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Dev Biol. 2017 Oct 15;430(2):385-396. doi: 10.1016/j.ydbio.2017.03.010. Epub 2017 Mar 18.

Imaging early embryonic calcium activity with GCaMP6s transgenic zebrafish.

Author information

1
Department of Developmental Biology, Washington University School of Medicine, St. Louis, MO 63110, USA.
2
Department of Anesthesiology, Division of Basic Research, Washington University School of Medicine, St. Louis, MO 63110, USA; Department of Biomedical Engineering, Washington University in St. Louis, St Louis, MO, 63105, USA.
3
Department of Anesthesiology, Division of Basic Research, Washington University School of Medicine, St. Louis, MO 63110, USA; Department of Biomedical Engineering, Washington University in St. Louis, St Louis, MO, 63105, USA; Department of Neuroscience, Washington University in St. Louis, St. Louis, MO, USA.
4
Department of Developmental Biology, Washington University School of Medicine, St. Louis, MO 63110, USA. Electronic address: solnical@wustl.edu.

Abstract

Intracellular Ca2+ 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.

KEYWORDS:

Calcium transients; Dorsal forerunner cells; Embryonic cleavages; Gastrulation; Nodal; β-catenin

PMID:
28322738
PMCID:
PMC5835148
DOI:
10.1016/j.ydbio.2017.03.010
[Indexed for MEDLINE]
Free PMC Article

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