Transgenic zebrafish expressing green fluorescent protein in dopaminergic neurons of the ventral diencephalon

Xi, Yanwei; Yu, Man; Godoy, Rafael; Hatch, Gary; Poitras, Luc; Ekker, Marc

doi:10.1002/dvdy.22742

Cited by 77 publications

(79 citation statements)

References 42 publications

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“…Although most neurons in the vDC expressed both EGFP and TH, we also detected cells that are EGFP-positive but TH-negative in the PT possibly due to the fact that these EGFP-positive neurons might be th2-positive neurons [8,16] due to the genome duplication during teleost evolution [26]. For this reason, the anti-TH antibody used in our study has a relatively lower affinity for the TH2 protein.…”

Section: Discussionmentioning

confidence: 72%

“…For this purpose, we used the dopaminergic neurotoxin, 6-OHDA. The effects of this drug, in contrast to MPTP [8,11], had not been studied in zebra fish expressing reporters in DA neurons. In our larvae, a concentration of 500 µM of 6-OHDA induced loss of EGFP-positive neurons, and elicited marked deficits in the motor activity of fish.…”

Section: Discussionmentioning

confidence: 99%

“…The teleost, zebra fish (Danio rerio) has proven to be an excellent model organism for developmental genetic studies and recently has emerged as a valuable model to study of different aspects associated with neuro degeneration [7,8]. As a vertebrate, the basic organization and divisions of the CNS are similar to those of other vertebrates and specialized neuronal populations, including the dopaminergic system also appear well conserved [9].…”

Section: Introductionmentioning

confidence: 99%

“…Despite the diverse role of DA cells in retinal functions, the understanding of DA cell function has been limited because they have a low density in the retina and cannot be identified in living retina by morphological characteristics. However, the use of other promoters (e.g., dat and vmat2) have contributed successfully for the understanding of the role of Volume 4 Issue 1 -2017 different monoaminergic neurons in the brain [11], including the dopaminergic neurons (DA) located in the diencenphalon [8,17]. In spite of this, the DA cell function is not completely understood and it could benefit from the development of other transgenic zebra fish models with increased labeling of DA neurons in the brain.…”

Section: Introductionmentioning

confidence: 99%

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Green Fluorescent Protein Labeling of Dopaminergic Neurons in Zebrafish for the Study of Parkinson’s Diseases

Outeiro¹

2017

JMEN

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The loss of dopaminergic neurons in the substantia nigra is the pathological hallmark of Parkinson's disease (PD) associated with the characteristic motor deficits. Despite all the recent progress in the identification of genes and pathways associated with PD, we still lack a complete understanding of the molecular mechanisms underpinning the disease. Our goals were to develop a new transgenic zebra fish line where tyrosine hydroxylase (TH)-positive neurons were labeled with green fluorescent protein (EGFP) under the control of the zebra fish tyrosine hydroxylase (th1) promoter and to demonstrate the usefulness of the model for the study of the mechanisms underlying cell death in a toxin induced model of PD.A 2.5 kb gene fragment of the th1 promoter was isolated and cloned into a miniTol2vector followed by EGFP and microinjected to originate the Tg (-2.5th: EGFP) zebra fish line. Tg (-2.5th: EGFP) were visualized by fluorescence microscopy for EGFP and TH expression during development. The levels of EGFP mRNA were detected by RT-qPCR and protein expression was examined by western blot analysis. The model of neurotoxicity was generated by treating transgenic zebra fish with 6-hydroxydopamine (6-OHDA), followed by the analysis of cell death markers and behavior.EGFP expression was detected mainly in the posterior tuberculum of the ventral diencephalon (vDC). In the vDC, TH-labeled neurons of groups 1-were correctly labeled with EGFP. Further, we demonstrated that these neurons in larvae and adults are sensitive to 6-OHDA treatment, resulting in cell loss and behavioral deficits. In conclusion, this transgenic line constitutes a powerful tool to visualize and study dopaminergic neurons throughout the zebra fish life-cycle and enables high-throughput screening of molecules of interest for PD.

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Section: Discussionmentioning

confidence: 72%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Green Fluorescent Protein Labeling of Dopaminergic Neurons in Zebrafish for the Study of Parkinson’s Diseases

Outeiro¹

2017

JMEN

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“…We added a floxed Neomycin resistance (Neo R ) cassette downstream of the mCherry coding sequence. We used essentially the same genomic region as in the Tg(dat:EGFP) transgenic line, 8 which is a 27-kb fragment (including *13 kb of the dat 5¢-flanking region). The targeting construct included two arms (Arm A and Arm B, about 250 bp each), which are sequences homologous to the region of the dat locus flanking the EGFP sequence in the dat:EGFP construct.…”

Section: Transgenic Construct and Zebrafish Transgenesismentioning

confidence: 99%

Transgenic Zebrafish Expressing mCherry in the Mitochondria of Dopaminergic Neurons

et al. 2015

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Genetic mutations and environmental toxins are known to affect mitochondrial health and have been implicated in the progressive degeneration of dopaminergic neurons in Parkinson's disease. To visualize mitochondria in dopaminergic neurons of live zebrafish, we used the regulatory elements of the dopamine transporter (dat) gene to target a reporter, mCherry, after fusion with the mitochondrial localizing signal (MLS) of Tom20. Immunoblot analysis of mitochondrial and cytosolic fractions from Tg(dat:tom20 MLS-mCherry) larvae shows that mCherry is efficiently targeted to the mitochondria. Confocal imaging of live fish was carried out from 1 day postfertilization (dpf) to 9 dpf. We also colocalized dat mRNA expression with the mCherry protein in the olfactory bulb (OB), subpallium (SP), pretectum (Pr), diencephalic clusters 2 and 3 (DC2/3), caudal hypothalamus (Hc), locus coeruleus (LC), anterior preoptic area (POa), retinal amacrine cells (RAC), caudal hypothalamus (Hc), and preoptic area (PO). Treating Tg(dat:tom20 MLS-mCherry) larvae with the dopaminergic neurotoxin MPTP (1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine) at 2 or 3 dpf resulted in a decrease in mCherry fluorescence in the pretectum, olfactory bulb, subpallium, diencephalic clusters 2 and 3, and the caudal hypothalamus. Labeling of mitochondria in nigrostriatal dopaminergic neurons of zebrafish could allow their visualization in vivo following genetic or pharmacological manipulations.

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Detection of Mitochondrial Toxicity Using Zebrafish

Chan

Williamson

2018

Mitochondrial Dysfunction Caused by Drugs and Environmental Toxicants

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Transgenic zebrafish expressing green fluorescent protein in dopaminergic neurons of the ventral diencephalon

Cited by 77 publications

References 42 publications

Green Fluorescent Protein Labeling of Dopaminergic Neurons in Zebrafish for the Study of Parkinson’s Diseases

Green Fluorescent Protein Labeling of Dopaminergic Neurons in Zebrafish for the Study of Parkinson’s Diseases

Transgenic Zebrafish Expressing mCherry in the Mitochondria of Dopaminergic Neurons

Detection of Mitochondrial Toxicity Using Zebrafish

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