Dataset Information

Targeted two-photon chemical apoptotic ablation of defined cell types in vivo.

ABSTRACT: A major bottleneck limiting understanding of mechanisms and consequences of cell death in complex organisms is the inability to induce and visualize this process with spatial and temporal precision in living animals. Here we report a technique termed two-photon chemical apoptotic targeted ablation (2Phatal) that uses focal illumination with a femtosecond-pulsed laser to bleach a nucleic acid-binding dye causing dose-dependent apoptosis of individual cells without collateral damage. Using 2Phatal, we achieve precise ablation of distinct populations of neurons, glia and pericytes in the mouse brain and in zebrafish. When combined with organelle-targeted fluorescent proteins and biosensors, we uncover previously unrecognized cell-type differences in patterns of apoptosis and associated dynamics of ribosomal disassembly, calcium overload and mitochondrial fission. 2Phatal provides a powerful and rapidly adoptable platform to investigate in vivo functional consequences and neural plasticity following cell death as well as apoptosis, cell clearance and tissue remodelling in diverse organs and species.

SUBMITTER: Hill RA

PROVIDER: S-EPMC5501159 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Targeted two-photon chemical apoptotic ablation of defined cell types in vivo.

Hill Robert A RA Damisah Eyiyemisi C EC Chen Fuyi F Kwan Alex C AC Grutzendler Jaime J

Nature communications 20170616

A major bottleneck limiting understanding of mechanisms and consequences of cell death in complex organisms is the inability to induce and visualize this process with spatial and temporal precision in living animals. Here we report a technique termed two-photon chemical apoptotic targeted ablation (2Phatal) that uses focal illumination with a femtosecond-pulsed laser to bleach a nucleic acid-binding dye causing dose-dependent apoptosis of individual cells without collateral damage. Using 2Phatal ...[more]

PMID: 28621306

Similar Datasets

Project description:BackgroundDevelopment of a functional retina depends on regulated differentiation of several types of neurons and generation of a highly complex network between the different types of neurons. In addition, each type of retinal neuron includes several distinct morphological types. Very little is known about the mechanisms responsible for generating this diversity of retinal neurons, which may also display specific patterns of regional distribution.ResultsIn a screen in zebrafish, using a trapping vector carrying an engineered yeast Gal4 transcription activator and a UAS:eGFP reporter cassette, we have identified two transgenic lines of zebrafish co-expressing eGFP and Gal4 in specific subsets of retinal bipolar cells. The eGFP-labelling facilitated analysis of axon terminals within the inner plexiform layer of the adult retina and showed that the fluorescent bipolar cells correspond to previously defined morphological types. Strong regional restriction of eGFP-positive bipolar cells to the central part of the retina surrounding the optic nerve was observed in adult zebrafish. Furthermore, we achieved specific ablation of the labelled bipolar cells in 5 days old larvae, using a bacterial nitroreductase gene under Gal4-UAS control in combination with the prodrug metronidazole. Following prodrug treatment, nitroreductase expressing bipolar cells were efficiently ablated without affecting surrounding retina architecture, and recovery occurred within a few days due to increased generation of new bipolar cells.ConclusionThis report shows that enhancer trapping can be applied to label distinct morphological types of bipolar cells in the zebrafish retina. The genetic labelling of these cells yielded co-expression of a modified Gal4 transcription activator and the fluorescent marker eGFP. Our work also demonstrates the potential utility of the Gal4-UAS system for induction of other transgenes, including a bacterial nitroreductase fusion gene, which can facilitate analysis of bipolar cell differentiation and how the retina recovers from specific ablation of these cells.

Dataset Information

Targeted two-photon chemical apoptotic ablation of defined cell types in vivo.

Publications

Targeted two-photon chemical apoptotic ablation of defined cell types in vivo.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets