarker of disease severity and exacerbations or for assessing the efficacy of chronotherapy for disease management. Here, we provide a comprehensive overview of clock-controlled cellular and molecular functions in the lungs and highlight the repercussions of clock disruption on the pathophysiology of chronic airway diseases and their exacerbations. Furthermore, we highlight the potential for the molecular clock as a novel chronopharmacological target for the management of lung pathophysiology. PMID:26361874

Optical atomic clocks

NASA Astrophysics Data System (ADS)

Poli, N.; Oates, C. W.; Gill, P.; Tino, G. M.

2013-12-01

In the last ten years extraordinary results in time and frequency metrology have been demonstrated. Frequency-stabilization techniques for continuous-wave lasers and femtosecond optical frequency combs have enabled a rapid development of frequency standards based on optical transitions in ultra-cold neutral atoms and trapped ions. As a result, today's best performing atomic clocks tick at an optical rate and allow scientists to perform high-resolution measurements with a precision approaching a few parts in 1018. This paper reviews the history and the state of the art in optical-clock research and addresses the implementation of optical clocks in a possible future redefinition of the SI second as well as in tests of fundamental physics.

Countdown Clock Ribbon Cutting

NASA Image and Video Library

2016-03-01

The newest display at the Kennedy Space Center Visitor Complex is the spaceport's historic countdown clock. It is now located at the entrance to the visitor complex. The clock was set up at the space center's Press Site and used from the launch of Apollo 12 on Nov. 14, 1969 to the final space shuttle mission, STS-135, launched on July 8, 2011. The old countdown clock was replaced in 2014 with a modern light emitting diode, or LED, display.

Anesthesia and euthanasia in zebrafish.

PubMed

Matthews, Monte; Varga, Zoltán M

2012-01-01

Because of the relative ease of embryonic manipulation and observation, the ability to produce a great number of genetic mutations, efficient screening methods, and the continued advance of molecular genetic tools, such as the progress in sequencing and mapping of the zebrafish genome, the use of zebrafish (Danio rerio) as a biomedical model organism continues to expand. However, studies involving zebrafish husbandry and veterinary care struggle to keep pace with scientific progress. This article outlines some of the current, acceptable methods for providing anesthesia and euthanasia and provides some examples of how performance-based approaches can be used to advance the relatively limited number of anesthetic and euthanizing techniques available for zebrafish.

Egyptian "Star Clocks"

NASA Astrophysics Data System (ADS)

Symons, Sarah

Diagonal, transit, and Ramesside star clocks are tables of astronomical information occasionally found in ancient Egyptian temples, tombs, and papyri. The tables represent the motions of selected stars (decans and hour stars) throughout the Egyptian civil year. Analysis of star clocks leads to greater understanding of ancient Egyptian constellations, ritual astronomical activities, observational practices, and pharaonic chronology.

Maintenance of Zebrafish Lines at the European Zebrafish Resource Center.

PubMed

Geisler, Robert; Borel, Nadine; Ferg, Marco; Maier, Jana Viktoria; Strähle, Uwe

2016-07-01

We have established a European Zebrafish Resource Center (EZRC) at the KIT. This center not only maintains and distributes a large number of existing mutant and transgenic zebrafish lines but also gives zebrafish researchers access to screening services and technologies such as imaging and high-throughput sequencing, provided by the Institute of Toxicology and Genetics (ITG). The EZRC maintains and distributes the stock collection of the Nüsslein-Volhard laboratory, comprising over 2000 publicly released mutations, as frozen sperm samples. Within the framework of the ZF-HEALTH EU project, the EZRC distributes over 10,000 knockout mutations from the Sanger Institute (United Kingdom), as well as over 100 mutant and transgenic lines from other sources. In this article, we detail the measures we have taken to ensure the health of our fish, including hygiene, quarantine, and veterinary inspections.

Single-ion, transportable optical atomic clocks

NASA Astrophysics Data System (ADS)

Delehaye, Marion; Lacroûte, Clément

2018-03-01

For the past 15 years, tremendous progress within the fields of laser stabilization, optical frequency combs and atom cooling and trapping have allowed the realization of optical atomic clocks with unrivaled performances. These instruments can perform frequency comparisons with fractional uncertainties well below ?, finding applications in fundamental physics tests, relativistic geodesy and time and frequency metrology. Even though most optical clocks are currently laboratory setups, several proposals for using these clocks for field measurements or within an optical clock network have been published, and most of time and frequency metrology institutes have started to develop transportable optical clocks. For the purpose of this special issue, we chose to focus on trapped-ion optical clocks. Even though their short-term fractional frequency stability is impaired by a lower signal-to-noise ratio, they offer a high potential for compactness: trapped ions demand low optical powers and simple loading schemes, and can be trapped in small vacuum chambers. We review recent advances on the clock key components, including ion trap and ultra-stable optical cavity, as well as existing projects and experiments which draw the picture of what future transportable, single-ion optical clocks may resemble.

Gigabit Ethernet Asynchronous Clock Compensation FIFO

NASA Technical Reports Server (NTRS)

Duhachek, Jeff

2012-01-01

Clock compensation for Gigabit Ethernet is necessary because the clock recovered from the 1.25 Gb/s serial data stream has the potential to be 200 ppm slower or faster than the system clock. The serial data is converted to 10-bit parallel data at a 125 MHz rate on a clock recovered from the serial data stream. This recovered data needs to be processed by a system clock that is also running at a nominal rate of 125 MHz, but not synchronous to the recovered clock. To cross clock domains, an asynchronous FIFO (first-in-first-out) is used, with the write pointer (wprt) in the recovered clock domain and the read pointer (rptr) in the system clock domain. Because the clocks are generated from separate sources, there is potential for FIFO overflow or underflow. Clock compensation in Gigabit Ethernet is possible by taking advantage of the protocol data stream features. There are two distinct data streams that occur in Gigabit Ethernet where identical data is transmitted for a period of time. The first is configuration, which happens during auto-negotiation. The second is idle, which occurs at the end of auto-negotiation and between every packet. The identical data in the FIFO can be repeated by decrementing the read pointer, thus compensating for a FIFO that is draining too fast. The identical data in the FIFO can also be skipped by incrementing the read pointer, which compensates for a FIFO draining too slowly. The unique and novel features of this FIFO are that it works in both the idle stream and the configuration streams. The increment or decrement of the read pointer is different in the idle and compensation streams to preserve disparity. Another unique feature is that the read pointer to write pointer difference range changes between compensation and idle to minimize FIFO latency during packet transmission.

The Circadian Clock Coordinates Ribosome Biogenesis

PubMed Central

Symul, Laura; Martin, Eva; Atger, Florian; Naef, Felix; Gachon, Frédéric

2013-01-01

Biological rhythms play a fundamental role in the physiology and behavior of most living organisms. Rhythmic circadian expression of clock-controlled genes is orchestrated by a molecular clock that relies on interconnected negative feedback loops of transcription regulators. Here we show that the circadian clock exerts its function also through the regulation of mRNA translation. Namely, the circadian clock influences the temporal translation of a subset of mRNAs involved in ribosome biogenesis by controlling the transcription of translation initiation factors as well as the clock-dependent rhythmic activation of signaling pathways involved in their regulation. Moreover, the circadian oscillator directly regulates the transcription of ribosomal protein mRNAs and ribosomal RNAs. Thus the circadian clock exerts a major role in coordinating transcription and translation steps underlying ribosome biogenesis. PMID:23300384

An analysis of clock-shift experiments: is scatter increased and deflection reduced in clock-shifted homing pigeons?

PubMed

Chappell

1997-01-01

Clock-shifting (altering the phase of the internal clock) in homing pigeons leads to a deflection in the vanishing bearing of the clock-shifted group relative to controls. However, two unexplained phenomena are common in clock-shift experiments: the vanishing bearings of the clock-shifted group are often more scattered (with a shorter vector length) than those of the control group, and the deflection of the mean bearing of the clock-shifted group from that of the controls is often smaller than expected theoretically. Here, an analysis of 55 clock-shift experiments performed in four countries over 21 years is reported. The bearings of the clock-shifted groups were significantly more scattered than those of controls and less deflected than expected, but these effects were not significantly different at familiar and unfamiliar sites. The possible causes of the effects are discussed and evaluated with reference to this analysis and other experiments. The most likely causes appear to be conflict between the directions indicated by the sun compass and either unshifted familiar visual landmarks (at familiar sites only) or the unshifted magnetic compass (possible at both familiar and unfamiliar sites).

Molecular cogs of the insect circadian clock.

PubMed

Shirasu, Naoto; Shimohigashi, Yasuyuki; Tominaga, Yoshiya; Shimohigashi, Miki

2003-08-01

During the last five years, enormous progress has been made in understanding the molecular basis of circadian systems, mainly by molecular genetic studies using the mouse and fly. Extensive evidence has revealed that the core clock machinery involves "clock genes" and "clock proteins" functioning as molecular cogs. These participate in transcriptional/translational feedback loops and many homologous clock-components in the fruit fly Drosophila are also expressed in mammalian clock tissues with circadian rhythms. Thus, the mechanisms of the central clock seem to be conserved across animal kingdom. However, some recent studies imply that the present widely accepted molecular models of circadian clocks may not always be supported by the experimental evidence.

High Performance Clocks and Gravity Field Determination

NASA Astrophysics Data System (ADS)

Müller, J.; Dirkx, D.; Kopeikin, S. M.; Lion, G.; Panet, I.; Petit, G.; Visser, P. N. A. M.

2018-02-01

Time measured by an ideal clock crucially depends on the gravitational potential and velocity of the clock according to general relativity. Technological advances in manufacturing high-precision atomic clocks have rapidly improved their accuracy and stability over the last decade that approached the level of 10^{-18}. This notable achievement along with the direct sensitivity of clocks to the strength of the gravitational field make them practically important for various geodetic applications that are addressed in the present paper. Based on a fully relativistic description of the background gravitational physics, we discuss the impact of those highly-precise clocks on the realization of reference frames and time scales used in geodesy. We discuss the current definitions of basic geodetic concepts and come to the conclusion that the advances in clocks and other metrological technologies will soon require the re-definition of time scales or, at least, clarification to ensure their continuity and consistent use in practice. The relative frequency shift between two clocks is directly related to the difference in the values of the gravity potential at the points of clock's localization. According to general relativity the relative accuracy of clocks in 10^{-18} is equivalent to measuring the gravitational red shift effect between two clocks with the height difference amounting to 1 cm. This makes the clocks an indispensable tool in high-precision geodesy in addition to laser ranging and space geodetic techniques. We show how clock measurements can provide geopotential numbers for the realization of gravity-field-related height systems and can resolve discrepancies in classically-determined height systems as well as between national height systems. Another application of clocks is the direct use of observed potential differences for the improved recovery of regional gravity field solutions. Finally, clock measurements for space-borne gravimetry are analyzed along with

Regulation of monoamine oxidase A by circadian-clock components implies clock influence on mood.

PubMed

Hampp, Gabriele; Ripperger, Jürgen A; Houben, Thijs; Schmutz, Isabelle; Blex, Christian; Perreau-Lenz, Stéphanie; Brunk, Irene; Spanagel, Rainer; Ahnert-Hilger, Gudrun; Meijer, Johanna H; Albrecht, Urs

2008-05-06

The circadian clock has been implicated in addiction and several forms of depression [1, 2], indicating interactions between the circadian and the reward systems in the brain [3-5]. Rewards such as food, sex, and drugs influence this system in part by modulating dopamine neurotransmission in the mesolimbic dopamine reward circuit, including the ventral tegmental area (VTA) and the ventral striatum (NAc). Hence, changes in dopamine levels in these brain areas are proposed to influence mood in humans and mice [6-10]. To establish a molecular link between the circadian-clock mechanism and dopamine metabolism, we analyzed the murine promoters of genes encoding key enzymes important in dopamine metabolism. We find that transcription of the monoamine oxidase A (Maoa) promoter is regulated by the clock components BMAL1, NPAS2, and PER2. A mutation in the clock gene Per2 in mice leads to reduced expression and activity of MAOA in the mesolimbic dopaminergic system. Furthermore, we observe increased levels of dopamine and altered neuronal activity in the striatum, and these results probably lead to behavioral alterations observed in Per2 mutant mice in despair-based tests. These findings suggest a role of circadian-clock components in dopamine metabolism highlighting a role of the clock in regulating mood-related behaviors.

Wnt-regulated dynamics of positional information in zebrafish somitogenesis

PubMed Central

Bajard, Lola; Morelli, Luis G.; Ares, Saúl; Pécréaux, Jacques; Jülicher, Frank; Oates, Andrew C.

2014-01-01

How signaling gradients supply positional information in a field of moving cells is an unsolved question in patterning and morphogenesis. Here, we ask how a Wnt signaling gradient regulates the dynamics of a wavefront of cellular change in a flow of cells during somitogenesis. Using time-controlled perturbations of Wnt signaling in the zebrafish embryo, we changed segment length without altering the rate of somite formation or embryonic elongation. This result implies specific Wnt regulation of the wavefront velocity. The observed Wnt signaling gradient dynamics and timing of downstream events support a model for wavefront regulation in which cell flow plays a dominant role in transporting positional information. PMID:24595291

Viral Diseases in Zebrafish: What Is Known and Unknown

PubMed Central

Crim, Marcus J.; Riley, Lela K.

2013-01-01

Naturally occurring viral infections have the potential to introduce confounding variability that leads to invalid and misinterpreted data. Whereas the viral diseases of research rodents are well characterized and closely monitored, no naturally occurring viral infections have been characterized for the laboratory zebrafish (Danio rerio), an increasingly important biomedical research model. Despite the ignorance about naturally occurring zebrafish viruses, zebrafish models are rapidly expanding in areas of biomedical research where the confounding effects of unknown infectious agents present a serious concern. In addition, many zebrafish research colonies remain linked to the ornamental (pet) zebrafish trade, which can contribute to the introduction of new pathogens into research colonies, whereas mice used for research are purpose bred, with no introduction of new mice from the pet industry. Identification, characterization, and monitoring of naturally occurring viruses in zebrafish are crucial to the improvement of zebrafish health, the reduction of unwanted variability, and the continued development of the zebrafish as a model organism. This article addresses the importance of identifying and characterizing the viral diseases of zebrafish as the scope of zebrafish models expands into new research areas and also briefly addresses zebrafish susceptibility to experimental viral infection and the utility of the zebrafish as an infection and immunology model. PMID:23382345

Segregation of Clock and Non-Clock Regulatory Functions of REV-ERB.

PubMed

Butler, Andrew A; Burris, Thomas P

2015-08-04

The molecular clock is a master controller of circadian cellular processes that affect growth, metabolic homeostasis, and behavior. A report in Science by Zhang et al. (2015) redefines our understanding of how Rev-erba acts as an internal feedback inhibitor that modulates activity of the core clock while simultaneously regulating tissue-specific metabolic processes. Copyright © 2015 Elsevier Inc. All rights reserved.

Tight real-time synchronization of a microwave clock to an optical clock across a turbulent air path

PubMed Central

Bergeron, Hugo; Sinclair, Laura C.; Swann, William C.; Nelson, Craig W.; Deschênes, Jean-Daniel; Baumann, Esther; Giorgetta, Fabrizio R.; Coddington, Ian; Newbury, Nathan R.

2018-01-01

The ability to distribute the precise time and frequency from an optical clock to remote platforms could enable future precise navigation and sensing systems. Here we demonstrate tight, real-time synchronization of a remote microwave clock to a master optical clock over a turbulent 4-km open air path via optical two-way time-frequency transfer. Once synchronized, the 10-GHz frequency signals generated at each site agree to 10−14 at one second and below 10−17 at 1000 seconds. In addition, the two clock times are synchronized to ±13 fs over an 8-hour period. The ability to phase-synchronize 10-GHz signals across platforms supports future distributed coherent sensing, while the ability to time-synchronize multiple microwave-based clocks to a high-performance master optical clock supports future precision navigation/timing systems. PMID:29607352

Tight real-time synchronization of a microwave clock to an optical clock across a turbulent air path.

PubMed

Bergeron, Hugo; Sinclair, Laura C; Swann, William C; Nelson, Craig W; Deschênes, Jean-Daniel; Baumann, Esther; Giorgetta, Fabrizio R; Coddington, Ian; Newbury, Nathan R

2016-04-01

The ability to distribute the precise time and frequency from an optical clock to remote platforms could enable future precise navigation and sensing systems. Here we demonstrate tight, real-time synchronization of a remote microwave clock to a master optical clock over a turbulent 4-km open air path via optical two-way time-frequency transfer. Once synchronized, the 10-GHz frequency signals generated at each site agree to 10 -14 at one second and below 10 -17 at 1000 seconds. In addition, the two clock times are synchronized to ±13 fs over an 8-hour period. The ability to phase-synchronize 10-GHz signals across platforms supports future distributed coherent sensing, while the ability to time-synchronize multiple microwave-based clocks to a high-performance master optical clock supports future precision navigation/timing systems.

A Role for Timely Nuclear Translocation of Clock Repressor Proteins in Setting Circadian Clock Speed

PubMed Central

Lee, Euna

2014-01-01

By means of a circadian clock system, all the living organisms on earth including human beings can anticipate the environmental rhythmic changes such as light/dark and warm/cold periods in a daily as well as in a yearly manner. Anticipating such environmental changes provide organisms with survival benefits via manifesting behavior and physiology at an advantageous time of the day and year. Cell-autonomous circadian oscillators, governed by transcriptional feedback loop composed of positive and negative elements, are organized into a hierarchical system throughout the organisms and generate an oscillatory expression of a clock gene by itself as well as clock controlled genes (ccgs) with a 24 hr periodicity. In the feedback loop, hetero-dimeric transcription factor complex induces the expression of negative regulatory proteins, which in turn represses the activity of transcription factors to inhibit their own transcription. Thus, for robust oscillatory rhythms of the expression of clock genes as well as ccgs, the precise control of subcellular localization and/or timely translocation of core clock protein are crucial. Here, we discuss how sub-cellular localization and nuclear translocation are controlled in a time-specific manner focusing on the negative regulatory clock proteins. PMID:25258565

Code-Phase Clock Bias and Frequency Offset in PPP Clock Solutions.

PubMed

Defraigne, Pascale; Sleewaegen, Jean-Marie

2016-07-01

Precise point positioning (PPP) is a zero-difference single-station technique that has proved to be very effective for time and frequency transfer, enabling the comparison of atomic clocks with a precision of a hundred picoseconds and a one-day stability below the 1e-15 level. It was, however, noted that for some receivers, a frequency difference is observed between the clock solution based on the code measurements and the clock solution based on the carrier-phase measurements. These observations reveal some inconsistency either between the code and carrier phases measured by the receiver or between the data analysis strategy of codes and carrier phases. One explanation for this discrepancy is the time offset that can exist for some receivers between the code and the carrier-phase latching. This paper explains how a code-phase bias in the receiver hardware can induce a frequency difference between the code and the carrier-phase clock solutions. The impact on PPP is then quantified. Finally, the possibility to determine this code-phase bias in the PPP modeling is investigated, and the first results are shown to be inappropriate due to the high level of code noise.

Zebrafish Melanoma.

PubMed

Kaufman, Charles K

2016-01-01

Melanoma skin cancer is a potentially deadly disease in humans and has remained extremely difficult to treat once it has metastasized. In just the last 10 years, a number of models of melanoma have been developed in the zebrafish that are biologically faithful to the human disease and have already yielded important insights into the fundamental biology of melanoma and offered new potential avenues for treatment. With the diversity and breadth of the molecular genetic tools available in the zebrafish, these melanoma models will continue to be refined and expanded upon to keep pace with the rapidly evolving field of melanoma biology.

Atomic clocks for geodesy.

PubMed

Mehlstäubler, Tanja E; Grosche, Gesine; Lisdat, Christian; Schmidt, Piet O; Denker, Heiner

2018-06-01

We review experimental progress on optical atomic clocks and frequency transfer, and consider the prospects of using these technologies for geodetic measurements. Today, optical atomic frequency standards have reached relative frequency inaccuracies below 10 -17 , opening new fields of fundamental and applied research. The dependence of atomic frequencies on the gravitational potential makes atomic clocks ideal candidates for the search for deviations in the predictions of Einstein's general relativity, tests of modern unifying theories and the development of new gravity field sensors. In this review, we introduce the concepts of optical atomic clocks and present the status of international clock development and comparison. Besides further improvement in stability and accuracy of today's best clocks, a large effort is put into increasing the reliability and technological readiness for applications outside of specialized laboratories with compact, portable devices. With relative frequency uncertainties of 10 -18 , comparisons of optical frequency standards are foreseen to contribute together with satellite and terrestrial data to the precise determination of fundamental height reference systems in geodesy with a resolution at the cm-level. The long-term stability of atomic standards will deliver excellent long-term height references for geodetic measurements and for the modelling and understanding of our Earth.

Atomic clocks for geodesy

NASA Astrophysics Data System (ADS)

Mehlstäubler, Tanja E.; Grosche, Gesine; Lisdat, Christian; Schmidt, Piet O.; Denker, Heiner

2018-06-01

We review experimental progress on optical atomic clocks and frequency transfer, and consider the prospects of using these technologies for geodetic measurements. Today, optical atomic frequency standards have reached relative frequency inaccuracies below 10‑17, opening new fields of fundamental and applied research. The dependence of atomic frequencies on the gravitational potential makes atomic clocks ideal candidates for the search for deviations in the predictions of Einstein’s general relativity, tests of modern unifying theories and the development of new gravity field sensors. In this review, we introduce the concepts of optical atomic clocks and present the status of international clock development and comparison. Besides further improvement in stability and accuracy of today’s best clocks, a large effort is put into increasing the reliability and technological readiness for applications outside of specialized laboratories with compact, portable devices. With relative frequency uncertainties of 10‑18, comparisons of optical frequency standards are foreseen to contribute together with satellite and terrestrial data to the precise determination of fundamental height reference systems in geodesy with a resolution at the cm-level. The long-term stability of atomic standards will deliver excellent long-term height references for geodetic measurements and for the modelling and understanding of our Earth.

Development of the zebrafish myoseptum with emphasis on the myotendinous junction.

PubMed

Charvet, Benjamin; Malbouyres, Marilyne; Pagnon-Minot, Aurélie; Ruggiero, Florence; Le Guellec, Dominique

2011-12-01

Zebrafish myosepta connect two adjacent muscle cells and transmit muscular forces to axial structures during swimming via the myotendinous junction (MTJ). The MTJ establishes transmembrane linkages system consisting of extracellular matrix molecules (ECM) surrounding the basement membrane, cytoskeletal elements anchored to sarcolema, and all intermediate proteins that link ECM to actin filaments. Using a series of zebrafish specimens aged between 24 h post-fertilization and 2 years old, the present paper describes at the transmission electron microscope level the development of extracellular and intracellular elements of the MTJ. The transverse myoseptum development starts during the segmentation period by deposition of sparse and loosely organized collagen fibrils. During the hatching period, a link between actin filaments and sarcolemma is established. The basal lamina underlining sarcolemma is well differentiated. Later, collagen fibrils display an orthogonal orientation and fibroblast-like cells invade the myoseptal stroma. A dense network of collagen fibrils is progressively formed that both anchor myoseptal fibroblasts and sarcolemmal basement membrane. The differentiation of a functional MTJ is achieved when sarcolemma interacts with both cytoskeletal filaments and extracellular components. This solid structural link between contractile apparatus and ECM leads to sarcolemma deformations resulting in the formation of regular invaginations, and allows force transmission during muscle contraction. This paper presents the first ultrastructural atlas of the zebrafish MTJ development, which represents an useful tool to analyse the mechanisms of the myotendinous system formation and their disruption in muscle disorders.

pitx2 Deficiency Results in Abnormal Ocular and Craniofacial Development in Zebrafish

PubMed Central

Liu, Yi; Semina, Elena V.

2012-01-01

Human PITX2 mutations are associated with Axenfeld-Rieger syndrome, an autosomal-dominant developmental disorder that involves ocular anterior segment defects, dental hypoplasia, craniofacial dysmorphism and umbilical abnormalities. Characterization of the PITX2 pathway and identification of the mechanisms underlying the anomalies associated with PITX2 deficiency is important for better understanding of normal development and disease; studies of pitx2 function in animal models can facilitate these analyses. A knockdown of pitx2 in zebrafish was generated using a morpholino that targeted all known alternative transcripts of the pitx2 gene; morphant embryos generated with the pitx2ex4/5 splicing-blocking oligomer produced abnormal transcripts predicted to encode truncated pitx2 proteins lacking the third (recognition) helix of the DNA-binding homeodomain. The morphological phenotype of pitx2ex4/5 morphants included small head and eyes, jaw abnormalities and pericardial edema; lethality was observed at ∼6–8-dpf. Cartilage staining revealed a reduction in size and an abnormal shape/position of the elements of the mandibular and hyoid pharyngeal arches; the ceratobranchial arches were also decreased in size. Histological and marker analyses of the misshapen eyes of the pitx2ex4/5 morphants identified anterior segment dysgenesis and disordered hyaloid vasculature. In summary, we demonstrate that pitx2 is essential for proper eye and craniofacial development in zebrafish and, therefore, that PITX2/pitx2 function is conserved in vertebrates. PMID:22303467

Polygenic Sex Determination System in Zebrafish

PubMed Central

Liew, Woei Chang; Bartfai, Richard; Lim, Zijie; Sreenivasan, Rajini; Siegfried, Kellee R.; Orban, Laszlo

2012-01-01

Background Despite the popularity of zebrafish as a research model, its sex determination (SD) mechanism is still unknown. Most cytogenetic studies failed to find dimorphic sex chromosomes and no primary sex determining switch has been identified even though the assembly of zebrafish genome sequence is near to completion and a high resolution genetic map is available. Recent publications suggest that environmental factors within the natural range have minimal impact on sex ratios of zebrafish populations. The primary aim of this study is to find out more about how sex is determined in zebrafish. Methodology/Principal Findings Using classical breeding experiments, we found that sex ratios across families were wide ranging (4.8% to 97.3% males). On the other hand, repeated single pair crossings produced broods of very similar sex ratios, indicating that parental genotypes have a role in the sex ratio of the offspring. Variation among family sex ratios was reduced after selection for breeding pairs with predominantly male or female offspring, another indication that zebrafish sex is regulated genetically. Further examinations by a PCR-based “blind assay" and array comparative genomic hybridization both failed to find universal sex-linked differences between the male and female genomes. Together with the ability to increase the sex bias of lines by selective breeding, these data suggest that zebrafish is unlikely to utilize a chromosomal sex determination (CSD) system. Conclusions/Significance Taken together, our study suggests that zebrafish sex is genetically determined with limited, secondary influences from the environment. As we have not found any sign for CSD in the species, we propose that the zebrafish has a polygenic sex determination system. PMID:22506019

What is the Thalamus in Zebrafish?

PubMed Central

Mueller, Thomas

2012-01-01

Current research on the thalamus and related structures in the zebrafish diencephalon identifies an increasing number of both neurological structures and ontogenetic processes as evolutionary conserved between teleosts and mammals. The patterning processes, for example, which during the embryonic development of zebrafish form the thalamus proper appear largely conserved. Yet also striking differences between zebrafish and other vertebrates have been observed, particularly when we look at mature and histologically differentiated brains. A case in point is the migrated preglomerular complex of zebrafish which evolved only within the lineage of ray-finned fish and has no counterpart in mammals or tetrapod vertebrates. Based on its function as a sensory relay station with projections to pallial zones, the preglomerular complex has been compared to specific thalamic nuclei in mammals. However, no thalamic projections to the zebrafish dorsal pallium, which corresponds topologically to the mammalian isocortex, have been identified. Merely one teleostean thalamic nucleus proper, the auditory nucleus, projects to a part of the dorsal telencephalon, the pallial amygdala. Studies on patterning mechanisms identify a rostral and caudal domain in the embryonic thalamus proper. In both, teleosts and mammals, the rostral domain gives rise to GABAergic neurons, whereas glutamatergic neurons originate in the caudal domain of the zebrafish thalamus. The distribution of GABAergic derivatives in the adult zebrafish brain, furthermore, revealed previously overlooked thalamic nuclei and redefined already established ones. These findings require some reconsideration regarding the topological origin of these adult structures. In what follows, I discuss how evolutionary conserved and newly acquired features of the developing and adult zebrafish thalamus can be compared to the mammalian situation. PMID:22586363

Circadian rhythms and light responsiveness of mammalian clock gene, Clock and BMAL1, transcripts in the rat retina.

PubMed

Namihira, M; Honma, S; Abe, H; Tanahashi, Y; Ikeda, M; Honma, K

1999-08-13

Circadian expression and light-responsiveness of the mammalian clock genes, Clock and BMAL1, in the rat retina were examined by in situ hydbribization under constant darkness. A small but significant daily variation was detected in the Clock transcript level, but not in BMAL1. Light increased the Clock and BMAL1 expressions significantly when examined 60 min after exposure. The light-induced gene expression was phase-dependent for Clock and peaked at ZT2, while rather constant throughout the day for BMAL1. These findings suggest that Clock and BMAL1 play different roles in the generation of circadian rhytm in the retina from those in the suprachiasmatic nucleus. Different roles are also suggested between the two genes in the photic signal transduction in the retina.

CLOCK regulates mammary epithelial cell growth and differentiation

PubMed Central

Crodian, Jennifer; Suárez-Trujillo, Aridany; Erickson, Emily; Weldon, Bethany; Crow, Kristi; Cummings, Shelby; Chen, Yulu; Shamay, Avi; Mabjeesh, Sameer J.; Plaut, Karen

2016-01-01

Circadian clocks influence virtually all physiological processes, including lactation. Here, we investigate the role of the CLOCK gene in regulation of mammary epithelial cell growth and differentiation. Comparison of mammary morphology in late-pregnant wild-type and ClockΔ19 mice, showed that gland development was negatively impacted by genetic loss of a functional timing system. To understand whether these effects were due, in part, to loss of CLOCK function in the gland, the mouse mammary epithelial cell line, HC11, was transfected with short hairpin RNA that targeted Clock (shClock). Cells transfected with shClock expressed 70% less Clock mRNA than wild-type (WT) HC11 cultures, which resulted in significantly depressed levels of CLOCK protein (P < 0.05). HC11 lines carrying shClock had four-fold higher growth rates (P < 0.05), and the percentage of cells in G1 phase was significantly higher (90.1 ± 1.1% of shClock vs. 71.3 ± 3.6% of WT-HC11) following serum starvation. Quantitative-PCR (qPCR) analysis showed shClock had significant effects (P < 0.0001) on relative expression levels of Ccnd1, Wee1, and Tp63. qPCR analysis of the effect of shClock on Fasn and Cdh1 expression in undifferentiated cultures and cultures treated 96 h with dexamethasone, insulin, and prolactin (differentiated) found levels were reduced by twofold and threefold, respectively (P < 0.05), in shClock line relative to WT cultures. Abundance of CDH1 and TP63 proteins were significantly reduced in cultures transfected with shClock. These data support how CLOCK plays a role in regulation of epithelial cell growth and differentiation in the mammary gland. PMID:27707717

Application of Zebrafish Model to Environmental Toxicology.

PubMed

Komoike, Yuta; Matsuoka, Masato

2016-01-01

Recently, a tropical freshwater fish, the zebrafish, has been generally used as a useful model organism in various fields of life science worldwide. The zebrafish model has also been applied to environmental toxicology; however, in Japan, it has not yet become widely used. In this review, we will introduce the biological and historical backgrounds of zebrafish as an animal model and their breeding. We then present the current status of toxicological experiments using zebrafish that were treated with some important environmental contaminants, including cadmium, organic mercury, 2,3,7,8-tetrachlorodibenzo-p-dioxin, and tributyltin. Finally, the future possible application of genetically modified zebrafish to the study of environmental toxicology is discussed.

Towards Self-Clocked Gated OCDMA Receiver

NASA Astrophysics Data System (ADS)

Idris, S.; Osadola, T.; Glesk, I.

2013-02-01

A novel incoherent OCDMA receiver with incorporated all-optical clock recovery for self-synchronization of a time gate for the multi access interferences (MAI) suppression and minimizing the effect of data time jitter in incoherent OCDMA system was successfully developed and demonstrated. The solution was implemented and tested in a multiuser environment in an out of the laboratory OCDMA testbed with two-dimensional wavelength-hopping time-spreading coding scheme and OC-48 (2.5 Gbp/s) data rate. The self-clocked all-optical time gate uses SOA-based fibre ring laser optical clock, recovered all-optically from the received OCDMA traffic to control its switching window for cleaning the autocorrelation peak from the surrounding MAI. A wider eye opening was achieved when the all-optically recovered clock from received data was used for synchronization if compared to a static approach with the RF clock being generated by a RF synthesizer. Clean eye diagram was also achieved when recovered clock is used to drive time gating.

Geopotential measurements with synchronously linked optical lattice clocks

NASA Astrophysics Data System (ADS)

Takano, Tetsushi; Takamoto, Masao; Ushijima, Ichiro; Ohmae, Noriaki; Akatsuka, Tomoya; Yamaguchi, Atsushi; Kuroishi, Yuki; Munekane, Hiroshi; Miyahara, Basara; Katori, Hidetoshi

2016-10-01

According to Einstein's theory of relativity, the passage of time changes in a gravitational field. On Earth, raising a clock by 1 cm increases its apparent tick rate by 1.1 parts in 1018, allowing chronometric levelling through comparison of optical clocks. Here, we demonstrate such geopotential measurements by determining the height difference of master and slave clocks separated by 15 km with an uncertainty of 5 cm. A subharmonic of the master clock laser is delivered through a telecom fibre to synchronously operate the distant clocks. Clocks operated under such phase coherence reject clock laser noise and facilitate proposals for linking clocks and interferometers. Taken over half a year, 11 measurements determine the fractional frequency difference between the two clocks to be 1,652.9(5.9) × 10-18, consistent with an independent measurement by levelling and gravimetry. Our system demonstrates a building block for an internet of clocks, which may constitute ‘quantum benchmarks’, serving as height references with dynamic responses.

Optical clocks and relativity.

PubMed

Chou, C W; Hume, D B; Rosenband, T; Wineland, D J

2010-09-24

Observers in relative motion or at different gravitational potentials measure disparate clock rates. These predictions of relativity have previously been observed with atomic clocks at high velocities and with large changes in elevation. We observed time dilation from relative speeds of less than 10 meters per second by comparing two optical atomic clocks connected by a 75-meter length of optical fiber. We can now also detect time dilation due to a change in height near Earth's surface of less than 1 meter. This technique may be extended to the field of geodesy, with applications in geophysics and hydrology as well as in space-based tests of fundamental physics.

Biological Clocks & Circadian Rhythms

ERIC Educational Resources Information Center

Robertson, Laura; Jones, M. Gail

2009-01-01

The study of biological clocks and circadian rhythms is an excellent way to address the inquiry strand in the National Science Education Standards (NSES) (NRC 1996). Students can study these everyday phenomena by designing experiments, gathering and analyzing data, and generating new experiments. As students explore biological clocks and circadian…

Circadian clock proteins and immunity.

PubMed

Curtis, Anne M; Bellet, Marina M; Sassone-Corsi, Paolo; O'Neill, Luke A J

2014-02-20

Immune parameters change with time of day and disruption of circadian rhythms has been linked to inflammatory pathologies. A circadian-clock-controlled immune system might allow an organism to anticipate daily changes in activity and feeding and the associated risk of infection or tissue damage to the host. Responses to bacteria have been shown to vary depending on time of infection, with mice being more at risk of sepsis when challenged ahead of their activity phase. Studies highlight the extent to which the molecular clock, most notably the core clock proteins BMAL1, CLOCK, and REV-ERBα, control fundamental aspects of the immune response. Examples include the BMAL1:CLOCK heterodimer regulating toll-like receptor 9 (TLR9) expression and repressing expression of the inflammatory monocyte chemokine ligand (CCL2) as well as REV-ERBα suppressing the induction of interleukin-6. Understanding the daily rhythm of the immune system could have implications for vaccinations and how we manage infectious and inflammatory diseases. Copyright © 2014 Elsevier Inc. All rights reserved.

Clock and trigger synchronization between several chassis of digital data acquisition modules

NASA Astrophysics Data System (ADS)

Hennig, W.; Tan, H.; Walby, M.; Grudberg, P.; Fallu-Labruyere, A.; Warburton, W. K.; Vaman, C.; Starosta, K.; Miller, D.

2007-08-01

In applications with segmented high purity Ge detectors or other detector arrays with tens or hundreds of channels, the high development cost and limited flexibility of application specific integrated circuits outweigh their benefits of low power and small size. The readout electronics typically consist of multi-channel data acquisition modules in a common chassis for power, clock and trigger distribution, and data readout. As arrays become larger and reach several hundred channels, the readout electronics have to be divided over several chassis, but still must maintain precise synchronization of clocks and trigger signals across all channels. This division becomes necessary not only because of limits given by the instrumentation standards on module size and chassis slot numbers, but also because data readout times increase when more modules share the same data bus and because power requirements approach the limits of readily available power supplies. In this paper, we present a method for distributing clocks and triggers between 4 PXI chassis containing DGF Pixie-16 modules with up to 226 acquisition channels per chassis. The data acquisition system is intended to instrument the over 600 channels of the SeGA detector array at the National Superconducting Cyclotron Laboratory. Our solution is designed to achieve synchronous acquisition of detector waveforms from all channels with a jitter of less than 1 ns, and can be extended to a larger number of chassis if desired.

Pitfalls of Insulin Pump Clocks

PubMed Central

Reed, Amy J.

2014-01-01

The objective was to raise awareness about the importance of ensuring that insulin pumps internal clocks are set up correctly at all times. This is a very important safety issue because all commercially available insulin pumps are not GPS-enabled (though this is controversial), nor equipped with automatically adjusting internal clocks. Special attention is paid to how basal and bolus dose errors can be introduced by daylight savings time changes, travel across time zones, and am-pm clock errors. Correct setting of insulin pump internal clock is crucial for appropriate insulin delivery. A comprehensive literature review is provided, as are illustrative cases. Incorrect setting can potentially result in incorrect insulin delivery, with potential harmful consequences, if too much or too little insulin is delivered. Daylight saving time changes may not significantly affect basal insulin delivery, given the triviality of the time difference. However, bolus insulin doses can be dramatically affected. Such problems may occur when pump wearers have large variations in their insulin to carb ratio, especially if they forget to change their pump clock in the spring. More worrisome than daylight saving time change is the am-pm clock setting. If this setting is set up incorrectly, both basal rates and bolus doses will be affected. Appropriate insulin delivery through insulin pumps requires correct correlation between dose settings and internal clock time settings. Because insulin pumps are not GPS-enabled or automatically time-adjusting, extra caution should be practiced by patients to ensure correct time settings at all times. Clinicians and diabetes educators should verify the date/time of insulin pumps during patients’ visits, and should remind their patients to always verify these settings. PMID:25355713

Morphometric analysis and neuroanatomical mapping of the zebrafish brain.

PubMed

Gupta, Tripti; Marquart, Gregory D; Horstick, Eric J; Tabor, Kathryn M; Pajevic, Sinisa; Burgess, Harold A

2018-06-21

Large-scale genomic studies have recently identified genetic variants causative for major neurodevelopmental disorders, such as intellectual disability and autism. However, determining how underlying developmental processes are affected by these mutations remains a significant challenge in the field. Zebrafish is an established model system in developmental neurogenetics that may be useful in uncovering the mechanisms of these mutations. Here we describe the use of voxel-intensity, deformation field, and volume-based morphometric techniques for the systematic and unbiased analysis of gene knock-down and environmental exposure-induced phenotypes in zebrafish. We first present a computational method for brain segmentation based on transgene expression patterns to create a comprehensive neuroanatomical map. This map allowed us to disclose statistically significant changes in brain microstructure and composition in neurodevelopmental models. We demonstrate the effectiveness of morphometric techniques in measuring changes in the relative size of neuroanatomical subdivisions in atoh7 morphant larvae and in identifying phenotypes in larvae treated with valproic acid, a chemical demonstrated to increase the risk of autism in humans. These tools enable rigorous evaluation of the effects of gene mutations and environmental exposures on neural development, providing an entry point for cellular and molecular analysis of basic developmental processes as well as neurodevelopmental and neurodegenerative disorders. Published by Elsevier Inc.

The importance of Zebrafish in biomedical research.

PubMed

Tavares, Bárbara; Santos Lopes, Susana

2013-01-01

Zebrafish (Danio rerio) is an ideal model organism for the study of vertebrate development. This is due to the large clutches that each couple produces, with up to 200 embryos every 7 days, and to the fact that the embryos and larvae are small, transparent and undergo rapid external development. Using scientific literature research tools available online and the keywords Zebrafish, biomedical research, human disease, and drug screening, we reviewed original studies and reviews indexed in PubMed. In this review we summarized work conducted with this model for the advancement of our knowledge related to several human diseases. We also focused on the biomedical research being performed in Portugal with the zebrafish model. Powerful live imaging and genetic tools are currently available for zebrafish making it a valuable model in biomedical research. The combination of these properties with the optimization of automated systems for drug screening has transformed the zebrafish into "a top model" in biomedical research, drug discovery and toxicity testing. Furthermore, with the optimization of xenografts technology it will be possible to use zebrafish to aide in the choice of the best therapy for each patient. Zebrafish is an excellent model organism in biomedical research, drug development and in clinical therapy.

Cost and Precision of Brownian Clocks

NASA Astrophysics Data System (ADS)

Barato, Andre C.; Seifert, Udo

2016-10-01

Brownian clocks are biomolecular networks that can count time. A paradigmatic example are proteins that go through a cycle, thus regulating some oscillatory behavior in a living system. Typically, such a cycle requires free energy often provided by ATP hydrolysis. We investigate the relation between the precision of such a clock and its thermodynamic costs. For clocks driven by a constant thermodynamic force, a given precision requires a minimal cost that diverges as the uncertainty of the clock vanishes. In marked contrast, we show that a clock driven by a periodic variation of an external protocol can achieve arbitrary precision at arbitrarily low cost. This result constitutes a fundamental difference between processes driven by a fixed thermodynamic force and those driven periodically. As a main technical tool, we map a periodically driven system with a deterministic protocol to one subject to an external protocol that changes in stochastic time intervals, which simplifies calculations significantly. In the nonequilibrium steady state of the resulting bipartite Markov process, the uncertainty of the clock can be deduced from the calculable dispersion of a corresponding current.

GPS satellite clock determination in case of inter-frequency clock biases for triple-frequency precise point positioning

NASA Astrophysics Data System (ADS)

Guo, Jiang; Geng, Jianghui

2017-12-01

Significant time-varying inter-frequency clock biases (IFCBs) within GPS observations prevent the application of the legacy L1/L2 ionosphere-free clock products on L5 signals. Conventional approaches overcoming this problem are to estimate L1/L5 ionosphere-free clocks in addition to their L1/L2 counterparts or to compute IFCBs between the L1/L2 and L1/L5 clocks which are later modeled through a harmonic analysis. In contrast, we start from the undifferenced uncombined GNSS model and propose an alternative approach where a second satellite clock parameter dedicated to the L5 signals is estimated along with the legacy L1/L2 clock. In this manner, we do not need to rely on the correlated L1/L2 and L1/L5 ionosphere-free observables which complicates triple-frequency GPS stochastic models, or account for the unfavorable time-varying hardware biases in undifferenced GPS functional models since they can be absorbed by the L5 clocks. An extra advantage over the ionosphere-free model is that external ionosphere constraints can potentially be introduced to improve PPP. With 27 days of triple-frequency GPS data from globally distributed stations, we find that the RMS of the positioning differences between our GPS model and all conventional models is below 1 mm for all east, north and up components, demonstrating the effectiveness of our model in addressing triple-frequency observations and time-varying IFCBs. Moreover, we can combine the L1/L2 and L5 clocks derived from our model to calculate precisely the L1/L5 clocks which in practice only depart from their legacy counterparts by less than 0.006 ns in RMS. Our triple-frequency GPS model proves convenient and efficient in combating time-varying IFCBs and can be generalized to more than three frequency signals for satellite clock determination.

Zebrafish neurobehavioral phenomics for aquatic neuropharmacology and toxicology research.

PubMed

Kalueff, Allan V; Echevarria, David J; Homechaudhuri, Sumit; Stewart, Adam Michael; Collier, Adam D; Kaluyeva, Aleksandra A; Li, Shaomin; Liu, Yingcong; Chen, Peirong; Wang, JiaJia; Yang, Lei; Mitra, Anisa; Pal, Subharthi; Chaudhuri, Adwitiya; Roy, Anwesha; Biswas, Missidona; Roy, Dola; Podder, Anupam; Poudel, Manoj K; Katare, Deepshikha P; Mani, Ruchi J; Kyzar, Evan J; Gaikwad, Siddharth; Nguyen, Michael; Song, Cai

2016-01-01

Zebrafish (Danio rerio) are rapidly emerging as an important model organism for aquatic neuropharmacology and toxicology research. The behavioral/phenotypic complexity of zebrafish allows for thorough dissection of complex human brain disorders and drug-evoked pathological states. As numerous zebrafish models become available with a wide spectrum of behavioral, genetic, and environmental methods to test novel drugs, here we discuss recent zebrafish phenomics methods to facilitate drug discovery, particularly in the field of biological psychiatry. Additionally, behavioral, neurological, and endocrine endpoints are becoming increasingly well-characterized in zebrafish, making them an inexpensive, robust and effective model for toxicology research and pharmacological screening. We also discuss zebrafish behavioral phenotypes, experimental considerations, pharmacological candidates and relevance of zebrafish neurophenomics to other 'omics' (e.g., genomic, proteomic) approaches. Finally, we critically evaluate the limitations of utilizing this model organism, and outline future strategies of research in the field of zebrafish phenomics. Copyright © 2015 Elsevier B.V. All rights reserved.

Mixtures, Metabolites, and Mechanisms: Understanding Toxicology Using Zebrafish.

PubMed

Gamse, Joshua T; Gorelick, Daniel A

2016-10-01

For more than 60 years, zebrafish have been used in toxicological studies. Due to their transparency, genetic tractability, and compatibility with high-throughput screens, zebrafish embryos are uniquely suited to study the effects of pharmaceuticals and environmental insults on embryonic development, organ formation and function, and reproductive success. This special issue of Zebrafish highlights the ways zebrafish are used to investigate the toxic effects of endocrine disruptors, pesticides, and heavy metals.

Using quasars as standard clocks for measuring cosmological redshift.

PubMed

Dai, De-Chang; Starkman, Glenn D; Stojkovic, Branislav; Stojkovic, Dejan; Weltman, Amanda

2012-06-08

We report hitherto unnoticed patterns in quasar light curves. We characterize segments of the quasar's light curves with the slopes of the straight lines fit through them. These slopes appear to be directly related to the quasars' redshifts. Alternatively, using only global shifts in time and flux, we are able to find significant overlaps between the light curves of different pairs of quasars by fitting the ratio of their redshifts. We are then able to reliably determine the redshift of one quasar from another. This implies that one can use quasars as standard clocks, as we explicitly demonstrate by constructing two independent methods of finding the redshift of a quasar from its light curve.

Mycobacteriosis in zebrafish colonies.

PubMed

Whipps, Christopher M; Lieggi, Christine; Wagner, Robert

2012-01-01

Mycobacteriosis, a chronic bacterial infection, has been associated with severe losses in some zebrafish facilities and low-level mortalities and unknown impacts in others. The occurrence of at least six different described species (Mycobacterium abscessus, M. chelonae, M. fortuitum, M. haemophilum, M. marinum, M. peregrinum) from zebrafish complicates diagnosis and control because each species is unique. As a generalization, mycobacteria are often considered opportunists, but M. haemophilum and M. marinum appear to be more virulent. Background genetics of zebrafish and environmental conditions influence the susceptibility of fish and progression of disease, emphasizing the importance of regular monitoring and good husbandry practices. A combined approach to diagnostics is ultimately the most informative, with histology as a first-level screen, polymerase chain reaction for rapid detection and species identification, and culture for strain differentiation. Occurrence of identical strains of Mycobacterium in both fish and biofilms in zebrafish systems suggests transmission can occur when fish feed on infected tissues or tank detritus containing mycobacteria. Within a facility, good husbandry practices and sentinel programs are essential for minimizing the impacts of mycobacteria. In addition, quarantine and screening of animals coming into a facility is important for eliminating the introduction of the more severe pathogens. Elimination of mycobacteria from an aquatic system is likely not feasible because these species readily establish biofilms on surfaces even in extremely low nutrient conditions. Risks associated with each commonly encountered species need to be identified and informed management plans developed. Basic research on the growth characteristics, disinfection, and pathogenesis of zebrafish mycobacteria is critical moving forward.

Clocked combustor can array

DOEpatents

Kim, Won-Wook; McMahan, Kevin Weston; Srinivasan, Shiva Kumar

2017-01-17

The present application provides a clocked combustor can array for coherence reduction in a gas turbine engine. The clocked combustor can array may include a number of combustor cans positioned in a circumferential array. A first set of the combustor cans may have a first orientation and a second set of the combustor cans may have a second orientation.

Derivation and experimental verification of clock synchronization theory

NASA Technical Reports Server (NTRS)

Palumbo, Daniel L.

1994-01-01

The objective of this work is to validate mathematically derived clock synchronization theories and their associated algorithms through experiment. Two theories are considered, the Interactive Convergence Clock Synchronization Algorithm and the Mid-Point Algorithm. Special clock circuitry was designed and built so that several operating conditions and failure modes (including malicious failures) could be tested. Both theories are shown to predict conservative upper bounds (i.e., measured values of clock skew were always less than the theory prediction). Insight gained during experimentation led to alternative derivations of the theories. These new theories accurately predict the clock system's behavior. It is found that a 100% penalty is paid to tolerate worst case failures. It is also shown that under optimal conditions (with minimum error and no failures) the clock skew can be as much as 3 clock ticks. Clock skew grows to 6 clock ticks when failures are present. Finally, it is concluded that one cannot rely solely on test procedures or theoretical analysis to predict worst case conditions. conditions.

Episodic-like memory in zebrafish.

PubMed

Hamilton, Trevor J; Myggland, Allison; Duperreault, Erika; May, Zacnicte; Gallup, Joshua; Powell, Russell A; Schalomon, Melike; Digweed, Shannon M

2016-11-01

Episodic-like memory tests often aid in determining an animal's ability to recall the what, where, and which (context) of an event. To date, this type of memory has been demonstrated in humans, wild chacma baboons, corvids (Scrub jays), humming birds, mice, rats, Yucatan minipigs, and cuttlefish. The potential for this type of memory in zebrafish remains unexplored even though they are quickly becoming an essential model organism for the study of a variety of human cognitive and mental disorders. Here we explore the episodic-like capabilities of zebrafish (Danio rerio) in a previously established mammalian memory paradigm. We demonstrate that when zebrafish were presented with a familiar object in a familiar context but a novel location within that context, they spend more time in the novel quadrant. Thus, zebrafish display episodic-like memory as they remember what object they saw, where they saw it (quadrant location), and on which occasion (yellow or blue walls) it was presented.

Mixtures, Metabolites, and Mechanisms: Understanding Toxicology Using Zebrafish

PubMed Central

Gamse, Joshua T.

2016-01-01

Abstract For more than 60 years, zebrafish have been used in toxicological studies. Due to their transparency, genetic tractability, and compatibility with high-throughput screens, zebrafish embryos are uniquely suited to study the effects of pharmaceuticals and environmental insults on embryonic development, organ formation and function, and reproductive success. This special issue of Zebrafish highlights the ways zebrafish are used to investigate the toxic effects of endocrine disruptors, pesticides, and heavy metals. PMID:27618129

The circadian clock in cancer development and therapy

USDA-ARS?s Scientific Manuscript database

Most aspects of mammalian function display circadian rhythms driven by an endogenous clock. The circadian clock is operated by genes and comprises a central clock in the brain that responds to environmental cues and controls subordinate clocks in peripheral tissues via circadian output pathways. The...

A quantum network of clocks

NASA Astrophysics Data System (ADS)

Kómár, P.; Kessler, E. M.; Bishof, M.; Jiang, L.; Sørensen, A. S.; Ye, J.; Lukin, M. D.

2014-08-01

The development of precise atomic clocks plays an increasingly important role in modern society. Shared timing information constitutes a key resource for navigation with a direct correspondence between timing accuracy and precision in applications such as the Global Positioning System. By combining precision metrology and quantum networks, we propose a quantum, cooperative protocol for operating a network of geographically remote optical atomic clocks. Using nonlocal entangled states, we demonstrate an optimal utilization of global resources, and show that such a network can be operated near the fundamental precision limit set by quantum theory. Furthermore, the internal structure of the network, combined with quantum communication techniques, guarantees security both from internal and external threats. Realization of such a global quantum network of clocks may allow construction of a real-time single international time scale (world clock) with unprecedented stability and accuracy.

Zebrafish model systems for developmental neurobehavioral toxicology.

PubMed

Bailey, Jordan; Oliveri, Anthony; Levin, Edward D

2013-03-01

Zebrafish offer many advantages that complement classic mammalian models for the study of normal development as well as for the teratogenic effects of exposure to hazardous compounds. The clear chorion and embryo of the zebrafish allow for continuous visualization of the anatomical changes associated with development, which, along with short maturation times and the capability of complex behavior, makes this model particularly useful for measuring changes to the developing nervous system. Moreover, the rich array of developmental, behavioral, and molecular benefits offered by the zebrafish have contributed to an increasing demand for the use of zebrafish in behavioral teratology. Essential for this endeavor has been the development of a battery of tests to evaluate a spectrum of behavior in zebrafish. Measures of sensorimotor plasticity, emotional function, cognition and social interaction have been used to characterize the persisting adverse effects of developmental exposure to a variety of chemicals including therapeutic drugs, drugs of abuse and environmental toxicants. In this review, we present and discuss such tests and data from a range of developmental neurobehavioral toxicology studies using zebrafish as a model. Zebrafish provide a key intermediate model between high throughput in vitro screens and the classic mammalian models as they have the accessibility of in vitro models and the complex functional capabilities of mammalian models. Copyright © 2013 Wiley Periodicals, Inc.

Zebrafish Model Systems for Developmental Neurobehavioral Toxicology

PubMed Central

Bailey, Jordan; Oliveri, Anthony; Levin, Edward D.

2014-01-01

Zebrafish offer many advantages that complement classic mammalian models for the study of normal development as well as for the teratogenic effects of exposure to hazardous compounds. The clear chorion and embryo of the zebrafish allow for continuous visualization of the anatomical changes associated with development, which, along with short maturation times and the capability of complex behavior, makes this model particularly useful for measuring changes to the developing nervous system. Moreover, the rich array of developmental, behavioral, and molecular benefits offered by the zebrafish have contributed to an increasing demand for the use of zebrafish in behavioral teratology. Essential for this endeavor has been the development of a battery of tests to evaluate a spectrum of behavior in zebrafish. Measures of sensorimotor plasticity, emotional function, cognition and social interaction have been used to characterize the persisting adverse effects of developmental exposure to a variety of chemicals including therapeutic drugs, drugs of abuse and environmental toxicants. In this review, we present and discuss such tests and data from a range of developmental neurobehavioral toxicology studies using zebrafish as a model. Zebrafish provide a key intermediate model between high throughput in vitro screens and the classic mammalian models as they have the accessibility of in vitro models and the complex functional capabilities of mammalian models. PMID:23723169

Clock gene variation in Tachycineta swallows

PubMed Central

Dor, Roi; Cooper, Caren B; Lovette, Irby J; Massoni, Viviana; Bulit, Flor; Liljesthrom, Marcela; Winkler, David W

2012-01-01

Many animals use photoperiod cues to synchronize reproduction with environmental conditions and thereby improve their reproductive success. The circadian clock, which creates endogenous behavioral and physiological rhythms typically entrained to photoperiod, is well characterized at the molecular level. Recent work provided evidence for an association between Clock poly-Q length polymorphism and latitude and, within a population, an association with the date of laying and the length of the incubation period. Despite relatively high overall breeding synchrony, the timing of clutch initiation has a large impact on the fitness of swallows in the genus Tachycineta. We compared length polymorphism in the Clock poly-Q region among five populations from five different Tachycineta species that breed across a hemisphere-wide latitudinal gradient (Fig. 1). Clock poly-Q variation was not associated with latitude; however, there was an association between Clock poly-Q allele diversity and the degree of clutch size decline within breeding seasons. We did not find evidence for an association between Clock poly-Q variation and date of clutch initiation in for any of the five Tachycineta species, nor did we found a relationship between incubation duration and Clock genotype. Thus, there is no general association between latitude, breeding phenology, and Clock polymorphism in this clade of closely related birds. Figure 1 Photos of Tachycineta swallows that were used in this study: A) T. bicolor from Ithaca, New York, B) T. leucorrhoa from Chascomús, Argentina, C) T. albilinea from Hill Bank, Belize, D) T. meyeni from Puerto Varas, Chile, and E) T. thalassina from Mono Lake, California, Photographers: B: Valentina Ferretti; A, C-E: David Winkler. PMID:22408729

Macrophage–Microbe Interactions: Lessons from the Zebrafish Model

PubMed Central

Yoshida, Nagisa; Frickel, Eva-Maria; Mostowy, Serge

2017-01-01

Macrophages provide front line defense against infections. The study of macrophage–microbe interplay is thus crucial for understanding pathogenesis and infection control. Zebrafish (Danio rerio) larvae provide a unique platform to study macrophage–microbe interactions in vivo, from the level of the single cell to the whole organism. Studies using zebrafish allow non-invasive, real-time visualization of macrophage recruitment and phagocytosis. Furthermore, the chemical and genetic tractability of zebrafish has been central to decipher the complex role of macrophages during infection. Here, we discuss the latest developments using zebrafish models of bacterial and fungal infection. We also review novel aspects of macrophage biology revealed by zebrafish, which can potentiate development of new therapeutic strategies for humans. PMID:29250076

Precise time dissemination via portable atomic clocks

NASA Technical Reports Server (NTRS)

Putkovich, K.

1982-01-01

The most precise operational method of time dissemination over long distances presently available to the Precise Time and Time Interval (PTTI) community of users is by means of portable atomic clocks. The Global Positioning System (GPS), the latest system showing promise of replacing portable clocks for global PTTI dissemination, was evaluated. Although GPS has the technical capability of providing superior world-wide dissemination, the question of present cost and future accessibility may require a continued reliance on portable clocks for a number of years. For these reasons a study of portable clock operations as they are carried out today was made. The portable clock system that was utilized by the U.S. Naval Observatory (NAVOBSY) in the global synchronization of clocks over the past 17 years is described and the concepts on which it is based are explained. Some of its capabilities and limitations are also discussed.

Clock-drawing test and unilateral spatial neglect.

PubMed

Ishiai, S; Sugishita, M; Ichikawa, T; Gono, S; Watabiki, S

1993-01-01

We investigated the ability of 25 patients with left unilateral spatial neglect to make a clock face by putting numbers inside a printed circle. Impairment seen in this clock-drawing test did not parallel neglect severity as judged by results of the line-cancellation and line-bisection tests, as well as the copying of a daisy. The score for clock drawing correlated highly with the verbal WAIS score. Most neglect patients with a verbal IQ of 87 or more could draw a clock face fairly well and used planning in placing the numbers 12, 3, 6, and 9 before the others. In clock drawing, verbal intelligence may compensate for left unilateral spatial neglect. We therefore recommend use of the line-cancellation and line-bisection tests, as well as the copying test, but do not recommend use of the clock-drawing test in the diagnosis of left unilateral spatial neglect.

An optical clock to go

NASA Astrophysics Data System (ADS)

Ludlow, Andrew D.

2018-05-01

Bringing next-generation atomic clocks out of the lab is not an easy task, but doing so will unlock many new possibilities. As a crucial first step, a portable atomic clock has now been deployed for relativistic geodesy measurements in the Alps.

Molecular targets for small-molecule modulators of circadian clocks

PubMed Central

He, Baokun; Chen, Zheng

2016-01-01

Background Circadian clocks are endogenous timing systems that regulate various aspects of mammalian metabolism, physiology and behavior. Traditional chronotherapy refers to the administration of drugs in a defined circadian time window to achieve optimal pharmacokinetic and therapeutic efficacies. In recent years, substantial efforts have been dedicated to developing novel small-molecule modulators of circadian clocks. Methods Here, we review the recent progress in the identification of molecular targets of small-molecule clock modulators and their efficacies in clock-related disorders. Specifically, we examine the clock components and regulatory factors as possible molecular targets of small molecules, and we review several key clock-related disorders as promising venues for testing the preventive/therapeutic efficacies of these small molecules. Finally, we also discuss circadian regulation of drug metabolism. Results Small molecules can modulate the period, phase and/or amplitude of the circadian cycle. Core clock proteins, nuclear hormone receptors, and clock-related kinases and other epigenetic regulators are promising molecular targets for small molecules. Through these targets small molecules exert protective effects against clock-related disorders including the metabolic syndrome, immune disorders, sleep disorders and cancer. Small molecules can also modulate circadian drug metabolism and response to existing therapeutics. Conclusion Small-molecule clock modulators target clock components or diverse cellular pathways that functionally impinge upon the clock. Target identification of new small-molecule modulators will deepen our understanding of key regulatory nodes in the circadian network. Studies of clock modulators will facilitate their therapeutic applications, alone or in combination, for clock-related diseases. PMID:26750111

Contextual Fear Conditioning in Zebrafish

ERIC Educational Resources Information Center

Kenney, Justin W.; Scott, Ian C.; Josselyn, Sheena A.; Frankland, Paul W.

2017-01-01

Zebrafish are a genetically tractable vertebrate that hold considerable promise for elucidating the molecular basis of behavior. Although numerous recent advances have been made in the ability to precisely manipulate the zebrafish genome, much less is known about many aspects of learning and memory in adult fish. Here, we describe the development…

Metabolic Plasticity Enables Circadian Adaptation to Acute Hypoxia in Zebrafish Cells.

PubMed

Sandbichler, Adolf M; Jansen, Bianca; Peer, Bettina A; Paulitsch, Monika; Pelster, Bernd; Egg, Margit

2018-01-01

Reduced oxygen availability, hypoxia, is frequently encountered by organisms, tissues and cells, in aquatic environments as well as in high altitude or under pathological conditions such as infarct, stroke or cancer. The hypoxic signaling pathway was found to be mutually intertwined with circadian timekeeping in vertebrates and, as reported recently, also in mammals. However, the impact of hypoxia on intracellular metabolic oscillations is still unknown. For determination of metabolites we used Multilabel Reader based fluorescence and luminescence assays, circadian levels of Hypoxia Inducible Factor 1 alpha and oxidized peroxiredoxins were semi quantified by Western blotting and ratiometric quantification of cytosolic and mitochondrial H2O2 was achieved with stable transfections of a redox sensitive green fluorescent protein sensor into zebrafish fibroblasts. Circadian oscillations of core clock gene mRNA´s were assessed using realtime qPCR with subsequent cosine wave fit analysis. Here we show that under normoxia primary metabolic activity of cells predominately occurs during day time and that after acute hypoxia of two hours, administrated immediately before each sampling point, steady state concentrations of glycolytic key metabolites such as glucose and lactate reveal to be highly rhythmic, following a circadian pattern with highest levels during the night periods and reflecting the circadian variation of the cellular response to hypoxia. Remarkably, rhythms in glycolysis are transferred to cellular energy states under normoxic conditions, so that ADP/ATP ratios oscillate as well, which is the first evidence for cycling ADP/ATP pools in a metazoan cell line to our knowledge. Furthermore, the hypoxia induced alterations in rhythms of glycolysis lead to the alignment of three major cellular redox systems, namely the circadian oscillations of NAD+/NADH and NADP+/NADPH ratios and of increased nocturnal levels of oxidized peroxiredoxins, resulting in a highly

Processing of visually presented clock times.

PubMed

Goolkasian, P; Park, D C

1980-11-01

The encoding and representation of visually presented clock times was investigated in three experiments utilizing a comparative judgment task. Experiment 1 explored the effects of comparing times presented in different formats (clock face, digit, or word), and Experiment 2 examined angular distance effects created by varying positions of the hands on clock faces. In Experiment 3, encoding and processing differences between clock faces and digitally presented times were directly measured. Same/different reactions to digitally presented times were faster than to times presented on a clock face, and this format effect was found to be a result of differences in processing that occurred after encoding. Angular separation also had a limited effect on processing. The findings are interpreted within the framework of theories that refer to the importance of representational codes. The applicability to the data of Bank's semantic-coding theory, Paivio's dual-coding theory, and the levels-of-processing view of memory are discussed.

Extra-hypothalamic brain clocks in songbirds: Photoperiodic state dependent clock gene oscillations in night-migratory blackheaded buntings, Emberiza melanocephala.

PubMed

Singh, Devraj; Kumar, Vinod

2017-04-01

The avian circadian pacemaker system is comprised of independent clocks in the retina, pineal and hypothalamus, as shown by daily and circadian oscillations of core clock genes (Per2, Cry1, Bmal1 and Clock) in several birds including migratory blackheaded buntings (Emberiza melanocephala). This study investigated the extra-hypothalamic brain circadian clocks in blackheaded buntings, and measured Per2, Cry1, Cry2, Bmal1 and Clock mRNA expressions at 4h intervals over 24h beginning 1h after light-on in the left and right telencephalon, optic tectum and cerebellum, the brain regions involved in several physiological and cognitive functions. Because of seasonal alterations in the circadian clock dependent brain functions, we measured daily clock gene oscillations in buntings photoperiod-induced with the non-migratory state under short days (SDnM), and the pre-migratory (LDpM), migratory (LDM) and post-migratory (refractory, LDR) states under long days. Daily Per2 oscillations were not altered with changes in the photoperiodic states, except for about 2-3h phase difference in the optic tectum between the SDnM and LDpM states. However, there were about 3-5h differences in the phase and 2 to 4 fold change in the amplitude of daily Bmal1 and Cry1 mRNA oscillations between the photoperiod-induced states. Further, Cry2 and Clock genes lacked a significant oscillation, except in Cb (Cry2) and TeO and Rt (Clock) under LDR state. Overall, these results show the presence of circadian clocks in extra-hypothalamic brain regions of blackheaded buntings, and suggest tissue-dependent alterations in the waveforms of mRNA oscillations with transitions in the photoperiod-induced seasonal states in a long-day species. Copyright © 2017 Elsevier B.V. All rights reserved.

A Novel Photonic Clock and Carrier Recovery Device

NASA Technical Reports Server (NTRS)

Yao, X. Steve; Lutes, George; Maleki, Lute

1996-01-01

As data communication rates climb toward ten Gb/s, clock recovery and synchronization become more difficult, if not impossible, using conventional electronic circuits. We present in this article experimental results of a high speed clock and carrier recovery using a novel device called a photonic oscillator that we recently developed in our laboratory. This device is capable of recovering clock signals up to 70 GHz. To recover the clock, the incoming data is injected into the photonic oscillator either through the optical injection port or the electrical injection port. The free running photonic oscillator is tuned to oscillate at a nominal frequency equal to the clock frequency of the incoming data. With the injection of the data, the photonic oscillator will be quickly locked to clock frequency of the data stream while rejecting other frequency components associated with the data. Consequently, the output of the locked photonic oscillator is a continuous periodical wave synchronized with the incoming data or simply the recovered clock. We have demonstrated a clock to spur ratio of more than 60 dB of the recovered clock using this technique. Similar to the clock recovery, the photonic oscillator can be used to recover a high frequency carrier degraded by noise and an improvement of about 50 dB in signal-to-noise ratio was demonstrated. The photonic oscillator has both electrical and optical inputs and outputs and can be directly interfaced with a photonic system without signal conversion. In addition to clock and carrier recovery, the photonic oscillator can also be used for (1) stable high frequency clock signal generation, (2) frequency multiplication, (3) square wave and comb frequency generation, and (4) photonic phase locked loop.

Clock is not a component of Z-bands.

PubMed

Wang, Jushuo; Dube, Dipak K; White, Jennifer; Fan, Yingli; Sanger, Jean M; Sanger, Joseph W

2012-12-01

The process of Z-band assembly begins with the formation of small Z-bodies composed of a complex of proteins rich in alpha-actinin. As additional proteins are added to nascent myofibrils, Z-bodies are transformed into continuous bands that form coherent discs of interacting proteins at the boundaries of sarcomeres. The steps controlling the transition of Z-bodies to Z-bands are not known. The report that a circadian protein, Clock, was localized in the Z-bands of neonatal rat cardiomyocytes raised the question whether this transcription factor could be involved in Z-band assembly. We found that the anti-Clock antibody used in the reported study also stained the Z-bands and Z-bodies of mouse and avian cardiac and skeletal muscle cells. YFP constructs of Clock that were assembled, however, did not localize to the Z-bands of muscle cells. Controls of Clock's activity showed that cotransfection of muscle cells with pYFP-Clock and pCeFP-BMAL1 led to the expected nuclear localization of YFP-Clock with its binding partner CeFP-BMAL1. Neither CeFP-BMAL1 nor antibodies directed against BMAL1 localized to Z-bands. A bimolecular fluorescence complementation assay (VC-BMAL1 and VN-Clock) confirmed the absence of Clock and BMAL1 from Z-bands, and their nuclear colocalization. A second anti-Clock antibody stained nuclei, but not Z-bands, of cells cotransfected with Clock and BMAL1 plasmids. Western blots of reactions of muscle extracts and purified alpha-actinins with the two anti-Clock antibodies showed that the original antibody cross-reacted with alpha-actinin and the second did not. These results cannot confirm Clock as an active component of Z-bands. © 2012 Wiley Periodicals, Inc. Copyright © 2012 Wiley Periodicals, Inc.

The role of the notochord in amniote vertebral column segmentation.

PubMed

Ward, Lizzy; Pang, Angel S W; Evans, Susan E; Stern, Claudio D

2018-07-01

The vertebral column is segmented, comprising an alternating series of vertebrae and intervertebral discs along the head-tail axis. The vertebrae and outer portion (annulus fibrosus) of the disc are derived from the sclerotome part of the somites, whereas the inner nucleus pulposus of the disc is derived from the notochord. Here we investigate the role of the notochord in vertebral patterning through a series of microsurgical experiments in chick embryos. Ablation of the notochord causes loss of segmentation of vertebral bodies and discs. However, the notochord cannot segment in the absence of the surrounding sclerotome. To test whether the notochord dictates sclerotome segmentation, we grafted an ectopic notochord. We find that the intrinsic segmentation of the sclerotome is dominant over any segmental information the notochord may possess, and no evidence that the chick notochord is intrinsically segmented. We propose that the segmental pattern of vertebral bodies and discs in chick is dictated by the sclerotome, which first signals to the notochord to ensure that the nucleus pulposus develops in register with the somite-derived annulus fibrosus. Later, the notochord is required for maintenance of sclerotome segmentation as the mature vertebral bodies and intervertebral discs form. These results highlight differences in vertebral development between amniotes and teleosts including zebrafish, where the notochord dictates the segmental pattern. The relative importance of the sclerotome and notochord in vertebral patterning has changed significantly during evolution. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Oscillator networks with tissue-specific circadian clocks in plants.

PubMed

Inoue, Keisuke; Araki, Takashi; Endo, Motomu

2017-09-08

Many organisms rely on circadian clocks to synchronize their biological processes with the 24-h rotation of the earth. In mammals, the circadian clock consists of a central clock in the suprachiasmatic nucleus and peripheral clocks in other tissues. The central clock is tightly coupled to synchronize rhythmicity and can organize peripheral clocks through neural and hormonal signals. In contrast to mammals, it has long been assumed that the circadian clocks in each plant cell is able to be entrained by external light, and they are only weakly coupled to each other. Recently, however, several reports have demonstrated that plants have unique oscillator networks with tissue-specific circadian clocks. Here, we introduce our current view regarding tissue-specific properties and oscillator networks of plant circadian clocks. Accumulating evidence suggests that plants have multiple oscillators, which show distinct properties and reside in different tissues. A direct tissue-isolation technique and micrografting have clearly demonstrated that plants have hierarchical oscillator networks consisting of multiple tissue-specific clocks. Copyright © 2017. Published by Elsevier Ltd.

Knockdown of prothrombin in zebrafish.

PubMed

Day, Kenneth; Krishnegowda, Naveen; Jagadeeswaran, Pudur

2004-01-01

Thrombin is a serine protease generated from its zymogen, prothrombin, and plays a central role in the coagulation cascade. It is also important for mammalian development. The zebrafish has now been established as an excellent genetic model for studies on mammalian hemostasis and development. In this report, we used prothrombin-specific antisense morpholinos to knock down the levels of prothrombin to characterize the effects of prothrombin deficiency in the zebrafish embryo. Prothrombin morpholino-injected zebrafish embryos yielded an early phenotype exhibiting severe abnormalities that later showed occasional bleeding. In a second late phenotype, the embryos had no observable morphological abnormalities in early stages, but showed occasional bleeding at later stages. These phenotypes resembled characteristics shown by prothrombin knockout mice. Laser-induced vascular injury on some of the normal appearing phenotypic larvae showed a prolonged time to occlusion, and recombinant zebrafish prothrombin injected into these larvae restored a normal time to occlusion thus showing the specificity of the morpholino effect. The system developed here should be useful for investigation of the role of thrombin in vertebrate development.

A Zebrafish Heart Failure Model for Assessing Therapeutic Agents.

PubMed

Zhu, Xiao-Yu; Wu, Si-Qi; Guo, Sheng-Ya; Yang, Hua; Xia, Bo; Li, Ping; Li, Chun-Qi

2018-03-20

Heart failure is a leading cause of death and the development of effective and safe therapeutic agents for heart failure has been proven challenging. In this study, taking advantage of larval zebrafish, we developed a zebrafish heart failure model for drug screening and efficacy assessment. Zebrafish at 2 dpf (days postfertilization) were treated with verapamil at a concentration of 200 μM for 30 min, which were determined as optimum conditions for model development. Tested drugs were administered into zebrafish either by direct soaking or circulation microinjection. After treatment, zebrafish were randomly selected and subjected to either visual observation and image acquisition or record videos under a Zebralab Blood Flow System. The therapeutic effects of drugs on zebrafish heart failure were quantified by calculating the efficiency of heart dilatation, venous congestion, cardiac output, and blood flow dynamics. All 8 human heart failure therapeutic drugs (LCZ696, digoxin, irbesartan, metoprolol, qiliqiangxin capsule, enalapril, shenmai injection, and hydrochlorothiazide) showed significant preventive and therapeutic effects on zebrafish heart failure (p < 0.05, p < 0.01, and p < 0.001) in the zebrafish model. The larval zebrafish heart failure model developed and validated in this study could be used for in vivo heart failure studies and for rapid screening and efficacy assessment of preventive and therapeutic drugs.

Global synchronization of parallel processors using clock pulse width modulation

DOEpatents

Chen, Dong; Ellavsky, Matthew R.; Franke, Ross L.; Gara, Alan; Gooding, Thomas M.; Haring, Rudolf A.; Jeanson, Mark J.; Kopcsay, Gerard V.; Liebsch, Thomas A.; Littrell, Daniel; Ohmacht, Martin; Reed, Don D.; Schenck, Brandon E.; Swetz, Richard A.

2013-04-02

A circuit generates a global clock signal with a pulse width modification to synchronize processors in a parallel computing system. The circuit may include a hardware module and a clock splitter. The hardware module may generate a clock signal and performs a pulse width modification on the clock signal. The pulse width modification changes a pulse width within a clock period in the clock signal. The clock splitter may distribute the pulse width modified clock signal to a plurality of processors in the parallel computing system.

Flat mount preparation for observation and analysis of zebrafish embryo specimens stained by whole mount in situ hybridization.

PubMed

Cheng, Christina N; Li, Yue; Marra, Amanda N; Verdun, Valerie; Wingert, Rebecca A

2014-07-17

The zebrafish embryo is now commonly used for basic and biomedical research to investigate the genetic control of developmental processes and to model congenital abnormalities. During the first day of life, the zebrafish embryo progresses through many developmental stages including fertilization, cleavage, gastrulation, segmentation, and the organogenesis of structures such as the kidney, heart, and central nervous system. The anatomy of a young zebrafish embryo presents several challenges for the visualization and analysis of the tissues involved in many of these events because the embryo develops in association with a round yolk mass. Thus, for accurate analysis and imaging of experimental phenotypes in fixed embryonic specimens between the tailbud and 20 somite stage (10 and 19 hours post fertilization (hpf), respectively), such as those stained using whole mount in situ hybridization (WISH), it is often desirable to remove the embryo from the yolk ball and to position it flat on a glass slide. However, performing a flat mount procedure can be tedious. Therefore, successful and efficient flat mount preparation is greatly facilitated through the visual demonstration of the dissection technique, and also helped by using reagents that assist in optimal tissue handling. Here, we provide our WISH protocol for one or two-color detection of gene expression in the zebrafish embryo, and demonstrate how the flat mounting procedure can be performed on this example of a stained fixed specimen. This flat mounting protocol is broadly applicable to the study of many embryonic structures that emerge during early zebrafish development, and can be implemented in conjunction with other staining methods performed on fixed embryo samples.

Flat Mount Preparation for Observation and Analysis of Zebrafish Embryo Specimens Stained by Whole Mount In situ Hybridization

PubMed Central

Cheng, Christina N.; Li, Yue; Marra, Amanda N.; Verdun, Valerie; Wingert, Rebecca A.

2014-01-01

The zebrafish embryo is now commonly used for basic and biomedical research to investigate the genetic control of developmental processes and to model congenital abnormalities. During the first day of life, the zebrafish embryo progresses through many developmental stages including fertilization, cleavage, gastrulation, segmentation, and the organogenesis of structures such as the kidney, heart, and central nervous system. The anatomy of a young zebrafish embryo presents several challenges for the visualization and analysis of the tissues involved in many of these events because the embryo develops in association with a round yolk mass. Thus, for accurate analysis and imaging of experimental phenotypes in fixed embryonic specimens between the tailbud and 20 somite stage (10 and 19 hours post fertilization (hpf), respectively), such as those stained using whole mount in situ hybridization (WISH), it is often desirable to remove the embryo from the yolk ball and to position it flat on a glass slide. However, performing a flat mount procedure can be tedious. Therefore, successful and efficient flat mount preparation is greatly facilitated through the visual demonstration of the dissection technique, and also helped by using reagents that assist in optimal tissue handling. Here, we provide our WISH protocol for one or two-color detection of gene expression in the zebrafish embryo, and demonstrate how the flat mounting procedure can be performed on this example of a stained fixed specimen. This flat mounting protocol is broadly applicable to the study of many embryonic structures that emerge during early zebrafish development, and can be implemented in conjunction with other staining methods performed on fixed embryo samples. PMID:25078510

The sympathy of two pendulum clocks: beyond Huygens' observations.

PubMed

Peña Ramirez, Jonatan; Olvera, Luis Alberto; Nijmeijer, Henk; Alvarez, Joaquin

2016-03-29

This paper introduces a modern version of the classical Huygens' experiment on synchronization of pendulum clocks. The version presented here consists of two monumental pendulum clocks--ad hoc designed and fabricated--which are coupled through a wooden structure. It is demonstrated that the coupled clocks exhibit 'sympathetic' motion, i.e. the pendula of the clocks oscillate in consonance and in the same direction. Interestingly, when the clocks are synchronized, the common oscillation frequency decreases, i.e. the clocks become slow and inaccurate. In order to rigorously explain these findings, a mathematical model for the coupled clocks is obtained by using well-established physical and mechanical laws and likewise, a theoretical analysis is conducted. Ultimately, the sympathy of two monumental pendulum clocks, interacting via a flexible coupling structure, is experimentally, numerically, and analytically demonstrated.

Zebrafish: an animal model for research in veterinary medicine.

PubMed

Nowik, N; Podlasz, P; Jakimiuk, A; Kasica, N; Sienkiewicz, W; Kaleczyc, J

2015-01-01

The zebrafish (Danio rerio) has become known as an excellent model organism for studies of vertebrate biology, vertebrate genetics, embryonal development, diseases and drug screening. Nevertheless, there is still lack of detailed reports about usage of the zebrafish as a model in veterinary medicine. Comparing to other vertebrates, they can lay hundreds of eggs at weekly intervals, externally fertilized zebrafish embryos are accessible to observation and manipulation at all stages of their development, which makes possible to simplify the research techniques such as fate mapping, fluorescent tracer time-lapse lineage analysis and single cell transplantation. Although zebrafish are only 2.5 cm long, they are easy to maintain. Intraperitoneal and intracerebroventricular injections, blood sampling and measurement of food intake are possible to be carry out in adult zebrafish. Danio rerio is a useful animal model for neurobiology, developmental biology, drug research, virology, microbiology and genetics. A lot of diseases, for which the zebrafish is a perfect model organism, affect aquatic animals. For a part of them, like those caused by Mycobacterium marinum or Pseudoloma neutrophila, Danio rerio is a natural host, but the zebrafish is also susceptible to the most of fish diseases including Itch, Spring viraemia of carp and Infectious spleen and kidney necrosis. The zebrafish is commonly used in research of bacterial virulence. The zebrafish embryo allows for rapid, non-invasive and real time analysis of bacterial infections in a vertebrate host. Plenty of common pathogens can be examined using zebrafish model: Streptococcus iniae, Vibrio anguillarum or Listeria monocytogenes. The steps are taken to use the zebrafish also in fungal research, especially that dealing with Candida albicans and Cryptococcus neoformans. Although, the zebrafish is used commonly as an animal model to study diseases caused by external agents, it is also useful in studies of metabolic

A quantum network of clocks

NASA Astrophysics Data System (ADS)

Komar, Peter; Kessler, Eric; Bishof, Michael; Jiang, Liang; Sorensen, Anders; Ye, Jun; Lukin, Mikhail

2014-05-01

Shared timing information constitutes a key resource for positioning and navigation with a direct correspondence between timing accuracy and precision in applications such as the Global Positioning System (GPS). By combining precision metrology and quantum networks, we propose here a quantum, cooperative protocol for the operation of a network consisting of geographically remote optical atomic clocks. Using non-local entangled states, we demonstrate an optimal utilization of the global network resources, and show that such a network can be operated near the fundamental limit set by quantum theory yielding an ultra-precise clock signal. Furthermore, the internal structure of the network, combined with basic techniques from quantum communication, guarantees security both from internal and external threats. Realization of such a global quantum network of clocks may allow construction of a real-time single international time scale (world clock) with unprecedented stability and accuracy. See also: Komar et al. arXiv:1310.6045 (2013) and Kessler et al. arXiv:1310.6043 (2013).

Circadian Clock genes Per2 and clock regulate steroid production, cell proliferation, and luteinizing hormone receptor transcription in ovarian granulosa cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shimizu, Takashi, E-mail: shimizut@obihiro.ac.jp; Hirai, Yuko; Murayama, Chiaki

2011-08-19

Highlights: {yields} Treatment with Per2 and Clock siRNAs decreased the number of granulosa cells and LHr expression. {yields}Per2 siRNA treatment did not stimulate the production of estradiol and expression of P450arom. {yields} Clock siRNA treatment inhibited the production of estradiol and expression of P450arom mRNA. {yields}Per2 and Clock siRNA treatment increased and unchanged, respectively, progesterone production in FSH-treated granulosa cells. {yields} The expression of StAR mRNA was increased by Per2 siRNA and unchanged by Clock siRNA. -- Abstract: Circadian Clock genes are associated with the estrous cycle in female animals. Treatment with Per2 and Clock siRNAs decreased the number ofmore » granulosa cells and LHr expression in follicle-stimulating hormone FSH-treated granulosa cells. Per2 siRNA treatment did not stimulate the production of estradiol and expression of P450arom, whereas Clock siRNA treatment inhibited the production of estradiol and expression of P450arom mRNA. Per2 and Clock siRNA treatment increased and unchanged, respectively, progesterone production in FSH-treated granulosa cells. Similarly, expression of StAR mRNA was increased by Per2 siRNA and unchanged by Clock siRNA. Our data provide a new insight that Per2 and Clock have different action on ovarian granulosa cell functions.« less

Zebrafish: A Versatile Animal Model for Fertility Research.

PubMed

Hoo, Jing Ying; Kumari, Yatinesh; Shaikh, Mohd Farooq; Hue, Seow Mun; Goh, Bey Hing

2016-01-01

The utilization of zebrafish in biomedical research is very common in the research world nowadays. Today, it has emerged as a favored vertebrate organism for the research in science of reproduction. There is a significant growth in amount numbers of scientific literature pertaining to research discoveries in reproductive sciences in zebrafish. It has implied the importance of zebrafish in this particular field of research. In essence, the current available literature has covered from the very specific brain region or neurons of zebrafish, which are responsible for reproductive regulation, until the gonadal level of the animal. The discoveries and findings have proven that this small animal is sharing a very close/similar reproductive system with mammals. More interestingly, the behavioral characteristics and along with the establishment of animal courtship behavior categorization in zebrafish have laid an even stronger foundation and firmer reason on the suitability of zebrafish utilization in research of reproductive sciences. In view of the immense importance of this small animal for the development of reproductive sciences, this review aimed at compiling and describing the proximate close similarity of reproductive regulation on zebrafish and human along with factors contributing to the infertility, showing its versatility and its potential usage for fertility research.

Zebrafish: A Versatile Animal Model for Fertility Research

PubMed Central

Hoo, Jing Ying; Kumari, Yatinesh; Shaikh, Mohd Farooq; Hue, Seow Mun

2016-01-01

The utilization of zebrafish in biomedical research is very common in the research world nowadays. Today, it has emerged as a favored vertebrate organism for the research in science of reproduction. There is a significant growth in amount numbers of scientific literature pertaining to research discoveries in reproductive sciences in zebrafish. It has implied the importance of zebrafish in this particular field of research. In essence, the current available literature has covered from the very specific brain region or neurons of zebrafish, which are responsible for reproductive regulation, until the gonadal level of the animal. The discoveries and findings have proven that this small animal is sharing a very close/similar reproductive system with mammals. More interestingly, the behavioral characteristics and along with the establishment of animal courtship behavior categorization in zebrafish have laid an even stronger foundation and firmer reason on the suitability of zebrafish utilization in research of reproductive sciences. In view of the immense importance of this small animal for the development of reproductive sciences, this review aimed at compiling and describing the proximate close similarity of reproductive regulation on zebrafish and human along with factors contributing to the infertility, showing its versatility and its potential usage for fertility research. PMID:27556045

The Vitamin C Clock Reaction.

ERIC Educational Resources Information Center

Wright, Stephen W.

2002-01-01

Describes an iodine clock reaction that produces an effect similar to the Landolt clock reaction. This reaction uses supermarket chemicals and avoids iodate, bisulfite, and mercury compounds. Ascorbic acid and tincture of iodine are the main reactants with alternate procedures provided for vitamin C tablets and orange juice. (DDR)

Geodesy and metrology with a transportable optical clock

NASA Astrophysics Data System (ADS)

Grotti, Jacopo; Koller, Silvio; Vogt, Stefan; Häfner, Sebastian; Sterr, Uwe; Lisdat, Christian; Denker, Heiner; Voigt, Christian; Timmen, Ludger; Rolland, Antoine; Baynes, Fred N.; Margolis, Helen S.; Zampaolo, Michel; Thoumany, Pierre; Pizzocaro, Marco; Rauf, Benjamin; Bregolin, Filippo; Tampellini, Anna; Barbieri, Piero; Zucco, Massimo; Costanzo, Giovanni A.; Clivati, Cecilia; Levi, Filippo; Calonico, Davide

2018-05-01

Optical atomic clocks, due to their unprecedented stability1-3 and uncertainty3-6, are already being used to test physical theories7,8 and herald a revision of the International System of Units9,10. However, to unlock their potential for cross-disciplinary applications such as relativistic geodesy11, a major challenge remains: their transformation from highly specialized instruments restricted to national metrology laboratories into flexible devices deployable in different locations12-14. Here, we report the first field measurement campaign with a transportable 87Sr optical lattice clock12. We use it to determine the gravity potential difference between the middle of a mountain and a location 90 km away, exploiting both local and remote clock comparisons to eliminate potential clock errors. A local comparison with a 171Yb lattice clock15 also serves as an important check on the international consistency of independently developed optical clocks. This campaign demonstrates the exciting prospects for transportable optical clocks.

PARCS-Primary Atomic Reference Clock in Space

NASA Astrophysics Data System (ADS)

Ashby, Neil

2000-04-01

The purpose of the PARCS project is to place an advanced Cesium clock on the International Space Station (ISS). The project has been approved by NASA at the level of Science Concept Review. Groups at the National Institute of Standards and Technology, Jet Propulsion Laboratory, University of Colorado, and Harvard-Smithsonian Astrophysical Observatory, University of Torino are collaborating on clock design and construction. The microgravity space environment allows laser-cooled Cs atoms to spend longer times in the beam, resulting in improved clock performance. Clock stabilities of 3 × 10-14 at one second and accuracies of 1 × 10-16 are projected. With improved clock performance, significant improvements in several fundamental special and general relativity experiments are expected. For an ISS orbit at 400 km altitude and eccentricity 0.02, the gravitational frequency shift should be measureable about 35 times better than the previous best, Gravity Probe A. Improvements in testing Local Position Invariance and in a Kennedy-Thorndike experiment are expected. Areas of technology such as world-wide timing and time transfer and navigation will also directly benefit from such a high-performance clock in space. This paper will briefly describe the PARCS clock. The principal limitations on performance of relativity experiments, scientific objectives and benefits, and projected outcomes, will be discussed.

Sample-Clock Phase-Control Feedback

NASA Technical Reports Server (NTRS)

Quirk, Kevin J.; Gin, Jonathan W.; Nguyen, Danh H.; Nguyen, Huy

2012-01-01

To demodulate a communication signal, a receiver must recover and synchronize to the symbol timing of a received waveform. In a system that utilizes digital sampling, the fidelity of synchronization is limited by the time between the symbol boundary and closest sample time location. To reduce this error, one typically uses a sample clock in excess of the symbol rate in order to provide multiple samples per symbol, thereby lowering the error limit to a fraction of a symbol time. For systems with a large modulation bandwidth, the required sample clock rate is prohibitive due to current technological barriers and processing complexity. With precise control of the phase of the sample clock, one can sample the received signal at times arbitrarily close to the symbol boundary, thus obviating the need, from a synchronization perspective, for multiple samples per symbol. Sample-clock phase-control feedback was developed for use in the demodulation of an optical communication signal, where multi-GHz modulation bandwidths would require prohibitively large sample clock frequencies for rates in excess of the symbol rate. A custom mixedsignal (RF/digital) offset phase-locked loop circuit was developed to control the phase of the 6.4-GHz clock that samples the photon-counting detector output. The offset phase-locked loop is driven by a feedback mechanism that continuously corrects for variation in the symbol time due to motion between the transmitter and receiver as well as oscillator instability. This innovation will allow significant improvements in receiver throughput; for example, the throughput of a pulse-position modulation (PPM) with 16 slots can increase from 188 Mb/s to 1.5 Gb/s.

