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Fig. 4 Tfap2e KD is implicated in structural brain anomalies in the early developing zfl. (A?C) 3D modelling of the brain of Tg(?3.1ngn1:GFP) zfl on Tfap2e KD at 2?days post fertilisation (dpf) and box plot graph of the forebrain hemisphere cleft measured as indicated (white brackets). (A) shows a schematic depiction of zfl brain in transverse (left) and coronary view (right) (modified from Dworschak et al, 2021).28 Pink: olfactory blub. Turquoise: mesencephalon. Purple: rhombencephalon. (B) The representative images were obtained by in vivo 2-photon z?stack imaging and subsequent modulation with IMARIS software. Transverse (left) and coronary view (right). (C) TB MO and e3i3 morphant zfl present with an enlarged hemisphere cleft consistent with a hydrocephalus. Note that only TB MO but not e3i3 MO treated zfl show a reduced brain volume (see online supplemental data 13). (D?E) Representative confocal in vivo images of Tg(?3.1ngn1:GFP) zfl at 2 dpf in transverse view after sulforhodamine SR101 injection (in red) into the diencephalic ventricle and box plot graph of ventricle diameters (white brackets) in relation to hemisphere diameter (dotted line). In TB MO zfl, both the diencephalic and the rhombencephalic ventricle are enlarged in the sense of a hydrocephalus. ***p<0.001, ****p<0.0001 (C: one-way ANOVA with Tukey?s multiple comparison; E: two-way ANOVA with Tukey?s multiple comparison). Number of independent experiments n=3 for all graphs, number of investigated zfl n as indicated. White scale bars: 200 µm. ANOVA, analysis of variance; DV, diencephalic ventricle; KD, knockdown; MO, morpholino oligonucleotide; ns, not significant; RV, rhombencephalic ventricle; TB, translation blocking; TV telencephalic ventricle; zfl, zebrafish larvae.