Generation of living color transgenic zebrafish to trace somatostatin-expressing cells and endocrine pancreas organization.

In the present study, both gfp and rfp transgenic zebrafish lines using a 2.5-kb zebrafish somatostatin2 (sst2) promoter were generated. During embryonic development, expression of GFP/RFP in the endocrine pancreas of transgenic embryos was initiated at approximately 20 hpf and the number of GFP/RFP positive cells in the pancreas increased in subsequent stages; thus, our newly generated Tg(sst2:gfp) and Tg(sst2:rfp) lines faithfully recapitulated sst2 expression in endocrine pancreatic cells and provided a useful tool in analyzing the development of Sst2-producing delta-cells in the pancreas. By crossing these new transgenic lines with previously available transgenic lines targeted in insulin (Ins)-producing beta-cells, Tg(ins:gfp) and Tg(ins:rfp), in combination with immunodetection of glucagon (Gcg)-producing alpha-cells and pancreatic polypeptide (PP)-producing PP-cells, the organization and composition of endocrine islets were investigated in both embryonic and adult pancreas. We found that there was always a big cluster of endocrine cells (principal islet) in the anterior-dorsal pancreas, followed by numerous smaller clusters (variable in size) of endocrine cells (secondary islets) along the anterior-posterior axis of the pancreas. All four types of endocrine cells were found in the principal islet, but secondary islets may or may not contain PP-cells. In addition, there were also discrete endocrine cells throughout the pancreas. In all co-localization experiments, we did not find any endocrine cells positive for more than one hormone markers, suggesting that these endocrine cells produce only a single hormone. In both principal and secondary islets, we found that beta-cells were generally located in the center and non-beta cells in the periphery; reminiscent of the "mantel-core" organization of islets of Langerhans in mammals where beta-cells form the core and non-beta-cells the mantel. In zebrafish primary islet, beta-cells constitute most of the mass (approximately 50%), followed by delta-cells and alpha-cells (20-25% each), and PP-cells (1-2%); this is also similar to the composition of mammalian islets.