scholarly journals Rotenone Neurotoxicity Causes Dopamine Neuron Loss in Zebrafish

2015 ◽  
Vol 5 (2) ◽  
pp. 16-21 ◽  
Author(s):  
Simon Martel ◽  
Jonathan Y Keow ◽  
Marc Ekker
Keyword(s):  
Oxidative Damage ◽  
Nous Avons ◽  
Fluorescent Protein ◽  
Regulatory Elements ◽  
Dopamine Neuron ◽  
Dopamine Neurons ◽  
Transgenic Zebrafish ◽  
Zebrafish Embryos ◽  
Maladie De Parkinson ◽  
Neuron Loss

ABSTRACT:Objectives: We sought to determine whether rotenone, a commonly used pesticide, exhibits neurotoxicity in zebrafish by causing dopamine neuron loss through rotenone-induced oxidative damage. Methods: We exposed transgenic zebrafish embryos expressing green fluorescent protein under the control of the cis-regulatory elements of dopamine transporter (dat) to rotenone to determine the neurotoxic effects of rotenone on dopamine neuron abundance and pattern distribution, as well as the presence of apoptotic markers. The oxidative stress potential of rotenone on embryos was assessed using a live MitoSOX Red assay, and behavioural testing on adult zebrafish was assessed using video recordings of midline crossing events. Results: Zebrafish embryos treated with rotenone displayed a 50% reduction in dopamine neurons in the ventral diencephalon when exposed to 30µM rotenone (n=6, p<0.001), and rotenone-exposed zebrafish raised to adulthood demonstrate an anxiety-like behaviour (n=5, p<0.01). Furthermore embryos exposed to rotenone also demonstrated a logarithmic increase in markers of oxidative damage (n=3, p<0.001) and apoptotic activity in their diencephalic neurons. Conclusions: These results show that rotenone can induce dopamine neuron loss in zebrafish, providing a useful model for studying the environmental causes of Parkinson’s disease. RÉSUMÉ:Objectif: Nous cherchons à déterminer si la roténone, un élément commun dans les pesticides, démontre de la neurotoxicité dans les poissons-zèbres en causant une perte de dopamine dans leurs neurones à travers le dommage oxydatif induit par la roténone. Méthode: Nous avons exposé des embryons de poissons-zèbres transgéniques qui expriment la protéine fluorescente verte sous le contrôle d’éléments cis-régulateurs des transporteurs sélectifs de dopamine (dat) à la roténone pour déterminer les effets neurotoxiques de ce dernier sur les niveaux dopaminergiques dans leurs neurones. De plus, nous avons évalué la présence de marqueurs apoptotiques. Le stress oxydatif potentiel de la roténone sur les embryons a été analysé par le « live MitoSOX Red assay » et les tests comportementaux sur les poissons-zèbres adultes furent analysés en utilisant des enregistrements vidéo. Résultats: Les embryons de poissons-zèbres qui ont été traités avec la roténone ont démontré une réduction de dopamine de 50% dans les neurones localisés dans le diencéphale ventral, quand exposés à 30µM de roténone (n=6, p<0.001). Ils ont également illustré une augmentation logarithmique dans les marqueurs de dommage oxydatif (n=3, p<0.001) et une activité apoptotique dans les neurones du diencéphale. Les poissons-zèbres exposés à de la roténone qui ont atteint l’âge adulte ont démontré des comportements d’anxiété (n=5, p<0.01). Conclusion: Les résultats démontrent que la roténone peut induire une perte dopaminergique dans les neurones des poissons-zèbres. Ces résultats s’avèrent utiles pour étudier davantage les causes environnementales reliées à la maladie de Parkinson.

scholarly journals An in vivo translation-reporter system for the study of protein synthesis in zebrafish embryos

2018 ◽  
Author(s):  
Inês Garcez Palha ◽  
Isabelle Anselme ◽  
Sylvie Schneider-Maunoury ◽  
François Giudicelli
Keyword(s):  
Protein Synthesis ◽  
Protease Inhibitor ◽  
Fluorescent Protein ◽  
Mrna Translation ◽  
Transgenic Zebrafish ◽  
Zebrafish Embryos ◽  
Reporter System ◽  
Control Of Gene Expression ◽  
A Cell

ABSTRACTControl of gene expression at the translation level is increasingly regarded as a key feature in many biological processes. Simple, inexpensive, and reliable procedures to visualise sites of protein production are required to allow observation of the spatiotemporal patterns of mRNA translation at subcellular resolution. We present a method, named SPoT (for Subcellular Patterns of Translation), developed upon the original TimeStamp technique (Lin et al., 2008), consisting in the expression of a fluorescent protein fused to a tagged, self-cleavable protease domain. Addition of a cell-permeable protease inhibitor instantly stabilizes newly produced, tagged protein allowing to distinguish recently synthesized protein from more ancient one. After a brief protease inhibitor treatment, the ratio of tagged vs non-tagged forms is highest at sites where proteins are the most recent, i.e. sites of synthesis. Therefore, by comparing tagged and non-tagged protein it is possible to spotlight sites of translation. By specifically expressing the SPoT cassette in neurons of transgenic zebrafish embryos, we reveal sites of neuronal protein synthesis in diverse cellular compartments during early development.

scholarly journals Pituitary Corticotroph Ontogeny and Regulation in Transgenic Zebrafish

2003 ◽  
Vol 17 (5) ◽  
pp. 959-966 ◽  
Author(s):  
Ning-Ai Liu ◽  
Haigen Huang ◽  
Zhongan Yang ◽  
Wiebke Herzog ◽  
Matthias Hammerschmidt ◽  
...  
Keyword(s):  
Fluorescent Protein ◽  
Cell Mass ◽  
Gene Promoter ◽  
Genetic System ◽  
Regulatory Elements ◽  
Transgenic Zebrafish ◽  
Posterior Pituitary ◽  
Gfp Expression ◽  
Green Fluorescent

Abstract We characterized zebrafish proopiomelanocortin (POMC) gene promoter, and sequence analysis revealed that the promoter contains regulatory elements conserved among vertebrate species. To monitor the ontogeny of the pituitary POMC lineage in living vertebrates, we generated transgenic zebrafish expressing green fluorescent protein (GFP) driven by the POMC promoter. Zebrafish POMC-GFP is first expressed asymmetrically as two bilateral groups of cells most anterior to the neural ridge midline at 18–20 h post fertilization (hpf). POMC-GFP-positive cells then fuse into a single-cell mass within the pituitary anlage after 24 hpf and subsequently organize as distinct anterior and posterior domains between 48 and 64 hpf. Immunohistochemical studies with ACTH and αMSH antisera showed that POMC-GFP was mainly targeted to both anterior and posterior pituitary corticotrophs, whereas posterior pituitary region melanotrophs did not express GFP. To determine in vivo zebrafish corticotroph responses, dexamethasone (10−5m) was added to live embryos, which selectively suppressed POMC-GFP expression in the anterior group of corticotrophs, suggesting a distinct domain that is responsive to glucocorticoid feedback. Transgenic zebrafish with specific POMC-GFP expression in pituitary corticotrophs offers a powerful genetic system for in vivo study of vertebrate corticotroph lineage development.

Requirements for green fluorescent protein detection in transgenic zebrafish embryos

Gene ◽  
1996 ◽  
Vol 173 (1) ◽  
pp. 99-103 ◽  
Author(s):  
Adam Amsterdam ◽  
Shuo Lin ◽  
Larry G. Moss ◽  
Nancy Hopkins
Keyword(s):  
Green Fluorescent Protein ◽  
Fluorescent Protein ◽  
Protein Detection ◽  
Transgenic Zebrafish ◽  
Zebrafish Embryos ◽  
Green Fluorescent

Pbx-dependent regulation of lbx gene expression in developing zebrafish embryos

Genome ◽  
2011 ◽  
Vol 54 (12) ◽  
pp. 973-985 ◽  
Author(s):  
Chris M. Lukowski ◽  
Danna Lynne Drummond ◽  
Andrew J. Waskiewicz
Keyword(s):  
Transcription Factors ◽  
Neural Tube ◽  
Muscle Development ◽  
Cell Types ◽  
Regulatory Elements ◽  
Transgenic Zebrafish ◽  
Zebrafish Embryos ◽  
Regulatory Interaction ◽  
Mrna In Situ Hybridization

Ladybird (Lbx) homeodomain transcription factors function in neural and muscle development—roles conserved from Drosophila to vertebrates. Lbx expression in mice specifies neural cell types, including dorsally located interneurons and association neurons, within the neural tube. Little, however, is known about the regulation of vertebrate lbx family genes. Here we describe the expression pattern of three zebrafish ladybird genes via mRNA in situ hybridization. Zebrafish lbx genes are expressed in distinct but overlapping regions within the developing neural tube, with strong expression within the hindbrain and spinal cord. The Hox family of transcription factors, in cooperation with cofactors such as Pbx and Meis, regulate hindbrain segmentation during embryogenesis. We have identified a novel regulatory interaction in which lbx1 genes are strongly downregulated in Pbx-depleted embryos. Further, we have produced a transgenic zebrafish line expressing dTomato and EGFP under the control of an lbx1b enhancer—a useful tool to acertain neuron location, migration, and morphology. Using this transgenic strain, we have identified a minimal neural lbx1b enhancer that contains key regulatory elements for expression of this transcription factor.

Live imaging of Runx1 expression in the dorsal aorta tracks the emergence of blood progenitors from endothelial cells

Blood ◽  
2010 ◽  
Vol 116 (6) ◽  
pp. 909-914 ◽  
Author(s):  
Enid Yi Ni Lam ◽  
Christopher J. Hall ◽  
Philip S. Crosier ◽  
Kathryn E. Crosier ◽  
Maria Vega Flores
Keyword(s):  
Endothelial Cells ◽  
Transcription Factor ◽  
Fluorescent Protein ◽  
Living Organism ◽  
Time Lapse ◽  
Dorsal Aorta ◽  
Transgenic Zebrafish ◽  
Hematopoietic Stem ◽  
Green Fluorescent

Abstract Blood cells of an adult vertebrate are continuously generated by hematopoietic stem cells (HSCs) that originate during embryonic life within the aorta-gonad-mesonephros region. There is now compelling in vivo evidence that HSCs are generated from aortic endothelial cells and that this process is critically regulated by the transcription factor Runx1. By time-lapse microscopy of Runx1-enhanced green fluorescent protein transgenic zebrafish embryos, we were able to capture a subset of cells within the ventral endothelium of the dorsal aorta, as they acquire hemogenic properties and directly emerge as presumptive HSCs. These nascent hematopoietic cells assume a rounded morphology, transiently occupy the subaortic space, and eventually enter the circulation via the caudal vein. Cell tracing showed that these cells subsequently populated the sites of definitive hematopoiesis (thymus and kidney), consistent with an HSC identity. HSC numbers depended on activity of the transcription factor Runx1, on blood flow, and on proper development of the dorsal aorta (features in common with mammals). This study captures the earliest events of the transition of endothelial cells to a hemogenic endothelium and demonstrates that embryonic hematopoietic progenitors directly differentiate from endothelial cells within a living organism.

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