PUBLICATION
Multicolor Time-lapse Imaging of Transgenic Zebrafish: Visualizing Retinal Stem Cells Activated by Targeted Neuronal Cell Ablation
- Authors
- Ariga, J., Walker, S.L., and Mumm, J.S.
- ID
- ZDB-PUB-101027-40
- Date
- 2010
- Source
- Journal of visualized experiments : JoVE (43): (Journal)
- Registered Authors
- Ariga, Junko, Mumm, Jeff, Walker, Steven
- Keywords
- none
- MeSH Terms
-
- Animals
- Animals, Genetically Modified
- Cell Line
- Fluorescent Dyes/chemistry
- Fluorescent Dyes/metabolism
- Image Processing, Computer-Assisted/methods*
- Nerve Regeneration
- Neurons/cytology*
- Neurons/physiology
- Retina/cytology*
- Retina/physiology
- Stem Cells/cytology*
- Stem Cells/physiology
- Zebrafish
- PubMed
- 20972390 Full text @ J. Vis. Exp.
Citation
Ariga, J., Walker, S.L., and Mumm, J.S. (2010) Multicolor Time-lapse Imaging of Transgenic Zebrafish: Visualizing Retinal Stem Cells Activated by Targeted Neuronal Cell Ablation. Journal of visualized experiments : JoVE. (43).
Abstract
High-resolution time-lapse imaging of living zebrafish larvae can be utilized to visualize how biological processes unfold (for review see (1)). Compound transgenic fish which express different fluorescent reporters in neighboring cell types provide a means of following cellular interactions (2) and/or tissue-level responses to experimental manipulations over time. In this video, we demonstrate methods that can be used for imaging multiple transgenically labeled cell types serially in individual fish over time courses that can span from minutes to several days. The techniques described are applicable to any study seeking to correlate the "behavior" of neighboring cells types over time, including: 1) serial 'catch and release' methods for imaging a large number of fish over successive days, 2) simplified approaches for separating fluorophores with overlapping excitation/emission profiles (e.g., GFP and YFP), 3) use of hypopigmented mutant lines to extend the time window available for high-resolution imaging into late larval stages of development, 4) use of membrane targeted fluorescent reporters to reveal fine morphological detail of individual cells as well as cellular details in larger populations of cells, and 5) a previously described method for chemically-induced ablation of transgenically targeted cell types; i.e., nitroreductase (NTR) mediated conversion of prodrug substrates, such as metronidazole (MTZ), to cytotoxic derivatives (3,5). As an example of these approaches, we will visualize the ablation and regeneration of a subtype of retinal bipolar neuron within individual fish over several days. Simultaneously we will monitor several other retinal cell types, including neighboring non-targeted bipolar cells and potential degeneration-stimulated retinal stem cells (i.e., MËller glia). This strategy is being applied in our lab to characterize cell- and tissue-level (e.g., stem cell niche) responses to the selective loss and regeneration of targeted neuronal cell types.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping