Diese Arbeit wurde vom NIH R01 HD050901 zu MCM und American Cancer Society Postdoctoral Fellowship # PF-05-041 bis 01-DDC zur TG unterstützt.

<ol><li> Ein männlicher Zebrafisch wird zuerst zerlegt werden, gefolgt von einer weiblichen Fische. Vor Beginn der Dissektion, betäuben ein Fisch in 0,2% Tricaine und dann einschläfern es durch Inkubation in Eiswasser für 15 Minuten. </li><li> Beginnen Sie mit dem leicht klopfen die Fische trocken auf einem Papiertuch und legt es auf dem Seziertisch Matte. Äußerlich haben Zebrafisch einzigen dorsalen, Schwanz-und Afterflossen und gepaart Brust-und Bauchflossen (Abbildung 1). <img src="/files/ftp_upload/1717/1717fig1.jpg" alt="Abbildung 1" /> Abbildung 1. Erwachsene männliche Fische mit Flossen gekennzeichnet. </li><li> Pin die Fische auf die Matte Sezieren durch den fleischigen Teil des Schwanzes und der ventrale Teil der Augenhöhle. </li><li> Snip die Haut an den Bauch des Fisches knapp vor der Afterflosse. Schneiden Sie die Haut und darunter liegende Muskulatur entlang der Bauch von der Afterflosse, die Operculum (die harte Abdeckung über der Kiemen) (Abbildung 2, Stufe 1). <img src="/files/ftp_upload/1717/1717fig2.jpg" alt="Abbildung 2" /> Abbildung 2. Steps bei der Beseitigung von Haut und Körper Wand Muskulatur der inneren Organe freizulegen. </li><li> Dann entfernen Sie das Operculum und die Brustflosse, einschließlich der Schultergürtel (die dicken, knöchernen Region an der Basis des fin) (Abbildung 2, Schritt 2). Schneiden Sie die Haut und darunter liegende Muskulatur zu Beginn von oben der nun freiliegenden Kiemen hinten an der Seite des Fisches und dann auf der Afterflosse (Abb. 2, Schritt 3). </li><li> Entfernen Sie vorsichtig die Haut und darunter liegende Muskulatur von der Seite des Fisches. Viele der inneren Organe sind jetzt sichtbar (Abbildung 3). <img src="/files/ftp_upload/1717/1717fig3.jpg" alt="Abbildung 3" /> Abbildung 3. Erwachsene männliche Fische mit Körperwand und Muskulatur entfernt ermöglicht die Visualisierung der inneren Organe. </li><li> Die Hoden sind lang, weiß, gepaart Organe, die an der dorsalen Körperwand befestigt sind. Entfernen eines Hodens und legen Sie sie in eine Schüssel mit PBS. Untersuchen Sie die Hoden mit reflektiertem Licht zu den Samenkanälchen (Abbildung 4), die Zysten enthalten mit verschiedenen Stadien der Entwicklung von Keimzellen aus spermatagonia zu Spermatiden (Leal et al., 2009) zu visualisieren. <img src="/files/ftp_upload/1717/1717fig4.jpg" alt="Abbildung 4" /> Abbildung 4. Dissection eines Hodens. Die weißen rundlichen Strukturen sind die Samenleiter. </li><li> Dann entfernen Sie den Magen-Darm-System aus der Körperhöhle des Fisches. Die Leber kann durch seine Größe, gelappt Morphologie, bräunlichen Farbe und umfangreiche Vaskularisierung identifiziert werden. Die Gallenblase, eine grüne durchscheinende Flüssigkeit gefüllten Sack, und die Milz, die leuchtend rot erscheint, sind in den Eingeweiden gefunden. </li><li> Separate den Darm aus dem Rest der Organe und auszustrecken. Die vorderen, mittleren und hinteren Regionen des Darms werden durch die Höhe der epithelialen Falten (Wallace et al., 2005) definiert. </li><li> Legen Sie ein Stück des Darms in PBS und beobachten Sie die epithelialen Falten mit Durchlicht. </li><li> Als nächstes untersuchen die Schwimmblase. Die Schwimmblase besteht aus einer hinteren Kammer, die an der Speiseröhre über den pneumatischen Kanal, und eine vordere Kammer, die an das Innenohr durch den Weberschen Apparat (Finney et al., 2006) ist in Verbindung steht. </li><li> Entfernen und Entsorgen der Schwimmblase. Unpin die Fische und re-pin es Bauchseite bis zu den Nieren, die sich entlang der dorsalen Körperwand befindet sezieren. Die Niere ist ein durchsichtiges Rosa-Struktur mit der dorsalen Aorta und pigmentierten Zellen assoziiert. Die Niere ist in Kopf, Körper und Schwanz Regionen (Abbildung 5) unterteilt. Dissect ein Stück der Niere und legen Sie sie in PBS. Tease neben dem Nierengewebe der Nierentubuli zu offenbaren. <img src="/files/ftp_upload/1717/1717fig5.jpg" alt="Abbildung 5" /> Abbildung 5. Lage der Kopf, Körper und Schwanz Nieren entlang der dorsalen Körperwand. </li><li> Sezieren einen weiblichen Fisch. Wie zuvor beschrieben, euthanize die Fische in Eiswasser und pat es trocken ist, bevor Pinning es dem Sezieren Matte. Entfernen Sie die Haut von der Seite des Fisches, wie zuvor gezeigt. Der Eierstock ist ein bilobed Struktur, die in der Leibeshöhle durch einen vaskularisierten Mesovarium ausgesetzt ist. </li><li> Nehmen Sie einen Lappen des Eierstocks, legen Sie sie in PBS, und prüfen Sie mit Durchlicht. Die Eizellen können neben gehänselt werden mit feinen Nadeln und dann inszeniert (Abbildung 6, Selman et al., 1993). Stage I Eizellen sind ca. 10 bis 150 Mikrometer groß und durchscheinend. Stage II-Oozyten sind etwa 150 bis 350 Mikrometer groß und definiert durch das Auftreten von kortikalen Granula. Stadium III Oozyten sind 350 bis 750 Mikrometer groß und undurchsichtig sind aufgrund der Ansammlung von Dotter. Reifen Stadium IV und V Eizellen sind lichtdurchlässig und in der Regel nicht in den Eierstöcken der Frauen, die vor kurzem gepaart gefunden haben. <img src="/files/ftp_upload/1717/1717fig6.jpg" alt="Abbildung 6" /> Abbildung 6. Live Stadium I, II, III und Eizellen unter einem beobachtetenBinokular mit Durchlicht. </li><li> Als nächstes zerlegen das Herz aus dem Fisch. Das Herz befindet sich posterior und ventral der Kiemen. Beginnen Sie mit dem Herausschneiden des Herzens und all das umliegende Gewebe und es in PBS. Sorgfältig sezieren weg das Gewebe rund um das Herz, darauf achten, daß die zarte Atrium Schäden. </li><li> Setzen Sie den sezierten Herzen in Ringer-Lösung, um den Herzschlag zu beobachten. Identifizieren Sie die Atrium, Ventrikel und Bulbus arteriosus (Abb. 7, Hu et al., 2001). <img src="/files/ftp_upload/1717/1717fig7.jpg" alt="Abbildung 7" /> Abbildung 7. Dissected erwachsenen Herzens. </li><li> Füllen Sie das Sezieren, indem das Gehirn aus dem Fisch. Beginnen Sie mit der unpinning die Fische und das Entfernen der Kopf mit einer Rasierklinge. Entfernen Sie so viel Weichgewebe wie möglich von der Bauchseite des Schädels mit einer Pinzette. Entfernen Sie die Augen mit kleinen Feder Schere. Brechen Sie den Schädel und entfernen Sie die Knochen aus dem ventralen Seite des Gehirns. Setzen Sie nun den Kopf in eine Schüssel mit PBS und die Haut abziehen und Schädelknochen von der dorsalen Seite des Gehirns. </li><li> Identifizieren Sie die Riechkolben, Telencephalon, Habenula, Tectum opticum, Cerebellum und Medulla (Abbildung 8, Wullimann et al, 1996;. Schilling, 2002). <img src="/files/ftp_upload/1717/1717fig8.jpg" alt="Abbildung 8" /> Abbildung 8. Dissected erwachsenen Zebrafisch Gehirn. Dorsalansicht, anterior nach oben. </li></ol>

<ol><li>Finney, J. L., Robertson, G. N., McGee, C. A., Smith, F. M., Croll, R. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Structure+and+Autonomic+Innervation+of+the+Swim+Bladder+in+the+Zebrafish+%28Danio+rerio%29%5BTitle%5D)+AND+%22J+Comp+Neurol%22%5BJournal%5D)">Structure and Autonomic Innervation of the Swim Bladder in the Zebrafish (Danio rerio).</a> J Comp Neurol. 495, 587-606 (2006).</li>
<li>Hu, N., Yost, H. J., Clark, E. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Cardiac+morphology+and+blood+pressure+in+the+adult+zebrafish%5BTitle%5D)+AND+%22Anat+Rec%22%5BJournal%5D)">Cardiac morphology and blood pressure in the adult zebrafish.</a> Anat Rec. 264, 1-12 (2001).</li>
<li>Leal, M. C., Cardoso, E. R., Nóbrega, R. H., Batlouni, S. R., Bogerd, J., França, L. R., Schulz, R. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Histological+and+stereological+evaluation+of+zebrafish+%28Danio+rerio%29+spermatogenesis+with+an+emphasis+on+spermatogonial+generations%5BTitle%5D)+AND+%22Biol+Reprod%22%5BJournal%5D)">Histological and stereological evaluation of zebrafish (Danio rerio) spermatogenesis with an emphasis on spermatogonial generations.</a> Biol Reprod. 81, 177-187 (2009).</li>
<li>Schilling, T. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+morphology+of+larval+and+adult+zebrafish%5BTitle%5D)+AND+%22Zebrafish%3A+A+Practical+Approach+%28The+Practical+Approach+Series%29%22%5BJournal%5D)">The morphology of larval and adult zebrafish.</a> Zebrafish: A Practical Approach (The Practical Approach Series). Nusslein-Volhard, C., Dahm, R. , Oxford University Press. (2002).</li>
<li>Selman, K., Wallace, R., Sarka, A., Qi, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Stages+of+oocyte+development+in+the+Zebrafish%2C+Brachydanio+rerio%5BTitle%5D)+AND+%22J.+Morphol%22%5BJournal%5D)">Stages of oocyte development in the Zebrafish, Brachydanio rerio.</a> J. Morphol. 218, 203-224 (1993).</li>
<li>Wallace, K. N., Akhter, S., Smith, E. M., Lorent, K., Pack, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Intestinal+growth+and+differentiation+in+zebrafish%5BTitle%5D)+AND+%22Mech+Dev%22%5BJournal%5D)">Intestinal growth and differentiation in zebrafish.</a> Mech Dev. 122, 157-173 (2005).</li>
<li>Wullimann, M. F., Rupp, B., Reichert, H. Neuroanatomy of the zebrafish brain : a topological atlas. , Birkhäuser Verlag. (1996).</li>
</ol>

Tierversuche wurden in Übereinstimmung mit den Richtlinien und Vorschriften her von der University of Pennsylvania Institutional Animal Care und Verwenden Ausschusses durchgeführt.

Reagenzien: 0,2% Tricaine (Ethyl-3-Aminobenzoesäure) 200 mg Pulver Tricaine 97,9 ml DD-Wasser ~ 1 ml 1 M Tris (pH 9) Der pH-Wert auf 7,0 Phosphat-gepufferte Kochsalzlösung (PBS) 4,0 g NaCl 0,1 g KCl 100 ml 0,1 M PO 4-Puffer, pH 7,3 150 ml dH 2 O Ringer-Lösung 6,7 g NaCl 0,2 g KCl 0,2 g CaCl 2 1,2 g Hepes 1 LH 2 O Der pH-Wert auf 7,2 Ausstattung: Dissecting Gericht mit Matte Electron Microscopy Sciences Katalog # 70540 Vannas Frühjahr Schere Feine Science Tools Katalog Nr. 91500-09

dissection of organs from the adult zebrafish

In den letzten 20 Jahren hat sich der Zebrafisch zu einem leistungsfähigen Modell-Organismus für das Verständnis der Entwicklung von Wirbeltieren und Krankheit. Obwohl experimentelle Analyse des Embryos und Larven ist sehr umfangreich und die Morphologie ist gut dokumentiert, sind die Beschreibungen der erwachsenen Zebrafisch Anatomie und Studien über die Entwicklung der erwachsenen Strukturen und Organe, zusammen mit Techniken für die Arbeit mit Erwachsenen fehlen. Die Organe der Larve eine bedeutende Veränderung in ihrer gesamten Struktur, Morphologie und anatomische Lage während des Larven zu erwachsenen Übergang. Äußerlich entwickelt die transparente Larve ihren charakteristischen Erwachsenen gestreiften Pigment Muster und gepaart Bauchflossen, während intern die Organe zu unterziehen massives Wachstum und Umbau. Darüber hinaus entwickelt die bipotentialen Geschlechtsdrüse Primordium entweder in Hoden oder Eierstock. Dieses Protokoll enthält viele Organe des Erwachsenen und zeigt Methoden für die Präparation des Gehirns, Gonaden, Magen-Darm-System, Herz und Niere des erwachsenen Zebrafisch. Die sezierten Organe können in-situ-Hybridisierung, Immunhistochemie, Histologie, RNA-Extraktion, Protein-Analyse und andere molekulare Techniken eingesetzt werden. Dieses Protokoll wird in die Erweiterung der Studien im Zebrafisch zu helfen, den Umbau des larvalen Organe, die Morphogenese von Organen spezifisch für die Erwachsenen und andere Untersuchungen der erwachsenen Organsysteme umfassen.

Watch this Scientific Journal Video about Dissection of Organs from the Adult Zebrafish at JoVE.com

Dissection of Organs from the Adult Zebrafish

Dieses Protokoll beschreibt ein Verfahren zur Identifizierung und seziert Organe von den erwachsenen Zebrafisch.

Dissection von Organen aus dem Adult Zebrafisch

Over the last 20 years, the zebrafish has become a powerful model organism for understanding vertebrate development and disease.  Although experimental ...

dissection-of-organs-from-the-adult-zebrafish

Research

JoVE Journal

Biology

104.7K Aufrufe.  University of Pennsylvania-School of Medicine. Dieses Protokoll beschreibt ein Verfahren zur Identifizierung und seziert Organe von den erwachsenen Zebrafisch.

Serie: Dissection of Organs from the Adult Zebrafish

Large-scale Zebrafish Embryonic Heart Dissection for Transcriptional Analysis

The zebrafish embryonic heart is composed of only a few hundred cells, representing only a small fraction of the entire embryo. Therefore, to prevent the cardiac transcriptome from being masked by the global embryonic transcriptome, it is necessary to collect sufficient numbers of hearts for further analyses. Furthermore, as zebrafish cardiac development proceeds rapidly, heart collection and RNA extraction methods need to be quick in order to ensure homogeneity of the samples. Here, we present a rapid manual dissection protocol for collecting functional/beating hearts from zebrafish embryos. This is an essential prerequisite for subsequent cardiac-specific RNA extraction to determine cardiac-specific gene expression levels by transcriptome analyses, such as quantitative real-time polymerase chain reaction (RT-qPCR). The method is based on differential adhesive properties of the zebrafish embryonic heart compared with other tissues; this allows for the rapid physical separation of cardiac from extracardiac tissue by a combination of fluidic shear force disruption, stepwise filtration and manual collection of transgenic fluorescently labeled hearts.

To analyse cardiac gene expression profiles during zebrafish heart development, total RNA has to be extracted from isolated hearts. Here, we present a protocol for collecting functional/beating hearts by rapid manual dissection from zebrafish embryos to obtain cardiac-specific mRNA.

Großzebrafisch Embryonale Herz Dissection für Transkriptionsanalyse

The zebrafish embryonic heart is composed of only a few hundred cells, representing only a small fraction of the entire embryo. Therefore, to prevent the ...

Forschung

Entwicklungsbiologie

Dissection of Larval Zebrafish Gonadal Tissue

Although wild zebrafish possess a ZZ/ZW sex-determination system, domesticated zebrafish have lost the sex chromosome. They utilize a polygenic sex determination system, where several genes distributed throughout the genome collectively determine the sex identities of individual fish. Currently, the genes involved in regulating gonad development and how they work remain elusive. Normally, isolating gonadal tissue is the first step to examine the sex developmental processes. Here, we present a procedure to isolate gonadal tissue from 17 dpf (days post fertilization) and 25 dpf zebrafish larvae. The isolated gonadal tissue may be subsequently examined by morphology and gene expression profiling.

Here, we present a protocol for isolating gonadal tissue of larval zebrafish, which will facilitate investigations of zebrafish sex differentiation and maintenance.

Dissection von Larval Zebrabärbling Gonadal Tissue

Although wild zebrafish possess a ZZ/ZW sex-determination system, domesticated zebrafish have lost the sex chromosome. They utilize a polygenic sex ...

