Fig. 3

ARQiv detection of cell loss and regeneration.

Inducible cell-type specific ablation in nitroreductase (NTR) expressing transgenic fish [29], [35] was used to test whether ARQiv could detect the loss and regeneration of fluorescently tagged cells: A–E) Tg(ins:PhiYFP-2A-nfsB, sst2:tagRFP)^lmc01 – targeting pancreatic beta cells, and F–J) Double transgenic Tg(pax6-DF4:gap43-CFP)^q01; Tg(-3.7rho:YFP-NTR)^gmc500 – targeting retinal rod photoreceptors. Beta cell targeting: A) Non-treated control larvae showing PhiYFP-NTR-expressing (i.e., targeted) beta cells (yellow) and neighboring tagRFP-expressing (i.e., non-targeted) delta cells (red) in the pancreas (arrow). B-B^*** series) Confocal time-lapse imaging shows reporter expression in beta and delta cells is stable in non-treated control larva from pre-treatment (B and B*, 3 dpf) through post-treatment (B^**, 4 dpf) and recovery (B^***, 10 dpf). C) MTZ-treated larvae showing loss YFP-NTR-expressing (i.e., targeted) pancreatic beta cells (arrow, absence of yellow). C–D series) Metronidazole (MTZ) treated larvae showing MTZ-induced loss of YFP-NTR-tagged beta cells (absence of yellow; C arrow, and D^**, 4 dpf) and subsequent regeneration after MTZ wash out and recovery (D^***, 10 dpf). Unperturbed reporter expression in neighboring tagRFP-tagged delta cells (red; C arrow, and D–D^***) demonstrates the specificity of the technique. E) ARQiv data demonstrating that beta cell regeneration kinetics can be monitored in late larval to juvenile stage fish (19 to 31 dpf). Paired t-test p-values: • = 0.01, • = 0.33, □ = 0.01. Rod photoreceptor targeting: F) Non-treated control larvae showing YFP-NTR-expressing rod cells (yellow) in eye. G–G^***) Confocal time-lapse series showing YFP reporter expression in rod cells (arrowheads) is stable over time in non-treated control larva from pre-treatment (G, 5 dpf) through post-treatment (G^*, 7 dpf) and recovery (G^***, 10 dpf). H-I series) Metronidazole (MTZ) treated larvae showing MTZ-induced loss of YFP-NTR-tagged rod cells (reduction of yellow in H, and I^**, 7 dpf) and subsequent regeneration after MTZ wash out and recovery (I^***, 10 dpf). J) ARQiv data demonstrating that rod cell regeneration kinetics can be monitored in larval to fish (5 to 10 dpf); paired t-test p-values: • = 0.001, • = 0.015, □ = 0.001. Data is presented as mean±standard error. All scale bars are 50 μm.