Name: use of the teton system to study molecular mechanisms of zebrafish regeneration
Uploaded: 2015-06-25T13:00:00.000+00:00
Duration: 10 min 22 s
Description: Watch this Scientific Journal Video about Use of the TetON System to Study Molecular Mechanisms of Zebrafish Regeneration at JoVE.com

Die Autoren danken Christa Haase, Doris Weber und Brigitte Korte für die technische Unterstützung. Arbeit im Labor Weidinger durch Zuschüsse von der Deutschen Forschungsgemeinschaft WE 4223 / 3-1 unterstützt, WE 4223 / 4-1 und von der Deutschen Gesellschaft für Kardiologie über eine Oskar-Lapp-Stipendium und ein Klaus-Georg-und-Sigrid- Hengstberger-Forschungsstipendium.

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S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Method+for+isolation+of+PCR-ready+genomic+DNA+from+zebrafish+tissues.">Method for isolation of PCR-ready genomic DNA from zebrafish tissues.</a> Biotechniques. 43 (5), 610 (2007).</li><li>Thermes, V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=I-SceI+meganuclease+mediates+highly+efficient+transgenesis+in+fish.">I-SceI meganuclease mediates highly efficient transgenesis in fish.</a> Mech Dev. 118 (1-2), 91-98 (2002).</li><li>Suster, M. L., Kikuta, H., Urasaki, A., Asakawa, K., Kawakami, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transgenesis+in+zebrafish+with+the+tol2+transposon+system.">Transgenesis in zebrafish with the tol2 transposon system.</a> Methods Mol Biol. , 56141-56163 (2009).</li><li>Grabher, C., Wittbrodt, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Meganuclease+and+transposon+mediated+transgenesis+in+medaka.">Meganuclease and transposon mediated transgenesis in medaka.</a> Genome Biol. 8, (2007).</li></ol>

Die Autoren haben nichts zu offenbaren.

Die erwachsenen Zebrafisch hat eine erstaunliche Fähigkeit, viele der inneren Organe und Anhänge erfolgreich zu regenerieren. Ein gründliches Verständnis der molekularen und zellulären Mechanismen erfordert gewebespezifische Analyse von Gen-Funktionen und Signalwege. Auf dem Weg zu diesem bietet die Teton-System ein effizientes Werkzeug für die raumzeitlich gesteuerte Genexpression in embryonalen und erwachsenen Zebrafisch. Die Teton System Konstrukte und Methoden in dieser Handschrift beschrieben wurden erfolgrei...

Der Zebrafisch ist ein gut etabliertes Wirbelmodellorganismus, um viele Aspekte der Entwicklung, der Physiologie, Krankheiten und Regeneration zu studieren. Mit der wachsenden Annahme der Zebrafisch als Modell für die post-embryonale biologische Prozesse, experimenteller Werkzeuge zur induzierbaren, gewebsspezifischen Manipulation der Genexpression oder Signalwege immer wichtiger geworden. Besonders haben Studien in Orgel und Anhängsel Regeneration im erwachsenen Zebrafisch aus einem Mangel an Tools für die Präparation der raum-zeitlichen Anforderungen von Signalwegen in diesen Regenerationsprozesse erlitten. Derzeit sind drei verschiedene Systeme verwendet worden, um bedingte, gewebespezifische Genexpression in regenerierenden Organen erwachsenen Zebrafisch zu erzielen: das Cre-lox-System, Mosaik Expression von Hitzeschock induzierbares Transposon-Transgenen mit vermittelte somatischen Transgenität und die TetOn System 1 -3. TetOn bezieht sich auf eine Variante eines Tetracyclin-contgewalzten Transkriptionsaktivierungssystems, wobei die Expression in Gegenwart des Antibiotikums Tetracyclin oder einem Derivat, beispielsweise Doxycyclin aktiviert. Die Cre-lox-System, wie es bisher in ausgewachsene Fische verwendet wurde, stützt sich auf eine Tamoxifen-gesteuert Cre-Rekombinase (CreERT2), deren Expression räumlich durch gewebespezifische regulatorische Elemente beschränkt. Cre gesteuerte Entnahme einer Stop-Kassette ermöglicht die Expression des Gens von Interesse von einem Promotor, der in allen Zelltypen 1 aktiv sein soll angetrieben. Transposon-vermittelte Erzeugung von mosaik ausgedrückt somatischen Transgenen stellt ein System für die induzierbare Expression des Transgens in den einzelnen Zelllinien. Die Injektion von Zebrafischembryonen mit einem Tol2 Transposon trägt ein Gen von Interesse unter der Transkriptionskontrolle eines Hitzeschockpromotor Ergebnisse in chimären Individuen in der Regel trägt das Transgen nur in diskreten Zelllinien aus einer regenerierenden Orgel 2. Während beide Systeme ermöglichen bedingte Gewebe spesche Genexpression ist das Cre-lox-System nicht reversibel, und die Strategie mit einem Transposon-beruhenden klonalen Kennzeichnung leidet ihre stochastische Natur. So haben wir und andere kürzlich transgenen Teton Systeme für den Einsatz in Zebrafisch, der zeitlich und räumlich gesteuerte Genexpression, die zusätzlich abstimmbaren und reversible 3-5 ist zu erleichtern angepasst. Die Teton System besteht hier aus einem transgenen Fahrer (TetActivator), in dem ein Doxycyclin (DOX) -induzierbaren Transkriptionsaktivator (verbesserte Umkehr Tetracyclin-Transaktivator, irtTA, kurze TETA) ist unter Kontrolle von gewebespezifischen genomischen regulatorischen Sequenzen. Zweitens bedarf es einer transgenen Responderlinie (TetResponder), die ein Gen von Interesse unter der Transkriptionskontrolle des Tetracyclin-Operator Häfen (Tet-Response-Element; Tetre) (Abbildung 1A). Somit ermöglicht die Verwendung von spezifischen Kombinationen von TetActivator und TetResponder Leitungen für bedingten gewebespezifischenFIC Manipulation der Genexpression. Wir haben vor kurzem nutzte die Teton-System für gewebespezifische Funktionen des Wnt / beta-Catenin-Signalweg in der erwachsenen Zebrafisch Regenerieren Schwanzflosse 3 zu untersuchen. In dem Protokoll hier skizzierten beschreiben wir einen Workflow für Aufbau und Nutzung der Teton-System in Zebrafisch, insbesondere für Studien der fin Regeneration. Dieses enthält detaillierte Anweisungen zum stabilen transgenen TetActivator und TetResponder Linien zu erzeugen, und ein Protokoll für die Induktion Transgen in Embryonen und erwachsenen Zebrafisch. Ferner beschreiben wir Verfahren zur Überprüfung der gewebespezifischen Genexpression in der Rippe Regenerat, einschließlich eines Protokolls zur Herstellung von Gefrierschnitten von adulten Zebrafischflossen. Zusätzlich diskutieren wir Überlegungen für den Entwurf des TetActivator Transgen, die Wahl eines Transgeneseverfahren und Nachweis von TetResponder Ausdruck. Daher ist das übergeordnete Ziel dieses Protokolls als Blaupause dienenfür den Aufbau einer funktionellen Teton Systems im Zebrafisch zu bedingten gewebespezifische Genexpression, die zu jedem Gewebe von Interesse angewendet werden kann, zu erreichen. Wir haben eine TetActivator der Vektor ermöglicht für I-SceI oder Tol2 vermittelte Erzeugung stabiler TetActivator Linien mit kurzen genomischen regulatorischen Sequenzen erzeugt (Enhancer Fragmente; Weidinger lab Plasmid Datenbank nicht 1247;. 1B). Dieses Konstrukt enthält eine TetActivator Kassette, bestehend aus dem M2 mutierten Variante der reversen Tet-Repressor-Domäne mit dem Herpes simplex-Virus VP16-Transaktivierungsdomäne-Derivat 3F [irtTAM2 (3F)] fusioniert. Expression des TetActivator (TETA) kann leicht überwacht werden, da es coexprimiert mit dem Fluorophor AmCyan aus demselben offenen Leserahmen; a p2a Peptid vermittelt ribosomalen Skipping, die bei der Herstellung von TETA und AmCyan als separate Proteine ​​bei einer 1 führen sollte: 1-Verhältnis 5,6. Das Konstrukt enthält auch eine poylinker 5 &#39;der TetActivator Kassette, um die Einführung der genomischen regulatorischen Sequenzen von Interesse unter Verwendung herkömmlicher Klonierungsverfahren zu erleichtern. , Der in ein bakterielles künstliches Chromosom rekombiniert werden können (BAC), die eine große genomische Region (; Zusätzlich haben wir ein Konstrukt, bestehend aus dem oben beschriebenen Kassetten TetActivator plus Kanamycin-Selektionskassette (. 1C Weidinger lab Plasmid Datenbank nicht 1180) erstellt in der Regel in dem Startcodon eines Gens, dessen Expressionsmuster ist durch das Transgen nachgeahmt werden). Beide Konstrukte sind von der Weidinger Labor auf Anfrage erhältlich.