Physical exercise improves learning in zebrafish, Danio rerio.

PubMed

Luchiari, Ana Carolina; Chacon, Diana Marques Martins

2013-11-01

Zebrafish is an ideal vertebrate model for neuroscience studies focusing on learning and memory. Although genetic manipulation of zebrafish is available, behavioral protocols are often lacking. In this study we tested whether physical activity can facilitate zebrafish's learning process in an associative conditioning task. Learning was inferred by the approach of the feeding area just after the conditioned stimulus (light). Unexercised zebrafish showed conditioning response from the 5th testing day while fish previously submitted to swim against the water current showed learning by the 3rd day of testing. It seems that physical activity may accelerate associative learning response in zebrafish, indicating the benefits of exercise for cognitive processes. We suggest that this preliminary work could be useful for high throughput screening. Copyright © 2013 Elsevier B.V. All rights reserved.

Quantum Algorithmic Readout in Multi-Ion Clocks.

PubMed

Schulte, M; Lörch, N; Leroux, I D; Schmidt, P O; Hammerer, K

2016-01-08

Optical clocks based on ensembles of trapped ions promise record frequency accuracy with good short-term stability. Most suitable ion species lack closed transitions, so the clock signal must be read out indirectly by transferring the quantum state of the clock ions to cotrapped logic ions of a different species. Existing methods of quantum logic readout require a linear overhead in either time or the number of logic ions. Here we describe a quantum algorithmic readout whose overhead scales logarithmically with the number of clock ions in both of these respects. The scheme allows a quantum nondemolition readout of the number of excited clock ions using a single multispecies gate operation which can also be used in other areas of ion trap technology such as quantum information processing, quantum simulations, metrology, and precision spectroscopy.

Normal anatomy and histology of the adult zebrafish.

PubMed

Menke, Aswin L; Spitsbergen, Jan M; Wolterbeek, Andre P M; Woutersen, Ruud A

2011-08-01

The zebrafish has been shown to be an excellent vertebrate model for studying the roles of specific genes and signaling pathways. The sequencing of its genome and the relative ease with which gene modifications can be performed have led to the creation of numerous human disease models that can be used for testing the potential and the toxicity of new pharmaceutical compounds. Many pharmaceutical companies already use the zebrafish for prescreening purposes. So far, the focus has been on ecotoxicity and the effects on embryonic development, but there is a trend to expand the use of the zebrafish with acute, subchronic, and chronic toxicity studies that are currently still carried out with the more conventional test animals such as rodents. However, before we can fully realize the potential of the zebrafish as an animal model for understanding human development, disease, and toxicology, we must first greatly advance our knowledge of normal zebrafish physiology, anatomy, and histology. To further this knowledge, we describe, in the present article, location and histology of the major zebrafish organ systems with a brief description of their function.

Making Waves: New Developments in Toxicology With the Zebrafish.

PubMed

Horzmann, Katharine A; Freeman, Jennifer L

2018-05-01

The laboratory zebrafish (Danio rerio) is now an accepted model in toxicologic research. The zebrafish model fills a niche between in vitro models and mammalian biomedical models. The developmental characteristics of the small fish are strategically being used by scientists to study topics ranging from high-throughput toxicity screens to toxicity in multi- and transgenerational studies. High-throughput technology has increased the utility of zebrafish embryonic toxicity assays in screening of chemicals and drugs for toxicity or effect. Additionally, advances in behavioral characterization and experimental methodology allow for observation of recognizable phenotypic changes after xenobiotic exposure. Future directions in zebrafish research are predicted to take advantage of CRISPR-Cas9 genome editing methods in creating models of disease and interrogating mechanisms of action with fluorescent reporters or tagged proteins. Zebrafish can also model developmental origins of health and disease and multi- and transgenerational toxicity. The zebrafish has many advantages as a toxicologic model and new methodologies and areas of study continue to expand the usefulness and application of the zebrafish.

Compact, Highly Stable Ion Atomic Clock

NASA Technical Reports Server (NTRS)

Prestage, John

2008-01-01

A mercury-ion clock now at the breadboard stage of development (see figure) has a stability comparable to that of a hydrogen-maser clock: In tests, the clock exhibited an Allan deviation of between 2 x 10(exp -13) and 3 x 10(exp -13) at a measurement time of 1 second, averaging to about 10(exp -15) at 1 day. However, the clock occupies a volume of only about 2 liters . about a hundredth of the volume of a hydrogen-maser clock. The ion-handling parts of the apparatus are housed in a sealed vacuum tube, wherein only a getter pump is used to maintain the vacuum. Hence, this apparatus is a prototype of a generation of small, potentially portable high-precision clocks for diverse ground- and space-based navigation and radio science applications. Furthermore, this new ion-clock technology is about 100 times more stable and precise than the rubidium atomic clocks currently in use in the NAV STAR GPS Earth-orbiting satellites. In this clock, mercury ions are shuttled between a quadrupole and a 16-pole linear radio-frequency trap. In the quadrupole trap, the ions are tightly confined and optical state selection from a Hg-202 radio-frequency-discharge ultraviolet lamp is carried out. In the 16-pole trap, the ions are more loosely confined and atomic transitions resonant at frequency of about 40.507 GHz are interrogated by use of a microwave beam at that frequency. The trapping of ions effectively eliminates the frequency pulling caused by wall collisions inherent to gas-cell clocks. The shuttling of the ions between the two traps enables separation of the state-selection process from the clock microwave- resonance process, so that each of these processes can be optimized independently of the other. The basic ion-shuttling, two-trap scheme as described thus far is not new: it has been the basis of designs of prior larger clocks. The novelty of the present development lies in major redesigns of its physics package (the ion traps and the vacuum and optical subsystems) to effect

Inexpensive programmable clock for a 12-bit computer

NASA Technical Reports Server (NTRS)

Vrancik, J. E.

1972-01-01

An inexpensive programmable clock was built for a digital PDP-12 computer. The instruction list includes skip on flag; clear the flag, clear the clock, and stop the clock; and preset the counter with the contents of the accumulator and start the clock. The clock counts at a rate determined by an external oscillator and causes an interrupt and sets a flag when a 12-bit overflow occurs. An overflow can occur after 1 to 4096 counts. The clock can be built for a total parts cost of less than $100 including power supply and I/O connector. Slight modification can be made to permit its use on larger machines (16 bit, 24 bit, etc.) and logic level shifting can be made to make it compatible with any computer.

Sound Clocks and Sonic Relativity

NASA Astrophysics Data System (ADS)

Todd, Scott L.; Menicucci, Nicolas C.

2017-10-01

Sound propagation within certain non-relativistic condensed matter models obeys a relativistic wave equation despite such systems admitting entirely non-relativistic descriptions. A natural question that arises upon consideration of this is, "do devices exist that will experience the relativity in these systems?" We describe a thought experiment in which `acoustic observers' possess devices called sound clocks that can be connected to form chains. Careful investigation shows that appropriately constructed chains of stationary and moving sound clocks are perceived by observers on the other chain as undergoing the relativistic phenomena of length contraction and time dilation by the Lorentz factor, γ , with c the speed of sound. Sound clocks within moving chains actually tick less frequently than stationary ones and must be separated by a shorter distance than when stationary to satisfy simultaneity conditions. Stationary sound clocks appear to be length contracted and time dilated to moving observers due to their misunderstanding of their own state of motion with respect to the laboratory. Observers restricted to using sound clocks describe a universe kinematically consistent with the theory of special relativity, despite the preferred frame of their universe in the laboratory. Such devices show promise in further probing analogue relativity models, for example in investigating phenomena that require careful consideration of the proper time elapsed for observers.

Identification and characterization of the zebrafish glutathione S-transferase Pi-1.

PubMed

Abunnaja, Maryam S; Kurogi, Katsuhisa; Mohammed, Yasir I; Sakakibara, Yoichi; Suiko, Masahito; Hassoun, Ezdihar A; Liu, Ming-Cheh

2017-10-01

Zebrafish has in recent years emerged as a popular vertebrate model for use in pharmacological and toxicological studies. While there have been sporadic studies on the zebrafish glutathione S-transferases (GSTs), the zebrafish GST gene superfamily still awaits to be fully elucidated. We report here the identification of 15 zebrafish cytosolic GST genes in NCBI GenBank database and the expression, purification, and enzymatic characterization of the zebrafish cytosolic GST Pi-1 (GSTP1). The cDNA encoding the zebrafish GSTP1 was cloned from a 3-month-old female zebrafish, expressed in Eschelichia coli host cells, and purified. Purified GSTP1 displayed glutathione-conjugating activity toward 1-chloro-2,4-dinitrobenzene as a representative substrate. The enzymatic characteristics of the zebrafish GSTP1, including pH-dependency, effects of metal cations, and kinetic parameters, were studied. Moreover, the expression of zebrafish GSTP1 at different developmental stages during embryogenesis, throughout larval development, onto maturity was examined. © 2017 Wiley Periodicals, Inc.

Automated measurement of zebrafish larval movement

PubMed Central

Cario, Clinton L; Farrell, Thomas C; Milanese, Chiara; Burton, Edward A

2011-01-01

Abstract The zebrafish is a powerful vertebrate model that is readily amenable to genetic, pharmacological and environmental manipulations to elucidate the molecular and cellular basis of movement and behaviour. We report software enabling automated analysis of zebrafish movement from video recordings captured with cameras ranging from a basic camcorder to more specialized equipment. The software, which is provided as open-source MATLAB functions, can be freely modified and distributed, and is compatible with multiwell plates under a wide range of experimental conditions. Automated measurement of zebrafish movement using this technique will be useful for multiple applications in neuroscience, pharmacology and neuropsychiatry. PMID:21646414

Automated measurement of zebrafish larval movement.

PubMed

Cario, Clinton L; Farrell, Thomas C; Milanese, Chiara; Burton, Edward A

2011-08-01

The zebrafish is a powerful vertebrate model that is readily amenable to genetic, pharmacological and environmental manipulations to elucidate the molecular and cellular basis of movement and behaviour. We report software enabling automated analysis of zebrafish movement from video recordings captured with cameras ranging from a basic camcorder to more specialized equipment. The software, which is provided as open-source MATLAB functions, can be freely modified and distributed, and is compatible with multiwell plates under a wide range of experimental conditions. Automated measurement of zebrafish movement using this technique will be useful for multiple applications in neuroscience, pharmacology and neuropsychiatry.

Learning and memory in zebrafish larvae

PubMed Central

Roberts, Adam C.; Bill, Brent R.; Glanzman, David L.

2013-01-01

Larval zebrafish possess several experimental advantages for investigating the molecular and neural bases of learning and memory. Despite this, neuroscientists have only recently begun to use these animals to study memory. However, in a relatively short period of time a number of forms of learning have been described in zebrafish larvae, and significant progress has been made toward their understanding. Here we provide a comprehensive review of this progress; we also describe several promising new experimental technologies currently being used in larval zebrafish that are likely to contribute major insights into the processes that underlie learning and memory. PMID:23935566

Clock recovery for high-speed optical communication

NASA Astrophysics Data System (ADS)

Pedrotti, Kenneth D.

1996-01-01

This paper reviews recent results for clock recovery circuits operating at speeds in excess of 1 Gbit/sec or realized as multichannel arrays. The emphasis is on synchronous optical network (SONET) type systems, their requirements, and the effect of the clock recovery circuits on system performance. Clock recovery approaches include filter based, phase-locked-loops, and all-optical methods.

Clock recovery for high-speed optical communication

NASA Astrophysics Data System (ADS)

Pedrotti, Ken

1996-01-01

This paper reviews recent results for clock recovery circuits operating at speeds in excess of 1 Gbit/sec or realized as multichannel arrays. The emphasis is on Synchronous Optical NETwork (SONET) type systems, their requirements, and the effect of the clock recovery circuits on system performance. Clock recovery approaches include filter based, phase-lockcd-loops, and all-optical methods.

Production of Androgenetic Zebrafish (Danio Rerio)

PubMed Central

Corley-Smith, G. E.; Lim, C. J.; Brandhorst, B. P.

1996-01-01

To help investigate the evolutionary origin of the imprinting (parent-of-origin mono-allelic expression) of paternal genes observed in mammals, we constructed haploid and diploid androgenetic zebrafish (Danio rerio). Haploid androgenotes were produced by fertilizing eggs that had been X-ray irradiated to eliminate the maternal genome. Subsequent inhibition of the first mitotic division of haploid androgenotes by heat shock produced diploid androgenotes. The lack of inheritance of maternal-specific DNA markers (RAPD and SSR) by putative diploid and haploid androgenotes confirmed the androgenetic origin of their genomes. Marker analysis was performed on 18 putative androgenotes (five diploids and 13 haploids) from six families. None of 157 maternal-specific RAPD markers analyzed, some of which were apparently homozygous, were passed on to any of these putative androgenotes. A mean of 7.7 maternal-specific markers were assessed per family. The survival of androgenetic zebrafish suggests that if paternal imprinting occurs in zebrafish, it does not result in essential genes being inactivated when their expression is required for development. Production of haploid androgenotes can be used to determine the meiotic recombination rate in male zebrafish. Androgenesis may also provide useful information about the mechanism of sex determination in zebrafish. PMID:8846903

Development of sensory systems in zebrafish (Danio rerio)

NASA Technical Reports Server (NTRS)

Moorman, S. J.

2001-01-01

Zebrafish possess all of the classic sensory modalities: taste, tactile, smell, balance, vision, and hearing. For each sensory system, this article provides a brief overview of the system in the adult zebrafish followed by a more detailed overview of the development of the system. By far the majority of studies performed in each of the sensory systems of the zebrafish have involved some aspect of molecular biology or genetics. Although molecular biology and genetics are not major foci of the paper, brief discussions of some of the mutant strains of zebrafish that have developmental defects in each specific sensory system are included. The development of the sensory systems is only a small sampling of the work being done using zebrafish and provides a mere glimpse of the potential of this model for the study of vertebrate development, physiology, and human disease.

UNUSUAL FINDINGS IN ZEBRAFISH, DANIO RERIO, FROM TOXICOLOGICAL STUDIES AND THE ZEBRAFISH INTERNATIONAL RESOURCE CENTER DIAGNOSTIC SERVICE

EPA Science Inventory

A number of interesting and unusual lesions have been diagnosed in zebrafish that have been evaluated from toxicological studies or submitted as cases to the Diagnostic Service at Oregon State University. Lesions were observed in various wild-type and mutant lines of zebrafish an...

The autoshaping procedure as a residual block clock

PubMed Central

Dinsmoor, James A.; Dougan, James D.; Pfister, John; Thiels, Edda

1992-01-01

In the first experiment, 4 pigeons were each presented with a recurring sequence of four key colors followed by the delivery of grain (block clock). Once the rate of pecking had stabilized, three of the colors were replaced, during different series of sessions, by a darkening of the key. The rate of pecking was reduced within those segments of the interval between deliveries of food during which the key was dark; when the key was dark during the final portion of the interval, rates were reduced throughout the entire interval. In the second experiment, 3 new pigeons were exposed to a different sequence of colors, and the final stimulus was replaced in successive conditions by a novel color, a darkened key, and a restoration of the original color. The data indicated that darkening the key had a more severe, more extensive, and more persistent effect than did a mere change in color. These results suggest that it may be fruitful to conceptualize the autoshaping procedure as a special version of the block clock in which pecking is suppressed throughout the greater part of the interval by darkening the key. In the final condition, the same stimulus appeared in each of the last three portions of the interval. The rate of pecking was lower during the last two portions than when distinctive colors were presented, with the peak rate now appearing in the fifth of seven equal temporal components. PMID:16812666

Contextual fear conditioning in zebrafish.

PubMed

Kenney, Justin W; Scott, Ian C; Josselyn, Sheena A; Frankland, Paul W

2017-10-01

Zebrafish are a genetically tractable vertebrate that hold considerable promise for elucidating the molecular basis of behavior. Although numerous recent advances have been made in the ability to precisely manipulate the zebrafish genome, much less is known about many aspects of learning and memory in adult fish. Here, we describe the development of a contextual fear conditioning paradigm using an electric shock as the aversive stimulus. We find that contextual fear conditioning is modulated by shock intensity, prevented by an established amnestic agent (MK-801), lasts at least 14 d, and exhibits extinction. Furthermore, fish of various background strains (AB, Tu, and TL) are able to acquire fear conditioning, but differ in fear extinction rates. Taken together, we find that contextual fear conditioning in zebrafish shares many similarities with the widely used contextual fear conditioning paradigm in rodents. Combined with the amenability of genetic manipulation in zebrafish, we anticipate that our paradigm will prove to be a useful complementary system in which to examine the molecular basis of vertebrate learning and memory. © 2017 Kenney et al.; Published by Cold Spring Harbor Laboratory Press.

Sleep loss reduces the DNA-binding of BMAL1, CLOCK, and NPAS2 to specific clock genes in the mouse cerebral cortex.

PubMed

Mongrain, Valérie; La Spada, Francesco; Curie, Thomas; Franken, Paul

2011-01-01

We have previously demonstrated that clock genes contribute to the homeostatic aspect of sleep regulation. Indeed, mutations in some clock genes modify the markers of sleep homeostasis and an increase in homeostatic sleep drive alters clock gene expression in the forebrain. Here, we investigate a possible mechanism by which sleep deprivation (SD) could alter clock gene expression by quantifying DNA-binding of the core-clock transcription factors CLOCK, NPAS2, and BMAL1 to the cis-regulatory sequences of target clock genes in mice. Using chromatin immunoprecipitation (ChIP), we first showed that, as reported for the liver, DNA-binding of CLOCK and BMAL1 to target clock genes changes in function of time-of-day in the cerebral cortex. Tissue extracts were collected at ZT0 (light onset), -6, -12, and -18, and DNA enrichment of E-box or E'-box containing sequences was measured by qPCR. CLOCK and BMAL1 binding to Cry1, Dbp, Per1, and Per2 depended on time-of-day, with maximum values reached at around ZT6. We then observed that SD, performed between ZT0 and -6, significantly decreased DNA-binding of CLOCK and BMAL1 to Dbp, consistent with the observed decrease in Dbp mRNA levels after SD. The DNA-binding of NPAS2 and BMAL1 to Per2 was also decreased by SD, although SD is known to increase Per2 expression in the cortex. DNA-binding to Per1 and Cry1 was not affected by SD. Our results show that the sleep-wake history can affect the clock molecular machinery directly at the level of chromatin binding thereby altering the cortical expression of Dbp and Per2 and likely other targets. Although the precise dynamics of the relationship between DNA-binding and mRNA expression, especially for Per2, remains elusive, the results also suggest that part of the reported circadian changes in DNA-binding of core clock components in tissues peripheral to the suprachiasmatic nuclei could, in fact, be sleep-wake driven.

Novel transcriptional networks regulated by CLOCK in human neurons.

PubMed

Fontenot, Miles R; Berto, Stefano; Liu, Yuxiang; Werthmann, Gordon; Douglas, Connor; Usui, Noriyoshi; Gleason, Kelly; Tamminga, Carol A; Takahashi, Joseph S; Konopka, Genevieve

2017-11-01

The molecular mechanisms underlying human brain evolution are not fully understood; however, previous work suggested that expression of the transcription factor CLOCK in the human cortex might be relevant to human cognition and disease. In this study, we investigated this novel transcriptional role for CLOCK in human neurons by performing chromatin immunoprecipitation sequencing for endogenous CLOCK in adult neocortices and RNA sequencing following CLOCK knockdown in differentiated human neurons in vitro. These data suggested that CLOCK regulates the expression of genes involved in neuronal migration, and a functional assay showed that CLOCK knockdown increased neuronal migratory distance. Furthermore, dysregulation of CLOCK disrupts coexpressed networks of genes implicated in neuropsychiatric disorders, and the expression of these networks is driven by hub genes with human-specific patterns of expression. These data support a role for CLOCK-regulated transcriptional cascades involved in human brain evolution and function. © 2017 Fontenot et al.; Published by Cold Spring Harbor Laboratory Press.

On-orbit frequency stability analysis of the GPS NAVSTAR-1 quartz clock and the NAVSTARs-6 and -8 rubidium clocks

NASA Technical Reports Server (NTRS)

Mccaskill, T. B.; Buisson, J. A.; Reid, W. G.

1984-01-01

An on-orbit frequency stability performance analysis of the GPS NAVSTAR-1 quartz clock and the NAVSTARs-6 and -8 rubidium clocks is presented. The clock offsets were obtained from measurements taken at the GPS monitor stations which use high performance cesium standards as a reference. Clock performance is characterized through the use of the Allan variance, which is evaluated for sample times of 15 minutes to two hours, and from one day to 10 days. The quartz and rubidium clocks' offsets were corrected for aging rate before computing the frequency stability. The effect of small errors in aging rate is presented for the NAVSTAR-8 rubidium clock's stability analysis. The analysis includes presentation of time and frequency residuals with respect to linear and quadratic models, which aid in obtaining aging rate values and identifying systematic and random effects. The frequency stability values were further processed with a time domain noise process analysis, which is used to classify random noise process and modulation type.

Countdown Clock Ribbon Cutting

NASA Image and Video Library

2016-03-01

Kennedy Space Center Director Bob Cabana speaks at the dedication of the newest display at the entrance to the center's visitor complex. The historic countdown clock was originally set up at the space center's Press Site and was used from the launch of Apollo 12 on Nov. 14, 1969 to the final space shuttle mission, STS-135, launched on July 8, 2011. The old countdown clock was replaced in 2014 with a modern light emitting diode, or LED, display.

Adult zebrafish intestine resection: a novel model of short bowel syndrome, adaptation, and intestinal stem cell regeneration.

PubMed

Schall, K A; Holoyda, K A; Grant, C N; Levin, D E; Torres, E R; Maxwell, A; Pollack, H A; Moats, R A; Frey, M R; Darehzereshki, A; Al Alam, D; Lien, C; Grikscheit, T C

2015-08-01

Loss of significant intestinal length from congenital anomaly or disease may lead to short bowel syndrome (SBS); intestinal failure may be partially offset by a gain in epithelial surface area, termed adaptation. Current in vivo models of SBS are costly and technically challenging. Operative times and survival rates have slowed extension to transgenic models. We created a new reproducible in vivo model of SBS in zebrafish, a tractable vertebrate model, to facilitate investigation of the mechanisms of intestinal adaptation. Proximal intestinal diversion at segment 1 (S1, equivalent to jejunum) was performed in adult male zebrafish. SBS fish emptied distal intestinal contents via stoma as in the human disease. After 2 wk, S1 was dilated compared with controls and villus ridges had increased complexity, contributing to greater villus epithelial perimeter. The number of intervillus pockets, the intestinal stem cell zone of the zebrafish increased and contained a higher number of bromodeoxyuridine (BrdU)-labeled cells after 2 wk of SBS. Egf receptor and a subset of its ligands, also drivers of adaptation, were upregulated in SBS fish. Igf has been reported as a driver of intestinal adaptation in other animal models, and SBS fish exposed to a pharmacological inhibitor of the Igf receptor failed to demonstrate signs of intestinal adaptation, such as increased inner epithelial perimeter and BrdU incorporation. We describe a technically feasible model of human SBS in the zebrafish, a faster and less expensive tool to investigate intestinal stem cell plasticity as well as the mechanisms that drive intestinal adaptation. Copyright © 2015 the American Physiological Society.

Adult zebrafish intestine resection: a novel model of short bowel syndrome, adaptation, and intestinal stem cell regeneration

PubMed Central

Schall, K. A.; Holoyda, K. A.; Grant, C. N.; Levin, D. E.; Torres, E. R.; Maxwell, A.; Pollack, H. A.; Moats, R. A.; Frey, M. R.; Darehzereshki, A.; Al Alam, D.; Lien, C.

2015-01-01

Loss of significant intestinal length from congenital anomaly or disease may lead to short bowel syndrome (SBS); intestinal failure may be partially offset by a gain in epithelial surface area, termed adaptation. Current in vivo models of SBS are costly and technically challenging. Operative times and survival rates have slowed extension to transgenic models. We created a new reproducible in vivo model of SBS in zebrafish, a tractable vertebrate model, to facilitate investigation of the mechanisms of intestinal adaptation. Proximal intestinal diversion at segment 1 (S1, equivalent to jejunum) was performed in adult male zebrafish. SBS fish emptied distal intestinal contents via stoma as in the human disease. After 2 wk, S1 was dilated compared with controls and villus ridges had increased complexity, contributing to greater villus epithelial perimeter. The number of intervillus pockets, the intestinal stem cell zone of the zebrafish increased and contained a higher number of bromodeoxyuridine (BrdU)-labeled cells after 2 wk of SBS. Egf receptor and a subset of its ligands, also drivers of adaptation, were upregulated in SBS fish. Igf has been reported as a driver of intestinal adaptation in other animal models, and SBS fish exposed to a pharmacological inhibitor of the Igf receptor failed to demonstrate signs of intestinal adaptation, such as increased inner epithelial perimeter and BrdU incorporation. We describe a technically feasible model of human SBS in the zebrafish, a faster and less expensive tool to investigate intestinal stem cell plasticity as well as the mechanisms that drive intestinal adaptation. PMID:26089336

Compensatory regulation of Na+ absorption by Na+/H+ exchanger and Na+-Cl- cotransporter in zebrafish (Danio rerio)

PubMed Central

2013-01-01

Introduction In mammals, internal Na+ homeostasis is maintained through Na+ reabsorption via a variety of Na+ transport proteins with mutually compensating functions, which are expressed in different segments of the nephrons. In zebrafish, Na+ homeostasis is achieved mainly through the skin/gill ionocytes, namely Na+/H+ exchanger (NHE3b)-expressing H+-ATPase rich (HR) cells and Na+-Cl- cotransporter (NCC)-expressing NCC cells, which are functionally homologous to mammalian proximal and distal convoluted tubular cells, respectively. The present study aimed to investigate whether or not the functions of HR and NCC ionocytes are differentially regulated to compensate for disruptions of internal Na+ homeostasis and if the cell differentiation of the ionocytes is involved in this regulation pathway. Results Translational knockdown of ncc caused an increase in HR cell number and a resulting augmentation of Na+ uptake in zebrafish larvae, while NHE3b loss-of-function caused an increase in NCC cell number with a concomitant recovery of Na+ absorption. Environmental acid stress suppressed nhe3b expression in HR cells and decreased Na+ content, which was followed by up-regulation of NCC cells accompanied by recovery of Na+ content. Moreover, knockdown of ncc resulted in a significant decrease of Na+ content in acid-acclimated zebrafish. Conclusions These results provide evidence that HR and NCC cells exhibit functional redundancy in Na+ absorption, similar to the regulatory mechanisms in mammalian kidney, and suggest this functional redundancy is a critical strategy used by zebrafish to survive in a harsh environment that disturbs body fluid Na+ homeostasis. PMID:23924428

Nephron proximal tubule patterning and corpuscles of Stannius formation are regulated by the sim1a transcription factor and retinoic acid in zebrafish.

PubMed

Cheng, Christina N; Wingert, Rebecca A

2015-03-01

The mechanisms that establish nephron segments are poorly understood. The zebrafish embryonic kidney, or pronephros, is a simplified yet conserved genetic model to study this renal development process because its nephrons contain segments akin to other vertebrates, including the proximal convoluted and straight tubules (PCT, PST). The zebrafish pronephros is also associated with the corpuscles of Stannius (CS), endocrine glands that regulate calcium and phosphate homeostasis, but whose ontogeny from renal progenitors is largely mysterious. Initial patterning of zebrafish renal progenitors in the intermediate mesoderm (IM) involves the formation of rostral and caudal domains, the former being reliant on retinoic acid (RA) signaling, and the latter being repressed by elevated RA levels. Here, using expression profiling to gain new insights into nephrogenesis, we discovered that the gene single minded family bHLH transcription factor 1a (sim1a) is dynamically expressed in the renal progenitors-first marking the caudal domain, then becoming restricted to the proximal segments, and finally exhibiting specific CS expression. In loss of function studies, sim1a knockdown expanded the PCT and abrogated both the PST and CS populations. Conversely, overexpression of sim1a modestly expanded the PST and CS, while it reduced the PCT. These results show that sim1a activity is necessary and partially sufficient to induce PST and CS fates, and suggest that sim1a may inhibit PCT fate and/or negotiate the PCT/PST boundary. Interestingly, the sim1a expression domain in renal progenitors is responsive to altered levels of RA, suggesting that RA regulates sim1a, directly or indirectly, during nephrogenesis. sim1a deficient embryos treated with exogenous RA formed nephrons that were predominantly composed of PCT segments, but lacked the enlarged PST observed in RA treated wild-types, indicating that RA is not sufficient to rescue the PST in the absence of sim1a expression. Alternately

Computerized image analysis for quantitative neuronal phenotyping in zebrafish.

PubMed

Liu, Tianming; Lu, Jianfeng; Wang, Ye; Campbell, William A; Huang, Ling; Zhu, Jinmin; Xia, Weiming; Wong, Stephen T C

2006-06-15

An integrated microscope image analysis pipeline is developed for automatic analysis and quantification of phenotypes in zebrafish with altered expression of Alzheimer's disease (AD)-linked genes. We hypothesize that a slight impairment of neuronal integrity in a large number of zebrafish carrying the mutant genotype can be detected through the computerized image analysis method. Key functionalities of our zebrafish image processing pipeline include quantification of neuron loss in zebrafish embryos due to knockdown of AD-linked genes, automatic detection of defective somites, and quantitative measurement of gene expression levels in zebrafish with altered expression of AD-linked genes or treatment with a chemical compound. These quantitative measurements enable the archival of analyzed results and relevant meta-data. The structured database is organized for statistical analysis and data modeling to better understand neuronal integrity and phenotypic changes of zebrafish under different perturbations. Our results show that the computerized analysis is comparable to manual counting with equivalent accuracy and improved efficacy and consistency. Development of such an automated data analysis pipeline represents a significant step forward to achieve accurate and reproducible quantification of neuronal phenotypes in large scale or high-throughput zebrafish imaging studies.

Designing and Testing of Self-Cleaning Recirculating Zebrafish Tanks.

PubMed

Nema, Shubham; Bhargava, Yogesh

2016-08-01

Maintenance of large number of zebrafish in captive conditions is a daunting task. This can be eased by the use of recirculating racks with self-cleaning zebrafish tanks. Commercially available systems are costly, and compatibility of intercompany products has never been investigated. Although various cost-effective designs and methods of construction of custom-made recirculating zebrafish racks are available in literature, the design of self-cleaning zebrafish tanks is still not available. In this study, we report the design and method of construction of the self-cleaning unit, which can be fitted in any zebrafish tank. We validated the design by investigating sediment cleaning process in rectangular and cylindrical tank geometries using time lapse imaging. Our results suggest that for both tank geometries, the tanks fitted with self-cleaning unit provided superior sediment cleaning than the tanks fitted with overflow-drain unit. Although the self-cleaning unit could clean the sediment completely from both geometries over prolonged period, the cleaning of sediments was faster in the cylindrical tank than the rectangular tank. In conclusion, cost and efforts of zebrafish maintenance could be significantly reduced through the installation of our self-cleaning unit in any custom-made zebrafish tank.

Genomic Approaches to Zebrafish Cancer

PubMed Central

2017-01-01

The zebrafish has emerged as an important model for studying cancer biology. Identification of DNA, RNA and chromatin abnormalities can give profound insight into the mechanisms of tumorigenesis and the there are many techniques for analyzing the genomes of these tumors. Here, I present an overview of the available technologies for analyzing tumor genomes in the zebrafish, including array based methods as well as next-generation sequencing technologies. I also discuss the ways in which zebrafish tumor genomes can be compared to human genomes using cross-species oncogenomics, which act to filter genomic noise and ultimately uncover central drivers of malignancy. Finally, I discuss downstream analytic tools, including network analysis, that can help to organize the alterations into coherent biological frameworks that can then be investigated further. PMID:27165352

Circadian clock proteins regulate neuronal redox homeostasis and neurodegeneration

PubMed Central

Musiek, Erik S.; Lim, Miranda M.; Yang, Guangrui; Bauer, Adam Q.; Qi, Laura; Lee, Yool; Roh, Jee Hoon; Ortiz-Gonzalez, Xilma; Dearborn, Joshua T.; Culver, Joseph P.; Herzog, Erik D.; Hogenesch, John B.; Wozniak, David F.; Dikranian, Krikor; Giasson, Benoit I.; Weaver, David R.; Holtzman, David M.; FitzGerald, Garret A.

2013-01-01

Brain aging is associated with diminished circadian clock output and decreased expression of the core clock proteins, which regulate many aspects of cellular biochemistry and metabolism. The genes encoding clock proteins are expressed throughout the brain, though it is unknown whether these proteins modulate brain homeostasis. We observed that deletion of circadian clock transcriptional activators aryl hydrocarbon receptor nuclear translocator–like (Bmal1) alone, or circadian locomotor output cycles kaput (Clock) in combination with neuronal PAS domain protein 2 (Npas2), induced severe age-dependent astrogliosis in the cortex and hippocampus. Mice lacking the clock gene repressors period circadian clock 1 (Per1) and period circadian clock 2 (Per2) had no observed astrogliosis. Bmal1 deletion caused the degeneration of synaptic terminals and impaired cortical functional connectivity, as well as neuronal oxidative damage and impaired expression of several redox defense genes. Targeted deletion of Bmal1 in neurons and glia caused similar neuropathology, despite the retention of intact circadian behavioral and sleep-wake rhythms. Reduction of Bmal1 expression promoted neuronal death in primary cultures and in mice treated with a chemical inducer of oxidative injury and striatal neurodegeneration. Our findings indicate that BMAL1 in a complex with CLOCK or NPAS2 regulates cerebral redox homeostasis and connects impaired clock gene function to neurodegeneration. PMID:24270424

Work Continues on Installing New Countdown Clock

NASA Image and Video Library

2014-11-26

At NASA's Kennedy Space Center in Florida, work continues to install 24 light emitting diode LED panels in the new countdown clock at the spaceport's Press Site. The modern, multimedia display is similar to the screens seen at sporting venues. The new screen will be nearly 26 feet wide by 7 feet high, a foot taller than the original clock. The historic countdown clock was designed by Kennedy engineers and built by space center technicians before Apollo 12 in 1969. NASA has requested to acquire the countdown clock from the agency’s Artifact Working Group at the agency's Headquarters for likely display at the Kennedy Space Center Visitor Complex.

Zebrafish tracking using convolutional neural networks.

PubMed

Xu, Zhiping; Cheng, Xi En

2017-02-17

Keeping identity for a long term after occlusion is still an open problem in the video tracking of zebrafish-like model animals, and accurate animal trajectories are the foundation of behaviour analysis. We utilize the highly accurate object recognition capability of a convolutional neural network (CNN) to distinguish fish of the same congener, even though these animals are indistinguishable to the human eye. We used data augmentation and an iterative CNN training method to optimize the accuracy for our classification task, achieving surprisingly accurate trajectories of zebrafish of different size and age zebrafish groups over different time spans. This work will make further behaviour analysis more reliable.

Zebrafish tracking using convolutional neural networks

NASA Astrophysics Data System (ADS)

Xu, Zhiping; Cheng, Xi En

2017-02-01

Keeping identity for a long term after occlusion is still an open problem in the video tracking of zebrafish-like model animals, and accurate animal trajectories are the foundation of behaviour analysis. We utilize the highly accurate object recognition capability of a convolutional neural network (CNN) to distinguish fish of the same congener, even though these animals are indistinguishable to the human eye. We used data augmentation and an iterative CNN training method to optimize the accuracy for our classification task, achieving surprisingly accurate trajectories of zebrafish of different size and age zebrafish groups over different time spans. This work will make further behaviour analysis more reliable.

Biological timing and the clock metaphor: oscillatory and hourglass mechanisms.

PubMed

Rensing, L; Meyer-Grahle, U; Ruoff, P

2001-05-01

Living organisms have developed a multitude of timing mechanisms--"biological clocks." Their mechanisms are based on either oscillations (oscillatory clocks) or unidirectional processes (hourglass clocks). Oscillatory clocks comprise circatidal, circalunidian, circadian, circalunar, and circannual oscillations--which keep time with environmental periodicities--as well as ultradian oscillations, ovarian cycles, and oscillations in development and in the brain, which keep time with biological timescales. These clocks mainly determine time points at specific phases of their oscillations. Hourglass clocks are predominantly found in development and aging and also in the brain. They determine time intervals (duration). More complex timing systems combine oscillatory and hourglass mechanisms, such as the case for cell cycle, sleep initiation, or brain clocks, whereas others combine external and internal periodicities (photoperiodism, seasonal reproduction). A definition of a biological clock may be derived from its control of functions external to its own processes and its use in determining temporal order (sequences of events) or durations. Biological and chemical oscillators are characterized by positive and negative feedback (or feedforward) mechanisms. During evolution, living organisms made use of the many existing oscillations for signal transmission, movement, and pump mechanisms, as well as for clocks. Some clocks, such as the circadian clock, that time with environmental periodicities are usually compensated (stabilized) against temperature, whereas other clocks, such as the cell cycle, that keep time with an organismic timescale are not compensated. This difference may be related to the predominance of negative feedback in the first class of clocks and a predominance of positive feedback (autocatalytic amplification) in the second class. The present knowledge of a compensated clock (the circadian oscillator) and an uncompensated clock (the cell cycle), as well

The sympathy of two pendulum clocks: beyond Huygens’ observations

PubMed Central

Peña Ramirez, Jonatan; Olvera, Luis Alberto; Nijmeijer, Henk; Alvarez, Joaquin

2016-01-01

This paper introduces a modern version of the classical Huygens’ experiment on synchronization of pendulum clocks. The version presented here consists of two monumental pendulum clocks—ad hoc designed and fabricated—which are coupled through a wooden structure. It is demonstrated that the coupled clocks exhibit ‘sympathetic’ motion, i.e. the pendula of the clocks oscillate in consonance and in the same direction. Interestingly, when the clocks are synchronized, the common oscillation frequency decreases, i.e. the clocks become slow and inaccurate. In order to rigorously explain these findings, a mathematical model for the coupled clocks is obtained by using well-established physical and mechanical laws and likewise, a theoretical analysis is conducted. Ultimately, the sympathy of two monumental pendulum clocks, interacting via a flexible coupling structure, is experimentally, numerically, and analytically demonstrated. PMID:27020903

The Square Light Clock and Special Relativity

ERIC Educational Resources Information Center

Galli, J. Ronald; Amiri, Farhang

2012-01-01

A thought experiment that includes a square light clock is similar to the traditional vertical light beam and mirror clock, except it is made up of four mirrors placed at a 45[degree] angle at each corner of a square of length L[subscript 0], shown in Fig. 1. Here we have shown the events as measured in the rest frame of the square light clock. By…

A precise clock distribution network for MRPC-based experiments

NASA Astrophysics Data System (ADS)

Wang, S.; Cao, P.; Shang, L.; An, Q.

2016-06-01

In high energy physics experiments, the MRPC (Multi-Gap Resistive Plate Chamber) detectors are widely used recently which can provide higher-resolution measurement for particle identification. However, the application of MRPC detectors leads to a series of challenges in electronics design with large number of front-end electronic channels, especially for distributing clock precisely. To deal with these challenges, this paper presents a universal scheme of clock transmission network for MRPC-based experiments with advantages of both precise clock distribution and global command synchronization. For precise clock distributing, the clock network is designed into a tree architecture with two stages: the first one has a point-to-multipoint long range bidirectional distribution with optical channels and the second one has a fan-out structure with copper link inside readout crates. To guarantee the precision of clock frequency or phase, the r-PTP (reduced Precision Time Protocol) and the DDMTD (digital Dual Mixer Time Difference) methods are used for frequency synthesis, phase measurement and adjustment, which is implemented by FPGA (Field Programmable Gate Array) in real-time. In addition, to synchronize global command execution, based upon this clock distribution network, synchronous signals are coded with clock for transmission. With technique of encoding/decoding and clock data recovery, signals such as global triggers or system control commands, can be distributed to all front-end channels synchronously, which greatly simplifies the system design. The experimental results show that both the clock jitter (RMS) and the clock skew can be less than 100 ps.

The Space Optical Clock project: status and perspectives

NASA Astrophysics Data System (ADS)

Schiller, Stephan; Tino, Guglielmo M.; Sterr, Uwe; Lemonde, Pierre; Görlitz, Axel; Salomon, Christophe

The Space Optical Clocks project aims at operating lattice clocks on the ISS for tests of funda-mental physics and for providing high-accuracy comparisons of future terrestrial optical clocks. A pre-phase-A study (2007-10), funded partially by ESA and DLR, includes the implementa-tion of several optical lattice clock systems using Strontium and Ytterbium as atomic systems and their characterization. Subcomponents of clock demonstrators with the added specification of transportability and using techniques that are suitable for later space use, such as all-solid-state lasers, low power consumption, and compact dimensions, have been developed and are being validated. The talk will give a brief overview over the achieved results and outline future developments.

Zebrafish as model organisms for studying drug-induced liver injury

PubMed Central

Vliegenthart, A D Bastiaan; Tucker, Carl S; Del Pozo, Jorge; Dear, James W

2014-01-01

Drug-induced liver injury (DILI) is a major challenge in clinical medicine and drug development. New models are needed for predicting which potential therapeutic compounds will cause DILI in humans, and new markers and mediators of DILI still need to be identified. This review highlights the strengths and weaknesses of using zebrafish as a high-throughput in vivo model for studying DILI. Although the zebrafish liver architecture is different from that of the mammalian liver, the main physiological processes remain similar. Zebrafish metabolize drugs using similar pathways to those in humans; they possess a wide range of cytochrome P450 enzymes that enable metabolic reactions including hydroxylation, conjugation, oxidation, demethylation and de-ethylation. Following exposure to a range of hepatotoxic drugs, the zebrafish liver develops histological patterns of injury comparable to those of mammalian liver, and biomarkers for liver injury can be quantified in the zebrafish circulation. The zebrafish immune system is similar to that of mammals, but the zebrafish inflammatory response to DILI is not yet defined. In order to quantify DILI in zebrafish, a wide variety of methods can be used, including visual assessment, quantification of serum enzymes and experimental serum biomarkers and scoring of histopathology. With further development, the zebrafish may be a model that complements rodents and may have value for the discovery of new disease pathways and translational biomarkers. PMID:24773296

Stable Kalman filters for processing clock measurement data

NASA Technical Reports Server (NTRS)

Clements, P. A.; Gibbs, B. P.; Vandergraft, J. S.

1989-01-01

Kalman filters have been used for some time to process clock measurement data. Due to instabilities in the standard Kalman filter algorithms, the results have been unreliable and difficult to obtain. During the past several years, stable forms of the Kalman filter have been developed, implemented, and used in many diverse applications. These algorithms, while algebraically equivalent to the standard Kalman filter, exhibit excellent numerical properties. Two of these stable algorithms, the Upper triangular-Diagonal (UD) filter and the Square Root Information Filter (SRIF), have been implemented to replace the standard Kalman filter used to process data from the Deep Space Network (DSN) hydrogen maser clocks. The data are time offsets between the clocks in the DSN, the timescale at the National Institute of Standards and Technology (NIST), and two geographically intermediate clocks. The measurements are made by using the GPS navigation satellites in mutual view between clocks. The filter programs allow the user to easily modify the clock models, the GPS satellite dependent biases, and the random noise levels in order to compare different modeling assumptions. The results of this study show the usefulness of such software for processing clock data. The UD filter is indeed a stable, efficient, and flexible method for obtaining optimal estimates of clock offsets, offset rates, and drift rates. A brief overview of the UD filter is also given.

Relativity theory and time perception: single or multiple clocks?

PubMed

Buhusi, Catalin V; Meck, Warren H

2009-07-22

Current theories of interval timing assume that humans and other animals time as if using a single, absolute stopwatch that can be stopped or reset on command. Here we evaluate the alternative view that psychological time is represented by multiple clocks, and that these clocks create separate temporal contexts by which duration is judged in a relative manner. Two predictions of the multiple-clock hypothesis were tested. First, that the multiple clocks can be manipulated (stopped and/or reset) independently. Second, that an event of a given physical duration would be perceived as having different durations in different temporal contexts, i.e., would be judged differently by each clock. Rats were trained to time three durations (e.g., 10, 30, and 90 s). When timing was interrupted by an unexpected gap in the signal, rats reset the clock used to time the "short" duration, stopped the "medium" duration clock, and continued to run the "long" duration clock. When the duration of the gap was manipulated, the rats reset these clocks in a hierarchical order, first the "short", then the "medium", and finally the "long" clock. Quantitative modeling assuming re-allocation of cognitive resources in proportion to the relative duration of the gap to the multiple, simultaneously timed event durations was used to account for the results. These results indicate that the three event durations were effectively timed by separate clocks operated independently, and that the same gap duration was judged relative to these three temporal contexts. Results suggest that the brain processes the duration of an event in a manner similar to Einstein's special relativity theory: A given time interval is registered differently by independent clocks dependent upon the context.

Heart Repair and Regeneration: Recent Insights from Zebrafish Studies

PubMed Central

Lien, Ching-Ling; Harrison, Michael R.; Tuan, Tai-Lan; Starnes, Vaughn A

2012-01-01

Cardiovascular disease is the leading cause of death in United States and worldwide. Failure to properly repair or regenerate damaged cardiac tissues after myocardial infarction is a major cause of heart failure. In contrast to humans and other mammals, zebrafish hearts regenerate after substantial injury or tissue damage. Here, we review recent progress in studying zebrafish heart regeneration, addressing the molecular and cellular responses in the three tissue layers of the heart: myocardium, epicardium, and endocardium. We also compare different injury models utilized to study zebrafish heart regeneration, and discuss the differences in responses to injury between mammalian and zebrafish hearts. By learning how zebrafish hearts regenerate naturally, we can better design therapeutic strategies for repairing human hearts after myocardial infarction. PMID:22818295

The zebrafish genome: a review and msx gene case study.

PubMed

Postlethwait, J H

2006-01-01

Zebrafish is one of several important teleost models for understanding principles of vertebrate developmental, molecular, organismal, genetic, evolutionary, and genomic biology. Efficient investigation of the molecular genetic basis of induced mutations depends on knowledge of the zebrafish genome. Principles of zebrafish genomic analysis, including gene mapping, ortholog identification, conservation of syntenies, genome duplication, and evolution of duplicate gene function are discussed here using as a case study the zebrafish msxa, msxb, msxc, msxd, and msxe genes, which together constitute zebrafish orthologs of tetrapod Msx1, Msx2, and Msx3. Genomic analysis suggests orthologs for this difficult to understand group of paralogs.

Sleep Loss Reduces the DNA-Binding of BMAL1, CLOCK, and NPAS2 to Specific Clock Genes in the Mouse Cerebral Cortex

PubMed Central

Curie, Thomas; Franken, Paul

2011-01-01

We have previously demonstrated that clock genes contribute to the homeostatic aspect of sleep regulation. Indeed, mutations in some clock genes modify the markers of sleep homeostasis and an increase in homeostatic sleep drive alters clock gene expression in the forebrain. Here, we investigate a possible mechanism by which sleep deprivation (SD) could alter clock gene expression by quantifying DNA-binding of the core-clock transcription factors CLOCK, NPAS2, and BMAL1 to the cis-regulatory sequences of target clock genes in mice. Using chromatin immunoprecipitation (ChIP), we first showed that, as reported for the liver, DNA-binding of CLOCK and BMAL1 to target clock genes changes in function of time-of-day in the cerebral cortex. Tissue extracts were collected at ZT0 (light onset), −6, −12, and −18, and DNA enrichment of E-box or E'-box containing sequences was measured by qPCR. CLOCK and BMAL1 binding to Cry1, Dbp, Per1, and Per2 depended on time-of-day, with maximum values reached at around ZT6. We then observed that SD, performed between ZT0 and −6, significantly decreased DNA-binding of CLOCK and BMAL1 to Dbp, consistent with the observed decrease in Dbp mRNA levels after SD. The DNA-binding of NPAS2 and BMAL1 to Per2 was also decreased by SD, although SD is known to increase Per2 expression in the cortex. DNA-binding to Per1 and Cry1 was not affected by SD. Our results show that the sleep-wake history can affect the clock molecular machinery directly at the level of chromatin binding thereby altering the cortical expression of Dbp and Per2 and likely other targets. Although the precise dynamics of the relationship between DNA-binding and mRNA expression, especially for Per2, remains elusive, the results also suggest that part of the reported circadian changes in DNA-binding of core clock components in tissues peripheral to the suprachiasmatic nuclei could, in fact, be sleep-wake driven. PMID:22039518

Physical Layer Ethernet Clock Synchronization

DTIC Science & Technology

2010-11-01

42 nd Annual Precise Time and Time Interval (PTTI) Meeting 77 PHYSICAL LAYER ETHERNET CLOCK SYNCHRONIZATION Reinhard Exel, Georg...oeaw.ac.at Nikolaus Kerö Oregano Systems, Mohsgasse 1, 1030 Wien, Austria E-mail: nikolaus.keroe@oregano.at Abstract Clock synchronization ...is a service widely used in distributed networks to coordinate data acquisition and actions. As the requirement to achieve tighter synchronization

Biosecurity and Health Monitoring at the Zebrafish International Resource Center

PubMed Central

Varga, Zoltán M.; Kent, Michael L.

2016-01-01

Abstract The Zebrafish International Resource Center (ZIRC) is a repository and distribution center for mutant, transgenic, and wild-type zebrafish. In recent years annual imports of new zebrafish lines to ZIRC have increased tremendously. In addition, after 15 years of research, we have identified some of the most virulent pathogens affecting zebrafish that should be avoided in large production facilities, such as ZIRC. Therefore, while importing a high volume of new lines we prioritize safeguarding the health of our in-house fish colony. Here, we describe the biosecurity and health-monitoring program implemented at ZIRC. This strategy was designed to prevent introduction of new zebrafish pathogens, minimize pathogens already present in the facility, and ensure a healthy zebrafish colony for in-house uses and shipment to customers. PMID:27031282

Laser controlled atom source for optical clocks.

PubMed

Kock, Ole; He, Wei; Świerad, Dariusz; Smith, Lyndsie; Hughes, Joshua; Bongs, Kai; Singh, Yeshpal

2016-11-18

Precision timekeeping has been a driving force in innovation, from defining agricultural seasons to atomic clocks enabling satellite navigation, broadband communication and high-speed trading. We are on the verge of a revolution in atomic timekeeping, where optical clocks promise an over thousand-fold improvement in stability and accuracy. However, complex setups and sensitivity to thermal radiation pose limitations to progress. Here we report on an atom source for a strontium optical lattice clock which circumvents these limitations. We demonstrate fast (sub 100 ms), cold and controlled emission of strontium atomic vapours from bulk strontium oxide irradiated by a simple low power diode laser. Our results demonstrate that millions of strontium atoms from the vapour can be captured in a magneto-optical trap (MOT). Our method enables over an order of magnitude reduction in scale of the apparatus. Future applications range from satellite clocks testing general relativity to portable clocks for inertial navigation systems and relativistic geodesy.

Laser controlled atom source for optical clocks

PubMed Central

Kock, Ole; He, Wei; Świerad, Dariusz; Smith, Lyndsie; Hughes, Joshua; Bongs, Kai; Singh, Yeshpal

2016-01-01

Precision timekeeping has been a driving force in innovation, from defining agricultural seasons to atomic clocks enabling satellite navigation, broadband communication and high-speed trading. We are on the verge of a revolution in atomic timekeeping, where optical clocks promise an over thousand-fold improvement in stability and accuracy. However, complex setups and sensitivity to thermal radiation pose limitations to progress. Here we report on an atom source for a strontium optical lattice clock which circumvents these limitations. We demonstrate fast (sub 100 ms), cold and controlled emission of strontium atomic vapours from bulk strontium oxide irradiated by a simple low power diode laser. Our results demonstrate that millions of strontium atoms from the vapour can be captured in a magneto-optical trap (MOT). Our method enables over an order of magnitude reduction in scale of the apparatus. Future applications range from satellite clocks testing general relativity to portable clocks for inertial navigation systems and relativistic geodesy. PMID:27857214

Applications of Clocks to Space Navigation & "Planetary GPS"

NASA Technical Reports Server (NTRS)

Lichten, Stephen M.

2004-01-01

The ability to fly atomic clocks on GPS satellites has profoundly defined the capabilities and limitations of GPS in near-Earth applications. It is likely that future infrastructure for Lunar and Mars applications will be constrained by financial factors. The development of a low cost, small, high performance space clock -- or ultrahigh performance space clocks -- could revolutionize and drive the entire approach to GPS-like systems at the Moon (or Mars), and possibly even change the future of GPS at Earth. Many system trade studies are required. The performance of future GPS-like tracking systems at the Moon or Mars will depend critically on clock performance, availability of inertial sensors, and constellation coverage. Example: present-day GPS carry 10(exp -13) clocks and require several updates per day. With 10(exp -15) clocks, a constellation at Mars could operate autonomously with updates just once per month. Use of GPS tracking at the Moon should be evaluated in a technical study.

A clock network for geodesy and fundamental science.

PubMed

Lisdat, C; Grosche, G; Quintin, N; Shi, C; Raupach, S M F; Grebing, C; Nicolodi, D; Stefani, F; Al-Masoudi, A; Dörscher, S; Häfner, S; Robyr, J-L; Chiodo, N; Bilicki, S; Bookjans, E; Koczwara, A; Koke, S; Kuhl, A; Wiotte, F; Meynadier, F; Camisard, E; Abgrall, M; Lours, M; Legero, T; Schnatz, H; Sterr, U; Denker, H; Chardonnet, C; Le Coq, Y; Santarelli, G; Amy-Klein, A; Le Targat, R; Lodewyck, J; Lopez, O; Pottie, P-E

2016-08-09

Leveraging the unrivalled performance of optical clocks as key tools for geo-science, for astronomy and for fundamental physics beyond the standard model requires comparing the frequency of distant optical clocks faithfully. Here, we report on the comparison and agreement of two strontium optical clocks at an uncertainty of 5 × 10(-17) via a newly established phase-coherent frequency link connecting Paris and Braunschweig using 1,415 km of telecom fibre. The remote comparison is limited only by the instability and uncertainty of the strontium lattice clocks themselves, with negligible contributions from the optical frequency transfer. A fractional precision of 3 × 10(-17) is reached after only 1,000 s averaging time, which is already 10 times better and more than four orders of magnitude faster than any previous long-distance clock comparison. The capability of performing high resolution international clock comparisons paves the way for a redefinition of the unit of time and an all-optical dissemination of the SI-second.

Advancing epilepsy treatment through personalized genetic zebrafish models.

PubMed

Griffin, A; Krasniak, C; Baraban, S C

2016-01-01

With an increase in the number of disease causing genetic mutations identified from epilepsy cohorts, zebrafish are proving to be an attractive vertebrate model for functional analysis of these allele variants. Not only do zebrafish have conserved gene functions, but larvae harboring mutations in identified human epileptic genes show spontaneous seizure activity and mimic the convulsive behavioral movements observed in humans. With zebrafish being compatible with medium to high-throughput screening, they are also proving to be a unique and powerful system for early preclinical drug screening, including novel target identification, pharmacology, and toxicology. Additionally, with recent advances in genomic engineering technologies, it is now possible to study the precise pathophysiology of patient-specific gene mutations in zebrafish. The following sections highlight how the unique attributes of zebrafish, in combination with genetic modifications, are continuing to transform our understanding of epilepsy and help identify personalized therapeutics for specific patient cohorts. © 2016 Elsevier B.V. All rights reserved.

Examination of a Palatogenic Gene Program in Zebrafish

PubMed Central

Swartz, Mary E.; Sheehan-Rooney, Kelly; Dixon, Michael J.; Eberhart, Johann K.

2011-01-01

Human palatal clefting is debilitating and difficult to rectify surgically. Animal models enhance our understanding of palatogenesis and are essential in strategies designed to ameliorate palatal malformations in humans. Recent studies have shown that the zebrafish palate, or anterior neurocranium, is under similar genetic control to the amniote palatal skeleton. We extensively analyzed palatogenesis in zebrafish to determine the similarity of gene expression and function across vertebrates. By 36 hpf palatogenic cranial neural crest cells reside in homologous regions of the developing face compared to amniote species. Transcription factors and signaling molecules regulating mouse palatogenesis are expressed in similar domains during palatogenesis in zebrafish. Functional investigation of a subset of these genes, fgf10a, tgfb2, pax9 and smad5 revealed their necessity in zebrafish palatogenesis. Collectively, these results suggest that the gene regulatory networks regulating palatogenesis may be conserved across vertebrate species, demonstrating the utility of zebrafish as a model for palatogenesis. PMID:22016187

Clock jitter generator with picoseconds resolution

NASA Astrophysics Data System (ADS)

Jovanović, Goran; Stojčev, Mile; Nikolić, Tatjana

2013-06-01

The clock is one of the most critical signals in any synchronous system. As CMOS technology has scaled, supply voltages have dropped chip power consumption has increased and the effects of jitter due to clock frequency increase have become critical and jitter budget has become tighter. This article describes design and development of low-cost mixed-signal programmable jitter generator with high resolution. The digital technique is used for coarse-grain and an analogue technique for fine-grain clock phase shifting. Its structure allows injection of various random and deterministic jitter components in a controllable and programmable fashion. Each jitter component can be switched on or off. The jitter generator can be used in jitter tolerance test and jitter transfer function measurement of high-speed synchronous digital circuits. At operating system clock frequency of 220 MHz, a jitter with 4 ps resolution can be injected.

Next Generation JPL Ultra-Stable Trapped Ion Atomic Clocks

NASA Technical Reports Server (NTRS)

Burt, Eric; Tucker, Blake; Larsen, Kameron; Hamell, Robert; Tjoelker, Robert

2013-01-01

Over the past decade, trapped ion atomic clock development at the Jet Propulsion Laboratory (JPL) has focused on two directions: 1) new atomic clock technology for space flight applications that require strict adherence to size, weight, and power requirements, and 2) ultra-stable atomic clocks, usually for terrestrial applications emphasizing ultimate performance. In this paper we present a new ultra-stable trapped ion clock designed, built, and tested in the second category. The first new standard, L10, will be delivered to the Naval Research Laboratory for use in characterizing DoD space clocks.

Systematic approaches to toxicology in the zebrafish.

PubMed

Peterson, Randall T; Macrae, Calum A

2012-01-01

As the current paradigms of drug discovery evolve, it has become clear that a more comprehensive understanding of the interactions between small molecules and organismal biology will be vital. The zebrafish is emerging as a complement to existing in vitro technologies and established preclinical in vivo models that can be scaled for high-throughput. In this review, we highlight the current status of zebrafish toxicology studies, identify potential future niches for the model in the drug development pipeline, and define the hurdles that must be overcome as zebrafish technologies are refined for systematic toxicology.

Thrombus segmentation by texture dynamics from microscopic image sequences

NASA Astrophysics Data System (ADS)

Brieu, Nicolas; Serbanovic-Canic, Jovana; Cvejic, Ana; Stemple, Derek; Ouwehand, Willem; Navab, Nassir; Groher, Martin

2010-03-01

The genetic factors of thrombosis are commonly explored by microscopically imaging the coagulation of blood cells induced by injuring a vessel of mice or of zebrafish mutants. The latter species is particularly interesting since skin transparency permits to non-invasively acquire microscopic images of the scene with a CCD camera and to estimate the parameters characterizing the thrombus development. These parameters are currently determined by manual outlining, which is both error prone and extremely time consuming. Even though a technique for automatic thrombus extraction would be highly valuable for gene analysts, little work can be found, which is mainly due to very low image contrast and spurious structures. In this work, we propose to semi-automatically segment the thrombus over time from microscopic image sequences of wild-type zebrafish larvae. To compensate the lack of valuable spatial information, our main idea consists of exploiting the temporal information by modeling the variations of the pixel intensities over successive temporal windows with a linear Markov-based dynamic texture formalization. We then derive an image from the estimated model parameters, which represents the probability of a pixel to belong to the thrombus. We employ this probability image to accurately estimate the thrombus position via an active contour segmentation incorporating also prior and spatial information of the underlying intensity images. The performance of our approach is tested on three microscopic image sequences. We show that the thrombus is accurately tracked over time in each sequence if the respective parameters controlling prior influence and contour stiffness are correctly chosen.

Circadian Rhythms, the Molecular Clock, and Skeletal Muscle

PubMed Central

Lefta, Mellani; Wolff, Gretchen; Esser, Karyn A.

2015-01-01

Almost all organisms ranging from single cell bacteria to humans exhibit a variety of behavioral, physiological, and biochemical rhythms. In mammals, circadian rhythms control the timing of many physiological processes over a 24-h period, including sleep-wake cycles, body temperature, feeding, and hormone production. This body of research has led to defined characteristics of circadian rhythms based on period length, phase, and amplitude. Underlying circadian behaviors is a molecular clock mechanism found in most, if not all, cell types including skeletal muscle. The mammalian molecular clock is a complex of multiple oscillating networks that are regulated through transcriptional mechanisms, timed protein turnover, and input from small molecules. At this time, very little is known about circadian aspects of skeletal muscle function/metabolism but some progress has been made on understanding the molecular clock in skeletal muscle. The goal of this chapter is to provide the basic terminology and concepts of circadian rhythms with a more detailed review of the current state of knowledge of the molecular clock, with reference to what is known in skeletal muscle. Research has demonstrated that the molecular clock is active in skeletal muscles and that the muscle-specific transcription factor, MyoD, is a direct target of the molecular clock. Skeletal muscle of clock-compromised mice, Bmal1−/− and ClockΔ19 mice, are weak and exhibit significant disruptions in expression of many genes required for adult muscle structure and metabolism. We suggest that the interaction between the molecular clock, MyoD, and metabolic factors, such as PGC-1, provide a potential system of feedback loops that may be critical for both maintenance and adaptation of skeletal muscle. PMID:21621073

Zebrafish models in neuropsychopharmacology and CNS drug discovery.

PubMed

Khan, Kanza M; Collier, Adam D; Meshalkina, Darya A; Kysil, Elana V; Khatsko, Sergey L; Kolesnikova, Tatyana; Morzherin, Yury Yu; Warnick, Jason E; Kalueff, Allan V; Echevarria, David J

2017-07-01

Despite the high prevalence of neuropsychiatric disorders, their aetiology and molecular mechanisms remain poorly understood. The zebrafish (Danio rerio) is increasingly utilized as a powerful animal model in neuropharmacology research and in vivo drug screening. Collectively, this makes zebrafish a useful tool for drug discovery and the identification of disordered molecular pathways. Here, we discuss zebrafish models of selected human neuropsychiatric disorders and drug-induced phenotypes. As well as covering a broad range of brain disorders (from anxiety and psychoses to neurodegeneration), we also summarize recent developments in zebrafish genetics and small molecule screening, which markedly enhance the disease modelling and the discovery of novel drug targets. © 2017 The British Pharmacological Society.

Silver nanoparticles induce developmental stage-specific embryonic phenotypes in zebrafish

NASA Astrophysics Data System (ADS)

Lee, Kerry J.; Browning, Lauren M.; Nallathamby, Prakash D.; Osgood, Christopher J.; Xu, Xiao-Hong Nancy

2013-11-01

Much is anticipated from the development and deployment of nanomaterials in biological organisms, but concerns remain regarding their biocompatibility and target specificity. Here we report our study of the transport, biocompatibility and toxicity of purified and stable silver nanoparticles (Ag NPs, 13.1 +/- 2.5 nm in diameter) upon the specific developmental stages of zebrafish embryos using single NP plasmonic spectroscopy. We find that single Ag NPs passively diffuse into five different developmental stages of embryos (cleavage, early-gastrula, early-segmentation, late-segmentation, and hatching stages), showing stage-independent diffusion modes and diffusion coefficients. Notably, the Ag NPs induce distinctive stage and dose-dependent phenotypes and nanotoxicity, upon their acute exposure to the Ag NPs (0-0.7 nM) for only 2 h. The late-segmentation embryos are most sensitive to the NPs with the lowest critical concentration (CNP,c << 0.02 nM) and highest percentages of cardiac abnormalities, followed by early-segmentation embryos (CNP,c < 0.02 nM), suggesting that disruption of cell differentiation by the NPs causes the most toxic effects on embryonic development. The cleavage-stage embryos treated with the NPs develop into a wide variety of phenotypes (abnormal finfold, tail/spinal cord flexure, cardiac malformation/edema, yolk sac edema, and acephaly). These organ structures are not yet developed in cleavage-stage embryos, suggesting that the earliest determinative events to create these structures are ongoing, and disrupted by NPs, which leads to the downstream effects. In contrast, the hatching embryos are most resistant to the Ag NPs, and majority of embryos (94%) develop normally, and none of them develop abnormally. Interestingly, early-gastrula embryos are less sensitive to the NPs than cleavage and segmentation stage embryos, and do not develop abnormally. These important findings suggest that the Ag NPs are not simple poisons, and they can target

Galanin regulates blood glucose level in the zebrafish: a morphological and functional study.

PubMed

Podlasz, P; Jakimiuk, A; Chmielewska-Krzesinska, M; Kasica, N; Nowik, N; Kaleczyc, J

2016-01-01

The present study has demonstrated the galaninergic innervation of the endocrine pancreas including sources of the galaninergic nerve fibers, and the influence of galanin receptor agonists on blood glucose level in the zebrafish. For the first time, a very abundant galaninergic innervation of the endocrine pancreas during development is shown, from the second day post-fertilization to adulthood. The fibers originated from ganglia consisting of galanin-IR, non-adrenergic (non-sensory) neurons located rostrally to the pancreatic tissue. The ganglia were found on the dorsal side of the initial part of the anterior intestinal segment, close to the intestinal branch of the vagus nerve. The galanin-IR neurons did not show immunoreactivity for applied antibodies against tyrosine hydroxylase, choline acetyltransferase, and vesicular acetylcholine transporter. Intraperitoneal injections of galanin analog NAX 5055 resulted in a statistically significant increase in the blood glucose level. Injections of another galanin receptor agonist, galnon, also caused a rise in blood glucose level; however, it was not statistically significant. The present findings suggest that, like in mammals, in the zebrafish galanin is involved in the regulation of blood glucose level. However, further studies are needed to elucidate the exact mechanism of the galanin action.

Incorporating zebrafish omics into chemical biology and toxicology.

PubMed

Sukardi, Hendrian; Ung, Choong Yong; Gong, Zhiyuan; Lam, Siew Hong

2010-03-01

In this communication, we describe the general aspects of omics approaches for analyses of transcriptome, proteome, and metabolome, and how they can be strategically incorporated into chemical screening and perturbation studies using the zebrafish system. Pharmacological efficacy and selectivity of chemicals can be evaluated based on chemical-induced phenotypic effects; however, phenotypic observation has limitations in identifying mechanistic action of chemicals. We suggest adapting gene-expression-based high-throughput screening as a complementary strategy to zebrafish-phenotype-based screening for mechanistic insights about the mode of action and toxicity of a chemical, large-scale predictive applications and comparative analysis of chemical-induced omics signatures, which are useful to identify conserved biological responses, signaling pathways, and biomarkers. The potential mechanistic, predictive, and comparative applications of omics approaches can be implemented in the zebrafish system. Examples of these using the omics approaches in zebrafish, including data of ours and others, are presented and discussed. Omics also facilitates the translatability of zebrafish studies across species through comparison of conserved chemical-induced responses. This review is intended to update interested readers with the current omics approaches that have been applied in chemical studies on zebrafish and their potential in enhancing discovery in chemical biology.

Mapping the zebrafish brain methylome using reduced representation bisulfite sequencing

PubMed Central

Chatterjee, Aniruddha; Ozaki, Yuichi; Stockwell, Peter A; Horsfield, Julia A; Morison, Ian M; Nakagawa, Shinichi

2013-01-01

Reduced representation bisulfite sequencing (RRBS) has been used to profile DNA methylation patterns in mammalian genomes such as human, mouse and rat. The methylome of the zebrafish, an important animal model, has not yet been characterized at base-pair resolution using RRBS. Therefore, we evaluated the technique of RRBS in this model organism by generating four single-nucleotide resolution DNA methylomes of adult zebrafish brain. We performed several simulations to show the distribution of fragments and enrichment of CpGs in different in silico reduced representation genomes of zebrafish. Four RRBS brain libraries generated 98 million sequenced reads and had higher frequencies of multiple mapping than equivalent human RRBS libraries. The zebrafish methylome indicates there is higher global DNA methylation in the zebrafish genome compared with its equivalent human methylome. This observation was confirmed by RRBS of zebrafish liver. High coverage CpG dinucleotides are enriched in CpG island shores more than in the CpG island core. We found that 45% of the mapped CpGs reside in gene bodies, and 7% in gene promoters. This analysis provides a roadmap for generating reproducible base-pair level methylomes for zebrafish using RRBS and our results provide the first evidence that RRBS is a suitable technique for global methylation analysis in zebrafish. PMID:23975027

Body weight, metabolism and clock genes

PubMed Central

2010-01-01

Biological rhythms are present in the lives of almost all organisms ranging from plants to more evolved creatures. These oscillations allow the anticipation of many physiological and behavioral mechanisms thus enabling coordination of rhythms in a timely manner, adaption to environmental changes and more efficient organization of the cellular processes responsible for survival of both the individual and the species. Many components of energy homeostasis exhibit circadian rhythms, which are regulated by central (suprachiasmatic nucleus) and peripheral (located in other tissues) circadian clocks. Adipocyte plays an important role in the regulation of energy homeostasis, the signaling of satiety and cellular differentiation and proliferation. Also, the adipocyte circadian clock is probably involved in the control of many of these functions. Thus, circadian clocks are implicated in the control of energy balance, feeding behavior and consequently in the regulation of body weight. In this regard, alterations in clock genes and rhythms can interfere with the complex mechanism of metabolic and hormonal anticipation, contributing to multifactorial diseases such as obesity and diabetes. The aim of this review was to define circadian clocks by describing their functioning and role in the whole body and in adipocyte metabolism, as well as their influence on body weight control and the development of obesity. PMID:20712885

Huygens’ clocks revisited

PubMed Central

Kitanov, Petko M.; Langford, William F.

2017-01-01

In 1665, Huygens observed that two identical pendulum clocks, weakly coupled through a heavy beam, soon synchronized with the same period and amplitude but with the two pendula swinging in opposite directions. This behaviour is now called anti-phase synchronization. This paper presents an analysis of the behaviour of a large class of coupled identical oscillators, including Huygens' clocks, using methods of equivariant bifurcation theory. The equivariant normal form for such systems is developed and the possible solutions are characterized. The transformation of the physical system parameters to the normal form parameters is given explicitly and applied to the physical values appropriate for Huygens' clocks, and to those of more recent studies. It is shown that Huygens' physical system could only exhibit anti-phase motion, explaining why Huygens observed exclusively this. By contrast, some more recent researchers have observed in-phase or other more complicated motion in their own experimental systems. Here, it is explained which physical characteristics of these systems allow for the existence of these other types of stable solutions. The present analysis not only accounts for these previously observed solutions in a unified framework, but also introduces behaviour not classified by other authors, such as a synchronized toroidal breather and a chaotic toroidal breather. PMID:28989780

GNSS Clock Error Impacts on Radio Occultation Retrievals

NASA Astrophysics Data System (ADS)

Weiss, Jan; Sokolovskiy, Sergey; Schreiner, Bill; Yoon, Yoke

2017-04-01

We assess the impacts of GPS and GLONASS clock errors on radio occultation retrieval of bending angle, refractivity, and temperature from low Earth orbit. The major contributing factor is the interpretation of GNSS clock offsets sampled at 30 sec or longer intervals. Using 1 Hz GNSS clock estimates as truth we apply several interpolation and fitting schemes to evaluate how they affect the accuracy of atmospheric retrieval products. The results are organized by GPS and GLONASS space vehicle and the GNSS clock interpolation/fitting scheme. We find that bending angle error is roughly similar for all current GPS transmitters (about 0.7 mcrad) but note some differences related to the type of atomic oscillator onboard the transmitter satellite. GLONASS bending angle errors show more variation over the constellation and are approximately two times larger than GPS. An investigation of the transmitter clock spectra reveals this is due to more power in periods between 2-10 sec. Retrieved refractivity and temperature products show clear differences between GNSS satellite generations, and indicate that GNSS clocks sampled at intervals smaller than 5 sec significantly improve accuracy, particularly for GLONASS. We conclude by summarizing the tested GNSS clock estimation and application strategies in the context of current and future radio occultation missions.

Accelerated Optical Projection Tomography Applied to In Vivo Imaging of Zebrafish

PubMed Central

Correia, Teresa; Yin, Jun; Ramel, Marie-Christine; Andrews, Natalie; Katan, Matilda; Bugeon, Laurence; Dallman, Margaret J.; McGinty, James; Frankel, Paul; French, Paul M. W.; Arridge, Simon

2015-01-01

Optical projection tomography (OPT) provides a non-invasive 3-D imaging modality that can be applied to longitudinal studies of live disease models, including in zebrafish. Current limitations include the requirement of a minimum number of angular projections for reconstruction of reasonable OPT images using filtered back projection (FBP), which is typically several hundred, leading to acquisition times of several minutes. It is highly desirable to decrease the number of required angular projections to decrease both the total acquisition time and the light dose to the sample. This is particularly important to enable longitudinal studies, which involve measurements of the same fish at different time points. In this work, we demonstrate that the use of an iterative algorithm to reconstruct sparsely sampled OPT data sets can provide useful 3-D images with 50 or fewer projections, thereby significantly decreasing the minimum acquisition time and light dose while maintaining image quality. A transgenic zebrafish embryo with fluorescent labelling of the vasculature was imaged to acquire densely sampled (800 projections) and under-sampled data sets of transmitted and fluorescence projection images. The under-sampled OPT data sets were reconstructed using an iterative total variation-based image reconstruction algorithm and compared against FBP reconstructions of the densely sampled data sets. To illustrate the potential for quantitative analysis following rapid OPT data acquisition, a Hessian-based method was applied to automatically segment the reconstructed images to select the vasculature network. Results showed that 3-D images of the zebrafish embryo and its vasculature of sufficient visual quality for quantitative analysis can be reconstructed using the iterative algorithm from only 32 projections—achieving up to 28 times improvement in imaging speed and leading to total acquisition times of a few seconds. PMID:26308086

Using Integer Clocks to Verify the Timing-Sync Sensor Network Protocol

NASA Technical Reports Server (NTRS)

Huang, Xiaowan; Singh, Anu; Smolka, Scott A.

2010-01-01

We use the UPPAAL model checker for Timed Automata to verify the Timing-Sync time-synchronization protocol for sensor networks (TPSN). The TPSN protocol seeks to provide network-wide synchronization of the distributed clocks in a sensor network. Clock-synchronization algorithms for sensor networks such as TPSN must be able to perform arithmetic on clock values to calculate clock drift and network propagation delays. They must be able to read the value of a local clock and assign it to another local clock. Such operations are not directly supported by the theory of Timed Automata. To overcome this formal-modeling obstacle, we augment the UPPAAL specification language with the integer clock derived type. Integer clocks, which are essentially integer variables that are periodically incremented by a global pulse generator, greatly facilitate the encoding of the operations required to synchronize clocks as in the TPSN protocol. With this integer-clock-based model of TPSN in hand, we use UPPAAL to verify that the protocol achieves network-wide time synchronization and is devoid of deadlock. We also use the UPPAAL Tracer tool to illustrate how integer clocks can be used to capture clock drift and resynchronization during protocol execution

Generation and characterization of Kctd15 mutations in zebrafish

PubMed Central

Heffer, Alison; Marquart, Gregory D.; Aquilina-Beck, Allisan; Saleem, Nabil; Burgess, Harold A.

2017-01-01

Potassium channel tetramerization domain containing 15 (Kctd15) was previously found to have a role in early neural crest (NC) patterning, specifically delimiting the region where NC markers are expressed via repression of transcription factor AP-2a and inhibition of Wnt signaling. We used transcription activator-like effector nucleases (TALENs) to generate null mutations in zebrafish kctd15a and kctd15b paralogs to study the in vivo role of Kctd15. We found that while deletions producing frame-shift mutations in each paralog showed no apparent phenotype, kctd15a/b double mutant zebrafish are smaller in size and show several phenotypes including some affecting the NC, such as expansion of the early NC domain, increased pigmentation, and craniofacial defects. Both melanophore and xanthophore pigment cell numbers and early markers are up-regulated in the double mutants. While we find no embryonic craniofacial defects, adult mutants have a deformed maxillary segment and missing barbels. By confocal imaging of mutant larval brains we found that the torus lateralis (TLa), a region implicated in gustatory networks in other fish, is absent. Ablation of this brain tissue in wild type larvae mimics some aspects of the mutant growth phenotype. Thus kctd15 mutants show deficits in the development of both neural crest derivatives, and specific regions within the central nervous system, leading to a strong reduction in normal growth rates. PMID:29216270

Cryptochrome Mediates Light-Dependent Magnetosensitivity of Drosophila's Circadian Clock

PubMed Central

Yoshii, Taishi; Ahmad, Margaret; Helfrich-Förster, Charlotte

2009-01-01

Since 1960, magnetic fields have been discussed as Zeitgebers for circadian clocks, but the mechanism by which clocks perceive and process magnetic information has remained unknown. Recently, the radical-pair model involving light-activated photoreceptors as magnetic field sensors has gained considerable support, and the blue-light photoreceptor cryptochrome (CRY) has been proposed as a suitable molecule to mediate such magnetosensitivity. Since CRY is expressed in the circadian clock neurons and acts as a critical photoreceptor of Drosophila's clock, we aimed to test the role of CRY in magnetosensitivity of the circadian clock. In response to light, CRY causes slowing of the clock, ultimately leading to arrhythmic behavior. We expected that in the presence of applied magnetic fields, the impact of CRY on clock rhythmicity should be altered. Furthermore, according to the radical-pair hypothesis this response should be dependent on wavelength and on the field strength applied. We tested the effect of applied static magnetic fields on the circadian clock and found that flies exposed to these fields indeed showed enhanced slowing of clock rhythms. This effect was maximal at 300 μT, and reduced at both higher and lower field strengths. Clock response to magnetic fields was present in blue light, but absent under red-light illumination, which does not activate CRY. Furthermore, cryb and cryOUT mutants did not show any response, and flies overexpressing CRY in the clock neurons exhibited an enhanced response to the field. We conclude that Drosophila's circadian clock is sensitive to magnetic fields and that this sensitivity depends on light activation of CRY and on the applied field strength, consistent with the radical pair mechanism. CRY is widespread throughout biological systems and has been suggested as receptor for magnetic compass orientation in migratory birds. The present data establish the circadian clock of Drosophila as a model system for CRY

Zebrafish models for the functional genomics of neurogenetic disorders.

PubMed

Kabashi, Edor; Brustein, Edna; Champagne, Nathalie; Drapeau, Pierre

2011-03-01

In this review, we consider recent work using zebrafish to validate and study the functional consequences of mutations of human genes implicated in a broad range of degenerative and developmental disorders of the brain and spinal cord. Also we present technical considerations for those wishing to study their own genes of interest by taking advantage of this easily manipulated and clinically relevant model organism. Zebrafish permit mutational analyses of genetic function (gain or loss of function) and the rapid validation of human variants as pathological mutations. In particular, neural degeneration can be characterized at genetic, cellular, functional, and behavioral levels. Zebrafish have been used to knock down or express mutations in zebrafish homologs of human genes and to directly express human genes bearing mutations related to neurodegenerative disorders such as spinal muscular atrophy, ataxia, hereditary spastic paraplegia, amyotrophic lateral sclerosis (ALS), epilepsy, Huntington's disease, Parkinson's disease, fronto-temporal dementia, and Alzheimer's disease. More recently, we have been using zebrafish to validate mutations of synaptic genes discovered by large-scale genomic approaches in developmental disorders such as autism, schizophrenia, and non-syndromic mental retardation. Advances in zebrafish genetics such as multigenic analyses and chemical genetics now offer a unique potential for disease research. Thus, zebrafish hold much promise for advancing the functional genomics of human diseases, the understanding of the genetics and cell biology of degenerative and developmental disorders, and the discovery of therapeutics. This article is part of a Special Issue entitled Zebrafish Models of Neurological Diseases. Copyright © 2010 Elsevier B.V. All rights reserved.

Morphologic analysis of the zebrafish digestive system.

PubMed

Trotter, Andrew J; Parslow, Adam C; Heath, Joan K

2009-01-01

The zebrafish provides an ideal model for the study of vertebrate organogenesis, including the formation of the digestive tract and its associated organs. Despite optical transparency of embryos, the internal position of the developing digestive system and its close juxtaposition with the yolk initially made morphological analysis relatively challenging, particularly during the first 3 d of development. However, methodologies have been successfully developed to address these problems and comprehensive morphologic analysis of the developing digestive system has now been achieved using a combination of light and fluorescence microscope approaches-including confocal analysis-to visualize wholemount and histological preparations of zebrafish embryos. Furthermore, the expanding number of antibodies that cross-react with zebrafish proteins and the generation of tissue-specific transgenic green fluorescent protein reporter lines that mark specific cell and tissue compartments have greatly enhanced our ability to successfully image the developing zebrafish digestive system.

Open-RAC: Open-Design, Recirculating and Auto-Cleaning Zebrafish Maintenance System.

PubMed

Nema, Shubham; Bhargava, Yogesh

2017-08-01

Zebrafish is a vertebrate animal model. Their maintenance in large number under laboratory conditions is a daunting task. Commercially available recirculating zebrafish maintenance systems are used to efficiently handle the tasks of automatic sediment cleaning from zebrafish tanks with minimal waste of water. Due to their compact nature, they also ensure the maximal use of available lab space. However, the high costs of commercial systems present a limitation to researchers with limited funds. A cost-effective zebrafish maintenance system with major features offered by commercially available systems is highly desirable. Here, we describe a compact and recirculating zebrafish maintenance system. Our system is composed of cost-effective components, which are available in local markets and/or can be procured via online vendors. Depending on the expertise of end users, the system can be assembled in 2 days. The system is completely customizable as it offers geometry independent zebrafish tanks that are capable of auto-cleaning the sediments. Due to these features, we called our setup as Open-RAC (Open-design, Recirculating and Auto-Cleaning zebrafish maintenance system). Open-RAC is a cost-effective and viable alternative to the currently available zebrafish maintenance systems. Thus, we believe that the use of Open-RAC could promote the zebrafish research by removing the cost barrier for researchers.

Inexpensive Clock for Displaying Planetary or Sidereal Time

NASA Technical Reports Server (NTRS)

Lux, James

2007-01-01

An inexpensive wall clock has been devised for displaying solar time or sidereal time as it would be perceived on a planet other than the Earth, or for displaying sidereal time on the Earth. The concept of a wall clock synchronized to a period other than the terrestrial mean solar day is not new in itself. What is new here is that the clock is realized through a relatively simple electronic modification of a common battery-powered, quartz-crystal-oscillator-driven wall clock. The essence of the modification is to shut off the internal oscillator of the clock and replace the internal-oscillator output signal with a signal of the required frequency generated by an external oscillator. The unmodified clock electronic circuitry includes a quartz crystal connected to an integrated circuit (IC) that includes, among other parts, a buffer amplifier that conditions the oscillator output. The modification is effected by removing the quartz crystal and connecting the output terminal of the external oscillator, via a capacitor, to the input terminal of the buffer amplifier

A clock network for geodesy and fundamental science

PubMed Central

Lisdat, C.; Grosche, G.; Quintin, N.; Shi, C.; Raupach, S.M.F.; Grebing, C.; Nicolodi, D.; Stefani, F.; Al-Masoudi, A.; Dörscher, S.; Häfner, S.; Robyr, J.-L.; Chiodo, N.; Bilicki, S.; Bookjans, E.; Koczwara, A.; Koke, S.; Kuhl, A.; Wiotte, F.; Meynadier, F.; Camisard, E.; Abgrall, M.; Lours, M.; Legero, T.; Schnatz, H.; Sterr, U.; Denker, H.; Chardonnet, C.; Le Coq, Y.; Santarelli, G.; Amy-Klein, A.; Le Targat, R.; Lodewyck, J.; Lopez, O; Pottie, P.-E.

2016-01-01

Leveraging the unrivalled performance of optical clocks as key tools for geo-science, for astronomy and for fundamental physics beyond the standard model requires comparing the frequency of distant optical clocks faithfully. Here, we report on the comparison and agreement of two strontium optical clocks at an uncertainty of 5 × 10−17 via a newly established phase-coherent frequency link connecting Paris and Braunschweig using 1,415 km of telecom fibre. The remote comparison is limited only by the instability and uncertainty of the strontium lattice clocks themselves, with negligible contributions from the optical frequency transfer. A fractional precision of 3 × 10−17 is reached after only 1,000 s averaging time, which is already 10 times better and more than four orders of magnitude faster than any previous long-distance clock comparison. The capability of performing high resolution international clock comparisons paves the way for a redefinition of the unit of time and an all-optical dissemination of the SI-second. PMID:27503795

A tunable artificial circadian clock in clock-defective mice

PubMed Central

D'Alessandro, Matthew; Beesley, Stephen; Kim, Jae Kyoung; Chen, Rongmin; Abich, Estela; Cheng, Wayne; Yi, Paul; Takahashi, Joseph S.; Lee, Choogon

2015-01-01

Self-sustaining oscillations are essential for diverse physiological functions such as the cell cycle, insulin secretion and circadian rhythms. Synthetic oscillators using biochemical feedback circuits have been generated in cell culture. These synthetic systems provide important insight into design principles for biological oscillators, but have limited similarity to physiological pathways. Here we report the generation of an artificial, mammalian circadian clock in vivo, capable of generating robust, tunable circadian rhythms. In mice deficient in Per1 and Per2 genes (thus lacking circadian rhythms), we artificially generate PER2 rhythms and restore circadian sleep/wake cycles with an inducible Per2 transgene. Our artificial clock is tunable as the period and phase of the rhythms can be modulated predictably. This feature, and other design principles of our work, might enhance the study and treatment of circadian dysfunction and broader aspects of physiology involving biological oscillators. PMID:26617050

Fast Clock Recovery for Digital Communications

NASA Technical Reports Server (NTRS)

Tell, R. G.

1985-01-01

Circuit extracts clock signal from random non-return-to-zero data stream, locking onto clock within one bit period at 1-gigabitper-second data rate. Circuit used for synchronization in opticalfiber communications. Derives speed from very short response time of gallium arsenide metal/semiconductor field-effect transistors (MESFET's).

Realistic Clocks for a Universe Without Time

NASA Astrophysics Data System (ADS)

Bryan, K. L. H.; Medved, A. J. M.

2018-01-01

There are a number of problematic features within the current treatment of time in physical theories, including the "timelessness" of the Universe as encapsulated by the Wheeler-DeWitt equation. This paper considers one particular investigation into resolving this issue; a conditional probability interpretation that was first proposed by Page and Wooters. Those authors addressed the apparent timelessness by subdividing a faux Universe into two entangled parts, "the clock" and "the remainder of the Universe", and then synchronizing the effective dynamics of the two subsystems by way of conditional probabilities. The current treatment focuses on the possibility of using a (somewhat) realistic clock system; namely, a coherent-state description of a damped harmonic oscillator. This clock proves to be consistent with the conditional probability interpretation; in particular, a standard evolution operator is identified with the position of the clock playing the role of time for the rest of the Universe. Restrictions on the damping factor are determined and, perhaps contrary to expectations, the optimal choice of clock is not necessarily one of minimal damping.

APOLLO clock performance and normal point corrections

NASA Astrophysics Data System (ADS)

Liang, Y.; Murphy, T. W., Jr.; Colmenares, N. R.; Battat, J. B. R.

2017-12-01

The Apache point observatory lunar laser-ranging operation (APOLLO) has produced a large volume of high-quality lunar laser ranging (LLR) data since it began operating in 2006. For most of this period, APOLLO has relied on a GPS-disciplined, high-stability quartz oscillator as its frequency and time standard. The recent addition of a cesium clock as part of a timing calibration system initiated a comparison campaign between the two clocks. This has allowed correction of APOLLO range measurements—called normal points—during the overlap period, but also revealed a mechanism to correct for systematic range offsets due to clock errors in historical APOLLO data. Drift of the GPS clock on  ∼1000 s timescales contributed typically 2.5 mm of range error to APOLLO measurements, and we find that this may be reduced to  ∼1.6 mm on average. We present here a characterization of APOLLO clock errors, the method by which we correct historical data, and the resulting statistics.

Optical Lattice Clocks with Weakly Bound Molecules

NASA Astrophysics Data System (ADS)

Borkowski, Mateusz

2018-02-01

Optical molecular clocks promise unparalleled sensitivity to the temporal variation of the electron-to-proton mass ratio and insight into possible new physics beyond the standard model. We propose to realize a molecular clock with bosonic 174Yb2 molecules, where the forbidden 1S0 →3P0 clock transition would be induced magnetically. The use of a bosonic species avoids possible complications due to the hyperfine structure present in fermionic species. While direct clock line photoassociation would be challenging, weakly bound ground state molecules could be produced by stimulated Raman adiabatic passage and used instead. The recent scattering measurements [L. Franchi, et al. New J. Phys. 19, 103037 (2017), 10.1088/1367-2630/aa8fb4] enable us to determine the positions of target 1S0 +3P0 vibrational levels and calculate the Franck-Condon factors for clock transitions between ground and excited molecular states. The resulting magnetically induced Rabi frequencies are similar to those for atoms hinting that an experimental realization is feasible. A successful observation could pave the way towards Hz-level molecular spectroscopy.

Transvection Arising from Transgene Interactions in Zebrafish.

PubMed

Keefe, Matthew D; Bonkowsky, Joshua L

2017-02-01

There has been a rapid expansion in use of transgenic technologies in zebrafish. We report a novel example of transinteractions of genetic elements, or transvection. This interaction led to a novel expression pattern and illustrates a precautionary example regarding use of transgenes in zebrafish.

Behavioural fever in zebrafish larvae.

PubMed

Rey, Sonia; Moiche, Visila; Boltaña, Sebastian; Teles, Mariana; MacKenzie, Simon

2017-02-01

Behavioural fever has been reported in different species of mobile ectotherms including the zebrafish, Danio rerio, in response to exogenous pyrogens. In this study we report, to our knowledge for the first time, upon the ontogenic onset of behavioural fever in zebrafish (Danio rerio) larvae. For this, zebrafish larvae (from first feeding to juveniles) were placed in a continuous thermal gradient providing the opportunity to select their preferred temperature. The novel thermal preference aquarium was based upon a continuous vertical column system and allows for non-invasive observation of larvae vertical distribution under isothermal (T R at 28 °C) and thermal gradient conditions (T CH : 28-32 °C). Larval thermal preference was assessed under both conditions with or without an immersion challenge, in order to detect the onset of the behavioural fever response. Our results defined the onset of the dsRNA induced behavioural fever at 18-20 days post fertilization (dpf). Significant differences were observed in dsRNA challenged larvae, which prefer higher temperatures (1-4 °C increase) throughout the experimental period as compared to non-challenged larvae. In parallel we measured the abundance of antiviral transcripts; viperin, gig2, irf7, trim25 and Mxb mRNAs in dsRNA challenged larvae under both thermal regimes: T R and T Ch . Significant increases in the abundance of all measured transcripts were recorded under thermal choice conditions signifying that thermo-coupling and the resultant enhancement of the immune response to dsRNA challenge occurs from 18 dpf onwards in the zebrafish. The results are of importance as they identify a key developmental stage where the neuro-immune interface matures in the zebrafish likely providing increased resistance to viral infection. Copyright © 2016 Elsevier Ltd. All rights reserved.

Zebrafish models for translational neuroscience research: from tank to bedside

PubMed Central

Stewart, Adam Michael; Braubach, Oliver; Spitsbergen, Jan; Gerlai, Robert; Kalueff, Allan V.

2014-01-01

The zebrafish (Danio rerio) is emerging as a new important species for studying mechanisms of brain function and dysfunction. Focusing on selected central nervous system (CNS) disorders (brain cancer, epilepsy, and anxiety) and using them as examples, we discuss the value of zebrafish models in translational neuroscience. We further evaluate the contribution of zebrafish to neuroimaging, circuit level, and drug discovery research. Outlining the role of zebrafish in modeling a wide range of human brain disorders, we also summarize recent applications and existing challenges in this field. Finally, we emphasize the potential of zebrafish models in behavioral phenomics and high-throughput genetic/small molecule screening, which is critical for CNS drug discovery and identifying novel candidate genes. PMID:24726051

Characterization of behavioral and endocrine effects of LSD on zebrafish.

PubMed

Grossman, Leah; Utterback, Eli; Stewart, Adam; Gaikwad, Siddharth; Chung, Kyung Min; Suciu, Christopher; Wong, Keith; Elegante, Marco; Elkhayat, Salem; Tan, Julia; Gilder, Thomas; Wu, Nadine; Dileo, John; Cachat, Jonathan; Kalueff, Allan V

2010-12-25

Lysergic acid diethylamide (LSD) is a potent hallucinogenic drug that strongly affects animal and human behavior. Although adult zebrafish (Danio rerio) are emerging as a promising neurobehavioral model, the effects of LSD on zebrafish have not been investigated previously. Several behavioral paradigms (the novel tank, observation cylinder, light-dark box, open field, T-maze, social preference and shoaling tests), as well as modern video-tracking tools and whole-body cortisol assay were used to characterize the effects of acute LSD in zebrafish. While lower doses (5-100 microg/L) did not affect zebrafish behavior, 250 microg/L LSD increased top dwelling and reduced freezing in the novel tank and observation cylinder tests, also affecting spatiotemporal patterns of activity (as assessed by 3D reconstruction of zebrafish traces and ethograms). LSD evoked mild thigmotaxis in the open field test, increased light behavior in the light-dark test, reduced the number of arm entries and freezing in the T-maze and social preference test, without affecting social preference. In contrast, LSD affected zebrafish shoaling (increasing the inter-fish distance in a group), and elevated whole-body cortisol levels. Overall, our findings show sensitivity of zebrafish to LSD action, and support the use of zebrafish models to study hallucinogenic drugs of abuse. Copyright (c) 2010 Elsevier B.V. All rights reserved.

Hourly Updated GNSS Orbit and Clock

NASA Astrophysics Data System (ADS)

Song, S.; Xue, J.

2016-12-01

With the development of the performance of GNSS, the hourly updated orbit and clock of GNSS are paid much more attention and used by more and more users because of the timeliness and high accuracy. The hourly GNSS orbit and clock are produced routinely in Shanghai Analysis Center(AC) of the International GNSS Monitoring and Assessment Service (iGMAS).In this article, the accuracy of hourly and 6-hourly updated ultra-rapid GPS,GLONASS,GALILEO,BDS orbit and clock (SHU1 and SHU6) are analyzed relative to the final production in detail. The analysis show that, in calculation session, there's no much difference between the mean SHU1 and SHU6 RMS and STD for GNSS orbit and clock. However, for BDS clock in prediction session, the RMS and STD of BDS SHU1 are 2.6ns and 0.5ns respectively, the RMS of BDS SHU6 increase from 2.7ns to 4.5ns from the 1st to the 6th hour prediction session, but there's no much changes of STD. For GPS clock in prediction session, the RMS and STD of GPS SHU1 is quite stable with 0.5ns and 0.2ns.The RMS of GPS SHU6 clock increase from 0.6ns to 1.0ns from the 1st to the 6th hour, but STD is stable at about 0.2ns.For the orbit in calculate session, the RMS of BDS SHU1 is a little less than that of SHU6,the RMS of GPS SHU1 and SHU6 orbit are approximately at the same level. In prediction session, the RMS of IGSO/MEO for BDS SHU1 is relative stable, but the RMS of SHU6 1st-6th hour prediction session increase from about 26.5cm to 32.7cm. The RMS of GPS SHU1 orbit's prediction session is about 3.4cm,but which increase from 3.3cm to 4.3cm for GPS SHU6 1st-6th hour prediction session.The comparison of GLONASS and GALILEO orbit and clock also will be described.The results show that the hourly update is more important for BDS at this stage.Moreover,some problems appearing in satellites and stations can be found earlier by 1 hourly updated frequency.

15 years of zebrafish chemical screening

PubMed Central

Rennekamp, Andrew J.; Peterson, Randall T.

2015-01-01

In 2000, the first chemical screen using living zebrafish in a multi-well plate was reported. Since then, more than 60 additional screens have been published describing whole-organism drug and pathway discovery projects in zebrafish. To investigate the scope of the work reported in the last 14 years and to identify trends in the field, we analyzed the discovery strategies of 64 primary research articles from the literature. We found that zebrafish screens have expanded beyond the use of developmental phenotypes to include behavioral, cardiac, metabolic, proliferative and regenerative endpoints. Additionally, many creative strategies have been used to uncover the mechanisms of action of new small molecules including chemical phenocopy, genetic phenocopy, mutant rescue, and spatial localization strategies. PMID:25461724

A clocking discipline for two-phase digital integrated circuits

NASA Astrophysics Data System (ADS)

Noice, D. C.

1983-09-01

Sooner or later a designer of digital circuits must face the problem of timing verification so he can avoid errors caused by clock skew, critical races, and hazards. Unlike previous verification methods, such as timing simulation and timing analysis, the approach presented here guarantees correct operation despite uncertainty about delays in the circuit. The result is a clocking discipline that deals with timing abstractions only. It is not based on delay calculations; it is only concerned with the correct, synchronous operation at some clock rate. Accordingly, it may be used earlier in the design cycle, which is particularly important to integrated circuit designs. The clocking discipline consists of a notation of clocking types, and composition rules for using the types. Together, the notation and rules define a formal theory of two phase clocking. The notation defines the names and exact characteristics for different signals that are used in a two phase digital system. The notation makes it possible to develop rules for propagating the clocking types through particular circuits.

Comparisons of mental clocks.

PubMed

Paivio, A

1978-02-01

Subjects in three experiments were presented with pairs of clock times and were required to choose the one in which the hour and minute hand formed the smaller angle. In Experiments 1 and 2, the times were presented digitally, necessitating a transformation into symbolic representations from which the angular size difference could be inferred. The results revealed orderly symbolic distance effects so that comparison reaction time increased as the angular size difference decreased. Moreover, subjects generally reported using imagery to make the judgment, and subjects scoring high on test of imagery ability were faster than those scoring low on such tests. Experiment 3 added a direct perceptual condition in which subjects compared angles between pairs of hands on two drawn (analog) clocks, as well as a mixed condition involving one digital and one analog clock time. The results showed comparable distance effects for all conditions. In addition, reaction time increased from the perceptual, to the mixed, to the pure-digital condition. These results are consistent with predictions from an image-based dual-coding theory.

Differences in Acute Alcohol-Induced Behavioral Responses Among Zebrafish Populations

PubMed Central

Gerlai, Robert; Ahmad, Fahad; Prajapati, Sonal

2009-01-01

Background With the arsenal of genetic tools available for zebrafish, this species has been successfully used to investigate the genetic aspects of human diseases from developmental disorders to cancer. Interest in the behavior and brain function of zebrafish is also increasing as CNS disorders may be modeled and studied with this species. Alcoholism and alcohol abuse are among the most devastating and costliest diseases. However, the mechanisms of these diseases are not fully understood. Zebrafish has been proposed as a model organism to study such mechanisms. Characterization of alcohol’s effects on zebrafish is a necessary step in this research. Methods Here, we compare the effects of acute alcohol (EtOH) administration on the behavior of zebrafish from 4 distinct laboratory-bred populations using automated as well as observation based behavioral quantification methods. Results Alcohol treatment resulted in significant dose-dependent behavioral changes but the dose–response trajectories differed among zebrafish populations. Conclusions The results demonstrate for the first time a genetic component in alcohol responses in adult zebrafish and also show the feasibility of high throughput behavioral screening. We discuss the exploration and exploitation of the genetic differences found. PMID:18652595

The Effect of Chronic Arsenic Exposure in Zebrafish

PubMed Central

Hallauer, Janell; Geng, Xiangrong; Yang, Hung-Chi; Shen, Jian; Tsai, Kan-Jen

2016-01-01

Abstract Arsenic is a prevalent environmental toxin and a Group one human carcinogenic agent. Chronic arsenic exposure has been associated with many human diseases. The aim of this study is to evaluate zebrafish as an animal model to assess arsenic toxicity in elevated long-term arsenic exposure. With prolonged exposure (6 months) to various concentrations of arsenic from 50 ppb to 300 ppb, effects of arsenic accumulation in zebrafish tissues, and phenotypes were investigated. Results showed that there are no significant changes of arsenic retention in zebrafish tissues, and zebrafish did not exhibit any visible tumor formation under arsenic exposure conditions. However, the zebrafish demonstrate a dysfunction in their neurological system, which is reflected by a reduction of locomotive activity. Moreover, elevated levels of the superoxide dismutase (SOD2) protein were detected in the eye and liver, suggesting increased oxidative stress. In addition, the progenies of arsenic-treated parents displayed a smaller biomass (four-fold reduction in body weight) compared with those from their parental controls. This result indicates that arsenic may induce genetic or epigenetic changes that are then passed on to the next generation. Overall, this study demonstrates that zebrafish is a convenient vertebrate model with advantages in the evaluation of arsenic-associated neurological disorders as well as its influences on the offspring. PMID:27140519

Zebrafish sex: a complicated affair

PubMed Central

Liew, Woei Chang

2014-01-01

In this review, we provide a detailed overview of studies on the elusive sex determination (SD) and gonad differentiation mechanisms of zebrafish (Danio rerio). We show that the data obtained from most studies are compatible with polygenic sex determination (PSD), where the decision is made by the allelic combinations of several loci. These loci are typically dispersed throughout the genome, but in some teleost species a few of them might be located on a preferential pair of (sex) chromosomes. The PSD system has a much higher level of variation of SD genotypes both at the level of gametes and the sexual genotype of individuals, than that of the chromosomal sex determination systems. The early sexual development of zebrafish males is a complicated process, as they first develop a ‘juvenile ovary’, that later undergoes a transformation to give way to a testis. To date, three major developmental pathways were shown to be involved with gonad differentiation through the modulation of programmed cell death. In our opinion, there are more pathways participating in the regulation of zebrafish gonad differentiation/transformation. Introduction of additional powerful large-scale genomic approaches into the analysis of zebrafish reproduction will result in further deepening of our knowledge as well as identification of additional pathways and genes associated with these processes in the near future. PMID:24148942

The Circadian Clock in Cancer Development and Therapy

PubMed Central

Fu, Loning; Kettner, Nicole M.

2014-01-01

Most aspects of mammalian function display circadian rhythms driven by an endogenous clock. The circadian clock is operated by genes and comprises a central clock in the brain that responds to environmental cues and controls subordinate clocks in peripheral tissues via circadian output pathways. The central and peripheral clocks coordinately generate rhythmic gene expression in a tissue-specific manner in vivo to couple diverse physiological and behavioral processes to periodic changes in the environment. However, as the world industrialized, activities that disrupt endogenous homeostasis with external circadian cues have increased. This change in lifestyle has been linked to increased risk of diseases in all aspects of human health, including cancer. Studies in humans and animal models have revealed that cancer development in vivo is closely associated with the loss of circadian homeostasis in energy balance, immune function and aging that are supported by cellular functions important for tumor suppression including cell proliferation, senescence, metabolism and DNA damage response. The clock controls these cellular functions both locally in cells of peripheral tissues and at the organismal level via extracellular signaling. Thus, the hierarchical mammalian circadian clock provides a unique system to study carcinogenesis as a deregulated physiological process in vivo. The asynchrony between host and malignant tissues in cell proliferation and metabolism also provides new and exciting options for novel anti-cancer therapies. PMID:23899600

Zebrafish Models for Human Acute Organophosphorus Poisoning.

PubMed

Faria, Melissa; Garcia-Reyero, Natàlia; Padrós, Francesc; Babin, Patrick J; Sebastián, David; Cachot, Jérôme; Prats, Eva; Arick Ii, Mark; Rial, Eduardo; Knoll-Gellida, Anja; Mathieu, Guilaine; Le Bihanic, Florane; Escalon, B Lynn; Zorzano, Antonio; Soares, Amadeu M V M; Raldúa, Demetrio

2015-10-22

Terrorist use of organophosphorus-based nerve agents and toxic industrial chemicals against civilian populations constitutes a real threat, as demonstrated by the terrorist attacks in Japan in the 1990 s or, even more recently, in the Syrian civil war. Thus, development of more effective countermeasures against acute organophosphorus poisoning is urgently needed. Here, we have generated and validated zebrafish models for mild, moderate and severe acute organophosphorus poisoning by exposing zebrafish larvae to different concentrations of the prototypic organophosphorus compound chlorpyrifos-oxon. Our results show that zebrafish models mimic most of the pathophysiological mechanisms behind this toxidrome in humans, including acetylcholinesterase inhibition, N-methyl-D-aspartate receptor activation, and calcium dysregulation as well as inflammatory and immune responses. The suitability of the zebrafish larvae to in vivo high-throughput screenings of small molecule libraries makes these models a valuable tool for identifying new drugs for multifunctional drug therapy against acute organophosphorus poisoning.

Live imaging of apoptotic cells in zebrafish

PubMed Central

van Ham, Tjakko J.; Mapes, James; Kokel, David; Peterson, Randall T.

2010-01-01

Many debilitating diseases, including neurodegenerative diseases, involve apoptosis. Several methods have been developed for visualizing apoptotic cells in vitro or in fixed tissues, but few tools are available for visualizing apoptotic cells in live animals. Here we describe a genetically encoded fluorescent reporter protein that labels apoptotic cells in live zebrafish embryos. During apoptosis, the phospholipid phosphatidylserine (PS) is exposed on the outer leaflet of the plasma membrane. The calcium-dependent protein Annexin V (A5) binds PS with high affinity, and biochemically purified, fluorescently labeled A5 probes have been widely used to detect apoptosis in vitro. Here we show that secreted A5 fused to yellow fluorescent protein specifically labels apoptotic cells in living zebrafish. We use this fluorescent probe to characterize patterns of apoptosis in living zebrafish larvae and to visualize neuronal cell death at single-cell resolution in vivo.—Van Ham, T. J., Mapes, J., Kokel, D., Peterson, R. T. Live imaging of apoptotic cells in zebrafish. PMID:20601526

Method and system for downhole clock synchronization

DOEpatents

Hall, David R.; Bartholomew, David B.; Johnson, Monte; Moon, Justin; Koehler, Roger O.

2006-11-28

A method and system for use in synchronizing at least two clocks in a downhole network are disclosed. The method comprises determining a total signal latency between a controlling processing element and at least one downhole processing element in a downhole network and sending a synchronizing time over the downhole network to the at least one downhole processing element adjusted for the signal latency. Electronic time stamps may be used to measure latency between processing elements. A system for electrically synchronizing at least two clocks connected to a downhole network comprises a controlling processing element connected to a synchronizing clock in communication over a downhole network with at least one downhole processing element comprising at least one downhole clock. Preferably, the downhole network is integrated into a downhole tool string.

Control of her1 expression during zebrafish somitogenesis by a Delta-dependent oscillator and an independent wave-front activity

PubMed Central

Holley, Scott A.; Geisler, Robert; Nüsslein-Volhard, Christiane

2000-01-01

Somitogenesis has been linked both to a molecular clock that controls the oscillation of gene expression in the presomitic mesoderm (PSM) and to Notch pathway signaling. The oscillator, or clock, is thought to create a prepattern of stripes of gene expression that regulates the activity of the Notch pathway that subsequently directs somite border formation. Here, we report that the zebrafish gene after eight (aei) that is required for both somitogenesis and neurogenesis encodes the Notch ligand DeltaD. Additional analysis revealed that stripes of her1 expression oscillate within the PSM and that aei/DeltaD signaling is required for this oscillation. aei/DeltaD expression does not oscillate, indicating that the activity of the Notch pathway upstream of her1 may function within the oscillator itself. Moreover, we found that her1 stripes are expressed in the anlage of consecutive somites, indicating that its expression pattern is not pair-rule. Analysis of her1 expression in aei/DeltaD, fused somites (fss), and aei;fss embryos uncovered a wave-front activity that is capable of continually inducing her1 expression de novo in the anterior PSM in the absence of the oscillation of her1. The wave-front activity, in reference to the clock and wave-front model, is defined as such because it interacts with the oscillator-derived pattern in the anterior PSM and is required for somite morphogenesis. This wave-front activity is blocked in embryos mutant for fss but not aei/DeltaD. Thus, our analysis indicates that the smooth sequence of formation, refinement, and fading of her1 stripes in the PSM is governed by two separate activities. PMID:10887161

HCV IRES-Mediated Core Expression in Zebrafish

PubMed Central

Zhang, Jing-Pu; Hu, Zhan-Ying; Tong, Jun-Wei; Ding, Cun-Bao; Peng, Zong-Gen; Zhao, Li-Xun; Song, Dan-Qing; Jiang, Jian-Dong

2013-01-01

The lack of small animal models for hepatitis C virus has impeded the discovery and development of anti-HCV drugs. HCV-IRES plays an important role in HCV gene expression, and is an attractive target for antiviral therapy. In this study, we report a zebrafish model with a biscistron expression construct that can co-transcribe GFP and HCV-core genes by human hepatic lipase promoter and zebrafish liver fatty acid binding protein enhancer. HCV core translation was designed mediated by HCV-IRES sequence and gfp was by a canonical cap-dependent mechanism. Results of fluorescence image and in situ hybridization indicate that expression of HCV core and GFP is liver-specific; RT-PCR and Western blotting show that both core and gfp expression are elevated in a time-dependent manner for both transcription and translation. It means that the HCV-IRES exerted its role in this zebrafish model. Furthermore, the liver-pathological impact associated with HCV-infection was detected by examination of gene markers and some of them were elevated, such as adiponectin receptor, heparanase, TGF-β, PDGF-α, etc. The model was used to evaluate three clinical drugs, ribavirin, IFNα-2b and vitamin B12. The results show that vitamin B12 inhibited core expression in mRNA and protein levels in dose-dependent manner, but failed to impact gfp expression. Also VB12 down-regulated some gene transcriptions involved in fat liver, liver fibrosis and HCV-associated pathological process in the larvae. It reveals that HCV-IRES responds to vitamin B12 sensitively in the zebrafish model. Ribavirin did not disturb core expression, hinting that HCV-IRES is not a target site of ribavirin. IFNα-2b was not active, which maybe resulted from its degradation in vivo for the long time. These findings demonstrate the feasibility of the zebrafish model for screening of anti-HCV drugs targeting to HCV-IRES. The zebrafish system provides a novel evidence of using zebrafish as a HCV model organism. PMID:23469178

Analysis of Lethality and Malformations During Zebrafish (Danio rerio) Development.

PubMed

Raghunath, Azhwar; Perumal, Ekambaram

2018-01-01

The versatility offered by zebrafish (Danio rerio) makes it a powerful and an attractive vertebrate model in developmental toxicity and teratogenicity assays. Apart from the newly introduced chemicals as drugs, xenobiotics also induce abnormal developmental abnormalities and congenital malformations in living organisms. Over the recent decades, zebrafish embryo/larva has emerged as a potential tool to test teratogenicity potential of these chemicals. Zebrafish responds to compounds as mammals do as they share similarities in their development, metabolism, physiology, and signaling pathways with that of mammals. The methodology used by the different scientists varies enormously in the zebrafish embryotoxicity test. In this chapter, we present methods to assess lethality and malformations during zebrafish development. We propose two major malformations scoring systems: binomial and relative morphological scoring systems to assess the malformations in zebrafish embryos/larvae. Based on the scoring of the malformations, the test compound can be classified as a teratogen or a nonteratogen and its teratogenic potential is evaluated.

Conserved gene regulation during acute inflammation between zebrafish and mammals

PubMed Central

Forn-Cuní, G.; Varela, M.; Pereiro, P.; Novoa, B.; Figueras, A.

2017-01-01

Zebrafish (Danio rerio), largely used as a model for studying developmental processes, has also emerged as a valuable system for modelling human inflammatory diseases. However, in a context where even mice have been questioned as a valid model for these analysis, a systematic study evaluating the reproducibility of human and mammalian inflammatory diseases in zebrafish is still lacking. In this report, we characterize the transcriptomic regulation to lipopolysaccharide in adult zebrafish kidney, liver, and muscle tissues using microarrays and demonstrate how the zebrafish genomic responses can effectively reproduce the mammalian inflammatory process induced by acute endotoxin stress. We provide evidence that immune signaling pathways and single gene expression is well conserved throughout evolution and that the zebrafish and mammal acute genomic responses after lipopolysaccharide stimulation are highly correlated despite the differential susceptibility between species to that compound. Therefore, we formally confirm that zebrafish inflammatory models are suited to study the basic mechanisms of inflammation in human inflammatory diseases, with great translational impact potential. PMID:28157230

Electromagnetic synchronisation of clocks with finite separation in a rotating system

NASA Astrophysics Data System (ADS)

Cohen, J. M.; Moses, H. E.; Rosenblum, A.

1984-11-01

For clocks on the vertices of a triangle, it is shown that clock synchronisation using electromagnetic signals between finitely spaced clocks in a rotating frame leads to the same synchronization error as a closely spaced band of clocks along the same light path. In addition, the above result is generalized to n equally spaced clocks.

Zebrafish as a model system to study toxicology.

PubMed

Dai, Yu-Jie; Jia, Yong-Fang; Chen, Na; Bian, Wan-Ping; Li, Qin-Kai; Ma, Yan-Bo; Chen, Yan-Ling; Pei, De-Sheng

2014-01-01

Monitoring and assessing the effects of contaminants in the aquatic eco-environment is critical in protecting human health and the environment. The zebrafish has been widely used as a prominent model organism in different fields because of its small size, low cost, diverse adaptability, short breeding cycle, high fecundity, and transparent embryos. Recent studies have demonstrated that zebrafish sensitivity can aid in monitoring environmental contaminants, especially with the application of transgenic technology in this area. The present review provides a brief overview of recent studies on wild-type and transgenic zebrafish as a model system to monitor toxic heavy metals, endocrine disruptors, and organic pollutants for toxicology. The authors address the new direction of developing high-throughput detection of genetically modified transparent zebrafish to open a new window for monitoring environmental pollutants. © 2013 SETAC.

Vane clocking effects in an embedded compressor stage

NASA Astrophysics Data System (ADS)

Key, Nicole Leanne

The objective of this research was to experimentally investigate the effects of vane clocking, the circumferential indexing of adjacent vane rows with similar vane counts, in an embedded compressor stage. Experiments were performed in the Purdue 3-Stage Compressor, which consists of an IGV followed by three stages. The IGV, Stator 1, and Stator 2 have identical vane counts of 44, and the effects of clocking were studied on Stage 2. The clocking configuration that located the upstream vane wake on the Stator 2 leading edge was identified with total pressure measurements at the inlet to Stator 2 and confirmed with measurements at the exit of Stator 2. For both loading conditions, the total temperature results showed that there was no measurable change associated with vane clocking in the amount of work done on the flow. At design loading, the change in stage efficiency with vane clocking was 0.27 points between the maximum and minimum efficiency clocking configurations. The maximum efficiency configuration was the case where the Stator 1 wake impinged on the Stator 2 leading edge. This condition produced a shallower and thinner Stator 2 wake compared to the clocking configuration that located the wake in the middle of the Stator 2 passage. By locating the Stator 1 wake at the leading edge, it dampened the Stator 2 boundary layer response to inlet fluctuations associated with the Rotor 2 wakes. At high loading, the change in Stage 2 efficiency increased to 1.07 points; however, the maximum efficiency clocking configuration was the case where the Stator 1 wake passed through the middle of the downstream vane passage. At high loading, the flow physics associated with vane clocking were different than at design loading because the location of the Stator 1 wake fluid on the Stator 2 leading edge triggered a boundary layer separation on the suction side of Stator 2 producing a wider and deeper wake. Vane clocking essentially affects the amount of interaction between the

Fast segmentation of stained nuclei in terabyte-scale, time resolved 3D microscopy image stacks.

PubMed

Stegmaier, Johannes; Otte, Jens C; Kobitski, Andrei; Bartschat, Andreas; Garcia, Ariel; Nienhaus, G Ulrich; Strähle, Uwe; Mikut, Ralf

2014-01-01

Automated analysis of multi-dimensional microscopy images has become an integral part of modern research in life science. Most available algorithms that provide sufficient segmentation quality, however, are infeasible for a large amount of data due to their high complexity. In this contribution we present a fast parallelized segmentation method that is especially suited for the extraction of stained nuclei from microscopy images, e.g., of developing zebrafish embryos. The idea is to transform the input image based on gradient and normal directions in the proximity of detected seed points such that it can be handled by straightforward global thresholding like Otsu's method. We evaluate the quality of the obtained segmentation results on a set of real and simulated benchmark images in 2D and 3D and show the algorithm's superior performance compared to other state-of-the-art algorithms. We achieve an up to ten-fold decrease in processing times, allowing us to process large data sets while still providing reasonable segmentation results.

Silver nanoparticles induce endoplasmatic reticulum stress response in zebrafish

DOE Office of Scientific and Technical Information (OSTI.GOV)

Christen, Verena; Capelle, Martinus; Fent, Karl, E-mail: karl.fent@fhnw.ch

2013-10-15

Silver nanoparticles (AgNPs) find increasing applications, and therefore humans and the environment are increasingly exposed to them. However, potential toxicological implications are not sufficiently known. Here we investigate effects of AgNPs (average size 120 nm) on zebrafish in vitro and in vivo, and compare them to human hepatoma cells (Huh7). AgNPs are incorporated in zebrafish liver cells (ZFL) and Huh7, and in zebrafish embryos. In ZFL cells AgNPs lead to induction of reactive oxygen species (ROS), endoplasmatic reticulum (ER) stress response, and TNF-α. Transcriptional alterations also occur in pro-apoptotic genes p53 and Bax. The transcriptional profile differed in ZFL andmore » Huh7 cells. In ZFL cells, the ER stress marker BiP is induced, concomitant with the ER stress marker ATF-6 and spliced XBP-1 after 6 h and 24 h exposure to 0.5 g/L and 0.05 g/L AgNPs, respectively. This indicates the induction of different pathways of the ER stress response. Moreover, AgNPs induce TNF-α. In zebrafish embryos exposed to 0.01, 0.1, 1 and 5 mg/L AgNPs hatching was affected and morphological defects occurred at high concentrations. ER stress related gene transcripts BiP and Synv are significantly up-regulated after 24 h at 0.1 and 5 mg/L AgNPs. Furthermore, transcriptional alterations occurred in the pro-apoptotic genes Noxa and p21. The ER stress response was strong in ZFL cells and occurred in zebrafish embryos as well. Our data demonstrate for the first time that AgNPs lead to induction of ER stress in zebrafish. The induction of ER stress can have several consequences including the activation of apoptotic and inflammatory pathways. - Highlights: • Effects of silver nanoparticles (120 nm AgNPs) are investigated in zebrafish. • AgNPs induce all ER stress reponses in vitro in zebrafish liver cells. • AgNPs induce weak ER stress in zebrafish embryos. • AgNPs induce oxidative stress and transcripts of pro-apoptosis genes.« less

New frontiers for zebrafish management.

PubMed

Lawrence, C

2016-01-01

The zebrafish (Danio rerio) is a preeminent model organism with a wide and expanding utility for numerous scientific disciplines. The same features that once endeared this small freshwater minnow to developmental biologists combined with its relatively high genetic similarity to mammals and the advent of new, more efficient methods for genome editing are now helping to spur expanded growth in its usage in various fields, including toxicology, drug discovery, transplant biology, disease modeling, and even aquaculture. Continued maturation and adoption of the zebrafish model system in these and other fields of science will require that methods and approaches for husbandry and management of these fish in controlled settings be refined and improved to the extent that, ultimately, zebrafish research becomes more reproducible, defined, cost-effective, and accessible to the masses. Knowledge and technology transfer from laboratory animal science and commercial aquaculture will be a necessary part of this development. Copyright © 2016 Elsevier Inc. All rights reserved.

Mayo Clinic Zebrafish Facility Overview.

PubMed

Leveque, Ryan E; Clark, Karl J; Ekker, Stephen C

2016-07-01

The zebrafish (Danio rerio) is a premier nonmammalian vertebrate model organism. This small aquatic fish is utilized in multiple disciplines in the Mayo Clinic community and by many laboratories around the world because of its biological similarity to humans, its advanced molecular genetics, the elucidation of its genome sequence, and the ever-expanding and outstanding new biological tools now available to the zebrafish researcher. The Mayo Clinic Zebrafish Facility (MCZF) houses ∼2,000 tanks annotated using an in-house, Internet cloud-based bar-coding system tied to our established zfishbook.org web infrastructure. Paramecia are the primary food source for larval fish rearing, using a simplified culture protocol described herein. The MCZF supports the specific ongoing research in a variety of laboratories, while also serving as a local hub for new scientists as they learn to tap into the potential of this model system for understanding normal development, disease, and as models of health.

Synthesizing genetic sequential logic circuit with clock pulse generator

PubMed Central

2014-01-01

Background Rhythmic clock widely occurs in biological systems which controls several aspects of cell physiology. For the different cell types, it is supplied with various rhythmic frequencies. How to synthesize a specific clock signal is a preliminary but a necessary step to further development of a biological computer in the future. Results This paper presents a genetic sequential logic circuit with a clock pulse generator based on a synthesized genetic oscillator, which generates a consecutive clock signal whose frequency is an inverse integer multiple to that of the genetic oscillator. An analogous electronic waveform-shaping circuit is constructed by a series of genetic buffers to shape logic high/low levels of an oscillation input in a basic sinusoidal cycle and generate a pulse-width-modulated (PWM) output with various duty cycles. By controlling the threshold level of the genetic buffer, a genetic clock pulse signal with its frequency consistent to the genetic oscillator is synthesized. A synchronous genetic counter circuit based on the topology of the digital sequential logic circuit is triggered by the clock pulse to synthesize the clock signal with an inverse multiple frequency to the genetic oscillator. The function acts like a frequency divider in electronic circuits which plays a key role in the sequential logic circuit with specific operational frequency. Conclusions A cascaded genetic logic circuit generating clock pulse signals is proposed. Based on analogous implement of digital sequential logic circuits, genetic sequential logic circuits can be constructed by the proposed approach to generate various clock signals from an oscillation signal. PMID:24884665

Synthesizing genetic sequential logic circuit with clock pulse generator.

PubMed

Chuang, Chia-Hua; Lin, Chun-Liang

2014-05-28

Rhythmic clock widely occurs in biological systems which controls several aspects of cell physiology. For the different cell types, it is supplied with various rhythmic frequencies. How to synthesize a specific clock signal is a preliminary but a necessary step to further development of a biological computer in the future. This paper presents a genetic sequential logic circuit with a clock pulse generator based on a synthesized genetic oscillator, which generates a consecutive clock signal whose frequency is an inverse integer multiple to that of the genetic oscillator. An analogous electronic waveform-shaping circuit is constructed by a series of genetic buffers to shape logic high/low levels of an oscillation input in a basic sinusoidal cycle and generate a pulse-width-modulated (PWM) output with various duty cycles. By controlling the threshold level of the genetic buffer, a genetic clock pulse signal with its frequency consistent to the genetic oscillator is synthesized. A synchronous genetic counter circuit based on the topology of the digital sequential logic circuit is triggered by the clock pulse to synthesize the clock signal with an inverse multiple frequency to the genetic oscillator. The function acts like a frequency divider in electronic circuits which plays a key role in the sequential logic circuit with specific operational frequency. A cascaded genetic logic circuit generating clock pulse signals is proposed. Based on analogous implement of digital sequential logic circuits, genetic sequential logic circuits can be constructed by the proposed approach to generate various clock signals from an oscillation signal.

Oxyntomodulin regulates resetting of the liver circadian clock by food

PubMed Central

Landgraf, Dominic; Tsang, Anthony H; Leliavski, Alexei; Koch, Christiane E; Barclay, Johanna L; Drucker, Daniel J; Oster, Henrik

2015-01-01

Circadian clocks coordinate 24-hr rhythms of behavior and physiology. In mammals, a master clock residing in the suprachiasmatic nucleus (SCN) is reset by the light–dark cycle, while timed food intake is a potent synchronizer of peripheral clocks such as the liver. Alterations in food intake rhythms can uncouple peripheral clocks from the SCN, resulting in internal desynchrony, which promotes obesity and metabolic disorders. Pancreas-derived hormones such as insulin and glucagon have been implicated in signaling mealtime to peripheral clocks. In this study, we identify a novel, more direct pathway of food-driven liver clock resetting involving oxyntomodulin (OXM). In mice, food intake stimulates OXM secretion from the gut, which resets liver transcription rhythms via induction of the core clock genes Per1 and 2. Inhibition of OXM signaling blocks food-mediated resetting of hepatocyte clocks. These data reveal a direct link between gastric filling with food and circadian rhythm phasing in metabolic tissues. DOI: http://dx.doi.org/10.7554/eLife.06253.001 PMID:25821984

An assay for lateral line regeneration in adult zebrafish.

PubMed

Pisano, Gina C; Mason, Samantha M; Dhliwayo, Nyembezi; Intine, Robert V; Sarras, Michael P

2014-04-08

Due to the clinical importance of hearing and balance disorders in man, model organisms such as the zebrafish have been used to study lateral line development and regeneration. The zebrafish is particularly attractive for such studies because of its rapid development time and its high regenerative capacity. To date, zebrafish studies of lateral line regeneration have mainly utilized fish of the embryonic and larval stages because of the lower number of neuromasts at these stages. This has made quantitative analysis of lateral line regeneration/and or development easier in the earlier developmental stages. Because many zebrafish models of neurological and non-neurological diseases are studied in the adult fish and not in the embryo/larvae, we focused on developing a quantitative lateral line regenerative assay in adult zebrafish so that an assay was available that could be applied to current adult zebrafish disease models. Building on previous studies by Van Trump et al. that described procedures for ablation of hair cells in adult Mexican blind cave fish and zebrafish (Danio rerio), our assay was designed to allow quantitative comparison between control and experimental groups. This was accomplished by developing a regenerative neuromast standard curve based on the percent of neuromast reappearance over a 24 hr time period following gentamicin-induced necrosis of hair cells in a defined region of the lateral line. The assay was also designed to allow extension of the analysis to the individual hair cell level when a higher level of resolution is required.

The circadian clock of Neurospora crassa.

PubMed

Baker, Christopher L; Loros, Jennifer J; Dunlap, Jay C

2012-01-01

Circadian clocks organize our inner physiology with respect to the external world, providing life with the ability to anticipate and thereby better prepare for major fluctuations in its environment. Circadian systems are widely represented in nearly all major branches of life, except archaebacteria, and within the eukaryotes, the filamentous fungus Neurospora crassa has served for nearly half a century as a durable model organism for uncovering the basic circadian physiology and molecular biology. Studies using Neurospora have clarified our fundamental understanding of the clock as nested positive and negative feedback loops regulated through transcriptional and post-transcriptional processes. These feedback loops are centered on a limited number of proteins that form molecular complexes, and their regulation provides a physical explanation for nearly all clock properties. This review will introduce the basics of circadian rhythms, the model filamentous fungus N. crassa, and provide an overview of the molecular components and regulation of the circadian clock. © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

The mammalian retina as a clock

NASA Technical Reports Server (NTRS)

Tosini, Gianluca; Fukuhara, Chiaki

2002-01-01

Many physiological, cellular, and biochemical parameters in the retina of vertebrates show daily rhythms that, in many cases, also persist under constant conditions. This demonstrates that they are driven by a circadian pacemaker. The presence of an autonomous circadian clock in the retina of vertebrates was first demonstrated in Xenopus laevis and then, several years later, in mammals. In X. laevis and in chicken, the retinal circadian pacemaker has been localized in the photoreceptor layer, whereas in mammals, such information is not yet available. Recent advances in molecular techniques have led to the identification of a group of genes that are believed to constitute the molecular core of the circadian clock. These genes are expressed in the retina, although with a slightly different 24-h profile from that observed in the central circadian pacemaker. This result suggests that some difference (at the molecular level) may exist between the retinal clock and the clock located in the suprachiasmatic nuclei of hypothalamus. The present review will focus on the current knowledge of the retinal rhythmicity and the mechanisms responsible for its control.

Biochemical basis for the biological clock

NASA Technical Reports Server (NTRS)

Morre, D. James; Chueh, Pin-Ju; Pletcher, Jake; Tang, Xiaoyu; Wu, Lian-Ying; Morre, Dorothy M.

2002-01-01

NADH oxidases at the external surface of plant and animal cells (ECTO-NOX proteins) exhibit stable and recurring patterns of oscillations with potentially clock-related, entrainable, and temperature-compensated period lengths of 24 min. To determine if ECTO-NOX proteins might represent the ultradian time keepers (pacemakers) of the biological clock, COS cells were transfected with cDNAs encoding tNOX proteins having a period length of 22 min or with C575A or C558A cysteine to alanine replacements having period lengths of 36 or 42 min. Here we demonstrate that such transfectants exhibited 22, 36, or 40 to 42 h circadian patterns in the activity of glyceraldehyde-3-phosphate dehydrogenase, a common clock-regulated protein, in addition to the endogenous 24 h circadian period length. The fact that the expression of a single oscillatory ECTO-NOX protein determines the period length of a circadian biochemical marker (60 X the ECTO-NOX period length) provides compelling evidence that ECTO-NOX proteins are the biochemical ultradian drivers of the cellular biological clock.

Clock Controller For Ac Self-Timing Analysis Of Logic System

DOEpatents

Lo, Tinchee; Flanagan, John D.

2004-05-18

A clock controller and clock generating method are provided for AC self-test timing analysis of a logic system. The controller includes latch circuitry which receives a DC input signal at a data input, and a pair of continuous out-of-phase clock signals at capture and launch clock inputs thereof. The latch circuitry outputs two overlapping pulses responsive to the DC input signal going high. The two overlapping pulses are provided to waveform shaper circuitry which produces therefrom two non-overlapping pulses at clock speed of the logic system to be tested. The two non-overlapping pulses are a single pair of clock pulses which facilitate AC self-test timing analysis of the logic system.

UPLC/MS MS data of testosterone metabolites in human and zebrafish liver microsomes and whole zebrafish larval microsomes.

PubMed

Saad, Moayad; Bijttebier, Sebastiaan; Matheeussen, An; Verbueken, Evy; Pype, Casper; Casteleyn, Christophe; Van Ginneken, Chris; Maes, Louis; Cos, Paul; Van Cruchten, Steven

2018-02-01

This article represents data regarding a study published in Toxicology in vitro entitled " in vitro CYP-mediated drug metabolism in the zebrafish (embryo) using human reference compounds" (Saad et al., 2017) [1]. Data were acquired with ultra-performance liquid chromatography - accurate mass mass spectrometry (UPLC-amMS). A full spectrum scan was conducted for the testosterone (TST) metabolites from the microsomal stability assay in zebrafish and humans. The microsomal proteins were extracted from adult zebrafish male (MLM) and female (FLM) livers, whole body homogenates of 96 h post fertilization larvae (EM) and a pool of human liver microsomes from 50 donors (HLM). Data are expressed as the abundance from the extracted ion chromatogram of the metabolites.

Transcriptional oscillation of canonical clock genes in mouse peripheral tissues

PubMed Central

Yamamoto, Takuro; Nakahata, Yasukazu; Soma, Haruhiko; Akashi, Makoto; Mamine, Takayoshi; Takumi, Toru

2004-01-01

Background The circadian rhythm of about 24 hours is a fundamental physiological function observed in almost all organisms from prokaryotes to humans. Identification of clock genes has allowed us to study the molecular bases for circadian behaviors and temporal physiological processes such as hormonal secretion, and has prompted the idea that molecular clocks reside not only in a central pacemaker, the suprachiasmatic nuclei (SCN) of hypothalamus in mammals, but also in peripheral tissues, even in immortalized cells. Furthermore, previous molecular dissection revealed that the mechanism of circadian oscillation at a molecular level is based on transcriptional regulation of clock and clock-controlled genes. Results We systematically analyzed the mRNA expression of clock and clock-controlled genes in mouse peripheral tissues. Eight genes (mBmal1, mNpas2, mRev-erbα, mDbp, mRev-erbβ, mPer3, mPer1 and mPer2; given in the temporal order of the rhythm peak) showed robust circadian expressions of mRNAs in all tissues except testis, suggesting that these genes are core molecules of the molecular biological clock. The bioinformatics analysis revealed that these genes have one or a combination of 3 transcriptional elements (RORE, DBPE, and E-box), which are conserved among human, mouse, and rat genome sequences, and indicated that these 3 elements may be responsible for the biological timing of expression of canonical clock genes. Conclusions The observation of oscillatory profiles of canonical clock genes is not only useful for physiological and pathological examination of the circadian clock in various organs but also important for systematic understanding of transcriptional regulation on a genome-wide basis. Our finding of the oscillatory expression of canonical clock genes with a temporal order provides us an interesting hypothesis, that cyclic timing of all clock and clock-controlled genes may be dependent on several transcriptional elements including 3 known elements

Fishing anti(lymph)angiogenic drugs with zebrafish.

PubMed

García-Caballero, Melissa; Quesada, Ana R; Medina, Miguel A; Marí-Beffa, Manuel

2018-02-01

Zebrafish, an amenable small teleost fish with a complex mammal-like circulatory system, is being increasingly used for drug screening and toxicity studies. It combines the biological complexity of in vivo models with a higher-throughput screening capability compared with other available animal models. Externally growing, transparent embryos, displaying well-defined blood and lymphatic vessels, allow the inexpensive, rapid, and automatable evaluation of drug candidates that are able to inhibit neovascularisation. Here, we briefly review zebrafish as a model for the screening of anti(lymph)angiogenic drugs, with emphasis on the advantages and limitations of the different zebrafish-based in vivo assays. Copyright © 2017 Elsevier Ltd. All rights reserved.

Optical Lattice Clocks with Weakly Bound Molecules.

PubMed

Borkowski, Mateusz

2018-02-23

Optical molecular clocks promise unparalleled sensitivity to the temporal variation of the electron-to-proton mass ratio and insight into possible new physics beyond the standard model. We propose to realize a molecular clock with bosonic ^{174}Yb_{2} molecules, where the forbidden ^{1}S_{0}→^{3}P_{0} clock transition would be induced magnetically. The use of a bosonic species avoids possible complications due to the hyperfine structure present in fermionic species. While direct clock line photoassociation would be challenging, weakly bound ground state molecules could be produced by stimulated Raman adiabatic passage and used instead. The recent scattering measurements [L. Franchi, et al. New J. Phys. 19, 103037 (2017)NJOPFM1367-263010.1088/1367-2630/aa8fb4] enable us to determine the positions of target ^{1}S_{0}+^{3}P_{0} vibrational levels and calculate the Franck-Condon factors for clock transitions between ground and excited molecular states. The resulting magnetically induced Rabi frequencies are similar to those for atoms hinting that an experimental realization is feasible. A successful observation could pave the way towards Hz-level molecular spectroscopy.

Using a transportable optical clock for chronometric levelling

NASA Astrophysics Data System (ADS)

Lisdat, Christian; Sterr, Uwe; Koller, Silvio; Grotti, Jacopo; Vogt, Stefan; Häfner, Sebastian; Herbers, Sofia; Al-Masoudi, Ali

2016-07-01

With their supreme accuracy and precision, optical clocks in combination with new methods of long-distance frequency transfer can be used to determine height differences by measuring the gravitational red shift between two clocks without accumulation of measurement errors, as in classical levelling. We are developing transportable optical clocks for this purpose that will also serve for the technology development regarding optical clocks in Space and for international comparisons between optical clocks that cannot be linked with sufficient accuracy otherwise. In this talk we will focus on the transportable strontium lattice clock that we are developing and its first evaluation. Presently, we achieve a fractional frequency instability of 3 × 10^{-17} after 1000 s averaging time, which is equivalent to a height resolution of 30 cm. The first uncertainty evaluation of the system yielded 7 × 10^{-17}. We expect rapid improvements to an uncertainty of a few parts in 10^{17}. The clock is now located within a car trailer, which requires compact and rugged lasers systems and physics package. Special care has been taken in the design of the ultra-frequency stable interrogation laser that has to achieve fractional frequency instabilities of considerably below 10^{-15}. Typical laboratory constructions of the reference resonator system used to pre-stabilize the laser frequency are not compatible with the requirement of transportability. In an actual levelling campaign, this clock will be connected via a stabilized optical fibre link with another, stationary frequency standard. The measured gravitational red shift will be compared with the ones calculated from potential differences derived with state of the art geodetic data and models. We will discuss the status of measurements of geodetic relevance with optical clocks and give an outlook on our next steps. This work is supported by QUEST, DFG (RTG 1729, CRC 1128), EU-FP7 (FACT) and EMRP (ITOC). The EMRP is jointly funded

Crosstalk of clock gene expression and autophagy in aging

PubMed Central

Kalfalah, Faiza; Janke, Linda; Schiavi, Alfonso; Tigges, Julia; Ix, Alexander; Ventura, Natascia; Boege, Fritz; Reinke, Hans

2016-01-01

Autophagy and the circadian clock counteract tissue degeneration and support longevity in many organisms. Accumulating evidence indicates that aging compromises both the circadian clock and autophagy but the mechanisms involved are unknown. Here we show that the expression levels of transcriptional repressor components of the circadian oscillator, most prominently the human Period homologue PER2, are strongly reduced in primary dermal fibroblasts from aged humans, while raising the expression of PER2 in the same cells partially restores diminished autophagy levels. The link between clock gene expression and autophagy is corroborated by the finding that the circadian clock drives cell-autonomous, rhythmic autophagy levels in immortalized murine fibroblasts, and that siRNA-mediated downregulation of PER2 decreases autophagy levels while leaving core clock oscillations intact. Moreover, the Period homologue lin-42 regulates autophagy and life span in the nematode Caenorhabditis elegans, suggesting an evolutionarily conserved role for Period proteins in autophagy control and aging. Taken together, this study identifies circadian clock proteins as set-point regulators of autophagy and puts forward a model, in which age-related changes of clock gene expression promote declining autophagy levels. PMID:27574892

Crosstalk of clock gene expression and autophagy in aging.

PubMed

Kalfalah, Faiza; Janke, Linda; Schiavi, Alfonso; Tigges, Julia; Ix, Alexander; Ventura, Natascia; Boege, Fritz; Reinke, Hans

2016-08-28

Autophagy and the circadian clock counteract tissue degeneration and support longevity in many organisms. Accumulating evidence indicates that aging compromises both the circadian clock and autophagy but the mechanisms involved are unknown. Here we show that the expression levels of transcriptional repressor components of the circadian oscillator, most prominently the human Period homologue PER2 , are strongly reduced in primary dermal fibroblasts from aged humans, while raising the expression of PER2 in the same cells partially restores diminished autophagy levels. The link between clock gene expression and autophagy is corroborated by the finding that the circadian clock drives cell-autonomous, rhythmic autophagy levels in immortalized murine fibroblasts, and that siRNA-mediated downregulation of PER2 decreases autophagy levels while leaving core clock oscillations intact. Moreover, the Period homologue lin-42 regulates autophagy and life span in the nematode Caenorhabditis elegans , suggesting an evolutionarily conserved role for Period proteins in autophagy control and aging. Taken together, this study identifies circadian clock proteins as set-point regulators of autophagy and puts forward a model, in which age-related changes of clock gene expression promote declining autophagy levels.

Central and peripheral clocks are coupled by a neuropeptide pathway in Drosophila

PubMed Central

Selcho, Mareike; Millán, Carola; Palacios-Muñoz, Angelina; Ruf, Franziska; Ubillo, Lilian; Chen, Jiangtian; Bergmann, Gregor; Ito, Chihiro; Silva, Valeria; Wegener, Christian; Ewer, John

2017-01-01

Animal circadian clocks consist of central and peripheral pacemakers, which are coordinated to produce daily rhythms in physiology and behaviour. Despite its importance for optimal performance and health, the mechanism of clock coordination is poorly understood. Here we dissect the pathway through which the circadian clock of Drosophila imposes daily rhythmicity to the pattern of adult emergence. Rhythmicity depends on the coupling between the brain clock and a peripheral clock in the prothoracic gland (PG), which produces the steroid hormone, ecdysone. Time information from the central clock is transmitted via the neuropeptide, sNPF, to non-clock neurons that produce the neuropeptide, PTTH. These secretory neurons then forward time information to the PG clock. We also show that the central clock exerts a dominant role on the peripheral clock. This use of two coupled clocks could serve as a paradigm to understand how daily steroid hormone rhythms are generated in animals. PMID:28555616

A hydrogen maser clock for space - Clocks in future possible and improbable applications

NASA Astrophysics Data System (ADS)

Vessot, Robert F. C.

The development of atomic-H maser clocks for space applications since 1967 is reviewed, with a focus on the 39-kg instrument built for a rocket-flight test of gravitational redshift in 1976. The stability of the oscillator and the instability of earth-space propagation in that test are described, and techniques for overcoming the latter effects are considered. More recent maser clocks employ an H sorption manifold rather than heavy ion pumps; their application to precise satellite position determination for space-based VLBI astronomy is discussed in detail. Extensive diagrams, drawings, and photographs are provided.

High-precision multi-node clock network distribution.

PubMed

Chen, Xing; Cui, Yifan; Lu, Xing; Ci, Cheng; Zhang, Xuesong; Liu, Bo; Wu, Hong; Tang, Tingsong; Shi, Kebin; Zhang, Zhigang

2017-10-01

A high precision multi-node clock network for multiple users was built following the precise frequency transmission and time synchronization of 120 km fiber. The network topology adopts a simple star-shaped network structure. The clock signal of a hydrogen maser (synchronized with UTC) was recovered from a 120 km telecommunication fiber link and then was distributed to 4 sub-stations. The fractional frequency instability of all substations is in the level of 10 -15 in a second and the clock offset instability is in sub-ps in root-mean-square average.

Stochastic models for atomic clocks

NASA Technical Reports Server (NTRS)

Barnes, J. A.; Jones, R. H.; Tryon, P. V.; Allan, D. W.

1983-01-01

For the atomic clocks used in the National Bureau of Standards Time Scales, an adequate model is the superposition of white FM, random walk FM, and linear frequency drift for times longer than about one minute. The model was tested on several clocks using maximum likelihood techniques for parameter estimation and the residuals were acceptably random. Conventional diagnostics indicate that additional model elements contribute no significant improvement to the model even at the expense of the added model complexity.

Natural Variation of the Circadian Clock in Neurospora.

PubMed

Koritala, Bala S C; Lee, Kwangwon

2017-01-01

Most living organisms on earth experience daily and expected changes from the rotation of the earth. For an organism, the ability to predict and prepare for incoming stresses or resources is a very important skill for survival. This cellular process of measuring daily time of the day is collectively called the circadian clock. Because of its fundamental role in survival in nature, there is a great interest in studying the natural variation of the circadian clock. However, characterizing the genetic and molecular mechanisms underlying natural variation of circadian clocks remains a challenging task. In this chapter, we will summarize the progress in studying natural variation of the circadian clock in the successful eukaryotic model Neurospora, which led to discovering many design principles of the molecular mechanisms of the eukaryotic circadian clock. Despite the success of the system in revealing the molecular mechanisms of the circadian clock, Neurospora has not been utilized to extensively study natural variation. We will review the challenges that hindered the natural variation studies in Neurospora, and how they were overcome. We will also review the advantages of Neurospora for natural variation studies. Since Neurospora is the model fungal species for circadian study, it represents over 5 million species of fungi on earth. These fungi play important roles in ecosystems on earth, and as such Neurospora could serve as an important model for understanding the ecological role of natural variation in fungal circadian clocks. © 2017 Elsevier Inc. All rights reserved.

In vivo loss of function study reveals the short stature homeobox-containing (shox) gene plays indispensable roles in early embryonic growth and bone formation in zebrafish.

PubMed

Sawada, Rie; Kamei, Hiroyasu; Hakuno, Fumihiko; Takahashi, Shin-Ichiro; Shimizu, Toshiaki

2015-02-01

Congenital loss of the SHOX gene is considered to be a genetic cause of short stature phenotype in Turner syndrome and Leri-Weill dyschondrosteosis patients. Though SHOX expression initiates during early fetal development, little is known about the embryonic roles of SHOX. The evolutionary conservation of the zebrafish shox gene and the convenience of the early developmental stages for analyses make zebrafish a preferred model. Here, we characterized structure, expression, and developmental roles of zebrafish shox through a loss-of-function approach. We found a previously undiscovered Shox protein that has both a homeodomain and an OAR-domain in zebrafish. The shox transcript emerged during the segmentation period and it increased in later stages. The predominant domains of shox expression were mandibular arch, pectoral fin, anterior notochord, rhombencephalon, and mesencephalon, suggesting that Shox is involved in bone and neural development. Translational blockade of Shox mRNA by an antisense morpholino oligo delayed embryonic growth, which was restored by the co-overexpression of morpholino-resistant Shox mRNA. At later stages, impaired Shox expression markedly delayed the calcification process in the anterior vertebral column and craniofacial bones. Our data demonstrate evolutionarily conserved Shox plays roles in early embryonic growth and in later bone formation. © 2014 Wiley Periodicals, Inc.

Effectiveness of recommended euthanasia methods in larval zebrafish (Danio rerio).

PubMed

Strykowski, Jennifer L; Schech, Joseph M

2015-01-01

The popularity of zebrafish and its use as a model organism in biomedical research including genetics, development, and toxicology, has increased over the past 20 y and continues to grow. However, guidelines for euthanasia remain vague, and the responsibility of creating appropriate euthanasia protocols essentially falls on individual facilities. To reduce variation in experimental results among labs, a standard method of euthanasia for zebrafish would be useful. Although various euthanasia methods have been compared, few studies focus on the effectiveness of euthanasia methods for larval zebrafish. In this study, we exposed larval zebrafish to each of 3 euthanasia agents (MS222, eugenol, and hypothermic shock) and assessed the recovery rate. Hypothermic shock appeared to be the most effective method for euthanizing zebrafish at 14 d after fertilization; however, this method may not be considered an efficient method for large numbers of larval zebrafish. Exposure to chemicals, such as MS222 and eugenol, were ineffective methods for euthanasia at this stage of development. When these agents are used, secondary measures should be taken to ensure death. Choosing a euthanasia method that is effective, efficient, and humane can be challenging. Determining a method of euthanasia that is suitable for fish of all stages will bring the zebrafish community closer to meeting this challenge.

Effectiveness of Recommended Euthanasia Methods in Larval Zebrafish (Danio rerio)

PubMed Central

Strykowski, Jennifer L; Schech, Joseph M

2015-01-01

The popularity of zebrafish and its use as a model organism in biomedical research including genetics, development, and toxicology, has increased over the past 20 y and continues to grow. However, guidelines for euthanasia remain vague, and the responsibility of creating appropriate euthanasia protocols essentially falls on individual facilities. To reduce variation in experimental results among labs, a standard method of euthanasia for zebrafish would be useful. Although various euthanasia methods have been compared, few studies focus on the effectiveness of euthanasia methods for larval zebrafish. In this study, we exposed larval zebrafish to each of 3 euthanasia agents (MS222, eugenol, and hypothermic shock) and assessed the recovery rate. Hypothermic shock appeared to be the most effective method for euthanizing zebrafish at 14 d after fertilization; however, this method may not be considered an efficient method for large numbers of larval zebrafish. Exposure to chemicals, such as MS222 and eugenol, were ineffective methods for euthanasia at this stage of development. When these agents are used, secondary measures should be taken to ensure death. Choosing a euthanasia method that is effective, efficient, and humane can be challenging. Determining a method of euthanasia that is suitable for fish of all stages will bring the zebrafish community closer to meeting this challenge. PMID:25651096

The zebrafish world of colors and shapes: preference and discrimination.

PubMed

Oliveira, Jessica; Silveira, Mayara; Chacon, Diana; Luchiari, Ana

2015-04-01

Natural environment imposes many challenges to animals, which have to use cognitive abilities to cope with and exploit it to enhance their fitness. Since zebrafish is a well-established model for cognitive studies and high-throughput screening for drugs and diseases that affect cognition, we tested their ability for ambient color preference and 3D objects discrimination to establish a protocol for memory evaluation. For the color preference test, zebrafish were observed in a multiple-chamber tank with different environmental color options. Zebrafish showed preference for blue and green, and avoided yellow and red. For the 3D objects discrimination, zebrafish were allowed to explore two equal objects and then observed in a one-trial test in which a new color, size, or shape of the object was presented. Zebrafish showed discrimination for color, shape, and color+shape combined, but not size. These results imply that zebrafish seem to use some categorical system to discriminate items, and distracters affect their ability for discrimination. The type of variables available (color and shape) may favor zebrafish objects perception and facilitate discrimination processing. We suggest that this easy and simple memory test could serve as a useful screening tool for cognitive dysfunction and neurotoxicological studies.

Zebrafish xenograft models of cancer and metastasis for drug discovery.

PubMed

Brown, Hannah K; Schiavone, Kristina; Tazzyman, Simon; Heymann, Dominique; Chico, Timothy Ja

2017-04-01

Patients with metastatic cancer suffer the highest rate of cancer-related death, but existing animal models of metastasis have disadvantages that limit our ability to understand this process. The zebrafish is increasingly used for cancer modelling, particularly xenografting of human cancer cell lines, and drug discovery, and may provide novel scientific and therapeutic insights. However, this model system remains underexploited. Areas covered: The authors discuss the advantages and disadvantages of the zebrafish xenograft model for the study of cancer, metastasis and drug discovery. They summarise previous work investigating the metastatic cascade, such as tumour-induced angiogenesis, intravasation, extravasation, dissemination and homing, invasion at secondary sites, assessing metastatic potential and evaluation of cancer stem cells in zebrafish. Expert opinion: The practical advantages of zebrafish for basic biological study and drug discovery are indisputable. However, their ability to sufficiently reproduce and predict the behaviour of human cancer and metastasis remains unproven. For this to be resolved, novel mechanisms must to be discovered in zebrafish that are subsequently validated in humans, and for therapeutic interventions that modulate cancer favourably in zebrafish to successfully translate to human clinical studies. In the meantime, more work is required to establish the most informative methods in zebrafish.

Electroretinogram analysis of the visual response in zebrafish larvae.

PubMed

Chrispell, Jared D; Rebrik, Tatiana I; Weiss, Ellen R

2015-03-16

The electroretinogram (ERG) is a noninvasive electrophysiological method for determining retinal function. Through the placement of an electrode on the surface of the cornea, electrical activity generated in response to light can be measured and used to assess the activity of retinal cells in vivo. This manuscript describes the use of the ERG to measure visual function in zebrafish. Zebrafish have long been utilized as a model for vertebrate development due to the ease of gene suppression by morpholino oligonucleotides and pharmacological manipulation. At 5-10 dpf, only cones are functional in the larval retina. Therefore, the zebrafish, unlike other animals, is a powerful model system for the study of cone visual function in vivo. This protocol uses standard anesthesia, micromanipulation and stereomicroscopy protocols that are common in laboratories that perform zebrafish research. The outlined methods make use of standard electrophysiology equipment and a low light camera to guide the placement of the recording microelectrode onto the larval cornea. Finally, we demonstrate how a commercially available ERG stimulator/recorder originally designed for use with mice can easily be adapted for use with zebrafish. ERG of larval zebrafish provides an excellent method of assaying cone visual function in animals that have been modified by morpholino oligonucleotide injection as well as newer genome engineering techniques such as Zinc Finger Nucleases (ZFNs), Transcription Activator-Like Effector Nucleases (TALENs), and Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9, all of which have greatly increased the efficiency and efficacy of gene targeting in zebrafish. In addition, we take advantage of the ability of pharmacological agents to penetrate zebrafish larvae to evaluate the molecular components that contribute to the photoresponse. This protocol outlines a setup that can be modified and used by researchers with various experimental goals.

Study on acute toxicity of amoxicillin wastewater to Zebrafish

NASA Astrophysics Data System (ADS)

Xie, Weifang; Shen, Hongyan

2017-12-01

The main research in this paper is to obtain the effect of pharmaceutical wastewater on the acute toxicity of Zebrafish. The experimental method of exposure is used in this research. Experiments were carried out with different groups of pharmaceutical wastewater. Zebrafish was cultivated in a five liter fish tank. In the experiment, according to mortality, initially a 96h preliminary test was carried out at exposure concentrations to determine if the amoxicillin wastewater was toxic and to define the concentration range (24h LC100, 96h LC0) to be employed in the definitive tests. Based on the half lethal concentration of Zebrafish, the acute toxicity of amoxicillin wastewater to Zebrafish was calculated and the toxicity grade of wastewater was determined. In the experiment, the Zebrafish was exposed with amoxicillin wastewater during 96h. The 24h, 48h, 72h and 96h LC50 of amoxicillin wastewater on the Zebrafish were 63.10%, 53.70%, 41.69% and 40.74%, respectively. At 96h, the test time is the longest, and the value of LC50 is the smallest. In the observation period of 96 hours, the LC50 of amoxicillin wastewater were in the range of 40% ~ 60% and the value of Tua is 1 ~ 2. It indicates amoxicillin wastewater is low toxic wastewater when the experimental time is shorter than 48h, amoxicillin wastewater is moderate toxicity wastewater when the experimental time is higher than 48h. According to the experimental data, with the exposure time and the volume percentage of amoxicillin wastewater increases, the mortality rate of Zebrafish is gradually increased and the toxicity of amoxicillin wastewater increases. It indicates that the toxicity of amoxicillin wastewater is the biggest and the effect of wastewater on Zebrafish is greatest. In some ways, the toxicity of amoxicillin wastewater can be affected by the test time.

Modeling Leukemogenesis in the Zebrafish Using Genetic and Xenograft Models.

PubMed

Rajan, Vinothkumar; Dellaire, Graham; Berman, Jason N

2016-01-01

The zebrafish is a widely accepted model to study leukemia. The major advantage of studying leukemogenesis in zebrafish is attributed to its short life cycle and superior imaging capacity. This chapter highlights using transgenic- and xenograft-based models in zebrafish to study a specific leukemogenic mutation and analyze therapeutic responses in vivo.

Influenza A Virus Infection Damages Zebrafish Skeletal Muscle and Exacerbates Disease in Zebrafish Modeling Duchenne Muscular Dystrophy

PubMed Central

Goody, Michelle; Jurczyszak, Denise; Kim, Carol; Henry, Clarissa

2017-01-01

INTRODUCTION: Both genetic and infectious diseases can result in skeletal muscle degeneration, inflammation, pain, and/or weakness. Duchenne muscular dystrophy (DMD) is the most common congenital muscle disease. DMD causes progressive muscle wasting due to mutations in Dystrophin. Influenza A and B viruses are frequently associated with muscle complications, especially in children. Infections activate an immune response and immunosuppressant drugs reduce DMD symptoms. These data suggest that the immune system may contribute to muscle pathology. However, roles of the immune response in DMD and Influenza muscle complications are not well understood. Zebrafish with dmd mutations are a well-characterized model in which to study the molecular and cellular mechanisms of DMD pathology. We recently showed that zebrafish can be infected by human Influenza A virus (IAV). Thus, the zebrafish is a powerful system with which to ask questions about the etiology and mechanisms of muscle damage due to genetic and/or infectious diseases. METHODS: We infected zebrafish with IAV and assayed muscle tissue structure, sarcolemma integrity, cell-extracellular matrix (ECM) attachment, and molecular and cellular markers of inflammation in response to IAV infection alone or in the context of DMD. RESULTS: We find that IAV-infected zebrafish display mild muscle degeneration with sarcolemma damage and compromised ECM adhesion. An innate immune response is elicited in muscle in IAV-infected zebrafish: NFkB signaling is activated, pro-inflammatory cytokine expression is upregulated, and neutrophils localize to sites of muscle damage. IAV-infected dmd mutants display more severe muscle damage than would be expected from an additive effect of dmd mutation and IAV infection, suggesting that muscle damage caused by Dystrophin-deficiency and IAV infection is synergistic. DISCUSSION: These data demonstrate the importance of preventing IAV infections in individuals with genetic muscle diseases

Influenza A Virus Infection Damages Zebrafish Skeletal Muscle and Exacerbates Disease in Zebrafish Modeling Duchenne Muscular Dystrophy.

PubMed

Goody, Michelle; Jurczyszak, Denise; Kim, Carol; Henry, Clarissa

2017-10-25

Both genetic and infectious diseases can result in skeletal muscle degeneration, inflammation, pain, and/or weakness. Duchenne muscular dystrophy (DMD) is the most common congenital muscle disease. DMD causes progressive muscle wasting due to mutations in Dystrophin. Influenza A and B viruses are frequently associated with muscle complications, especially in children. Infections activate an immune response and immunosuppressant drugs reduce DMD symptoms. These data suggest that the immune system may contribute to muscle pathology. However, roles of the immune response in DMD and Influenza muscle complications are not well understood. Zebrafish with dmd mutations are a well-characterized model in which to study the molecular and cellular mechanisms of DMD pathology. We recently showed that zebrafish can be infected by human Influenza A virus (IAV). Thus, the zebrafish is a powerful system with which to ask questions about the etiology and mechanisms of muscle damage due to genetic and/or infectious diseases. We infected zebrafish with IAV and assayed muscle tissue structure, sarcolemma integrity, cell-extracellular matrix (ECM) attachment, and molecular and cellular markers of inflammation in response to IAV infection alone or in the context of DMD. We find that IAV-infected zebrafish display mild muscle degeneration with sarcolemma damage and compromised ECM adhesion. An innate immune response is elicited in muscle in IAV-infected zebrafish: NFkB signaling is activated, pro-inflammatory cytokine expression is upregulated, and neutrophils localize to sites of muscle damage. IAV-infected dmd mutants display more severe muscle damage than would be expected from an additive effect of dmd mutation and IAV infection, suggesting that muscle damage caused by Dystrophin-deficiency and IAV infection is synergistic. These data demonstrate the importance of preventing IAV infections in individuals with genetic muscle diseases. Elucidating the mechanisms of immune

Zebrafish Craniofacial Development: A Window into Early Patterning

PubMed Central

Mork, Lindsey; Crump, Gage

2016-01-01

The formation of the face and skull involves a complex series of developmental events mediated by cells derived from the neural crest, endoderm, mesoderm, and ectoderm. Although vertebrates boast an enormous diversity of adult facial morphologies, the fundamental signaling pathways and cellular events that sculpt the nascent craniofacial skeleton in the embryo have proven to be highly conserved from fish to man. The zebrafish Danio rerio, a small freshwater cyprinid fish from eastern India, has served as a popular model of craniofacial development since the 1990s. Unique strengths of the zebrafish model include a simplified skeleton during larval stages, access to rapidly developing embryos for live imaging, and amenability to transgenesis and complex genetics. In this chapter, we describe the anatomy of the zebrafish craniofacial skeleton; its applications as models for the mammalian jaw, middle ear, palate, and cranial sutures; the superior imaging technology available in fish that has provided unprecedented insights into the dynamics of facial morphogenesis; the use of the zebrafish to decipher the genetic underpinnings of craniofacial biology; and finally a glimpse into the most promising future applications of zebrafish craniofacial research. PMID:26589928

Zebrafish Axenic Larvae Colonization with Human Intestinal Microbiota.

PubMed

Arias-Jayo, Nerea; Alonso-Saez, Laura; Ramirez-Garcia, Andoni; Pardo, Miguel A

2018-04-01

The human intestine hosts a vast and complex microbial community that is vital for maintaining several functions related with host health. The processes that determine the gut microbiome composition are poorly understood, being the interaction between species, the external environment, and the relationship with the host the most feasible. Animal models offer the opportunity to understand the interactions between the host and the microbiota. There are different gnotobiotic mice or rat models colonized with the human microbiota, however, to our knowledge, there are no reports on the colonization of germ-free zebrafish with a complex human intestinal microbiota. In the present study, we have successfully colonized 5 days postfertilization germ-free zebrafish larvae with the human intestinal microbiota previously extracted from a donor and analyzed by high-throughput sequencing the composition of the transferred microbial communities that established inside the zebrafish gut. Thus, we describe for first time which human bacteria phylotypes are able to colonize the zebrafish digestive tract. Species with relevant interest because of their linkage to dysbiosis in different human diseases, such as Akkermansia muciniphila, Eubacterium rectale, Faecalibacterium prausnitzii, Prevotella spp., or Roseburia spp. have been successfully transferred inside the zebrafish digestive tract.

Pacemaker-neuron–dependent disturbance of the molecular clockwork by a Drosophila CLOCK mutant homologous to the mouse Clock mutation

PubMed Central

Lee, Euna; Cho, Eunjoo; Kang, Doo Hyun; Jeong, Eun Hee; Chen, Zheng; Yoo, Seung-Hee; Kim, Eun Young

2016-01-01

Circadian clocks are composed of transcriptional/translational feedback loops (TTFLs) at the cellular level. In Drosophila TTFLs, the transcription factor dCLOCK (dCLK)/CYCLE (CYC) activates clock target gene expression, which is repressed by the physical interaction with PERIOD (PER). Here, we show that amino acids (AA) 657–707 of dCLK, a region that is homologous to the mouse Clock exon 19-encoded region, is crucial for PER binding and E-box–dependent transactivation in S2 cells. Consistently, in transgenic flies expressing dCLK with an AA657–707 deletion in the Clock (Clkout) genetic background (p{dClk-Δ};Clkout), oscillation of core clock genes’ mRNAs displayed diminished amplitude compared with control flies, and the highly abundant dCLKΔ657–707 showed significantly decreased binding to PER. Behaviorally, the p{dClk-Δ};Clkout flies exhibited arrhythmic locomotor behavior in the photic entrainment condition but showed anticipatory activities of temperature transition and improved free-running rhythms in the temperature entrainment condition. Surprisingly, p{dClk-Δ};Clkout flies showed pacemaker-neuron–dependent alterations in molecular rhythms; the abundance of dCLK target clock proteins was reduced in ventral lateral neurons (LNvs) but not in dorsal neurons (DNs) in both entrainment conditions. In p{dClk-Δ};Clkout flies, however, strong but delayed molecular oscillations in temperature cycle-sensitive pacemaker neurons, such as DN1s and DN2s, were correlated with delayed anticipatory activities of temperature transition. Taken together, our study reveals that the LNv molecular clockwork is more sensitive than the clockwork of DNs to dysregulation of dCLK by AA657–707 deletion. Therefore, we propose that the dCLK/CYC-controlled TTFL operates differently in subsets of pacemaker neurons, which may contribute to their specific functions. PMID:27489346

Loss of col8a1a Function during Zebrafish Embryogenesis Results in Congenital Vertebral Malformations

PubMed Central

Gray, Ryan S.; Wilm, Thomas; Smith, Jeff; Bagnat, Michel; Dale, Rodney M.; Topczewski, Jacek; Johnson, Stephen L.; Solnica-Krezel, Lilianna

2014-01-01

Congenital vertebral malformations (CVM) occur in 1 in 1,000 live births and in many cases can cause spinal deformities, such as scoliosis, and result in disability and distress of affected individuals. Many severe forms of the disease, such as spondylocostal dystostosis, are recessive monogenic traits affecting somitogenesis, however the etiologies of the majority of CVM cases remain undetermined. Here we demonstrate that morphological defects of the notochord in zebrafish can generate congenital-type spine defects. We characterize three recessive zebrafish leviathan/col8a1a mutant alleles (m531, vu41, vu105) that disrupt collagen type VIII alpha1a (col8a1a), and cause folding of the embryonic notochord and consequently adult vertebral column malformations. Furthermore, we provide evidence that a transient loss of col8a1a function or inhibition of Lysyl oxidases with drugs during embryogenesis was sufficient to generate vertebral fusions and scoliosis in the adult spine. Using periodic imaging of individual zebrafish, we correlate focal notochord defects of the embryo with vertebral malformations (VM) in the adult. Finally, we show that bends and kinks in the notochord can lead to aberrant apposition of osteoblasts normally confined to well-segmented areas of the developing vertebral bodies. Our results afford a novel mechanism for the formation of VM, independent of defects of somitogenesis, resulting from aberrant bone deposition at regions of misshapen notochord tissue. PMID:24333517

Loss of col8a1a function during zebrafish embryogenesis results in congenital vertebral malformations.

PubMed

Gray, Ryan S; Wilm, Thomas P; Smith, Jeff; Bagnat, Michel; Dale, Rodney M; Topczewski, Jacek; Johnson, Stephen L; Solnica-Krezel, Lilianna

2014-02-01

Congenital vertebral malformations (CVM) occur in 1 in 1000 live births and in many cases can cause spinal deformities, such as scoliosis, and result in disability and distress of affected individuals. Many severe forms of the disease, such as spondylocostal dystostosis, are recessive monogenic traits affecting somitogenesis, however the etiologies of the majority of CVM cases remain undetermined. Here we demonstrate that morphological defects of the notochord in zebrafish can generate congenital-type spine defects. We characterize three recessive zebrafish leviathan/col8a1a mutant alleles ((m531, vu41, vu105)) that disrupt collagen type VIII alpha1a (col8a1a), and cause folding of the embryonic notochord and consequently adult vertebral column malformations. Furthermore, we provide evidence that a transient loss of col8a1a function or inhibition of Lysyl oxidases with drugs during embryogenesis was sufficient to generate vertebral fusions and scoliosis in the adult spine. Using periodic imaging of individual zebrafish, we correlate focal notochord defects of the embryo with vertebral malformations (VM) in the adult. Finally, we show that bends and kinks in the notochord can lead to aberrant apposition of osteoblasts normally confined to well-segmented areas of the developing vertebral bodies. Our results afford a novel mechanism for the formation of VM, independent of defects of somitogenesis, resulting from aberrant bone deposition at regions of misshapen notochord tissue. Copyright © 2013 Elsevier Inc. All rights reserved.

RNA-seq analysis of Drosophila clock and non-clock neurons reveals neuron-specific cycling and novel candidate neuropeptides.

PubMed

Abruzzi, Katharine C; Zadina, Abigail; Luo, Weifei; Wiyanto, Evelyn; Rahman, Reazur; Guo, Fang; Shafer, Orie; Rosbash, Michael

2017-02-01

Locomotor activity rhythms are controlled by a network of ~150 circadian neurons within the adult Drosophila brain. They are subdivided based on their anatomical locations and properties. We profiled transcripts "around the clock" from three key groups of circadian neurons with different functions. We also profiled a non-circadian outgroup, dopaminergic (TH) neurons. They have cycling transcripts but fewer than clock neurons as well as low expression and poor cycling of clock gene transcripts. This suggests that TH neurons do not have a canonical circadian clock and that their gene expression cycling is driven by brain systemic cues. The three circadian groups are surprisingly diverse in their cycling transcripts and overall gene expression patterns, which include known and putative novel neuropeptides. Even the overall phase distributions of cycling transcripts are distinct, indicating that different regulatory principles govern transcript oscillations. This surprising cell-type diversity parallels the functional heterogeneity of the different neurons.

[Elevated expression of CLOCK is associated with poor prognosis in hepatocellular carcinoma].

PubMed

Li, Bo; Yang, Xiliang; Li, Jiaqi; Yang, Yi; Yan, Zhaoyong; Zhang, Hongxin; Mu, Jiao

2018-02-01

Objective To evaluate the expression of circadian locomotor output cycles kaput (CLOCK) and its effects on cell growth in hepatocellular carcinoma (HCC). Methods The expression of CLOCK in 158 pairs of human HCC tissues and matched noncancerous samples was detected by immunohistochemical (IHC) staining. The expression of CLOCK in HCC patients was also verified using the data from GEO and TCGA (a total of 356 cases). The relationship between CLOCK expression and clinicopathological features of HCC patients was analyzed by single factor statistical analysis. Kaplan-Meier survival curves of HCC patients were drawn to study the relationship between the expression level of CLOCK and the survival state. The effect of CLOCK on the growth of HepG2 cells was detected by MTS assay. Results The expression of CLOCK in HCC tissues was significantly higher than that in the adjacent tissues, and the up-regulation of CLOCK expression in HCC tissue was also confirmed in the public data of HCC (356 cases). HCC patients were divided into low CLOCK expression group and high CLOCK expression group. Univariate analysis showed that the expression of CLOCK was related to tumor size, TNM stage, and portal vein invasion in HCC patients. HCC patients with low CLOCK expression had longer overall survival time and relapse-free survival time than those with high CLOCK expression. The proliferation of cells significantly decreased after the expression of CLOCK was knocked down in HepG2 cells. Conclusion The expression of CLOCK in HCC tissues was much higher than that in normal liver tissues, and the high expression of CLOCK indicated the poor prognosis. The knockdown of CLOCK in HCC cells could inhibit the proliferation of HepG2 cells.

Method for somatic cell nuclear transfer in zebrafish.

PubMed

Siripattarapravat, Kannika; Cibelli, Jose B

2011-01-01

Somatic cell nuclear transfer (SCNT) has been a well-known technique for decades and widely applied to generate identical animals, including ones with genetic alterations. The system has been demonstrated successfully in zebrafish. The elaborated requirements of SCNT, however, limit reproducibility of the established model to a few groups in zebrafish research community. In this chapter, we meticulously outline each step of the published protocol as well as preparations of equipments and reagents used in zebrafish SCNT. All describable detailed-tips are elaborated in texts and figures. Copyright © 2011 Elsevier Inc. All rights reserved.

HNF1β Is Essential for Nephron Segmentation during Nephrogenesis

PubMed Central

Naylor, Richard W.; Przepiorski, Aneta; Ren, Qun; Yu, Jing

2012-01-01

Nephrons comprise a blood filter and an epithelial tubule that is subdivided into proximal and distal segments, but what directs this patterning during kidney organogenesis is not well understood. Using zebrafish, we found that the HNF1β paralogues hnf1ba and hnf1bb, which encode homeodomain transcription factors, are essential for normal segmentation of nephrons. Embryos deficient in hnf1ba and hnf1bb did not express proximal and distal segment markers, yet still developed an epithelial tubule. Initiating hnf1ba/b expression required Pax2a and Pax8, but hnf1ba/b-deficient embryos did not exhibit the expected downregulation of pax2a and pax8 at later stages of development, suggesting complex regulatory loops involving these molecules. Embryos deficient in hnf1ba/b also did not express the irx3b transcription factor, which is responsible for differentiation of the first distal tubule segment. Reciprocally, embryos deficient in irx3b exhibited downregulation of hnf1ba/b transcripts in the distal early segment, suggesting a segment-specific regulatory circuit. Deficiency of hnf1ba/b also led to ectopic expansion of podocytes into the proximal tubule domain. Epistasis experiments showed that the formation of podocytes required wt1a, which encodes the Wilms’ tumor suppressor-1 transcription factor, and rbpj, which encodes a mediator of canonical Notch signaling, downstream or parallel to hnf1ba/b. Taken together, these results suggest that Hnf1β factors are essential for normal segmentation of nephrons during kidney organogenesis. PMID:23160512

HNF1β is essential for nephron segmentation during nephrogenesis.

PubMed

Naylor, Richard W; Przepiorski, Aneta; Ren, Qun; Yu, Jing; Davidson, Alan J

2013-01-01

Nephrons comprise a blood filter and an epithelial tubule that is subdivided into proximal and distal segments, but what directs this patterning during kidney organogenesis is not well understood. Using zebrafish, we found that the HNF1β paralogues hnf1ba and hnf1bb, which encode homeodomain transcription factors, are essential for normal segmentation of nephrons. Embryos deficient in hnf1ba and hnf1bb did not express proximal and distal segment markers, yet still developed an epithelial tubule. Initiating hnf1ba/b expression required Pax2a and Pax8, but hnf1ba/b-deficient embryos did not exhibit the expected downregulation of pax2a and pax8 at later stages of development, suggesting complex regulatory loops involving these molecules. Embryos deficient in hnf1ba/b also did not express the irx3b transcription factor, which is responsible for differentiation of the first distal tubule segment. Reciprocally, embryos deficient in irx3b exhibited downregulation of hnf1ba/b transcripts in the distal early segment, suggesting a segment-specific regulatory circuit. Deficiency of hnf1ba/b also led to ectopic expansion of podocytes into the proximal tubule domain. Epistasis experiments showed that the formation of podocytes required wt1a, which encodes the Wilms' tumor suppressor-1 transcription factor, and rbpj, which encodes a mediator of canonical Notch signaling, downstream or parallel to hnf1ba/b. Taken together, these results suggest that Hnf1β factors are essential for normal segmentation of nephrons during kidney organogenesis.

Feedback repression is required for mammalian circadian clock function.

PubMed

Sato, Trey K; Yamada, Rikuhiro G; Ukai, Hideki; Baggs, Julie E; Miraglia, Loren J; Kobayashi, Tetsuya J; Welsh, David K; Kay, Steve A; Ueda, Hiroki R; Hogenesch, John B

2006-03-01

Direct evidence for the requirement of transcriptional feedback repression in circadian clock function has been elusive. Here, we developed a molecular genetic screen in mammalian cells to identify mutants of the circadian transcriptional activators CLOCK and BMAL1, which were uncoupled from CRYPTOCHROME (CRY)-mediated transcriptional repression. Notably, mutations in the PER-ARNT-SIM domain of CLOCK and the C terminus of BMAL1 resulted in synergistic insensitivity through reduced physical interactions with CRY. Coexpression of these mutant proteins in cultured fibroblasts caused arrhythmic phenotypes in population and single-cell assays. These data demonstrate that CRY-mediated repression of the CLOCK/BMAL1 complex activity is required for maintenance of circadian rhythmicity and provide formal proof that transcriptional feedback is required for mammalian clock function.

Anxiogenic-like effects of chronic nicotine exposure in zebrafish.

PubMed

Stewart, Adam Michael; Grossman, Leah; Collier, Adam D; Echevarria, David J; Kalueff, Allan V

2015-12-01

Nicotine is one of the most widely used and abused legal drugs. Although its pharmacological profile has been extensively investigated in humans and rodents, nicotine CNS action remains poorly understood. The importance of finding evolutionarily conserved signaling pathways, and the need to apply high-throughput in vivo screens for CNS drug discovery, necessitate novel efficient experimental models for nicotine research. Zebrafish (Danio rerio) are rapidly emerging as an excellent organism for studying drug abuse, neuropharmacology and toxicology and have recently been applied to testing nicotine. Anxiolytic, rewarding and memory-modulating effects of acute nicotine treatment in zebrafish are consistently reported in the literature. However, while nicotine abuse is more relevant to long-term exposure models, little is known about chronic effects of nicotine on zebrafish behavior. In the present study, chronic 4-day exposure to 1-2mg/L nicotine mildly increased adult zebrafish shoaling but did not alter baseline cortisol levels. We also found that chronic exposure to nicotine evokes robust anxiogenic behavioral responses in zebrafish tested in the novel tank test paradigm. Generally paralleling clinical and rodent data on anxiogenic effects of chronic nicotine, our study supports the developing utility of zebrafish for nicotine research. Copyright © 2015 Elsevier Inc. All rights reserved.

Zebrafish Models of Human Leukemia: Technological Advances and Mechanistic Insights.

PubMed

Harrison, Nicholas R; Laroche, Fabrice J F; Gutierrez, Alejandro; Feng, Hui

2016-01-01

Insights concerning leukemic pathophysiology have been acquired in various animal models and further efforts to understand the mechanisms underlying leukemic treatment resistance and disease relapse promise to improve therapeutic strategies. The zebrafish (Danio rerio) is a vertebrate organism with a conserved hematopoietic program and unique experimental strengths suiting it for the investigation of human leukemia. Recent technological advances in zebrafish research including efficient transgenesis, precise genome editing, and straightforward transplantation techniques have led to the generation of a number of leukemia models. The transparency of the zebrafish when coupled with improved lineage-tracing and imaging techniques has revealed exquisite details of leukemic initiation, progression, and regression. With these advantages, the zebrafish represents a unique experimental system for leukemic research and additionally, advances in zebrafish-based high-throughput drug screening promise to hasten the discovery of novel leukemia therapeutics. To date, investigators have accumulated knowledge of the genetic underpinnings critical to leukemic transformation and treatment resistance and without doubt, zebrafish are rapidly expanding our understanding of disease mechanisms and helping to shape therapeutic strategies for improved outcomes in leukemic patients.

Zebrafish Models of Human Leukemia: Technological Advances and Mechanistic Insights

PubMed Central

Harrison, Nicholas R.; Laroche, Fabrice J.F.; Gutierrez, Alejandro

2016-01-01

Insights concerning leukemic pathophysiology have been acquired in various animal models and further efforts to understand the mechanisms underlying leukemic treatment resistance and disease relapse promise to improve therapeutic strategies. The zebrafish (Danio rerio) is a vertebrate organism with a conserved hematopoietic program and unique experimental strengths suiting it for the investigation of human leukemia. Recent technological advances in zebrafish research including efficient transgenesis, precise genome editing, and straightforward transplantation techniques have led to the generation of a number of leukemia models. The transparency of the zebrafish when coupled with improved lineage-tracing and imaging techniques has revealed exquisite details of leukemic initiation, progression, and regression. With these advantages, the zebrafish represents a unique experimental system for leukemic research and additionally, advances in zebrafish-based high-throughput drug screening promise to hasten the discovery of novel leukemia therapeutics. To date, investigators have accumulated knowledge of the genetic underpinnings critical to leukemic transformation and treatment resistance and without doubt, zebrafish are rapidly expanding our understanding of disease mechanisms and helping to shape therapeutic strategies for improved outcomes in leukemic patients. PMID:27165361

Making the clock tick: the transcriptional landscape of the plant circadian clock.

PubMed

Ronald, James; Davis, Seth J

2017-01-01

Circadian clocks are molecular timekeepers that synchronise internal physiological processes with the external environment by integrating light and temperature stimuli. As in other eukaryotic organisms, circadian rhythms in plants are largely generated by an array of nuclear transcriptional regulators and associated co-regulators that are arranged into a series of interconnected molecular loops. These transcriptional regulators recruit chromatin-modifying enzymes that adjust the structure of the nucleosome to promote or inhibit DNA accessibility and thus guide transcription rates. In this review, we discuss the recent advances made in understanding the architecture of the Arabidopsis oscillator and the chromatin dynamics that regulate the generation of rhythmic patterns of gene expression within the circadian clock.

Production of zebrafish cardiospheres and cardiac progenitor cells in vitro and three-dimensional culture of adult zebrafish cardiac tissue in scaffolds.

PubMed

Zeng, Wendy R; Beh, Siew-Joo; Bryson-Richardson, Robert J; Doran, Pauline M

2017-09-01

The hearts of adult zebrafish (Danio rerio) are capable of complete regeneration in vivo even after major injury, making this species of particular interest for understanding the growth and differentiation processes required for cardiac tissue engineering. To date, little research has been carried out on in vitro culture of adult zebrafish cardiac cells. In this work, progenitor-rich cardiospheres suitable for cardiomyocyte differentiation and myocardial regeneration were produced from adult zebrafish hearts. The cardiospheres contained a mixed population of c-kit + and Mef2c + cells; proliferative peripheral cells of possible mesenchymal lineage were also observed. Cellular outgrowth from cardiac explants and cardiospheres was enhanced significantly using conditioned medium harvested from cultures of a rainbow trout cell line, suggesting that fish-specific trophic factors are required for zebrafish cardiac cell expansion. Three-dimensional culture of zebrafish heart cells in fibrous polyglycolic acid (PGA) scaffolds was carried out under dynamic fluid flow conditions. High levels of cell viability and cardiomyocyte differentiation were maintained within the scaffolds. Expression of cardiac troponin T, a marker of differentiated cardiomyocytes, increased during the first 7 days of scaffold culture; after 15 days, premature disintegration of the biodegradable scaffolds led to cell detachment and a decline in differentiation status. This work expands our technical capabilities for three-dimensional zebrafish cardiac cell culture with potential applications in tissue engineering, drug and toxicology screening, and ontogeny research. Biotechnol. Bioeng. 2017;114: 2142-2148. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

The development of a Kalman filter clock predictor

NASA Technical Reports Server (NTRS)

Davis, John A.; Greenhall, Charles A.; Boudjemaa, Redoane

2005-01-01

A Kalman filter based clock predictor is developed, and its performance evaluated using both simulated and real data. The clock predictor is shown to possess a neat to optimal Prediction Error Variance (PEV) when the underlying noise consists of one of the power law noise processes commonly encountered in time and frequency measurements. The predictor's performance is the presence of multiple noise processes is also examined. The relationship between the PEV obtained in the presence of multiple noise processes and those obtained for the individual component noise processes is examined. Comparisons are made with a simple linear clock predictor. The clock predictor is used to predict future values of the time offset between pairs of NPL's active hydrogen masers.

A transportable optical clock for chronometric levelling

NASA Astrophysics Data System (ADS)

Lisdat, C.; Koller, S. B.; Grotti, J.; Vogt, S.; Al-Masoudi, A.; Dörscher, S.; Herbers, S.; Häfner, S.; Sterr, U.

2016-12-01

With their supreme accuracy and precision, optical clocks in combination with new methods of long-distance frequency transfer can be used to determine height differences by measuring the gravitational red shift between two clocks without accumulation of measurement errors, as in classical levelling. We are developing transportable optical clocks for this purpose that will also serve for the technology development regarding optical clocks in Space and for international comparisons between optical clocks that cannot be linked with sufficient accuracy otherwise.In this talk we will focus on the transportable strontium lattice clock that we are developing and its first evaluation. Presently, we achieve a fractional frequency instability of 3×10-17 after 1000 s averaging time, which is equivalent to a height resolution of 30 cm. The first uncertainty evaluation of the system yielded 7×10-17. We expect rapid improvements to an uncertainty of a few parts in 1017.The clock then placed within a car trailer, which requires compact and rugged lasers systems and physics package. Special care has been taken in the design of the ultra-frequency stable interrogation laser that has to achieve fractional frequency instabilities of considerably below 10-15. Typical laboratory constructions of the reference resonator system used to pre-stabilize the laser frequency are not compatible with the requirement of transportability.In an actual levelling campaign, this clock will be connected e.g. via a stabilized optical fibre link with another, stationary frequency standard. The measured gravitational red shift can be compared to the ones calculated from potential differences derived with state of the art geodetic data and models. A first campaign has been completed in cooperation with colleagues from the Italian and UK metrology institutes INRIM and NPL, respectively, and the Institut für Erdmessung (IfE), Leibniz University Hannover. We will discuss the status of the evaluation and

Sub-nanosecond clock synchronization and precision deep space tracking

NASA Technical Reports Server (NTRS)

Dunn, Charles; Lichten, Stephen; Jefferson, David; Border, James S.

1992-01-01

Interferometric spacecraft tracking is accomplished at the NASA Deep Space Network (DSN) by comparing the arrival time of electromagnetic spacecraft signals to ground antennas separated by baselines on the order of 8000 km. Clock synchronization errors within and between DSN stations directly impact the attainable tracking accuracy, with a 0.3 ns error in clock synchronization resulting in an 11 nrad angular position error. This level of synchronization is currently achieved by observing a quasar which is angularly close to the spacecraft just after the spacecraft observations. By determining the differential arrival times of the random quasar signal at the stations, clock synchronization and propagation delays within the atmosphere and within the DSN stations are calibrated. Recent developments in time transfer techniques may allow medium accuracy (50-100 nrad) spacecraft observations without near-simultaneous quasar-based calibrations. Solutions are presented for a global network of GPS receivers in which the formal errors in clock offset parameters are less than 0.5 ns. Comparisons of clock rate offsets derived from GPS measurements and from very long baseline interferometry and the examination of clock closure suggest that these formal errors are a realistic measure of GPS-based clock offset precision and accuracy. Incorporating GPS-based clock synchronization measurements into a spacecraft differential ranging system would allow tracking without near-simultaneous quasar observations. The impact on individual spacecraft navigation error sources due to elimination of quasar-based calibrations is presented. System implementation, including calibration of station electronic delays, is discussed.

[10]-Gingerdiols as the major metabolites of [10]-gingerol in zebrafish embryos and in humans and their hematopoietic effects in zebrafish embryos

PubMed Central

Chen, Huadong; Soroka, Dominique N.; Haider, Jamil; Ferri-Lagneau, Karine F.; Leung, TinChung; Sang, Shengmin

2013-01-01

Gingerols are a series of major constituents in fresh ginger with the most abundant being [6]-, [8]-, and [10]-gingerols (6G, 8G, and 10G). We previously found that ginger extract and its purified components, especially 10G, potentially stimulate both the primitive and definitive waves of hematopoiesis (blood cell formation) in zebrafish embryos. However, it is still unclear if the metabolites of 10G retain the efficacy of the parent compound towards pathological anemia treatment. In the present study, we first investigated the metabolism of 10G in zebrafish embryos, and then explored the biotransformation of 10G in humans. Our results show that 10G was extensively metabolized in both zebrafish embryos and in humans, in which two major metabolites, (3S,5S)-[10]-gingerdiol and (3R,5S)-[10]-gingerdiol, were identified by analysis of the MSn spectra and comparison to authentic standards that we synthesized. After 24 hours of treatment of zebrafish embryos, 10G was mostly converted to its metabolites. Our results clearly indicate the reductive pathway is a major metabolic route for 10G in both zebrafish embryos and in humans. Furthermore, we investigated the hematopoietic effect of 10G and its two metabolites, which show similar hematopoietic effects as 10G in zebrafish embryos. PMID:23701129

Crosstalk between the Circadian Clock and Innate Immunity in Arabidopsis

PubMed Central

Zhang, Chong; Xie, Qiguang; Anderson, Ryan G.; Ng, Gina; Seitz, Nicholas C.; Peterson, Thomas; McClung, C. Robertson; McDowell, John M.; Kong, Dongdong; Kwak, June M.; Lu, Hua

2013-01-01

The circadian clock integrates temporal information with environmental cues in regulating plant development and physiology. Recently, the circadian clock has been shown to affect plant responses to biotic cues. To further examine this role of the circadian clock, we tested disease resistance in mutants disrupted in CCA1 and LHY, which act synergistically to regulate clock activity. We found that cca1 and lhy mutants also synergistically affect basal and resistance gene-mediated defense against Pseudomonas syringae and Hyaloperonospora arabidopsidis. Disrupting the circadian clock caused by overexpression of CCA1 or LHY also resulted in severe susceptibility to P. syringae. We identified a downstream target of CCA1 and LHY, GRP7, a key constituent of a slave oscillator regulated by the circadian clock and previously shown to influence plant defense and stomatal activity. We show that the defense role of CCA1 and LHY against P. syringae is at least partially through circadian control of stomatal aperture but is independent of defense mediated by salicylic acid. Furthermore, we found defense activation by P. syringae infection and treatment with the elicitor flg22 can feedback-regulate clock activity. Together this data strongly supports a direct role of the circadian clock in defense control and reveal for the first time crosstalk between the circadian clock and plant innate immunity. PMID:23754942

Zebrafish for the Study of the Biological Effects of Nicotine

PubMed Central

Klee, Eric W.; Schneider, Henning; Hurt, Richard D.; Ekker, Stephen C.

2011-01-01

Introduction: Zebrafish are emerging as a powerful animal model for studying the molecular and physiological effects of nicotine exposure. The zebrafish have many advantageous physical characteristics, including small size, high fecundity rates, and externally developing transparent embryos. When combined with a battery of molecular–genetic tools and behavioral assays, these attributes enable studies to be conducted that are not practical using traditional animal models. Methods: We reviewed the literature on the application of the zebrafish model as a preclinical model to study the biological effects of nicotine exposure. Results: The identified studies used zebrafish to examine the effects of nicotine exposure on early development, addiction, anxiety, and learning. The methods used included green fluorescent protein–labeled proteins to track in vivo nicotine-altered neuron development, nicotine-conditioned place preference, and locomotive sensitization linked with high-throughput molecular and genetic screens and behavioral models of learning and stress response to nicotine. Data are presented on the complete homology of all known human neural nicotinic acetylcholine receptors in zebrafish and on the biological similarity of human and zebrafish dopaminergic signaling. Conclusions: Tobacco dependence remains a major health problem worldwide. Further understanding of the molecular effects of nicotine exposure and genetic contributions to dependence may lead to improvement in patient treatment strategies. While there are limitations to the use of zebrafish as a preclinical model, it should provide a valuable tool to complement existing model systems. The reviewed studies demonstrate the enormous opportunity zebrafish have to advance the science of nicotine and tobacco research. PMID:21385906

A Fermi-degenerate three-dimentional optical lattice clock

NASA Astrophysics Data System (ADS)

Goban, Akihisa; Campbell, Sara; Hutson, Ross; Marti, G. Edward; Sonderhouse, Lindsay; Robinson, John; Zhang, Wei; Ye, Jun

2017-04-01

The pursuit of better atomic clocks has advanced many research areas, providing better quantum state control, tighter limits on fundamental constant variation, and improved tests of relativity. Recent progress in optical lattice clock to the accuracy of 2E-18 has benefited from the understanding of atomic interactions. Also the precision of clock spectroscopy has been applied to explore many-body interactions including SU(N) symmetry. In our previous 1D optical lattice, atomic interactions cause suppression and broadening of the atomic resonance, limiting the clock stability. To overcome this limitation, we demonstrate a scalable solution that takes advantage of the high density of a degenerate Fermi gas in a three-dimensional optical lattice to protect against on-site interaction shifts. Using an ultrastable laser, we achieve an unprecedented level of atom-light coherence, reaching a spectroscopic quality factor 5.2E15. We investigate clock systematics unique to this design; on-site interactions are resolved so that their contribution to clock shifts is orders of magnitude suppressed compared to the 1D optical lattice experiments. Also, we measure the combined scalar and tensor magic wavelengths for state-independent trapping along all three lattice axes. We acknowledge support from NIST, DARPA and the NSF JILA Physics Frontier Center.

Atomic clocks and the continuous-time random-walk

NASA Astrophysics Data System (ADS)

Formichella, Valerio; Camparo, James; Tavella, Patrizia

2017-11-01

Atomic clocks play a fundamental role in many fields, most notably they generate Universal Coordinated Time and are at the heart of all global navigation satellite systems. Notwithstanding their excellent timekeeping performance, their output frequency does vary: it can display deterministic frequency drift; diverse continuous noise processes result in nonstationary clock noise (e.g., random-walk frequency noise, modelled as a Wiener process), and the clock frequency may display sudden changes (i.e., "jumps"). Typically, the clock's frequency instability is evaluated by the Allan or Hadamard variances, whose functional forms can identify the different operative noise processes. Here, we show that the Allan and Hadamard variances of a particular continuous-time random-walk, the compound Poisson process, have the same functional form as for a Wiener process with drift. The compound Poisson process, introduced as a model for observed frequency jumps, is an alternative to the Wiener process for modelling random walk frequency noise. This alternate model fits well the behavior of the rubidium clocks flying on GPS Block-IIR satellites. Further, starting from jump statistics, the model can be improved by considering a more general form of continuous-time random-walk, and this could bring new insights into the physics of atomic clocks.

Shining a light on the Arabidopsis circadian clock.

PubMed

Oakenfull, Rachael J; Davis, Seth J

2017-11-01

The circadian clock provides essential timing information to ensure optimal growth to prevailing external environmental conditions. A major time-setting mechanism (zeitgeber) in clock synchronization is light. Differing light wavelengths, intensities, and photoperiodic duration are processed for the clock-setting mechanism. Many studies on light-input pathways to the clock have focused on Arabidopsis thaliana. Photoreceptors are specific chromic proteins that detect light signals and transmit this information to the central circadian oscillator through a number of different signalling mechanisms. The most well-characterized clock-mediating photoreceptors are cryptochromes and phytochromes, detecting blue, red, and far-red wavelengths of light. Ultraviolet and shaded light are also processed signals to the oscillator. Notably, the clock reciprocally generates rhythms of photoreceptor action leading to so-called gating of light responses. Intermediate proteins, such as Phytochrome interacting factors (PIFs), constitutive photomorphogenic 1 (COP1) and EARLY FLOWERING 3 (ELF3), have been established in signalling pathways downstream of photoreceptor activation. However, the precise details for these signalling mechanisms are not fully established. This review highlights both historical and recent efforts made to understand overall light input to the oscillator, first looking at how each wavelength of light is detected, this is then related to known input mechanisms and their interactions. © 2017 John Wiley & Sons Ltd.

The mammalian circadian clock and its entrainment by stress and exercise.

PubMed

Tahara, Yu; Aoyama, Shinya; Shibata, Shigenobu

2017-01-01

The mammalian circadian clock regulates day-night fluctuations in various physiological processes. The circadian clock consists of the central clock in the suprachiasmatic nucleus of the hypothalamus and peripheral clocks in peripheral tissues. External environmental cues, including light/dark cycles, food intake, stress, and exercise, provide important information for adjusting clock phases. This review focuses on stress and exercise as potent entrainment signals for both central and peripheral clocks, especially in regard to the timing of stimuli, types of stressors/exercises, and differences in the responses of rodents and humans. We suggest that the common signaling pathways of clock entrainment by stress and exercise involve sympathetic nervous activation and glucocorticoid release. Furthermore, we demonstrate that physiological responses to stress and exercise depend on time of day. Therefore, using exercise to maintain the circadian clock at an appropriate phase and amplitude might be effective for preventing obesity, diabetes, and cardiovascular disease.

SRC: Smart Reminder Clock

NASA Astrophysics Data System (ADS)

Kasim, Shahreen; Hafit, Hanayanti; Leong, Tan Hua; Hashim, Rathiah; Ruslai, Husni; Jahidin, Kamaruzzaman; Syafwan Arshad, Mohammad

2016-11-01

Nowadays, some people facing the problem to wake up in the morning. This was result to absence of the classes, meetings, and even exams. The aim of this project is to develop an android application that can force the user to wake up. The method used in this application are pedometer and Short Message Service (SMS) function. This application need the user to take their smartphone and walk about 10 steps to disable it, when the alarm clock is activated. After that, when the alarm clock was rang, this alarm application has automatically send a message to the users’ friends or parents phone to wake them up.

In vivo physiological recording from the lateral line of juvenile zebrafish

PubMed Central

Olt, Jennifer; Allen, Claire E.

2016-01-01

Key points Zebrafish provide a unique opportunity to investigate in vivo sensory transduction in mature hair cells.We have developed a method for studying the biophysical properties of mature hair cells from the lateral line of juvenile zebrafish.The method involves application of the anaesthetic benzocaine and intubation to maintain ventilation and oxygenation through the gills.The same approach could be used for in vivo functional studies in other sensory and non‐sensory systems from juvenile and adult zebrafish. Abstract Hair cells are sensory receptors responsible for transducing auditory and vestibular information into electrical signals, which are then transmitted with remarkable precision to afferent neurons. The zebrafish lateral line is emerging as an excellent in vivo model for genetic and physiological analysis of hair cells and neurons. However, research has been limited to larval stages because zebrafish become protected from the time of independent feeding under European law (from 5.2 days post‐fertilization (dpf) at 28.5°C). In larval zebrafish, the functional properties of most of hair cells, as well as those of other excitable cells, are still immature. We have developed an experimental protocol to record electrophysiological properties from hair cells of the lateral line in juvenile zebrafish. We found that the anaesthetic benzocaine at 50 mg l−1 was an effective and safe anaesthetic to use on juvenile zebrafish. Concentrations up to 300 mg l−1 did not affect the electrical properties or synaptic vesicle release of juvenile hair cells, unlike the commonly used anaesthetic MS‐222, which reduces the size of basolateral membrane K+ currents. Additionally, we implemented a method to maintain gill movement, and as such respiration and blood oxygenation, via the intubation of > 21 dpf zebrafish. The combination of benzocaine and intubation provides an experimental platform to investigate the physiology of mature hair cells from live

The circadian clock network in the brain of different Drosophila species.

PubMed

Hermann, Christiane; Saccon, Rachele; Senthilan, Pingkalai R; Domnik, Lilith; Dircksen, Heinrich; Yoshii, Taishi; Helfrich-Förster, Charlotte

2013-02-01

Comparative studies on cellular and molecular clock mechanisms have revealed striking similarities in the organization of the clocks among different animal groups. To gain evolutionary insight into the properties of the clock network within the Drosophila genus, we analyzed sequence identities and similarities of clock protein homologues and immunostained brains of 10 different Drosophila species using antibodies against vrille (VRI), PAR-protein domain1 (PDP1), and cryptochrome (CRY). We found that the clock network of both subgenera Sophophora and Drosophila consists of all lateral and dorsal clock neuron clusters that were previously described in Drosophila melanogaster. Immunostaining against CRY and the neuropeptide pigment-dispersing factor (PDF), however, revealed species-specific differences. All species of the Drosophila subgenus and D. pseudoobscura of the Sophophora subgenus completely lacked CRY in the large ventrolateral clock neurons (lLN(v) s) and showed reduced PDF immunostaining in the small ventrolateral clock neurons (sLN(v) s). In contrast, we found the expression of the ion transport peptide (ITP) to be consistent within the fifth sLN(v) and one dorsolateral clock neuron (LN(d) ) in all investigated species, suggesting a conserved putative function of this neuropeptide in the clock. We conclude that the general anatomy of the clock network is highly conserved throughout the Drosophila genus, although there is variation in PDF and CRY expression. Our comparative study is a first step toward understanding the organization of the circadian clock in Drosophila species adapted to different habitats. Copyright © 2012 Wiley Periodicals, Inc.

A review of monoaminergic neuropsychopharmacology in zebrafish.

PubMed

Maximino, Caio; Herculano, Anderson Manoel

2010-12-01

Monoamine neurotransmitters are the major regulatory mechanisms in the vertebrate brain, involved in the adjustment of motivation, emotion, and cognition. The chemical anatomy of these systems is thought to be highly conserved in the brain of all vertebrates, including zebrafish. Recently, the development of behavioral assays in zebrafish allowed the neuropsychopharmacological investigation of these circuits and its functions. Here we review neuroanatomical, genetic, neurochemical, and psychopharmacological evidence regarding the roles of histaminergic, dopaminergic, noradrenergic, serotonergic, and melatonergic systems in this species. We conclude that, in spite of species differences, zebrafish are suitable for the investigation of neuropsychopharmacology of drugs that affect theses systems; nonetheless, more thorough validation of behavioral methods is still needed.

Ground control system for the midcourse space experiment UTC clock

NASA Technical Reports Server (NTRS)

Dragonette, Richard

1994-01-01

One goal of the Midcourse Space Experiment (MSX) spacecraft Operations Planning Center is to maintain the onboard satellite UTC clock (UTC(MSX)) to within 1 millisecond of UTC(APL) (the program requirement is 10 msec). The UTC(MSX) clock employs as its time base an APL built 5 MHz quartz oscillator, which is expected to have frequency instabilities (aging rate + drift rate + frequency offset) that will cause the clock to drift approximately two to ten milliseconds per day. The UTC(MSX) clock can be advanced or retarded by the APL MSX satellite ground control center by integer multiples of 1 millisecond. The MSX Operations Planning Center is developing software which records the drift of UTC(MSX) relative to UTC(APL) and which schedules the time of day and magnitude of UTC(MSX) clock updates up to 48 hours in advance. Because of the manner in which MSX spacecraft activities are scheduled, MSX clock updates are planned 24 to 48 hours in advance, and stored in the satellite's computer controller for later execution. Data will be collected on the drift of UTC(MSX) relative to UTC(APL) over a three to five day period. Approximately six times per day, the time offset between UTC(MSX) and UTC(APL) will be measured by APL with a resolution of less than 100 microseconds. From this data a second order analytical model of the clock's drift will be derived. This model will be used to extrapolate the offset of the MSX clock in time from the present to 48 hours in the future. MSX clock updates will be placed on the spacecraft's daily schedule whenever the predicted clock offset exceeds 0.5 milliseconds. The paper includes a discussion of how the empirical model of the MSX clock is derived from satellite telemetry data, as well as the algorithm used to schedule MSX clock updates based on the model.

Iodine Clock Reaction.

ERIC Educational Resources Information Center

Mitchell, Richard S.

1996-01-01

Describes a combination of solutions that can be used in the study of kinetics using the iodine clock reaction. The combination slows down degradation of the prepared solutions and can be used successfully for several weeks. (JRH)

Transcriptional Control of Antioxidant Defense by the Circadian Clock

PubMed Central

Patel, Sonal A.; Velingkaar, Nikkhil S.

2014-01-01

Abstract Significance: The circadian clock, an internal timekeeping system, is implicated in the regulation of metabolism and physiology, and circadian dysfunctions are associated with pathological changes in model organisms and increased risk of some diseases in humans. Recent Advances: Data obtained in different organisms, including humans, have established a tight connection between the clock and cellular redox signaling making it among the major candidates for a link between the circadian system and physiological processes. Critical Issues: In spite of the recent progress in understanding the importance of the circadian clock in the regulation of reactive oxygen species homeostasis, molecular mechanisms and key regulators are mostly unknown. Future Directions: Here we review, with an emphasis on transcriptional control, the circadian-clock-dependent control of oxidative stress response system as a potential mechanism in age-associated diseases. We will discuss the roles of the core clock components such as brain and muscle ARNT-like 1, Circadian Locomotor Output Cycles Kaput, the circadian-clock-controlled transcriptional factors such as nuclear factor erythroid-2-related factor, and peroxisome proliferator-activated receptor and circadian clock control chromatin modifying enzymes from sirtuin family in the regulation of cellular and organism antioxidant defense. Antioxid. Redox Signal. 20, 2997–3006. PMID:24111970

Synchronous clock stopper for microprocessor

NASA Technical Reports Server (NTRS)

Kitchin, David A. (Inventor)

1985-01-01

A synchronous clock stopper circuit for inhibiting clock pulses to a microprocessor in response to a stop request signal, and for reinstating the clock pulses in response to a start request signal thereby to conserve power consumption of the microprocessor when used in an environment of limited power. The stopping and starting of the microprocessor is synchronized, by a phase tracker, with the occurrences of a predetermined phase in the instruction cycle of the microprocessor in which the I/O data and address lines of the microprocessor are of high impedance so that a shared memory connected to the I/O lines may be accessed by other peripheral devices. The starting and stopping occur when the microprocessor initiates and completes, respectively, an instruction, as well as before and after transferring data with a memory. Also, the phase tracker transmits phase information signals over a bus to other peripheral devices which signals identify the current operational phase of the microprocessor.

An atomic clock with 10(-18) instability.

PubMed

Hinkley, N; Sherman, J A; Phillips, N B; Schioppo, M; Lemke, N D; Beloy, K; Pizzocaro, M; Oates, C W; Ludlow, A D

2013-09-13

Atomic clocks have been instrumental in science and technology, leading to innovations such as global positioning, advanced communications, and tests of fundamental constant variation. Timekeeping precision at 1 part in 10(18) enables new timing applications in relativistic geodesy, enhanced Earth- and space-based navigation and telescopy, and new tests of physics beyond the standard model. Here, we describe the development and operation of two optical lattice clocks, both using spin-polarized, ultracold atomic ytterbium. A measurement comparing these systems demonstrates an unprecedented atomic clock instability of 1.6 × 10(-18) after only 7 hours of averaging.

Molecular Mechanisms Regulating Temperature Compensation of the Circadian Clock.

PubMed

Narasimamurthy, Rajesh; Virshup, David M

2017-01-01

An approximately 24-h biological timekeeping mechanism called the circadian clock is present in virtually all light-sensitive organisms from cyanobacteria to humans. The clock system regulates our sleep-wake cycle, feeding-fasting, hormonal secretion, body temperature, and many other physiological functions. Signals from the master circadian oscillator entrain peripheral clocks using a variety of neural and hormonal signals. Even centrally controlled internal temperature fluctuations can entrain the peripheral circadian clocks. But, unlike other chemical reactions, the output of the clock system remains nearly constant with fluctuations in ambient temperature, a phenomenon known as temperature compensation. In this brief review, we focus on recent advances in our understanding of the posttranslational modifications, especially a phosphoswitch mechanism controlling the stability of PER2 and its implications for the regulation of temperature compensation.

Understanding behavioral and physiological phenotypes of stress and anxiety in zebrafish.

PubMed

Egan, Rupert J; Bergner, Carisa L; Hart, Peter C; Cachat, Jonathan M; Canavello, Peter R; Elegante, Marco F; Elkhayat, Salem I; Bartels, Brett K; Tien, Anna K; Tien, David H; Mohnot, Sopan; Beeson, Esther; Glasgow, Eric; Amri, Hakima; Zukowska, Zofia; Kalueff, Allan V

2009-12-14

The zebrafish (Danio rerio) is emerging as a promising model organism for experimental studies of stress and anxiety. Here we further validate zebrafish models of stress by analyzing how environmental and pharmacological manipulations affect their behavioral and physiological phenotypes. Experimental manipulations included exposure to alarm pheromone, chronic exposure to fluoxetine, acute exposure to caffeine, as well as acute and chronic exposure to ethanol. Acute (but not chronic) alarm pheromone and acute caffeine produced robust anxiogenic effects, including reduced exploration, increased erratic movements and freezing behavior in zebrafish tested in the novel tank diving test. In contrast, ethanol and fluoxetine had robust anxiolytic effects, including increased exploration and reduced erratic movements. The behavior of several zebrafish strains was also quantified to ascertain differences in their behavioral profiles, revealing high-anxiety (leopard, albino) and low-anxiety (wild type) strains. We also used LocoScan (CleverSys Inc.) video-tracking tool to quantify anxiety-related behaviors in zebrafish, and dissect anxiety-related phenotypes from locomotor activity. Finally, we developed a simple and effective method of measuring zebrafish physiological stress responses (based on a human salivary cortisol assay), and showed that alterations in whole-body cortisol levels in zebrafish parallel behavioral indices of anxiety. Collectively, our results confirm zebrafish as a valid, reliable, and high-throughput model of stress and affective disorders.

Using zebrafish in systems toxicology for developmental toxicity testing.

PubMed

Nishimura, Yuhei; Inoue, Atsuto; Sasagawa, Shota; Koiwa, Junko; Kawaguchi, Koki; Kawase, Reiko; Maruyama, Toru; Kim, Soonih; Tanaka, Toshio

2016-01-01

With the high cost and the long-term assessment of developmental toxicity testing in mammals, the vertebrate zebrafish has become a useful alternative model organism for high-throughput developmental toxicity testing. Zebrafish is also very favorable for the 3R perspective in toxicology; however, the methodologies used by research groups vary greatly, posing considerable challenges to integrative analysis. In this review, we discuss zebrafish developmental toxicity testing, focusing on the methods of chemical exposure, the assessment of morphological abnormalities, housing conditions and their effects on the production of healthy embryos, and future directions. Zebrafish as a systems toxicology model has the potential to elucidate developmental toxicity pathways, and to provide a sound basis for human health risk assessments. © 2015 Japanese Teratology Society.

Social learning of an associative foraging task in zebrafish

NASA Astrophysics Data System (ADS)

Zala, Sarah M.; Määttänen, Ilmari

2013-05-01

The zebrafish ( Danio rerio) is increasingly becoming an important model species for studies on the genetic and neural mechanisms controlling behaviour and cognition. Here, we utilized a conditioned place preference (CPP) paradigm to study social learning in zebrafish. We tested whether social interactions with conditioned demonstrators enhance the ability of focal naïve individuals to learn an associative foraging task. We found that the presence of conditioned demonstrators improved focal fish foraging behaviour through the process of social transmission, whereas the presence of inexperienced demonstrators interfered with the learning of the control focal fish. Our results indicate that zebrafish use social learning for finding food and that this CPP paradigm is an efficient assay to study social learning and memory in zebrafish.

Tracking zebrafish larvae in group – Status and perspectives☆

PubMed Central

Martineau, Pierre R.; Mourrain, Philippe

2013-01-01

Video processing is increasingly becoming a standard procedure in zebrafish behavior investigations as it enables higher research throughput and new or better measures. This trend, fostered by the ever increasing performance-to-price ratio of the required recording and processing equipment, should be expected to continue in the foreseeable future, with video-processing based methods permeating more and more experiments and, as a result, expanding the very role of behavioral studies in zebrafish research. To assess whether the routine video tracking of zebrafish larvae directly in the Petri dish is a capability that can be expected in the near future, the key processing concepts are discussed and illustrated on published zebrafish studies when available or other animals when not. PMID:23707495

Resetting Biological Clocks

ERIC Educational Resources Information Center

Winfree, Arthur T.

1975-01-01

Reports on experiments conducted on two biological clocks, in organisms in the plant and animal kingdoms, which indicate that biological oscillation can be arrested by a single stimulus of a definite strength delivered at the proper time. (GS)

Temperature compensation and temperature sensation in the circadian clock

PubMed Central

Kidd, Philip B.; Young, Michael W.; Siggia, Eric D.

2015-01-01

All known circadian clocks have an endogenous period that is remarkably insensitive to temperature, a property known as temperature compensation, while at the same time being readily entrained by a diurnal temperature oscillation. Although temperature compensation and entrainment are defining features of circadian clocks, their mechanisms remain poorly understood. Most models presume that multiple steps in the circadian cycle are temperature-dependent, thus facilitating temperature entrainment, but then insist that the effect of changes around the cycle sums to zero to enforce temperature compensation. An alternative theory proposes that the circadian oscillator evolved from an adaptive temperature sensor: a gene circuit that responds only to temperature changes. This theory implies that temperature changes should linearly rescale the amplitudes of clock component oscillations but leave phase relationships and shapes unchanged. We show using timeless luciferase reporter measurements and Western blots against TIMELESS protein that this prediction is satisfied by the Drosophila circadian clock. We also review evidence for pathways that couple temperature to the circadian clock, and show previously unidentified evidence for coupling between the Drosophila clock and the heat-shock pathway. PMID:26578788

Kidney organogenesis in the zebrafish: insights into vertebrate nephrogenesis and regeneration

PubMed Central

Gerlach, Gary F.; Wingert, Rebecca A.

2012-01-01

Vertebrates form a progressive series of up to three kidney organs during development—the pronephros, mesonephros, and metanephros. Each kidney derives from the intermediate mesoderm and is comprised of conserved excretory units called nephrons. The zebrafish is a powerful model for vertebrate developmental genetics, and recent studies have illustrated that zebrafish and mammals share numerous similarities in nephron composition and physiology. The zebrafish embryo forms an architecturally simple pronephros that has two nephrons, and these eventually become a scaffold onto which a mesonephros of several hundred nephrons is constructed during larval stages. In adult zebrafish, the mesonephros exhibits ongoing nephrogenesis, generating new nephrons from a local pool of renal progenitors during periods of growth or following kidney injury. The characteristics of the zebrafish pronephros and mesonephros make them genetically tractable kidney systems in which to study the functions of renal genes and address outstanding questions about the mechanisms of nephrogenesis. Here, we provide an overview of the formation and composition of these zebrafish kidney organs, and discuss how various zebrafish mutants, gene knockdowns, and transgenic models have created frameworks in which to further delineate nephrogenesis pathways. PMID:24014448

Cardiac Ca2+ signalling in zebrafish: Translation of findings to man.

PubMed

van Opbergen, Chantal J M; van der Voorn, Stephanie M; Vos, Marc A; de Boer, Teun P; van Veen, Toon A B

2018-05-07

Sudden cardiac death is a leading cause of death worldwide, mainly caused by highly disturbed electrical activation patterns in the heart. Currently, murine models are the most popular model to study underlying molecular mechanisms of inherited or acquired cardiac electrical abnormalities, although the numerous electrophysiological discrepancies between mouse and human raise the question whether mice are the optimal model to study cardiac rhythm disorders. Recently it has been uncovered that the zebrafish cardiac electrophysiology seems surprisingly similar to the human heart, mainly because the zebrafish AP contains a clear plateau phase and ECG characteristics show alignment with the human ECG. Although, before using zebrafish as a model to study cardiac arrhythmogenesis, however, it is very important to gain a better insight into the electrophysiological characteristics of the zebrafish heart. In this review we outline the electrophysiological machinery of the zebrafish cardiomyocytes, with a special focus on the intracellular Ca 2+ dynamics and excitation-contraction coupling. We debate the potential of zebrafish as a model to study human cardiovascular diseases and postulate steps to employ zebrafish into a more 'humanized' model. Copyright © 2018 Elsevier Ltd. All rights reserved.

The ontogeny of sleep-wake cycles in zebrafish: a comparison to humans

PubMed Central

Sorribes, Amanda; Þorsteinsson, Haraldur; Arnardóttir, Hrönn; Jóhannesdóttir, Ingibjörg Þ.; Sigurgeirsson, Benjamín; de Polavieja, Gonzalo G.; Karlsson, Karl Æ.

2013-01-01

Zebrafish (Danio rerio) are used extensively in sleep research; both to further understanding of sleep in general and also as a model of human sleep. To date, sleep studies have been performed in larval and adult zebrafish but no efforts have been made to document the ontogeny of zebrafish sleep–wake cycles. Because sleep differs across phylogeny and ontogeny it is important to validate the use of zebrafish in elucidating the neural substrates of sleep. Here we describe the development of sleep and wake across the zebrafish lifespan and how it compares to humans. We find power-law distributions to best fit wake bout data but demonstrate that exponential distributions, previously used to describe sleep bout distributions, fail to adequately account for the data in either species. Regardless, the data reveal remarkable similarities in the ontogeny of sleep cycles in zebrafish and humans. Moreover, as seen in other organisms, zebrafish sleep levels are highest early in ontogeny and sleep and wake bouts gradually consolidate to form the adult sleep pattern. Finally, sleep percentage, bout duration, bout number, and sleep fragmentation are shown to allow for meaningful comparisons between zebrafish and human sleep. PMID:24312015

47 CFR 80.865 - Radiotelephone station clock.

Code of Federal Regulations, 2010 CFR

2010-10-01

....865 Telecommunication FEDERAL COMMUNICATIONS COMMISSION (CONTINUED) SAFETY AND SPECIAL RADIO SERVICES STATIONS IN THE MARITIME SERVICES Technical Equipment Requirements for Cargo Vessels Not Subject to Subpart W § 80.865 Radiotelephone station clock. A clock having a face of at least 12.7 cm (5 in.) in...

Direct Visualization of DNA Replication Dynamics in Zebrafish Cells.

PubMed

Kuriya, Kenji; Higashiyama, Eriko; Avşar-Ban, Eriko; Tamaru, Yutaka; Ogata, Shin; Takebayashi, Shin-ichiro; Ogata, Masato; Okumura, Katsuzumi

2015-12-01

Spatiotemporal regulation of DNA replication in the S-phase nucleus has been extensively studied in mammalian cells because it is tightly coupled with the regulation of other nuclear processes such as transcription. However, little is known about the replication dynamics in nonmammalian cells. Here, we analyzed the DNA replication processes of zebrafish (Danio rerio) cells through the direct visualization of replicating DNA in the nucleus and on DNA fiber molecules isolated from the nucleus. We found that zebrafish chromosomal DNA at the nuclear interior was replicated first, followed by replication of DNA at the nuclear periphery, which is reminiscent of the spatiotemporal regulation of mammalian DNA replication. However, the relative duration of interior DNA replication in zebrafish cells was longer compared to mammalian cells, possibly reflecting zebrafish-specific genomic organization. The rate of replication fork progression and ori-to-ori distance measured by the DNA combing technique were ∼ 1.4 kb/min and 100 kb, respectively, which are comparable to those in mammalian cells. To our knowledge, this is a first report that measures replication dynamics in zebrafish cells.

Myomaker mediates fusion of fast myocytes in zebrafish embryos

DOE Office of Scientific and Technical Information (OSTI.GOV)

Landemaine, Aurélie; Rescan, Pierre-Yves; Gabillard, Jean-Charles, E-mail: Jean-charles.gabillard@rennes.inra.fr

2014-09-05

Highlights: • Myomaker is transiently expressed in fast myocytes during embryonic myogenesis. • Myomaker is essential for fast myocyte fusion in zebrafish. • The function of myomaker is conserved among Teleostomi. - Abstract: Myomaker (also called Tmem8c), a new membrane activator of myocyte fusion was recently discovered in mice. Using whole mount in situ hybridization on zebrafish embryos at different stages of embryonic development, we show that myomaker is transiently expressed in fast myocytes forming the bulk of zebrafish myotome. Zebrafish embryos injected with morpholino targeted against myomaker were alive after yolk resorption and appeared morphologically normal, but they weremore » unable to swim, even under effect of a tactile stimulation. Confocal observations showed a marked phenotype characterized by the persistence of mononucleated muscle cells in the fast myotome at developmental stages where these cells normally fuse to form multinucleated myotubes. This indicates that myomaker is essential for myocyte fusion in zebrafish. Thus, there is an evolutionary conservation of myomaker expression and function among Teleostomi.« less

Cellular and molecular responses of adult zebrafish after exposure to CuO nanoparticles or ionic copper.

PubMed

Vicario-Parés, Unai; Lacave, Jose M; Reip, Paul; Cajaraville, Miren P; Orbea, Amaia

2018-01-01

Due to their antimicrobial, electrical and magnetic properties, copper nanoparticles (NPs) are suitable for a vast array of applications. Copper can be toxic to biota, making it necessary to assess the potential hazard of copper nanomaterials. Zebrafish (Danio rerio) were exposed to 10 µg Cu/L of CuO NPs of ≈100 nm (CuO-poly) or ionic copper to compare the effects provoked after 3 and 21 days of exposure and at 6 months post-exposure (mpe). At 21 days, significant copper accumulation was only detected in fish exposed to ionic copper. Exposure to both copper forms caused histopathological alterations that could reduce gill functionality, more markedly in the case of ionic copper. Nevertheless, at 6 mpe higher prevalences of gill lesions were detected in fish previously exposed to CuO-poly NPs. No relevant histological alterations were detected in liver, but the lysosomal membrane stability test showed significantly impaired general health status after exposure to both metal forms that lasted up to 6 mpe. 69 transcripts appeared regulated after 3 days of exposure to CuO-poly NPs, suggesting that NPs could produce oxidative stress and reduce metabolism and transport processes. Thirty transcripts were regulated after 21 days of exposure to ionic copper, indicating possible DNA damage. Genes of the circadian clock were identified as the key genes involved in time-dependent differences between the two copper forms. In conclusion, each copper form showed a distinct pattern of liver transcriptome regulation, but both caused gill histopathological alterations and long lasting impaired health status in adult zebrafish.

Reduced Kalman Filters for Clock Ensembles

NASA Technical Reports Server (NTRS)

Greenhall, Charles A.

2011-01-01

This paper summarizes the author's work ontimescales based on Kalman filters that act upon the clock comparisons. The natural Kalman timescale algorithm tends to optimize long-term timescale stability at the expense of short-term stability. By subjecting each post-measurement error covariance matrix to a non-transparent reduction operation, one obtains corrected clocks with improved short-term stability and little sacrifice of long-term stability.

Differential expression of neuroligin genes in the nervous system of zebrafish.

PubMed

Davey, Crystal; Tallafuss, Alexandra; Washbourne, Philip

2010-02-01

The establishment and maturation of appropriate synaptic connections is crucial in the development of neuronal circuits. Cellular adhesion is believed to play a central role in this process. Neuroligins are neuronal cell adhesion molecules that are hypothesized to act in the initial formation and maturation of synaptic connections. In order to establish the zebrafish as a model to investigate the in vivo role of Neuroligin proteins in nervous system development, we identified the zebrafish orthologs of neuroligin family members and characterized their expression. Zebrafish possess seven neuroligin genes. Synteny analysis and sequence comparisons show that NLGN2, NLGN3, and NLGN4X are duplicated in zebrafish, but NLGN1 has a single zebrafish ortholog. All seven zebrafish neuroligins are expressed in complex patterns in the developing nervous system and in the adult brain. The spatial and temporal expression patterns of these genes suggest that they occupy a role in nervous system development and maintenance.

Genetic differences in human circadian clock genes among worldwide populations.

PubMed

Ciarleglio, Christopher M; Ryckman, Kelli K; Servick, Stein V; Hida, Akiko; Robbins, Sam; Wells, Nancy; Hicks, Jennifer; Larson, Sydney A; Wiedermann, Joshua P; Carver, Krista; Hamilton, Nalo; Kidd, Kenneth K; Kidd, Judith R; Smith, Jeffrey R; Friedlaender, Jonathan; McMahon, Douglas G; Williams, Scott M; Summar, Marshall L; Johnson, Carl Hirschie

2008-08-01

The daily biological clock regulates the timing of sleep and physiological processes that are of fundamental importance to human health, performance, and well-being. Environmental parameters of relevance to biological clocks include (1) daily fluctuations in light intensity and temperature, and (2) seasonal changes in photoperiod (day length) and temperature; these parameters vary dramatically as a function of latitude and locale. In wide-ranging species other than humans, natural selection has genetically optimized adaptiveness along latitudinal clines. Is there evidence for selection of clock gene alleles along latitudinal/photoperiod clines in humans? A number of polymorphisms in the human clock genes Per2, Per3, Clock, and AANAT have been reported as alleles that could be subject to selection. In addition, this investigation discovered several novel polymorphisms in the human Arntl and Arntl2 genes that may have functional impact upon the expression of these clock transcriptional factors. The frequency distribution of these clock gene polymorphisms is reported for diverse populations of African Americans, European Americans, Ghanaians, Han Chinese, and Papua New Guineans (including 5 subpopulations within Papua New Guinea). There are significant differences in the frequency distribution of clock gene alleles among these populations. Population genetic analyses indicate that these differences are likely to arise from genetic drift rather than from natural selection.

Molecular clock integration of brown adipose tissue formation and function

PubMed Central

Nam, Deokhwa; Yechoor, Vijay K.; Ma, Ke

2016-01-01

Abstract The circadian clock is an essential time-keeping mechanism that entrains internal physiology to environmental cues. Despite the well-established link between the molecular clock and metabolic homeostasis, an intimate interplay between the clock machinery and the metabolically active brown adipose tissue (BAT) is only emerging. Recently, we came to appreciate that the formation and metabolic functions of BAT, a key organ for body temperature maintenance, are under an orchestrated circadian clock regulation. Two complementary studies from our group uncover that the cell-intrinsic clock machinery exerts concerted control of brown adipogenesis with consequent impacts on adaptive thermogenesis, which adds a previously unappreciated temporal dimension to the regulatory mechanisms governing BAT development and function. The essential clock transcriptional activator, Bmal1, suppresses adipocyte lineage commitment and differentiation, whereas the clock repressor, Rev-erbα, promotes these processes. This newly discovered temporal mechanism in fine-tuning BAT thermogenic capacity may enable energy utilization and body temperature regulation in accordance with external timing signals during development and functional recruitment. Given the important role of BAT in whole-body metabolic homeostasis, pharmacological interventions targeting the BAT-modulatory activities of the clock circuit may offer new avenues for the prevention and treatment of metabolic disorders, particularly those associated with circadian dysregulation. PMID:27385482

Clock face drawing test performance in children with ADHD.

PubMed

Ghanizadeh, Ahmad; Safavi, Salar; Berk, Michael

2013-01-01

The utility and discriminatory pattern of the clock face drawing test in ADHD is unclear. This study therefore compared Clock Face Drawing test performance in children with ADHD and controls. 95 school children with ADHD and 191 other children were matched for gender ratio and age. ADHD symptoms severities were assessed using DSM-IV ADHD checklist and their intellectual functioning was assessed. The participants completed three clock-drawing tasks, and the following four functions were assessed: Contour score, Numbers score, Hands setting score, and Center score. All the subscales scores of the three clock drawing tests of the ADHD group were lower than that of the control group. In ADHD children, inattention and hyperactivity/ impulsivity scores were not related to free drawn clock test scores. When pre-drawn contour test was performed, inattentiveness score was statistically associated with Number score while none of the other variables of age, gender, intellectual functioning, and hand use preference were associated with that kind of score. In pre-drawn clock, no association of ADHD symptoms with any CDT subscales found significant. In addition, more errors are observed with free drawn clock and Pre-drawn contour than pre-drawn clock. Putting Numbers and Hands setting are more sensitive measures to screen ADHD than Contour and Center drawing. Test performance, except Hands setting, may have already reached a developmental plateau. It is probable that Hand setting deficit in children with ADHD may not decrease from age 8 to 14 years. Performance of children with ADHD is associated with complexity of CDT.

Zebrafish: an important tool for liver disease research.

PubMed

Goessling, Wolfram; Sadler, Kirsten C

2015-11-01

As the incidence of hepatobiliary diseases increases, we must improve our understanding of the molecular, cellular, and physiological factors that contribute to the pathogenesis of liver disease. Animal models help us identify disease mechanisms that might be targeted therapeutically. Zebrafish (Danio rerio) have traditionally been used to study embryonic development but are also important to the study of liver disease. Zebrafish embryos develop rapidly; all of their digestive organs are mature in larvae by 5 days of age. At this stage, they can develop hepatobiliary diseases caused by developmental defects or toxin- or ethanol-induced injury and manifest premalignant changes within weeks. Zebrafish are similar to humans in hepatic cellular composition, function, signaling, and response to injury as well as the cellular processes that mediate liver diseases. Genes are highly conserved between humans and zebrafish, making them a useful system to study the basic mechanisms of liver disease. We can perform genetic screens to identify novel genes involved in specific disease processes and chemical screens to identify pathways and compounds that act on specific processes. We review how studies of zebrafish have advanced our understanding of inherited and acquired liver diseases as well as liver cancer and regeneration. Copyright © 2015 AGA Institute. Published by Elsevier Inc. All rights reserved.

Zebrafish: An Important Tool for Liver Disease Research

PubMed Central

Goessling, Wolfram; Sadler, Kirsten C.

2016-01-01

As the incidence of hepatobiliary diseases increases, we must improve our understanding of the molecular, cellular, and physiological factors that contribute to the pathogenesis of liver disease. Animal models help us identify disease mechanisms that might be targeted therapeutically. Zebrafish (Danio rerio) have traditionally been used to study embryonic development but are also important to the study of liver disease. Zebrafish embryos develop rapidly; all of their digestive organs are mature in larvae by 5 days of age. At this stage, they can develop hepatobiliary diseases caused by developmental defects or toxin- or ethanol-induced injury and manifest premalignant changes within weeks. Zebrafish are similar to humans in hepatic cellular composition, function, signaling, and response to injury as well as the cellular processes that mediate liver diseases. Genes are highly conserved between humans and zebrafish, making them a useful system to study the basic mechanisms of liver disease. We can perform genetic screens to identify novel genes involved in specific disease processes and chemical screens to identify pathways and compounds that act on specific processes. We review how studies of zebrafish have advanced our understanding of inherited and acquired liver diseases as well as liver cancer and regeneration. PMID:26319012

Heart-specific expression of laminopathic mutations in transgenic zebrafish.

PubMed

Verma, Ajay D; Parnaik, Veena K

2017-07-01

Lamins are key determinants of nuclear organization and function in the metazoan nucleus. Mutations in human lamin A cause a spectrum of genetic diseases that affect cardiac muscle and skeletal muscle as well as other tissues. A few laminopathies have been modeled using the mouse. As zebrafish is a well established model for the study of cardiac development and disease, we have investigated the effects of heart-specific lamin A mutations in transgenic zebrafish. We have developed transgenic lines of zebrafish expressing conserved lamin A mutations that cause cardiac dysfunction in humans. Expression of zlamin A mutations Q291P and M368K in the heart was driven by the zebrafish cardiac troponin T2 promoter. Homozygous mutant embryos displayed nuclear abnormalities in cardiomyocyte nuclei. Expression analysis showed the upregulation of genes involved in heart regeneration in transgenic mutant embryos and a cell proliferation marker was increased in adult heart tissue. At the physiological level, there was deviation of up to 20% from normal heart rate in transgenic embryos expressing mutant lamins. Adult homozygous zebrafish were fertile and did not show signs of early mortality. Our results suggest that transgenic zebrafish models of heart-specific laminopathies show cardiac regeneration and moderate deviations in heart rate during embryonic development. © 2017 International Federation for Cell Biology.

Whole-body and multispectral photoacoustic imaging of adult zebrafish

NASA Astrophysics Data System (ADS)

Huang, Na; Xi, Lei

2016-10-01

Zebrafish is a top vertebrate model to study developmental biology and genetics, and it is becoming increasingly popular for studying human diseases due to its high genome similarity to that of humans and the optical transparency in embryonic stages. However, it becomes difficult for pure optical imaging techniques to volumetric visualize the internal organs and structures of wild-type zebrafish in juvenile and adult stages with excellent resolution and penetration depth. Even with the establishment of mutant lines which remain transparent over the life cycle, it is still a challenge for pure optical imaging modalities to image the whole body of adult zebrafish with micro-scale resolution. However, the method called photoacoustic imaging that combines all the advantages of the optical imaging and ultrasonic imaging provides a new way to image the whole body of the zebrafish. In this work, we developed a non-invasive photoacoustic imaging system with optimized near-infrared illumination and cylindrical scanning to image the zebrafish. The lateral and axial resolution yield to 80 μm and 600 μm, respectively. Multispectral strategy with wavelengths from 690 nm to 930 nm was employed to image various organs inside the zebrafish. From the reconstructed images, most major organs and structures inside the body can be precisely imaged. Quantitative and statistical analysis of absorption for organs under illumination with different wavelengths were carried out.

Sleep–wake regulation and hypocretin–melatonin interaction in zebrafish

PubMed Central

Appelbaum, Lior; Wang, Gordon X.; Maro, Geraldine S.; Mori, Rotem; Tovin, Adi; Marin, Wilfredo; Yokogawa, Tohei; Kawakami, Koichi; Smith, Stephen J.; Gothilf, Yoav; Mignot, Emmanuel; Mourrain, Philippe

2009-01-01

In mammals, hypocretin/orexin (HCRT) neuropeptides are important sleep–wake regulators and HCRT deficiency causes narcolepsy. In addition to fragmented wakefulness, narcoleptic mammals also display sleep fragmentation, a less understood phenotype recapitulated in the zebrafish HCRT receptor mutant (hcrtr−/−). We therefore used zebrafish to study the potential mediators of HCRT-mediated sleep consolidation. Similar to mammals, zebrafish HCRT neurons express vesicular glutamate transporters indicating conservation of the excitatory phenotype. Visualization of the entire HCRT circuit in zebrafish stably expressing hcrt:EGFP revealed parallels with established mammalian HCRT neuroanatomy, including projections to the pineal gland, where hcrtr mRNA is expressed. As pineal-produced melatonin is a major sleep-inducing hormone in zebrafish, we further studied how the HCRT and melatonin systems interact functionally. mRNA level of arylalkylamine-N-acetyltransferase (AANAT2), a key enzyme of melatonin synthesis, is reduced in hcrtr−/− pineal gland during the night. Moreover, HCRT perfusion of cultured zebrafish pineal glands induces melatonin release. Together these data indicate that HCRT can modulate melatonin production at night. Furthermore, hcrtr−/− fish are hypersensitive to melatonin, but not other hypnotic compounds. Subthreshold doses of melatonin increased the amount of sleep and consolidated sleep in hcrtr−/− fish, but not in the wild-type siblings. These results demonstrate the existence of a functional HCRT neurons-pineal gland circuit able to modulate melatonin production and sleep consolidation. PMID:19966231

Ionic channels underlying the ventricular action potential in zebrafish embryo.

PubMed

Alday, Aintzane; Alonso, Hiart; Gallego, Monica; Urrutia, Janire; Letamendia, Ainhoa; Callol, Carles; Casis, Oscar

2014-06-01

Over the last years zebrafish has become a popular model in the study of cardiac physiology, pathology and pharmacology. Recently, the application of the 3Rs regulation and the characteristics of the embryo have reduced the use of adult zebrafish use in many studies. However, the zebrafish embryo cardiac physiology is poorly characterized since most works have used indirect techniques and direct recordings of cardiac action potential and ionic currents are scarce. In order to optimize the zebrafish embryo model, we used electrophysiological, pharmacological and immunofluorescence tools to identify the characteristics and the ionic channels involved in the ventricular action potentials of zebrafish embryos. The application of Na(+) or T-type Ca(+2) channel blockers eliminated the cardiac electrical activity, indicating that the action potential upstroke depends on Na(+) and T-type Ca(+2) currents. The plateau phase depends on L-type Ca(+2) channels since it is abolished by specific blockade. The direct channel blockade indicates that the action potential repolarization and diastolic potential depends on ERG K(+) channels. The presence in the embryonic heart of the Nav1.5, Cav1.2, Cav3.2 and ERG channels was also confirmed by immunofluorescence, while the absence of effect of specific blockers and immunostaining indicate that two K(+) repolarizing currents present in human heart, Ito and IKs, are absent in the embryonic zebrafish heart. Our results describe the ionic channels present and its role in the zebrafish embryo heart and support the use of zebrafish embryos to study human diseases and their use for drug testing. Copyright © 2014 Elsevier Ltd. All rights reserved.

The Shroud is Put Over the New Countdown Clock

NASA Image and Video Library

2014-12-09

At NASA's Kennedy Space Center in Florida, a shade is placed around the new countdown clock at the spaceport's Press Site. The modern, multimedia display is similar to the screens seen at sporting venues. The new screen is nearly 26 feet wide by 7 feet high, a foot taller than the original clock. The historic countdown clock was designed by Kennedy engineers and built by space center technicians before Apollo 12 in 1969. NASA has acquired the countdown clock from the agency’s Artifact Working Group at the agency's Headquarters for display at the Kennedy Space Center Visitor Complex.

Dynamics of statistical distance: Quantum limits for two-level clocks

DOE Office of Scientific and Technical Information (OSTI.GOV)

Braunstein, S.L.; Milburn, G.J.

1995-03-01

We study the evolution of statistical distance on the Bloch sphere under unitary and nonunitary dynamics. This corresponds to studying the limits to clock precision for a clock constructed from a two-state system. We find that the initial motion away from pure states under nonunitary dynamics yields the greatest accuracy for a one-tick'' clock; in this case the clock's precision is not limited by the largest frequency of the system.

Dihydroartemisinin promotes angiogenesis during the early embryonic development of zebrafish

PubMed Central

Ba, Qian; Duan, Juan; Tian, Jia-qiang; Wang, Zi-liang; Chen, Tao; Li, Xiao-guang; Chen, Pei-zhan; Wu, Song-jie; Xiang, Li; Li, Jing-quan; Chu, Rui-ai; Wang, Hui

2013-01-01

Aim: To investigate the embryotoxicity of dihydroartemisinin (DHA), the main active metabolite of artemisinin, in zebrafish, and explore the corresponding mechanisms. Methods: The embryos of wild type and TG (flk1:GFP) transgenic zebrafish were exposed to DHA. Developmental phenotypes of the embryos were observed. Development of blood vessels was directly observed in living embryos of TG (flk1:GFP) transgenic zebrafish under fluorescence microscope. The expression of angiogenesis marker genes vegfa, flk1, and flt1 in the embryos was detected using real-time PCR and RNA in situ hybridization assays. Results: Exposure to DHA (1–10 mg/L) dose-dependently caused abnormal zebrafish embryonic phenotypes in the early developmental stage. Furthermore, exposure to DHA (10 mg/L) resulted in more pronounced embryonic angiogenesis in TG (flk1:GFP) zebrafish line. Exposure to DHA (10 mg/L) significantly increased the mRNA expression of vegfa, flk1, and flt1 in the embryos. Knockdown of the flk1 protein partially blocked the effects of DHA on embryogenesis. Conclusion: DHA causes abnormal embryonic phenotypes and promotes angiogenesis in zebrafish early embryonic development, demonstrating the potential embryotoxicity of DHA. PMID:23708556

Cryptochrome and Period Proteins Are Regulated by the CLOCK/BMAL1 Gene: Crosstalk between the PPARs/RXRα-Regulated and CLOCK/BMAL1-Regulated Systems

PubMed Central

Nakamura, Koh-ichi; Inoue, Ikuo; Takahashi, Seiichiro; Komoda, Tsugikazu; Katayama, Shigehiro

2008-01-01

Feeding and the circadian system regulate lipid absorption and metabolism, and the expression of enzymes involved in lipid metabolism is believed to be directly controlled by the clock system. To investigate the interaction between the lipid metabolism system and the circadian system, we analyzed the effect of a CLOCK/BMAL1 heterodimer on the transcriptional regulation of PPAR-controlled genes through PPAR response elements (PPREs). Transcription of acyl-CoA oxidase, cellular retinol binding protein II (CRBPII), and 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) synthase was altered by CLOCK/BMAL1, and transcriptional activity via PPRE by PPARs/RXRα was enhanced by CLOCK/BMAL1 and/or by PPARs ligand/activators. We also found that CLOCK/BMAL1-mediated transcription of period (PER) and cryptochrome (CRY) was modulated by PPARα/RXRα. These results suggest that there may be crosstalk between the PPARs/RXRα-regulated system and the CLOCK/BMAL1-regulated system. PMID:18317514

Biologically inspired robots elicit a robust fear response in zebrafish

NASA Astrophysics Data System (ADS)

Ladu, Fabrizio; Bartolini, Tiziana; Panitz, Sarah G.; Butail, Sachit; Macrı, Simone; Porfiri, Maurizio

2015-03-01

We investigate the behavioral response of zebrafish to three fear-evoking stimuli. In a binary choice test, zebrafish are exposed to a live allopatric predator, a biologically-inspired robot, and a computer-animated image of the live predator. A target tracking algorithm is developed to score zebrafish behavior. Unlike computer-animated images, the robotic and live predator elicit a robust avoidance response. Importantly, the robotic stimulus elicits more consistent inter-individual responses than the live predator. Results from this effort are expected to aid in hypothesis-driven studies on zebrafish fear response, by offering a valuable approach to maximize data-throughput and minimize animal subjects.

Combinatorial Wnt control of zebrafish midbrain-hindbrain boundary formation.

PubMed

Buckles, Gerri R; Thorpe, Christopher J; Ramel, Marie-Christine; Lekven, Arne C

2004-05-01

Wnt signaling is known to be required for the normal development of the vertebrate midbrain and hindbrain, but genetic loss of function analyses in the mouse and zebrafish yield differing results regarding the relative importance of specific Wnt loci. In the zebrafish, Wnt1 and Wnt10b functionally overlap in their control of gene expression in the ventral midbrain-hindbrain boundary (MHB), but they are not required for the formation of the MHB constriction. Whether other wnt loci are involved in zebrafish MHB development is unclear, although the expression of at least two wnts, wnt3a and wnt8b, is maintained in wnt1/wnt10b mutants. In order to address the role of wnt3a in zebrafish, we have isolated a full length cDNA and examined its expression and function via knockdown by morpholino antisense oligonucleotide (MO)-mediated knockdown. The expression pattern of wnt3a appears to be evolutionarily conserved between zebrafish and mouse, and MO knockdown shows that Wnt3a, while not uniquely required for MHB development, is required in the absence of Wnt1 and Wnt10b for the formation of the MHB constriction. In zebrafish embryos lacking Wnt3a, Wnt1 and Wnt10b, the expression of engrailed orthologs, pax2a and fgf8 is not maintained after mid-somitogenesis. In contrast to acerebellar and no isthmus mutants, in which midbrain and hindbrain cells acquire new fates but cell number is not significantly affected until late in embryogenesis, zebrafish embryos lacking Wnt3a, Wnt1 and Wnt10b undergo extensive apoptosis in the midbrain and cerebellum anlagen beginning in mid-somitogenesis, which results in the absence of a significant portion of the midbrain and cerebellum. Thus, the requirement for Wnt signaling in forming the MHB constriction is evolutionarily conserved in vertebrates and it is possible in zebrafish to dissect the relative impact of multiple Wnt loci in midbrain and hindbrain development.

The neurogenetic frontier--lessons from misbehaving zebrafish.

PubMed

Burgess, Harold A; Granato, Michael

2008-11-01

One of the central questions in neuroscience is how refined patterns of connectivity in the brain generate and monitor behavior. Genetic mutations can influence neural circuits by disrupting differentiation or maintenance of component neuronal cells or by altering functional patterns of nervous system connectivity. Mutagenesis screens therefore have the potential to reveal not only the molecular underpinnings of brain development and function, but to illuminate the cellular basis of behavior. Practical considerations make the zebrafish an organism of choice for undertaking forward genetic analysis of behavior. The powerful array of experimental tools at the disposal of the zebrafish researcher makes it possible to link molecular function to neuronal properties that underlie behavior. This review focuses on specific challenges to isolating and analyzing behavioral mutants in zebrafish.

Automated image-based phenotypic analysis in zebrafish embryos

PubMed Central

Vogt, Andreas; Cholewinski, Andrzej; Shen, Xiaoqiang; Nelson, Scott; Lazo, John S.; Tsang, Michael; Hukriede, Neil A.

2009-01-01

Presently, the zebrafish is the only vertebrate model compatible with contemporary paradigms of drug discovery. Zebrafish embryos are amenable to automation necessary for high-throughput chemical screens, and optical transparency makes them potentially suited for image-based screening. However, the lack of tools for automated analysis of complex images presents an obstacle to utilizing the zebrafish as a high-throughput screening model. We have developed an automated system for imaging and analyzing zebrafish embryos in multi-well plates regardless of embryo orientation and without user intervention. Images of fluorescent embryos were acquired on a high-content reader and analyzed using an artificial intelligence-based image analysis method termed Cognition Network Technology (CNT). CNT reliably detected transgenic fluorescent embryos (Tg(fli1:EGFP)y1) arrayed in 96-well plates and quantified intersegmental blood vessel development in embryos treated with small molecule inhibitors of anigiogenesis. The results demonstrate it is feasible to adapt image-based high-content screening methodology to measure complex whole organism phenotypes. PMID:19235725

Functional inhibition of UQCRB suppresses angiogenesis in zebrafish

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cho, Yoon Sun; Jung, Hye Jin; Seok, Seung Hyeok

2013-04-19

Highlights: ► This is the first functional characterization of UQCRB in vivo model. ► Angiogenesis is inhibited with UQCRB loss of function in zebrafish. ► UQCRB is introduced as a prognostic marker for mitochondria- and angiogenesis-related diseases. -- Abstract: As a subunit of mitochondrial complex III, UQCRB plays an important role in complex III stability, electron transport, and cellular oxygen sensing. Herein, we report UQCRB function regarding angiogenesis in vivo with the zebrafish (Danio rerio). UQCRB knockdown inhibited angiogenesis in zebrafish leading to the suppression of VEGF expression. Moreover, the UQCRB-targeting small molecule terpestacin also inhibited angiogenesis and VEGF levelsmore » in zebrafish, supporting the role of UQCRB in angiogenesis. Collectively, UQCRB loss of function by either genetic and pharmacological means inhibited angiogenesis, indicating that UQCRB plays a key role in this process and can be a prognostic marker of angiogenesis- and mitochondria-related diseases.« less

Evolution of complexity in the zebrafish synapse proteome

PubMed Central

Bayés, Àlex; Collins, Mark O.; Reig-Viader, Rita; Gou, Gemma; Goulding, David; Izquierdo, Abril; Choudhary, Jyoti S.; Emes, Richard D.; Grant, Seth G. N.

2017-01-01

The proteome of human brain synapses is highly complex and is mutated in over 130 diseases. This complexity arose from two whole-genome duplications early in the vertebrate lineage. Zebrafish are used in modelling human diseases; however, its synapse proteome is uncharacterized, and whether the teleost-specific genome duplication (TSGD) influenced complexity is unknown. We report the characterization of the proteomes and ultrastructure of central synapses in zebrafish and analyse the importance of the TSGD. While the TSGD increases overall synapse proteome complexity, the postsynaptic density (PSD) proteome of zebrafish has lower complexity than mammals. A highly conserved set of ∼1,000 proteins is shared across vertebrates. PSD ultrastructural features are also conserved. Lineage-specific proteome differences indicate that vertebrate species evolved distinct synapse types and functions. The data sets are a resource for a wide range of studies and have important implications for the use of zebrafish in modelling human synaptic diseases. PMID:28252024

Zebrafish (Danio rerio): A Potential Model for Toxinological Studies.

PubMed

Vargas, Rafael Antonio; Sarmiento, Karen; Vásquez, Isabel Cristina

2015-10-01

Zebrafish are an emerging basic biomedical research model that has multiple advantages compared with other research models. Given that biotoxins, such as toxins, poisons, and venoms, represent health hazards to animals and humans, a low-cost biological model that is highly sensitive to biotoxins is useful to understand the damage caused by such agents and to develop biological tests to prevent and reduce the risk of poisoning in potential cases of bioterrorism or food contamination. In this article, a narrative review of the general aspects of zebrafish as a model in basic biomedical research and various studies in the field of toxinology that have used zebrafish as a biological model are presented. This information will provide useful material to beginner students and researchers who are interested in developing toxinological studies with the zebrafish model.

The zebrafish as a model for complex tissue regeneration

PubMed Central

Gemberling, Matthew; Bailey, Travis J.; Hyde, David R.; Poss, Kenneth D.

2013-01-01

For centuries, philosophers and scientists have been fascinated by the principles and implications of regeneration in lower vertebrate species. Two features have made zebrafish an informative model system for determining mechanisms of regenerative events. First, they are highly regenerative, able to regrow amputated fins, as well as a lesioned brain, retina, spinal cord, heart, and other tissues. Second, they are amenable to both forward and reverse genetic approaches, with a research toolset regularly updated by an expanding community of zebrafish researchers. Zebrafish studies have helped identify new mechanistic underpinnings of regeneration in multiple tissues, and in some cases have served as a guide for contemplating regenerative strategies in mammals. Here, we review the recent history of zebrafish as a genetic model system for understanding how and why tissue regeneration occurs. PMID:23927865

Improved Short-Term Clock Prediction Method for Real-Time Positioning.

PubMed

Lv, Yifei; Dai, Zhiqiang; Zhao, Qile; Yang, Sheng; Zhou, Jinning; Liu, Jingnan

2017-06-06

The application of real-time precise point positioning (PPP) requires real-time precise orbit and clock products that should be predicted within a short time to compensate for the communication delay or data gap. Unlike orbit correction, clock correction is difficult to model and predict. The widely used linear model hardly fits long periodic trends with a small data set and exhibits significant accuracy degradation in real-time prediction when a large data set is used. This study proposes a new prediction model for maintaining short-term satellite clocks to meet the high-precision requirements of real-time clocks and provide clock extrapolation without interrupting the real-time data stream. Fast Fourier transform (FFT) is used to analyze the linear prediction residuals of real-time clocks. The periodic terms obtained through FFT are adopted in the sliding window prediction to achieve a significant improvement in short-term prediction accuracy. This study also analyzes and compares the accuracy of short-term forecasts (less than 3 h) by using different length observations. Experimental results obtained from International GNSS Service (IGS) final products and our own real-time clocks show that the 3-h prediction accuracy is better than 0.85 ns. The new model can replace IGS ultra-rapid products in the application of real-time PPP. It is also found that there is a positive correlation between the prediction accuracy and the short-term stability of on-board clocks. Compared with the accuracy of the traditional linear model, the accuracy of the static PPP using the new model of the 2-h prediction clock in N, E, and U directions is improved by about 50%. Furthermore, the static PPP accuracy of 2-h clock products is better than 0.1 m. When an interruption occurs in the real-time model, the accuracy of the kinematic PPP solution using 1-h clock prediction product is better than 0.2 m, without significant accuracy degradation. This model is of practical significance

Husbandry stress exacerbates mycobacterial infections in adult zebrafish, Danio rerio (Hamilton)

USGS Publications Warehouse

Ramsay, J.M.; Watral, Virginia G.; Schreck, C.B.; Kent, M.L.

2009-01-01

Mycobacteria are significant pathogens of laboratory zebrafish, Danio rerio (Hamilton). Stress is often implicated in clinical disease and morbidity associated with mycobacterial infections but has yet to be examined with zebrafish. The aim of this study was to examine the effects of husbandry stressors on zebrafish infected with mycobacteria. Adult zebrafish were exposed to Mycobacterium marinum or Mycobacterium chelonae, two species that have been associated with disease in zebrafish. Infected fish and controls were then subjected to chronic crowding and handling stressors and examined over an 8-week period. Whole-body cortisol was significantly elevated in stressed fish compared to non-stressed fish. Fish infected with M. marinum ATCC 927 and subjected to husbandry stressors had 14% cumulative mortality while no mortality occurred among infected fish not subjected to husbandry stressors. Stressed fish, infected with M. chelonae H1E2 from zebrafish, were 15-fold more likely to be infected than non-stressed fish at week 8 post-injection. Sub-acute, diffuse infections were more common among stressed fish infected with M. marinum or M. chelonae than non-stressed fish. This is the first study to demonstrate an effect of stress and elevated cortisol on the morbidity, prevalence, clinical disease and histological presentation associated with mycobacterial infections in zebrafish. Minimizing husbandry stress may be effective at reducing the severity of outbreaks of clinical mycobacteriosis in zebrafish facilities. ?? 2009 Blackwell Publishing Ltd.

The Effects of Clock Drift on the Mars Exploration Rovers

NASA Technical Reports Server (NTRS)

Ali, Khaled S.; Vanelli, C. Anthony

2012-01-01

All clocks drift by some amount, and the mission clock on the Mars Exploration Rovers (MER) is no exception. The mission clock on both MER rovers drifted significantly since the rovers were launched, and it is still drifting on the Opportunity rover. The drift rate is temperature dependent. Clock drift causes problems for onboard behaviors and spacecraft operations, such as attitude estimation, driving, operation of the robotic arm, pointing for imaging, power analysis, and telecom analysis. The MER operations team has techniques to deal with some of these problems. There are a few techniques for reducing and eliminating the clock drift, but each has drawbacks. This paper presents an explanation of what is meant by clock drift on the rovers, its relationship to temperature, how we measure it, what problems it causes, how we deal with those problems, and techniques for reducing the drift.

A review of atomic clock technology, the performance capability of present spaceborne and terrestrial atomic clocks, and a look toward the future

NASA Technical Reports Server (NTRS)

Vessot, Robert F. C.

1989-01-01

Clocks have played a strong role in the development of general relativity. The concept of the proper clock is presently best realized by atomic clocks, whose development as precision instruments has evolved very rapidly in the last decades. To put a historical prospective on this progress since the year AD 1000, the time stability of various clocks expressed in terms of seconds of time error over one day of operation is shown. This stability of operation must not be confused with accuracy. Stability refers to the constancy of a clock operation as compared to that of some other clocks that serve as time references. Accuracy, on the other hand, is the ability to reproduce a previously defined frequency. The issues are outlined that must be considered when accuracy and stability of clocks and oscillators are studied. In general, the most widely used resonances result from the hyperfine interaction of the nuclear magnetic dipole moment and that of the outermost electron, which is characteristic of hydrogen and the alkali atoms. During the past decade hyperfine resonances of ions have also been used. The principal reason for both the accuracy and the stability of atomic clocks is the ability of obtaining very narrow hyperfine transition resonances by isolating the atom in some way so that only the applied stimulating microwave magnetic field is a significant source of perturbation. It is also important to make resonance transitions among hyperfine magnetic sublevels where separation is independent, at least to first order, of the magnetic field. In the case of ions stored in traps operating at high magnetic fields, one selects the trapping field to be consistent with a field-independent transition of the trapped atoms.

In vivo physiological recording from the lateral line of juvenile zebrafish.

PubMed

Olt, Jennifer; Allen, Claire E; Marcotti, Walter

2016-10-01

Zebrafish provide a unique opportunity to investigate in vivo sensory transduction in mature hair cells. We have developed a method for studying the biophysical properties of mature hair cells from the lateral line of juvenile zebrafish. The method involves application of the anaesthetic benzocaine and intubation to maintain ventilation and oxygenation through the gills. The same approach could be used for in vivo functional studies in other sensory and non-sensory systems from juvenile and adult zebrafish. Hair cells are sensory receptors responsible for transducing auditory and vestibular information into electrical signals, which are then transmitted with remarkable precision to afferent neurons. The zebrafish lateral line is emerging as an excellent in vivo model for genetic and physiological analysis of hair cells and neurons. However, research has been limited to larval stages because zebrafish become protected from the time of independent feeding under European law (from 5.2 days post-fertilization (dpf) at 28.5°C). In larval zebrafish, the functional properties of most of hair cells, as well as those of other excitable cells, are still immature. We have developed an experimental protocol to record electrophysiological properties from hair cells of the lateral line in juvenile zebrafish. We found that the anaesthetic benzocaine at 50 mg l(-1) was an effective and safe anaesthetic to use on juvenile zebrafish. Concentrations up to 300 mg l(-1) did not affect the electrical properties or synaptic vesicle release of juvenile hair cells, unlike the commonly used anaesthetic MS-222, which reduces the size of basolateral membrane K(+) currents. Additionally, we implemented a method to maintain gill movement, and as such respiration and blood oxygenation, via the intubation of > 21 dpf zebrafish. The combination of benzocaine and intubation provides an experimental platform to investigate the physiology of mature hair cells from live zebrafish. More

Zebrafish in Toxicology and Environmental Health.

PubMed

Bambino, Kathryn; Chu, Jaime

2017-01-01

As manufacturing processes and development of new synthetic compounds increase to keep pace with the expanding global demand, environmental health, and the effects of toxicant exposure are emerging as critical public health concerns. Additionally, chemicals that naturally occur in the environment, such as metals, have profound effects on human and animal health. Many of these compounds are in the news: lead, arsenic, and endocrine disruptors such as bisphenol A have all been widely publicized as causing disease or damage to humans and wildlife in recent years. Despite the widespread appreciation that environmental toxins can be harmful, there is limited understanding of how many toxins cause disease. Zebrafish are at the forefront of toxicology research; this system has been widely used as a tool to detect toxins in water samples and to investigate the mechanisms of action of environmental toxins and their related diseases. The benefits of zebrafish for studying vertebrate development are equally useful for studying teratogens. Here, we review how zebrafish are being used both to detect the presence of some toxins as well as to identify how environmental exposures affect human health and disease. We focus on areas where zebrafish have been most effectively used in ecotoxicology and in environmental health, including investigation of exposures to endocrine disruptors, industrial waste byproducts, and arsenic. © 2017 Elsevier Inc. All rights reserved.

Axonal regeneration in zebrafish spinal cord

PubMed Central

Hui, Subhra Prakash

2018-01-01

Abstract In the present review we discuss two interrelated events—axonal damage and repair—known to occur after spinal cord injury (SCI) in the zebrafish. Adult zebrafish are capable of regenerating axonal tracts and can restore full functionality after SCI. Unlike fish, axon regeneration in the adult mammalian central nervous system is extremely limited. As a consequence of an injury there is very little repair of disengaged axons and therefore functional deficit persists after SCI in adult mammals. In contrast, peripheral nervous system axons readily regenerate following injury and hence allow functional recovery both in mammals and fish. A better mechanistic understanding of these three scenarios could provide a more comprehensive insight into the success or failure of axonal regeneration after SCI. This review summarizes the present understanding of the cellular and molecular basis of axonal regeneration, in both the peripheral nervous system and the central nervous system, and large scale gene expression analysis is used to focus on different events during regeneration. The discovery and identification of genes involved in zebrafish spinal cord regeneration and subsequent functional experimentation will provide more insight into the endogenous mechanism of myelination and remyelination. Furthermore, precise knowledge of the mechanism underlying the extraordinary axonal regeneration process in zebrafish will also allow us to unravel the potential therapeutic strategies to be implemented for enhancing regrowth and remyelination of axons in mammals. PMID:29721326

Zebrafish in Toxicology and Environmental Health

PubMed Central

Bambino, Kathryn; Chu, Jaime

2018-01-01

As manufacturing processes and development of new synthetic compounds increase to keep pace with the expanding global demand, environmental health, and the effects of toxicant exposure are emerging as critical public health concerns. Additionally, chemicals that naturally occur in the environment, such as metals, have profound effects on human and animal health. Many of these compounds are in the news: lead, arsenic, and endocrine disruptors such as bisphenol A have all been widely publicized as causing disease or damage to humans and wildlife in recent years. Despite the widespread appreciation that environmental toxins can be harmful, there is limited understanding of how many toxins cause disease. Zebrafish are at the forefront of toxicology research; this system has been widely used as a tool to detect toxins in water samples and to investigate the mechanisms of action of environmental toxins and their related diseases. The benefits of zebrafish for studying vertebrate development are equally useful for studying teratogens. Here, we review how zebrafish are being used both to detect the presence of some toxins as well as to identify how environmental exposures affect human health and disease. We focus on areas where zebrafish have been most effectively used in ecotoxicology and in environmental health, including investigation of exposures to endocrine disruptors, industrial waste byproducts, and arsenic. PMID:28335863

Axonal regeneration in zebrafish spinal cord.

PubMed

Ghosh, Sukla; Hui, Subhra Prakash

2018-03-01

In the present review we discuss two interrelated events-axonal damage and repair-known to occur after spinal cord injury (SCI) in the zebrafish. Adult zebrafish are capable of regenerating axonal tracts and can restore full functionality after SCI. Unlike fish, axon regeneration in the adult mammalian central nervous system is extremely limited. As a consequence of an injury there is very little repair of disengaged axons and therefore functional deficit persists after SCI in adult mammals. In contrast, peripheral nervous system axons readily regenerate following injury and hence allow functional recovery both in mammals and fish. A better mechanistic understanding of these three scenarios could provide a more comprehensive insight into the success or failure of axonal regeneration after SCI. This review summarizes the present understanding of the cellular and molecular basis of axonal regeneration, in both the peripheral nervous system and the central nervous system, and large scale gene expression analysis is used to focus on different events during regeneration. The discovery and identification of genes involved in zebrafish spinal cord regeneration and subsequent functional experimentation will provide more insight into the endogenous mechanism of myelination and remyelination. Furthermore, precise knowledge of the mechanism underlying the extraordinary axonal regeneration process in zebrafish will also allow us to unravel the potential therapeutic strategies to be implemented for enhancing regrowth and remyelination of axons in mammals.

High-speed clock recovery unit based on a phase aligner

NASA Astrophysics Data System (ADS)

Tejera, Efrain; Esper-Chain, Roberto; Tobajas, Felix; De Armas, Valentin; Sarmiento, Roberto

2003-04-01

Nowadays clock recovery units are key elements in high speed digital communication systems. For an efficient operation, this units should generate a low jitter clock based on the NRZ received data, and be tolerant to long absence of transitions. Architectures based on Hogge phase detectors have been widely used, nevertheless, they are very sensitive to jitter of the received data and they have a limited tolerance to the absence of transitions. This paper shows a novel high speed clock recovery unit based on a phase aligner. The system allows a very fast clock recovery with a low jitter, moreover, it is very resistant to absence of transitions. The design is based on eight phases obtained from a reference clock running at the nominal frequency of the received signal. This high speed reference clock is generated using a crystal and a clock multiplier unit. The phase alignment system chooses, as starting point, the two phases closest to the data phase. This allows a maximum error of 45 degrees between the clock and data signal phases. Furthermore, the system includes a feed-back loop that interpolates the chosen phases to reduce the phase error to zero. Due to the high stability and reduced tolerance of the local reference clock, the jitter obtained is highly reduced and the system becomes able to operate under long absence of transitions. This performances make this design suitable for systems such as high speed serial link technologies. This system has been designed for CMOS 0.25μm at 1.25GHz and has been verified through HSpice simulations.

Machine Learning Helps Identify CHRONO as a Circadian Clock Component

PubMed Central

Venkataraman, Anand; Ramanathan, Chidambaram; Kavakli, Ibrahim H.; Hughes, Michael E.; Baggs, Julie E.; Growe, Jacqueline; Liu, Andrew C.; Kim, Junhyong; Hogenesch, John B.

2014-01-01

Over the last decades, researchers have characterized a set of “clock genes” that drive daily rhythms in physiology and behavior. This arduous work has yielded results with far-reaching consequences in metabolic, psychiatric, and neoplastic disorders. Recent attempts to expand our understanding of circadian regulation have moved beyond the mutagenesis screens that identified the first clock components, employing higher throughput genomic and proteomic techniques. In order to further accelerate clock gene discovery, we utilized a computer-assisted approach to identify and prioritize candidate clock components. We used a simple form of probabilistic machine learning to integrate biologically relevant, genome-scale data and ranked genes on their similarity to known clock components. We then used a secondary experimental screen to characterize the top candidates. We found that several physically interact with known clock components in a mammalian two-hybrid screen and modulate in vitro cellular rhythms in an immortalized mouse fibroblast line (NIH 3T3). One candidate, Gene Model 129, interacts with BMAL1 and functionally represses the key driver of molecular rhythms, the BMAL1/CLOCK transcriptional complex. Given these results, we have renamed the gene CHRONO (computationally highlighted repressor of the network oscillator). Bi-molecular fluorescence complementation and co-immunoprecipitation demonstrate that CHRONO represses by abrogating the binding of BMAL1 to its transcriptional co-activator CBP. Most importantly, CHRONO knockout mice display a prolonged free-running circadian period similar to, or more drastic than, six other clock components. We conclude that CHRONO is a functional clock component providing a new layer of control on circadian molecular dynamics. PMID:24737000

Quantification of birefringence readily measures the level of muscle damage in zebrafish

DOE Office of Scientific and Technical Information (OSTI.GOV)

Berger, Joachim, E-mail: Joachim.Berger@Monash.edu; Sztal, Tamar; Currie, Peter D.

2012-07-13

Highlights: Black-Right-Pointing-Pointer Report of an unbiased quantification of the birefringence of muscle of fish larvae. Black-Right-Pointing-Pointer Quantification method readily identifies level of overall muscle damage. Black-Right-Pointing-Pointer Compare zebrafish muscle mutants for level of phenotype severity. Black-Right-Pointing-Pointer Proposed tool to survey treatments that aim to ameliorate muscular dystrophy. -- Abstract: Muscular dystrophies are a group of genetic disorders that progressively weaken and degenerate muscle. Many zebrafish models for human muscular dystrophies have been generated and analysed, including dystrophin-deficient zebrafish mutants dmd that model Duchenne Muscular Dystrophy. Under polarised light the zebrafish muscle can be detected as a bright area in anmore » otherwise dark background. This light effect, called birefringence, results from the diffraction of polarised light through the pseudo-crystalline array of the muscle sarcomeres. Muscle damage, as seen in zebrafish models for muscular dystrophies, can readily be detected by a reduction in the birefringence. Therefore, birefringence is a very sensitive indicator of overall muscle integrity within larval zebrafish. Unbiased documentation of the birefringence followed by densitometric measurement enables the quantification of the birefringence of zebrafish larvae. Thereby, the overall level of muscle integrity can be detected, allowing the identification and categorisation of zebrafish muscle mutants. In addition, we propose that the establish protocol can be used to analyse treatments aimed at ameliorating dystrophic zebrafish models.« less

[Effect of rhynchophylline on behaviors of methamphetamine-dependent zebrafish and the mechanism].

PubMed

Chen, Yi-Fei; Peng, Ju; Fang, Miao; Liu, Yi; Nie, Ling-Hui; Mo, Zhi-Xian; Zhu, Ling-Ling

2016-11-20

To observe the effect of rhynchophylline on methamphetamine-dependent zebrafish and explore the possible mechanism. Zebrafish were divided into control group, amphetamine group, low- (50 mg/kg) and high (100 mg/kg)-dose rhynchophylline groups, and ketamine (150 mg/kg) group. Conditioned place preference (CPP) was induced in zebrafish with methamphetamine, and the staying time in the drug box and the tracking map of the zebrafish were observed with Noldus Ethovision XT system. The protein expressions of TH, NR2B and GLUR2 in the brain of zebrafish with CPP were detected with Western blotting. Compared with the control group, zebrafish in methamphetamine group showed significant variations in the staying time and swimming distance in the drug box after conditioning (P<0.05) with obvious alterations of NR2B, TH and GLUR2 expressions in the brain (P<0.05). Treatment of methamphetamine-dependent zebrafish with high-dose rhynchophylline significantly reduced the variations in the staying time and swimming distance in the drug box (P<0.05) and in the expressions of NR2B, TH and GLUR2 in the brain (P<0.05). Rhynchophylline can inhibit methamphetamine dependence in zebrafish, the mechanism of which may involve the expressions of TH, NR2B and GLUR2 proteins in the brain.

Normal vision can compensate for the loss of the circadian clock

PubMed Central

Schlichting, Matthias; Menegazzi, Pamela; Helfrich-Förster, Charlotte

2015-01-01

Circadian clocks are thought to be essential for timing the daily activity of animals, and consequently increase fitness. This view was recently challenged for clock-less fruit flies and mice that exhibited astonishingly normal activity rhythms under outdoor conditions. Compensatory mechanisms appear to enable even clock mutants to live a normal life in nature. Here, we show that gradual daily increases/decreases of light in the laboratory suffice to provoke normally timed sharp morning (M) and evening (E) activity peaks in clock-less flies. We also show that the compound eyes, but not Cryptochrome (CRY), mediate the precise timing of M and E peaks under natural-like conditions, as CRY-less flies do and eyeless flies do not show these sharp peaks independently of a functional clock. Nevertheless, the circadian clock appears critical for anticipating dusk, as well as for inhibiting sharp activity peaks during midnight. Clock-less flies only increase E activity after dusk and not before the beginning of dusk, and respond strongly to twilight exposure in the middle of the night. Furthermore, the circadian clock responds to natural-like light cycles, by slightly broadening Timeless (TIM) abundance in the clock neurons, and this effect is mediated by CRY. PMID:26378222

Drosophila Ionotropic Receptor 25a mediates circadian clock resetting by temperature.

PubMed

Chen, Chenghao; Buhl, Edgar; Xu, Min; Croset, Vincent; Rees, Johanna S; Lilley, Kathryn S; Benton, Richard; Hodge, James J L; Stanewsky, Ralf

2015-11-26

Circadian clocks are endogenous timers adjusting behaviour and physiology with the solar day. Synchronized circadian clocks improve fitness and are crucial for our physical and mental well-being. Visual and non-visual photoreceptors are responsible for synchronizing circadian clocks to light, but clock-resetting is also achieved by alternating day and night temperatures with only 2-4 °C difference. This temperature sensitivity is remarkable considering that the circadian clock period (~24 h) is largely independent of surrounding ambient temperatures. Here we show that Drosophila Ionotropic Receptor 25a (IR25a) is required for behavioural synchronization to low-amplitude temperature cycles. This channel is expressed in sensory neurons of internal stretch receptors previously implicated in temperature synchronization of the circadian clock. IR25a is required for temperature-synchronized clock protein oscillations in subsets of central clock neurons. Extracellular leg nerve recordings reveal temperature- and IR25a-dependent sensory responses, and IR25a misexpression confers temperature-dependent firing of heterologous neurons. We propose that IR25a is part of an input pathway to the circadian clock that detects small temperature differences. This pathway operates in the absence of known 'hot' and 'cold' sensors in the Drosophila antenna, revealing the existence of novel periphery-to-brain temperature signalling channels.

Initial atomic coherences and Ramsey frequency pulling in fountain clocks

NASA Astrophysics Data System (ADS)

Gerginov, Vladislav; Nemitz, Nils; Weyers, Stefan

2014-09-01

In the uncertainty budget of primary atomic cesium fountain clocks, evaluations of frequency-pulling shifts of the hyperfine clock transition caused by unintentional excitation of its nearby transitions (Rabi and Ramsey pulling) have been based so far on an approach developed for cesium beam clocks. We re-evaluate this type of frequency pulling in fountain clocks and pay particular attention to the effect of initial coherent atomic states. We find significantly enhanced frequency shifts caused by Ramsey pulling due to sublevel population imbalance and corresponding coherences within the state-selected hyperfine component of the initial atom ground state. Such shifts are experimentally investigated in an atomic fountain clock and quantitative agreement with the predictions of the model is demonstrated.

Oceans of Opportunity: Exploring Vertebrate Hematopoiesis in Zebrafish

PubMed Central

Carroll, Kelli J.; North, Trista E.

2015-01-01

Exploitation of the zebrafish model in hematology research has surged in recent years, becoming one of the most useful and tractable systems for understanding regulation of hematopoietic development, homeostasis, and malignancy. Despite the evolutionary distance between zebrafish and humans, remarkable genetic and phenotypic conservation in the hematopoietic system has enabled significant advancements in our understanding of blood stem and progenitor cell (HSPC) biology. The strengths of zebrafish in hematology research lie in the ability to perform real-time in vivo observations of hematopoietic stem, progenitor and effector cell emergence, expansion and function, as well as the ease with which novel genetic and chemical modifiers of specific hematopoietic processes or cell-types can be identified and characterized. Further, a myriad of transgenic lines have been developed including fluorescent reporter systems to aid in the visualization and quantification of specified cell types of interest and cell-lineage relationships, as well as effector lines that can be used to implement a wide range of experimental manipulations. As our understanding of the complex nature of HSPC biology during development, in response to infection or injury, or in the setting of hematological malignancy, continues to deepen, zebrafish will remain essential for exploring the spatio-temporal organization and integration of these fundamental processes, as well as the identification of efficacious small molecule modifiers of hematopoietic activity. In this review, we discuss the biology of the zebrafish hematopoietic system, including similarities and differences from mammals, and highlight important tools currently utilized in zebrafish embryos and adults to enhance our understanding of vertebrate hematology, with emphasis on findings that have impacted our understanding of the onset or treatment of human hematologic disorders and disease. PMID:24816275

BMP signaling modulates hepcidin expression in zebrafish embryos independent of hemojuvelin.

PubMed

Gibert, Yann; Lattanzi, Victoria J; Zhen, Aileen W; Vedder, Lea; Brunet, Frédéric; Faasse, Sarah A; Babitt, Jodie L; Lin, Herbert Y; Hammerschmidt, Matthias; Fraenkel, Paula G

2011-01-21

Hemojuvelin (Hjv), a member of the repulsive-guidance molecule (RGM) family, upregulates transcription of the iron regulatory hormone hepcidin by activating the bone morphogenetic protein (BMP) signaling pathway in mammalian cells. Mammalian models have identified furin, neogenin, and matriptase-2 as modifiers of Hjv's function. Using the zebrafish model, we evaluated the effects of hjv and its interacting proteins on hepcidin expression during embryonic development. We found that hjv is strongly expressed in the notochord and somites of the zebrafish embryo and that morpholino knockdown of hjv impaired the development of these structures. Knockdown of hjv or other hjv-related genes, including zebrafish orthologs of furin or neogenin, however, failed to decrease hepcidin expression relative to liver size. In contrast, overexpression of bmp2b or knockdown of matriptase-2 enhanced the intensity and extent of hepcidin expression in zebrafish embryos, but this occurred in an hjv-independent manner. Furthermore, we demonstrated that zebrafish hjv can activate the human hepcidin promoter and enhance BMP responsive gene expression in vitro, but is expressed at low levels in the zebrafish embryonic liver. Taken together, these data support an alternative mechanism for hepcidin regulation during zebrafish embryonic development, which is independent of hjv.

The HDAC Inhibitor TSA Ameliorates a Zebrafish Model of Duchenne Muscular Dystrophy.

PubMed

Johnson, Nathan M; Farr, Gist H; Maves, Lisa

2013-09-17

Zebrafish are an excellent model for Duchenne muscular dystrophy. In particular, zebrafish provide a system for rapid, easy, and low-cost screening of small molecules that can ameliorate muscle damage in dystrophic larvae. Here we identify an optimal anti-sense morpholino cocktail that robustly knocks down zebrafish Dystrophin (dmd-MO). We use two approaches, muscle birefringence and muscle actin expression, to quantify muscle damage and show that the dmd-MO dystrophic phenotype closely resembles the zebrafish dmd mutant phenotype. We then show that the histone deacetylase (HDAC) inhibitor TSA, which has been shown to ameliorate the mdx mouse Duchenne model, can rescue muscle fiber damage in both dmd-MO and dmd mutant larvae. Our study identifies optimal morpholino and phenotypic scoring approaches for dystrophic zebrafish, further enhancing the zebrafish dmd model for rapid and cost-effective small molecule screening.

The Central Clock Neurons Regulate Lipid Storage in Drosophila

PubMed Central

DiAngelo, Justin R.; Erion, Renske; Crocker, Amanda; Sehgal, Amita