Dissection of the Adult Zebrafish Kidney

Researchers working in the burgeoning field of adult stem cell biology seek to understand the signals that regulate the behavior and function of stem cells during normal homeostasis and disease states. The understanding of adult stem cells has broad reaching implications for the future of regenerative medicine1. For example, better knowledge about adult stem cell biology can facilitate the design of therapeutic strategies in which organs are triggered to heal themselves or even the creation of methods for growing organs in vitro that can be transplanted into humans1. The zebrafish has become a powerful animal model for the study of vertebrate cell biology2. There has been extensive documentation and analysis of embryonic development in the zebrafish3. Only recently have scientists sought to document adult anatomy and surgical dissection techniques4, as there has been a progressive movement within the zebrafish community to broaden the applications of this research organism to adult studies. For example, there are expanding interests in using zebrafish to investigate the biology of adult stem cell populations and make sophisticated adult models of diseases such as cancer5. Historically, isolation of the zebrafish adult kidney has been instrumental for studying hematopoiesis, as the kidney is the anatomical location of blood cell production in fish6,7. The kidney is composed of nephron functional units found in arborized arrangements, surrounded by hematopoietic tissue that is dispersed throughout the intervening spaces. The hematopoietic component consists of hematopoietic stem cells (HSCs) and their progeny that inhabit the kidney until they terminally differentiate8. In addition, it is now appreciated that a group of renal stem/progenitor cells (RPCs) also inhabit the zebrafish kidney organ and enable both kidney regeneration and growth, as observed in other fish species9-11. In light of this new discovery, the zebrafish kidney is one organ that houses the location of two exciting opportunities for adult stem cell biology studies. It is clear that many outstanding questions could be well served with this experimental system. To encourage expansion of this field, it is beneficial to document detailed methods of visualizing and then isolating the adult zebrafish kidney organ. This protocol details our procedure for dissection of the adult kidney from both unfixed and fixed animals.
Dissection of the kidney organ can be used to isolate and characterize hematopoietic and renal stem cells and their offspring using established techniques such as histology, fluorescence activated cell sorting (FACS)11,12, expression profiling13,14, and transplantation11,15. 
We hope that dissemination of this protocol will provide researchers with the knowledge to implement broader use of zebrafish studies that ultimately can be translated for human application.

The zebrafish kidney is home to both renal and hematopoietic adult stem/progenitor cells, and represents an outstanding opportunity to study these cell types and their progeny in a vertebrate model organism. Here, we demonstrate a detailed dissection procedure that enables the researcher to identify and surgically remove the adult zebrafish kidney, which can be used for applications such as cell isolation, transplantation, and expression studies of kidney and/or blood cell populations.

Präparation des Adult Zebrafisch Kidney

Researchers working in the burgeoning field of adult stem cell biology seek to understand the signals that regulate the behavior and function of stem ...

Biologie

Heart Dissection in Larval, Juvenile and Adult Zebrafish, Danio rerio

Zebrafish have become a beneficial and practical model organism for the study of embryonic heart development (see recent reviews1-6), however, work examining post-embryonic through adult cardiac development has been limited7-10. Examining the changing morphology of the maturing and aging heart are restricted by the lack of techniques available for staging and isolating juvenile and adult hearts. In order to analyze heart development over the fish's lifespan, we dissect zebrafish hearts at numerous stages and photograph them for further analysis11. The morphological features of the heart can easily be quantified and individual hearts can be further analyzed by a host of standard methods. Zebrafish grow at variable rates and maturation correlates better with fish size than age, thus, post-fixation, we photograph and measure fish length as a gauge of fish maturation. This protocol explains two distinct, size dependent dissection techniques for zebrafish, ranging from larvae 3.5mm standard length (SL) with hearts of 100&mu;m ventricle length (VL), to adults, with SL of 30mm and VL 1mm or larger. Larval and adult fish have quite distinct body and organ morphology. Larvae are not only significantly smaller, they have less pigment and each organ is visually very difficult to identify. For this reason, we use distinct dissection techniques.
We used pre-dissection fixation procedures, as we discovered that hearts dissected directly after euthanization have a more variable morphology, with very loose and balloon like atria compared with hearts removed following fixation. The fish fixed prior to dissection, retain in vivo morphology and chamber position (data not shown). In addition, for demonstration purposes, we take advantage of the heart (myocardial) specific GFP transgenic Tg(myl7:GFP)twu34 (12), which allows us to visualize the entire heart and is particularly useful at early stages in development when the cardiac morphology is less distinct from surrounding tissues. Dissection of the heart makes further analysis of the cell and molecular biology underlying heart development and maturation using in situ hybridization, immunohistochemistry, RNA extraction or other analytical methods easier in post-embryonic zebrafish. This protocol will provide a valuable technique for the study of cardiac development maturation and aging.

Heart Dissection in Larval, Juvenile and Adult Zebrafish, Danio rerio

A clear, standardized method for dissection and isolation of the zebrafish heart at multiple developmental stages are described. Annotation and quantification techniques are also discussed.

Herz Dissection in Larven, juvenile und adulte Zebrafisch, Danio rerio</em

Zebrafish have become a beneficial and practical model organism for the study of embryonic heart development (see recent reviews1-6), however, work ...

Analysis of Nephron Composition and Function in the Adult Zebrafish Kidney

The zebrafish model has emerged as a relevant system to study kidney development, regeneration and disease. Both the embryonic and adult zebrafish kidneys are composed of functional units known as nephrons, which are highly conserved with other vertebrates, including mammals. Research in zebrafish has recently demonstrated that two distinctive phenomena transpire after adult nephrons incur damage: first, there is robust regeneration within existing nephrons that replaces the destroyed tubule epithelial cells; second, entirely new nephrons are produced from renal progenitors in a process known as neonephrogenesis. In contrast, humans and other mammals seem to have only a limited ability for nephron epithelial regeneration. To date, the mechanisms responsible for these kidney regeneration phenomena remain poorly understood. Since adult zebrafish kidneys undergo both nephron epithelial regeneration and neonephrogenesis, they provide an outstanding experimental paradigm to study these events. Further, there is a wide range of genetic and pharmacological tools available in the zebrafish model that can be used to delineate the cellular and molecular mechanisms that regulate renal regeneration. One essential aspect of such research is the evaluation of nephron structure and function. This protocol describes a set of labeling techniques that can be used to gauge renal composition and test nephron functionality in the adult zebrafish kidney. Thus, these methods are widely applicable to the future phenotypic characterization of adult zebrafish kidney injury paradigms, which include but are not limited to, nephrotoxicant exposure regimes or genetic methods of targeted cell death such as the nitroreductase mediated cell ablation technique. Further, these methods could be used to study genetic perturbations in adult kidney formation and could also be applied to assess renal status during chronic disease modeling.

The zebrafish adult kidney is an excellent system for renal regeneration and disease studies. An essential aspect of such research is the assessment of nephron structure and function. This protocol describes several methodologies that can be implemented to assess nephron tubule composition and to evaluate renal reabsorption.

Analyse der Nephron Zusammensetzung und Funktion im erwachsenen Zebrafisch Nieren

The zebrafish model has emerged as a relevant system to study kidney development, regeneration and disease. Both the embryonic and adult zebrafish kidneys ...

Recovery of Adult Zebrafish Hearts for High-throughput Applications

Use of the zebrafish model system for studying development, regeneration, and disease is expanding toward use of adult hearts for cell dissociation and purification of RNA, DNA, and proteins. All of these applications demand the rapid recovery of significant numbers of zebrafish hearts to avoid gene regulatory, metabolic, and other changes that begin after death. Adult zebrafish hearts are also required for studying heart structure for a variety of mutants and for studying heart regeneration. However, the traditional zebrafish heart dissection is slow and difficult and requires specialized tools, making large-scale dissection of adult zebrafish hearts tedious. Traditional methods also harbor the risk of damaging the heart during the dissection. Here, we describe a method for dissection of adult zebrafish hearts that is fast, reproducible, and preserves heart architecture. Furthermore, this method does not require specialized tools, is painless for the zebrafish, can be performed on fresh or fixed specimens, and can be performed on zebrafish as young as one month old. The approach described expands the use of adult zebrafish for cardiovascular research.

Use of zebrafish for cardiovascular research is expanding towards research on adult hearts. For these applications, quick and simple isolation of cardiac tissues is key to avoid post-mortem changes and to obtain an adequate number of samples. Here, we describe a fast and reproducible method for dissecting adult zebrafish hearts.

Rückgewinnung von Adult Zebrafisch-Herzen für Hochdurchsatz-Anwendungen

Use of the zebrafish model system for studying development, regeneration, and disease is expanding toward use of adult hearts for cell dissociation and ...

