Caspase-mediated apoptosis induction in zebrafish cerebellar Purkinje neurons

Abstract

The zebrafish is a well-established model organism in which to study in vivo mechanisms of cell communication, differentiation and function. Existing cell ablation methods are either invasive or they rely on the cellular expression of prokaryotic enzymes and the use of antibiotic drugs as cell death-inducing compounds. We have recently established a novel inducible genetic cell ablation system based on tamoxifen-inducible Caspase 8 activity, thereby exploiting mechanisms of cell death intrinsic to most cell types. Here, we prove its suitability in vivo by monitoring the ablation of cerebellar Purkinje cells (PCs) in transgenic zebrafish that co-express the inducible caspase and a fluorescent reporter. Incubation of larvae in tamoxifen for 8 h activated endogenous Caspase 3 and cell death, whereas incubation for 16 h led to the near-complete loss of PCs by apoptosis. We observed synchronous cell death autonomous to the PC population and phagocytosing microglia in the cerebellum, reminiscent of developmental apoptosis in the forebrain. Thus, induction of apoptosis through targeted activation of caspase by tamoxifen (ATTAC TM ) further expands the repertoire of genetic tools for conditional interrogation of cellular functions.

Microglia in the developing retina couple phagocytosis with the progression of apoptosis via P2RY12 signaling

“…We chose this timing for imaging because it coincides with the initial peak of developmental apoptosis in the retina and microglial colonization of the retina has begun . When complexed with DNA, AO displays green fluorescence and can be used to visualize apoptotic cells in whole tissue . In particular, AO has been used to image apoptotic cells in live zebrafish …”

“…When complexed with DNA, AO displays green fluorescence and can be used to visualize apoptotic cells in whole tissue . In particular, AO has been used to image apoptotic cells in live zebrafish …”

Modeling Neurodegenerative Spinocerebellar Ataxia Type 13 in Zebrafish Using a Purkinje Neuron Specific Tunable Coexpression System

“…A prerequisite though for studying cell biological and molecular mechanisms of cerebellar neuronal differentiation, function, homeostasis, plasticity, degeneration, and regeneration, is the specific accessibility of distinct cerebellar neuronal cell types for genetic manipulation (Distel et al, 2010;Matsui et al, 2014;Weber et al, 2016;Theisen et al, 2018). In zebrafish, a PC specific regulatory element of the zebrafish aldolase c gene encoding the ZebrinII antigen has been identified previously, which has been very useful in establishing PC reporter strains (Tanabe et al, 2010).…”

Molecular Characterization of Caspase8 in Grass Carp (Ctenopharyngodon Idella)

“…Generally, caspase8 was a crucial effector for downstream signal transduction of apoptosis in vertebrates. Given that several other genes involved in apoptosis were identified in a few fish [14,17], such as zebrafish and murrel, the characterization of caspase8 in grass carp appears to be particularly important. This study will provide new insights into the information of fish caspases, and an important theoretical basic for subsquently clarifying the role of caspasse8 in fish.…”