<table><tbody><tr><td>Breeding boxes</td><td>Aqua Schwarz</td><td>AquaBox 1</td><td></td></tr><tr><td>Compound fluorescent microscope</td><td>e.g., Leica, Zeiss</td><td>varies with the manufacturer</td><td>to image fluorescent tissue sections</td></tr><tr><td>Confocal microscope</td><td>e.g., Leica, Zeiss</td><td>varies with the manufacturer</td><td>to image fluorescent tissue sections</td></tr><tr><td>Cryostat</td><td>e.g., Leica, Thermo-Scientific</td><td>varies with the manufacturer</td><td>for cryosectioning</td></tr><tr><td>4&amp;rsquo;, 6- diamidino-2-phenylinodole (Dapi)</td><td>Sigma-Aldrich</td><td>D9542</td><td>use 1/5,000 in PBS&lt;br/&gt; for visualization of nuclei</td></tr><tr><td>Doxycycline</td><td>Sigma-Aldrich</td><td>D9891</td><td>prepare stocks in 50% EtOH at 50 mg/ml (97 mM)&lt;br/&gt; for TetResponder induction</td></tr><tr><td>Paraformaldehyde (PFA)</td><td>Sigma-Aldrich</td><td>P6148</td><td>4% (w/v) paraformaldehyde in PBS, pH 7.5&lt;br/&gt; for fixation</td></tr><tr><td>1x Phosphat-buffer saline (PBS)</td><td></td><td></td><td>1.7 mM KH&lt;sub&gt;2&lt;/sub&gt;PO&lt;sub&gt;4&lt;/sub&gt;, 5.2 mM Na&lt;sub&gt;2&lt;/sub&gt;HPO&lt;sub&gt;4&lt;/sub&gt;, 150 mM NaCl, pH 7.5</td></tr><tr><td>1x Phosphat-buffer saline + Tween 20 (PBT)</td><td></td><td></td><td>1x PBS with 0.1% Tween 20</td></tr><tr><td>Superfrost Ultra Plus adhesion microscope slides</td><td>Thermo Scientific&amp;nbsp;</td><td>1014356190</td><td>for collection of tissue sections</td></tr><tr><td>Stereo fluorescent microscope</td><td>e.g., Leica, Zeiss</td><td>varies with the manufacturer</td><td>for fluorescence-based genotyping</td></tr><tr><td>Thermocycler</td><td>e.g., Biorad, Applied Biosystems</td><td>varies with the manufacturer</td><td>for PCR-based genotyping</td></tr><tr><td>Tissue freezing medium (TFM)</td><td>Triangel Biomedical Sciences</td><td>TFM-C</td><td>for embedding of tissue samples</td></tr><tr><td>Tricaine (L-Ethyl-m-amino-benzoate-methane sulfonate/MS-222)</td><td>Sigma-Aldrich</td><td>E10521</td><td>for anesthesia&lt;br/&gt; use at 1 mg/ml in E3 embryo medium</td></tr></tbody></table>

use of the teton system to study molecular mechanisms of zebrafish regeneration

The zebrafish has become a very important model organism for studying vertebrate development, physiology, disease, and tissue regeneration. A thorough understanding of the molecular and cellular mechanisms involved requires experimental tools that allow for inducible, tissue-specific manipulation of gene expression or signaling pathways. Therefore, we and others have recently adapted the TetON system for use in zebrafish. The TetON system facilitates temporally and spatially-controlled gene expression and we have recently used this tool to probe for tissue-specific functions of Wnt/beta&#8211;catenin signaling during zebrafish tail fin regeneration. Here we describe the workflow for using the TetON system to achieve inducible, tissue-specific gene expression in the adult regenerating zebrafish tail fin. This includes the generation of stable transgenic TetActivator and TetResponder lines, transgene induction and techniques for verification of tissue-specific gene expression in the fin regenerate. Thus, this protocol serves as blueprint for setting up a functional TetON system in zebrafish and its subsequent use, in particular for studying fin regeneration.

Um eine Funktions Teton System für gewebespezifische induzierbaren Genexpression, transgene TetActivator und TetResponder Linien etablieren müssen generiert werden (Abbildung 1A). Dies wird durch Mikroinjektion TetActivator (1B - C) erreicht oder TetResponder (1E) Konstrukte in frühen Zebrafischembryonen und anschließender Keimbahn-Integration. Funktions TetActivator Konstrukte können entweder durch Klonen von kurzen Regulationssequenzen (Enhancer-...

Watch this Scientific Journal Video about Use of the TetON System to Study Molecular Mechanisms of Zebrafish Regeneration at JoVE.com

Use of the TetON System to Study Molecular Mechanisms of Zebrafish Regeneration

Here we outline the workflow for using the TetON system to achieve tissue-specific gene expression in the adult regenerating zebrafish tail fin.

Die Nutzung der Teton-System, um molekulare Mechanismen bei Zebrafisch Regeneration Studieren

The zebrafish has become a very important model organism for studying vertebrate development, physiology, disease, and tissue regeneration. A thorough ...

use-teton-system-to-study-molecular-mechanisms-zebrafish

Research

JoVE Journal

Developmental Biology

10.6K Aufrufe.  Ulm University. Here we outline the workflow for using the TetON system to achieve tissue-specific gene expression in the adult regenerating zebrafish tail fin.

Serie: Use of the TetON System to Study Molecular Mechanisms of Zebrafish Regeneration

Kidney Regeneration in Adult Zebrafish by Gentamicin Induced Injury

The kidney is essential for fluid homeostasis, blood pressure regulation and filtration of waste from the body. The fundamental unit of kidney function is the nephron. Mammals are able to repair existing nephrons after injury, but lose the ability to form new nephrons soon after birth. In contrast to mammals, adult fish produce new nephrons (neonephrogenesis) throughout their lives in response to growth requirements or injury. Recently, lhx1a has been shown to mark nephron progenitor cells in the adult zebrafish kidney, however mechanisms controlling the formation of new nephrons after injury remain unknown. Here we show our method for robust and reproducible injury in the adult zebrafish kidney by intraperitoneal (i.p.) injection of gentamicin, which uses a noninvasive visual screening process to select for fish with strong but nonlethal injury. Using this method, we can determine optimal gentamicin dosages for injury and go on to demonstrate the effect of higher temperatures on kidney regeneration in zebrafish.

Here we present a reliable method to study adult kidney regeneration by inducing acute kidney injury by gentamicin injection. We show that injury is dependent on gentamicin dosage and environmental temperature using in situ hybridization to label lhx1a+ developing new nephrons.

Nieren-Regeneration im erwachsenen Zebrafisch durch Gentamicin induzierte Schädigung

The kidney is essential for fluid homeostasis, blood pressure regulation and filtration of waste from the body. The fundamental unit of kidney function is ...

Forschung

Entwicklungsbiologie

Capturing Tissue Repair in Zebrafish Larvae with Time-lapse Brightfield Stereomicroscopy

The zebrafish larval tail fin is ideal for studying tissue regeneration due to the simple architecture of the larval fin-fold, which comprises of two layers of skin that enclose undifferentiated mesenchyme, and because the larval tail fin regenerates rapidly within 2-3 days. Using this system, we demonstrate a method for capturing the repair dynamics of the amputated tail fin with time-lapse video brightfield stereomicroscopy. We demonstrate that fin amputation triggers a contraction of the amputation wound and extrusion of cells around the wound margin, leading to their subsequent clearance. Fin regeneration proceeds from proximal to distal direction after a short delay. In addition, developmental growth of the larva can be observed during all stages. The presented method provides an opportunity for observing and analyzing whole tissue-scale behaviors such as fin development and growth in a simple microscope setting, which is easily adaptable to any stereomicroscope with time-lapse capabilities.

We present a protocol for capturing the dynamics of zebrafish larval tail fin regeneration on a whole-tissue scale using brightfield-based stereomicroscopy. This technique enables capturing the regeneration dynamics with single cell resolution. This methodology can be adapted to any stereomicroscope equipped with a CCD camera and time-lapse software.

Erfassen von Gewebereparatur in Zebrafisch-Larven mit Zeitrafferhellstereomikroskopie

The zebrafish larval tail fin is ideal for studying tissue regeneration due to the simple architecture of the larval fin-fold, which comprises of two ...

Nanoparticle-mediated siRNA Gene-silencing in Adult Zebrafish Heart

Mammals have a very limited capacity to regenerate the heart after myocardial infarction. On the other hand, the adult zebrafish regenerates its heart after apex resection or cryoinjury, making it an important model organism for heart regeneration study. However, the lack of loss-of-function methods for adult organs has restricted insights into the mechanisms underlying heart regeneration. RNA interference via different delivery systems is a powerful tool for silencing genes in mammalian cells and model organisms. We have previously reported that siRNA-encapsulated nanoparticles successfully enter cells and result in a remarkable gene-specific knockdown in the regenerating adult zebrafish heart. Here, we present a simple, rapid, and efficient protocol for the dendrimer-mediated siRNA delivery and gene-silencing in the regenerating adult zebrafish heart. This method provides an alternative approach for determining gene functions in adult organs in zebrafish and can be extended to other model organisms as well.

It remains a major challenge to develop conditional gene-knockout or effective gene-knockdown in adult zebrafish organs. Here we report a protocol for performing nanoparticle-mediated siRNA gene-silencing in adult zebrafish heart, thus providing a new loss-of-function method for studying adult organs in zebrafish and other model organisms.

Nanopartikel-vermittelten SiRNA Gen-silencing in Erwachsenen Zebrafisch-Herz

Mammals have a very limited capacity to regenerate the heart after myocardial infarction. On the other hand, the adult zebrafish regenerates its heart ...

Spinal Cord Transection in the Larval Zebrafish

Mammals fail in sensory and motor recovery following spinal cord injury due to lack of axonal regrowth below the level of injury as well as an inability to reinitiate spinal neurogenesis. However, some anamniotes including the zebrafish Danio rerio exhibit both sensory and functional recovery even after complete transection of the spinal cord. The adult zebrafish is an established model organism for studying regeneration following spinal cord injury, with sensory and motor recovery by 6&#160;weeks post-injury. To take advantage of in vivo analysis of the regenerative process available in the transparent larval zebrafish as well as genetic tools not accessible in the adult, we use the larval zebrafish to study regeneration after spinal cord transection. Here we demonstrate a method for reproducibly and verifiably transecting the larval spinal cord. After transection, our data shows sensory recovery beginning at 2&#160;days post-injury (dpi), with the C-bend movement detectable by 3 dpi and resumption of free swimming by 5 dpi. Thus we propose the larval zebrafish as a companion tool to the adult zebrafish for the study of recovery after spinal cord injury.

After spinal transection, adult zebrafish have functional recovery by six weeks post-injury. To take advantage of larval transparency and faster recovery, we present a method for transecting the larval spinal cord. After transection, we observe sensory recovery beginning at 2&#160;days post-injury, and C-bend movement by 3 days post-injury.

Rückenmarksdurchtrennung in der Zebrafisch-Larven

Mammals fail in sensory and motor recovery following spinal cord injury due to lack of axonal regrowth below the level of injury as well as an inability ...

Neurowissenschaften

Stab Wound Injury of the Zebrafish Adult Telencephalon: A Method to Investigate Vertebrate Brain Neurogenesis and Regeneration

Adult zebrafish have an amazing capacity to regenerate their central nervous system after injury. To investigate the cellular response and the molecular mechanisms involved in zebrafish adult central nervous system (CNS) regeneration and repair, we developed a zebrafish model of adult telencephalic injury.
In this approach, we manually generate an injury by pushing an insulin syringe needle into the zebrafish adult telencephalon. At different post injury days, fish are sacrificed, their brains are dissected out and stained by immunohistochemistry and/or in situ hybridization (ISH) with appropriate markers to observe cell proliferation, gliogenesis, and neurogenesis. The contralateral unlesioned hemisphere serves as an internal control. This method combined for example with RNA deep sequencing can help to screen for new genes with a role in zebrafish adult telencephalon neurogenesis, regeneration, and repair.

To shed light on the cellular and molecular mechanisms of zebrafish adult neurogenesis and regeneration, we developed a protocol for invasive surgery causing mechanical injuries in the zebrafish adult telencephalon and subsequent monitoring of changes in the stabbed hemisphere by immunohistochemistry or in situ hybridization.

Stichwunde Verletzung des Zebrafisch Erwachsene Telen: eine Methode, um Wirbel Gehirn Neurogenese und Regeneration Untersuchen

Adult zebrafish have an amazing capacity to regenerate their central nervous system after injury. To investigate the cellular response and the molecular ...

Examining Muscle Regeneration in Zebrafish Models of Muscle Disease

Skeletal muscle has a remarkable ability to regenerate following injury, which is driven by obligate tissue resident muscle stem cells. Following injury, the muscle stem cell is activated and undergoes cell proliferation to generate a pool of myoblasts, which subsequently differentiate to form new muscle fibers. In many muscle wasting conditions, including muscular dystrophy and ageing, this process is impaired resulting in the inability of muscle to regenerate. The process of muscle regeneration in zebrafish is highly conserved with mammalian systems providing an excellent system to study muscle stem cell function and regeneration, in muscle wasting conditions such as muscular dystrophy. Here, we present a method to examine muscle regeneration in zebrafish models of muscle disease. The first step involves the use of a genotyping platform that allows the determination of the genotype of the larvae prior to eliciting an injury. Having determined the genotype, the muscle is injured using a needle stab, following which polarizing light microscopy is used to determine the extent of muscle regeneration. We therefore provide a high throughput pipeline which allows the examination of muscle regeneration in zebrafish models of muscle disease.

Skeletal muscle regeneration is driven by tissue resident muscle stem cells, which are impaired in many muscle diseases such as muscular dystrophy, and this results in the inability of muscle to regenerate. Here, we describe a protocol that allows the examination of muscle regeneration in zebrafish models of muscle disease.

Untersuchung der Muskelregeneration in Zebrafischmodellen von Muskelerkrankungen

Skeletal muscle has a remarkable ability to regenerate following injury, which is driven by obligate tissue resident muscle stem cells. Following injury, ...

