Commonly targeted cell types for ablation studies in zebrafish.

See S.Table 1 for more complete list. Image of a 5 days post-fertilization (dpf) Casper zebrafish larval59 with approximate position of cell type ablated. Name of cell, and transgene provided along with reference. Orange highlights indicate transgenic models used to study human pathologies.

NTR/MTZ-based screening platforms in zebrafish.

Overview of the integrated chemogenetic screening workflow using nitroreductase (NTR)-mediated ablation. (A) Experimental design showing transgenic zebrafish expressing NTR in target tissues, baseline imaging, and subsequent metronidazole (MTZ) treatment to induce cell-type-specific ablation. The use of parallel transgenic controls and multiwell plate layout enables quantitative assessment of tissue loss and recovery. (B) High-content chemical screening pipeline integrating automated imaging, hit identification, and pathway-level analysis using standardized statistical metrics. (C) Genetic screening framework coupling sgRNA-based mutagenesis with imaging-based phenotype scoring to uncover modifiers of cell loss or regeneration. (D) Behavioral assays to quantify functional recovery or pharmacological response.

Schematic of bipartite systems to drive robust levels of NTR.

On left, driver lines lead to expression (dashed arrows) of transactivators (Gal4 or QF, blue, purple spheres) under the control of a regulatory element of interest (REI). These transactivators bind their upstream activating sequences (either UAS or QUAS, green boxes) to achieve controlled and amplified NTR expression (tan spheres) in target cells. NTR expression can be monitored by co-production of mCherry (red spheres) either as a fusion protein with NTR or as separate proteins due to P2A dependent ribosome ‘skipping’.[139].

Live imaging of cell-death kinetics

(A) Schematic of 6 dpf larvae showing region of fish imaged in C-N where the pancreatic islet is located (red/yellow). (B) Diagram of the two transgenes (ins:Hmgb1-GFP, ins:mCherry-2a-NTR2.0) in the fish in C-N. The insulin promoter (grey box) drives expression of the following: (B-above) an Hmgb1-GFP fusion protein and (B-below) NTR2.0 and mCherry [presence of the P2A (blue box) induces ribosomal skipping producing separate proteins]. C-N Confocal images of the islet of in three larval fish over a time course from 6 dpf to 7 dpf (times along the X axis). C-F negative control – no MTZ (0). G-J fish treated with high MTZ dose (1mM). K-N fish treated with a low dose MTZ (10μM). G-N Dying β cells first lose red fluorescence, revealing green nuclei (arrow heads). A higher dose shows appearance of green nuclei (H) earlier than the lower dose (N). J 24 hrs in 1mM and no debris remains.