lh3 expression is upregulated in the optic chiasm during early regeneration. (A) Timeline of induced Lh3 expression with Tg(hsp70:lh3-myc) during zebrafish development as well as regeneration (hpf, hours post-fertilization; dpf, days post-fertilization; hpt, hours post-transection). (B,C) At 72 hpt in lh3 siblings with no heat-shock induction, RGC axons cross the midline to innervate the optic tecta, while RGC axons in lh3 mutants turn away from the midline and project aberrantly (C, magenta arrowheads). (D,E) In lh3 siblings and mutants that received a heat shock, RGC axons project across the midline (E, white arrowheads) and innervate the optic tecta. Dashed lines indicate the outline of the eyes. Scale bar: 50 µm. (F) Quantification of the percentage of nerves at 72 hpt that grew across the midline to re-innervate the optic tectum, did not grow across the midline and instead grew along aberrant trajectories, or showed stalled axonal growth in Tg(isl2b:GFP) larvae with and without a regeneration heat shock. Numbers indicate total number of transected nerves. ****P<0.0001, *P=0.0279; n.s., P=0.4804 (Fisher's exact test). (G-I) lh3 mRNA expression in Tg(isl2b:GFP) larvae pre-transection and at 2-6 hpt. (G) Outlined areas of the retina and the optic chiasm are shown in H and I, respectively. Scale bars: 50 µm in G,I; 20 µm in H. Axons in the chiasm region at 2-6 hpt have not yet fully degenerated following transection. Dashed lines indicate the outline of the eyes in I. Representative images across timepoints are of different larvae.
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