Stab Wound Injury Model of the Adult Optic Tectum Using Zebrafish and Medaka for the Comparative Analysis of Regenerative Capacity

While zebrafish have a superior capacity to regenerate their central nervous system (CNS), medaka has a lower CNS regenerative capacity. A brain injury model was developed in the adult optic tectum of zebrafish and medaka and comparative histological and molecular analyses were performed to elucidate the molecular mechanisms regulating the high regenerative capacity of this tissue across these fish species. Here a stab wound injury model is presented for the adult optic tectum using a needle and histological analyses for proliferation and differentiation of the neural stem cells (NSCs). A needle was manually inserted into the central region of the optic tectum, and then the fish were intracardially perfused, and their brains were dissected. These tissues were then cryosectioned and evaluated using immunostaining against the appropriate NSC proliferation and differentiation markers. This tectum injury model provides robust and reproducible results in both zebrafish and medaka, allowing for comparing NSC responses after injury. This method is available for small teleosts, including zebrafish, medaka, and African killifish, and enables us to compare their regenerative capacity and investigate unique molecular mechanisms.

A mechanical brain injury model in the adult zebrafish is described to investigate the molecular mechanisms regulating their high regenerative capacity. The method explains to create a stab wound injury in the optic tectum of multiple species of small fish to evaluate the regenerative responses using fluorescent immunostaining.

Stichwundenverletzungsmodell des adulten optischen Tektums unter Verwendung von Zebrafisch und Medaka zur vergleichenden Analyse der Regenerationsfähigkeit

While zebrafish have a superior capacity to regenerate their central nervous system (CNS), medaka has a lower CNS regenerative capacity. A brain injury ...

<meta charset="utf8"></head><body>Robuste Zebrafischzelltransplantation für die Regenerations- und Entwicklungsforschung

High-resolution Cell Transplantation in Embryonic and Larval Zebrafish

Development and regeneration occur by a process of genetically encoded spatiotemporally dynamic cellular interactions. The use of cell transplantation between animals to track cell fate and to induce mismatches in the genetic, spatial, or temporal properties of donor and host cells is a powerful means of examining the nature of these interactions. Organisms such as chick and amphibians have made crucial contributions to our understanding of development and regeneration, respectively, in large part because of their amenability to transplantation. The power of these models, however, has been limited by low genetic tractability. Likewise, the major genetic model organisms have lower amenability to transplantation. 
The zebrafish is a major genetic model for development and regeneration, and while cell transplantation is common in zebrafish, it is generally limited to the transfer of undifferentiated cells at the early blastula and gastrula stages of development. In this article, we present a simple and robust method that extends the zebrafish transplantation window to any embryonic or larval stage between at least 1 and 7 days post fertilization. The precision of this approach allows for the transplantation of as little as one cell with near-perfect spatial and temporal resolution in both donor and host animals. While we highlight here the transplantation of embryonic and larval neurons for the study of nerve development and regeneration, respectively, this approach is applicable to a wide range of progenitor and differentiated cell types and research questions.

<meta charset="utf8"></head><body>Autor im Rampenlicht: Kartierung der zellulären Konnektivität im Nervensystem des Zebrafisches während der Entwicklung und Regeneration

Here we present a protocol to transplant cells with high spatial and temporal resolution in zebrafish embryos and larvae at any stage between at least 1 and 7 days post fertilization.

Hochauflösende Zelltransplantation in embryonalen und larvalen Zebrafischen

Development and regeneration occur by a process of genetically encoded spatiotemporally dynamic cellular interactions. The use of cell transplantation ...

Die Nutzung der Teton-System, um molekulare Mechanismen bei Zebrafisch Regeneration Studieren

Zusammenfassung

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Stichwundenverletzungsmodell des adulten optischen Tektums unter Verwendung von Zebrafisch und Medaka zur vergleichenden Analyse der Regenerationsfähigkeit

Hochauflösende Zelltransplantation in embryonalen und larvalen Zebrafischen

Hochauflösende Zelltransplantation in embryonalen und larvalen Zebrafischen

Hochauflösende Zelltransplantation in embryonalen und larvalen Zebrafischen