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Fig. 2
The sequence of cellular events preceding pancreatic islet parasympathetic innervation is revealed with single-cell resolution time-lapse confocal imaging.
(A) Tg(elavl3:Gal4-VP16); Tg(UAS:EGFP-CAAX); Tg(sst2:RFP) zebrafish mounted in 0.5% agarose containing 0.017% tricaine were imaged with laser scanning confocal microscopy at 20 min time intervals. Maximum intensity projections of selected timeframes are presented (A, anterior; D, dorsal). Yellow arrows point to the detachment of neurons from the pancreatic islet. (B) Whole mount immunostaining at 34 hpf for GFP (neurons), RFP (delta cells), Insulin (beta cells), and Glucagon (alpha cells) after 10 hr time-lapse imaging. The detached neurons are not positive for endocrine cell markers. (C-E) Confocal imaging of Tg(elavl3:Gal4-VP16); Tg(UAS:EGFP-CAAX); Tg(sst2:RFP) zebrafish at 20–30 min time intervals. Maximum intensity projections of selected timeframes are presented. Yellow arrows point to the cellular events of interest. (F) Quantification of the time when the indicated cellular events were observed for individual fish (mean ± SEM). (G) Schematic of the sequence of cellular events preceding parasympathetic innervation of the pancreatic islet. V, vagus nerve; E, enteric nerve.