Fig. 4

Fig. 4

a The light-illumination paradigm of CaPs. The spinal cord of Tg[SAIG213A] Tg[UAS:opTDP-43z] Tg[UAS:EGFP] fish at the segment 13–18 level were illuminated with a blue laser, and CaPs were subjected to morphological analysis at 48–50 hpf. A single CaP was analyzed from dorsal (28, 31 hpf) and lateral (48 hpf) views. The fluorescence intensity along the longest inner diameter (dashed magenta arrow) is plotted at each time point. Blue arrows indicate the presumptive cytoplasmic area, where the opTDP-43z signal is faint. b Cytoplasmic shift of opTDP-43 is evaluated as a relative value of minimal (F(min), cytoplasm) and maximal (F(max), nuclear) fluorescence intensity along the longest inner diameter (dashed magenta line). The results were obtained from 32 cells (28, 31 hpf) and 17 cells (48 hpf) in three independent fish. *p < 0.0001, **p < 0.0001 (unpaired t-test, two-tailed). c Axons of CaPs expressing opTDP-43z with (BL, middle) or without (Dark, left) blue light stimulation and mRFP1-CRY2olig with blue light stimulation (right). d, e The total axon length and branching frequency of CaP axons in Tg[SAIG213A] Tg[UAS:EGFP] fish raised under normal laboratory light-dark cycle (L/D, the same data sets in Fig. 1e, f), Tg[SAIG213A] Tg[UAS:opTDP-43z] Tg[UAS:EGFP] fish with (BL, 15 cells, 4 animals) or without (Dark, 15 cells, 4 animals) blue light stimulation, and Tg[SAIG213A] Tg[UAS:mRFP1-CRY2olig] Tg[UAS:EGFP] fish with the stimulation (7 cells, 2 animals). ***p = 0.0068. f, g CaPs (arrowhead) and other mnr2b-positive motor neurons in the segment 14 (f) and 13–17 (g) of Tg[SAIG213A] Tg[UAS:opTDP-43z] Tg[mnr2b-hs:EGFP-TDP43z] fish that was illuminated with a blue light during 28–32 hpf. h, i Cytoplasmic shift of opTDP-43z and EGFP-TDP-43z in the CaP in (f). The fluorescence intensities of opTDP-43z (magenta) and EGFP-TDP-43z (green) were plotted along the blue dashed arrows (h). Images shown are enhanced to identify soma outline. i The relative intensity of cytoplasmic signal (F(min)/F(max)) for opTDP-43z (magenta) and EGFP-TDP-43z (green) in each spinal segment. The bars indicate 5 µm (a, b), 10 µm (f, h), 20 µm (c), and 30 µm (g). Error bars show SD.