High‐resolution analysis of central nervous system expression patterns in zebrafish Gal4 enhancer‐trap lines

Abstract

Background The application of the Gal4/UAS system to enhancer and gene trapping screens in zebrafish has greatly increased the ability to label and manipulate cell populations in multiple tissues, including the central nervous system (CNS). However the ability to select existing lines for specific applications has been limited by the lack of detailed expression analysis. Results We describe a Gal4 enhancer trap screen in which we used advanced image analysis, including 3D confocal reconstructions and documentation of expression patterns at multiple developmental timepoints. In all, we have created and annotated 98 lines exhibiting a wide range of expression patterns, most of which include CNS expression. Expression was also observed in non-neural tissues such as muscle, skin epithelium, vasculature, and neural crest derivatives. All lines and data are publicly available from the Zebrafish International Research Center (ZIRC) from the Zebrafish Model Organism Database (ZFIN). Conclusions Our detailed documentation of expression patterns, combined with the public availability of images and fish lines, provides a valuable resource for researchers wishing to study CNS development and function in zebrafish. Our data also suggest that many existing enhancer trap lines may have previously uncharacterized expression in multiple tissues and cell types.

Motor Behavior Mediated by Continuously Generated Dopaminergic Neurons in the Zebrafish Hypothalamus Recovers after Cell Ablation

“…We therefore took a genetic approach to ablate th2 + cells and examine the resulting effects on behavior. We generated a th2:Gal4 transgenic line [ Tg(th2:Gal-VP16) zd202 ] which was crossed to Tg(UAS-E1b:NTR-mCherry) c264 fish [18] to drive Nitroreductase (NTR) expression in th2 + neurons (Figure S1A) and enable chemical ablation [19, 20]. While the hypothalamic posterior recess of untreated brains had an average of 128.7 ± 20.6 mCherry+ cells (S.E.M., n=7 brains), after addition of 5 mM metronidazole (MTZ) to double transgenic embryos from 5–7 dpf, we observed a reduction to 18.1 ± 2.4 cells/brain (S.E.M., n=10 brains), a decrease of 86% (Figure 3A–B).…”

“…Ablation of th2+ neurons significantly reduced both the bout frequency and total amount of time spent swimming, but not the amplitude of individual swimming bouts (Figure 3C–E, Figure S1C, Movies S1–S2, p=0.009 for swim frequency; p=0.020 for swim time by 2-way ANOVA). In contrast treating non-transgenic siblings with MTZ (Figure 3C–E), and ablation of hypothalamic radial glia using a separate transgenic line [18] (Figure S1D–E), did not affect swimming behavior. Decreased frequency of swimming was also observed following MTZ treatment of an independent transgenic line, expressing NTR-GFP directly under the control of the th2 enhancer (Figure 3F–I and Figure S1B).…”

Hypothalamic radial glia function as self-renewing neural progenitors in the absence of Wnt/ß-catenin signaling

“…To test whether hypothalamic radial glia show a regenerative response similar to other neural stem cell populations, we used the Et(Gal4-VP16,myl7:gfp) zc1066a enhancer trap line in combination with the Tg(UAS-E1b:NTR-mCherry) jh17 effector line to express Nitroreductase (NTR) specifically in radial glia of the posterior recess (Otsuna et al, 2015), and thus ablate cells using metronidazole (MTZ) (Davison et al, 2007;Pisharath et al, 2007). Consistent with previous studies, incubation of nontransgenic larvae in 1 mM MTZ did not significantly affect cell proliferation or cell death (data not shown).…”

A toolbox to study epidermal cell types in zebrafish

“…To identify promoters driving expression in diverse cell types of the developing zebrafish epidermis, we screened a collection of Gal4 enhancer trap lines from a large-scale non-biased screen focused on identifying novel neural-specific lines in Danio rerio (Otsuna et al, 2015). The developing zebrafish epidermis is a bilayer during embryonic and larval development that comprises cells similar in function and markers to those comprising mammalian bronchial epithelia ( Fig.…”

“…To generate the enhancer-trap lines, the Et(Gal4-VP16;myl7:GFP) plasmid in the Tol2 backbone and transposase mRNA was injected into 1-cell-stage developing zebrafish embryos, as described in Otsuna et al (2015). Briefly, F0 embryos were raised to maturity and then crossed to a Tg(UAS-E1b:nfsBmCherry) c264 reporter line.…”