1. Imaging early embryonic calcium activity with GCaMP6s transgenic zebrafish.
- Author
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Chen J, Xia L, Bruchas MR, and Solnica-Krezel L
- Subjects
- Actins genetics, Animals, Animals, Genetically Modified, Blastomeres chemistry, Blastomeres ultrastructure, Blastula chemistry, Blastula ultrastructure, Body Patterning, Calmodulin genetics, Embryo, Nonmammalian chemistry, Embryo, Nonmammalian metabolism, Embryo, Nonmammalian ultrastructure, Genes, Reporter, Green Fluorescent Proteins analysis, Green Fluorescent Proteins genetics, Peptide Fragments genetics, Peptides genetics, Promoter Regions, Genetic, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins metabolism, Ubiquitin genetics, Blastomeres metabolism, Calcium analysis, Calcium Signaling physiology, Recombinant Fusion Proteins analysis, Zebrafish embryology
- Abstract
Intracellular Ca
2+ signaling regulates cellular activities during embryogenesis and in adult organisms. We generated stable Tg[βactin2:GCaMP6s]stl351 and Tg[ubi:GCaMP6s]stl352 transgenic lines that combine the ubiquitously-expressed Ca2+ indicator GCaMP6s with the transparent characteristics of zebrafish embryos to achieve superior in vivo Ca2+ imaging. Using the Tg[βactin2:GCaMP6s]stl351 line featuring strong GCaMP6s expression from cleavage through gastrula stages, we detected higher frequency of Ca2+ transients in the superficial blastomeres during the blastula stages preceding the midblastula transition. Additionally, GCaMP6s also revealed that dorsal-biased Ca2+ signaling that follows the midblastula transition persisted longer during gastrulation, compared with earlier studies. We observed that dorsal-biased Ca2+ signaling is diminished in ventralized ichabod/β-catenin2 mutant embryos and ectopically induced in embryos dorsalized by excess β-catenin. During gastrulation, we directly visualized Ca2+ signaling in the dorsal forerunner cells, which form in a Nodal signaling dependent manner and later give rise to the laterality organ. We found that excess Nodal increases the number and the duration of Ca2+ transients specifically in the dorsal forerunner cells. The GCaMP6s transgenic lines described here enable unprecedented visualization of dynamic Ca2+ events from embryogenesis through adulthood, augmenting the zebrafish toolbox., (Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.)- Published
- 2017
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