Partial Hepatectomy in Adult Zebrafish

Liver failure is one of the leading causes of death worldwide, and mortality from chronic liver disease is rising sharply in the United States. Healthy livers are capable of regenerating from toxic damage, but in advanced liver disease, the natural ability of the liver to regenerate is impaired. Zebrafish have emerged as a powerful experimental system for studying regeneration. They are an ideal model for studying liver regeneration from partial hepatectomy, a procedure with direct clinical relevance in which part of the liver is surgically removed, leaving the rest intact. There is no standard protocol for partial hepatectomy; previous studies using this model have used slightly different protocols and reported disparate results. Described here is an efficient, reproducible protocol for performing a partial hepatectomy in adult zebrafish. We use this technique to demonstrate that zebrafish are capable of epimorphic regeneration of the resected lobe. This protocol can be used to further interrogate the mechanisms required for liver regeneration in zebrafish.

This protocol describes the procedure for removing the ventral lobe of the liver in adult zebrafish to enable the study of liver regeneration.

Partielle Hepatektomie bei erwachsenen Zebrafischen

Liver failure is one of the leading causes of death worldwide, and mortality from chronic liver disease is rising sharply in the United States. Healthy ...

Isolation of Cardiac and Vascular Smooth Muscle Cells from Adult, Juvenile, Larval and Embryonic Zebrafish for Electrophysiological Studies

Zebrafish have long been used as a model vertebrate organism in cardiovascular research. The technical difficulties of isolating individual cells from the zebrafish cardiovascular tissues have been limiting in studying their electrophysiological properties. Previous methods have been described for dissection of zebrafish hearts and isolation of ventricular cardiac myocytes. However, the isolation of zebrafish atrial and vascular myocytes for electrophysiological characterization was not detailed. This work describes new and modified enzymatic protocols that routinely provide isolated juvenile and adult zebrafish ventricular and atrial cardiomyocytes, as well as vascular smooth muscle (VSM) cells from the bulbous arteriosus, suitable for patch-clamp experiments. There has been no literary evidence of electrophysiological studies on zebrafish cardiovascular tissues isolated at embryonic and larval stages of development. Partial dissociation techniques that allow patch-clamp experiments on individual cells from larval and embryonic hearts are demonstrated.

The present protocol describes the acute isolation of viable cardiac and vascular smooth muscle cells from adult, juvenile, larval, and embryonic zebrafish (Danio rerio), suitable for electrophysiological studies.

Isolierung von kardialen und vaskulären glatten Muskelzellen aus adulten, juvenilen, Larven und embryonalen Zebrafischen für elektrophysiologische Studien

Zebrafish have long been used as a model vertebrate organism in cardiovascular research. The technical difficulties of isolating individual cells from the ...

Adult Zebrafish Intestine Dissection: A Technique to Analyze Compound Absorption

Source: Jie J. et. al., <a href="https://www-jove-com.remotexs.ntu.edu.sg/v/58366/" target="_blank">Oral Intubation of Adult Zebrafish: A Model for Evaluating Intestinal Uptake of Bioactive Compounds</a>. J. Vis. Exp. (2018).
This video describes the technique of isolating the intestine to evaluate the intestinal uptake of bioactive compounds. In this sample, we will isolate the intestine from an adult zebrafish to analyze the uptake of immunostimulants.

Darmdissektion bei adulten Zebrafischen: Eine Technik zur Analyse der Absorption von Verbindungen

Source: Jie J. et. al., Oral Intubation of Adult Zebrafish: A Model for Evaluating Intestinal Uptake of Bioactive Compounds. J. Vis. Exp. (2018).
This ...

JoVE Encyclopedia of Experiments

Danio rerio (Zebrafisch)

Adult

Heart Dissection: A Method to Observe Zebrafish Cardiac Development

Source: Singleman C. et. al., <a href="https://www-jove-com.remotexs.ntu.edu.sg/v/3165/" target="_blank">Heart Dissection in Larval, Juvenile and Adult Zebrafish, Danio rerio</a>. J. Vis. Exp. (2011).
This video describes a technique of isolating the heart from a zebrafish at multiple developmental stages to study cardiac development, maturation, and ageing.

Herzdissektion: Eine Methode zur Beobachtung der Herzentwicklung von Zebrafischen

Source: Singleman C. et. al., Heart Dissection in Larval, Juvenile and Adult Zebrafish, Danio rerio. J. Vis. Exp. (2011).
This video describes a technique ...