Localized calcium signals in early zebrafish development.

Activation of the phosphoinositide (PI) pathway has been shown to be involved in the compaction of blastomeres in mouse embryos and in embryonic axis formation in Xenopus and in zebrafish embryos. Here we investigate Ca2+ signals in individual blastomeres of zebrafish embryos with the goal to better understand the role of PI and Ca2+ signaling for early vertebrate embryogenesis. Initial studies showed that the inositol 1,4,5-trisphosphate (IP3) concentration increases after the 32-cell stage of development, suggesting that IP3-mediated Ca2+ signals may be present during the blastula stage. Ca2+ signals were measured by identifying individual cells using confocal imaging of a nuclear localized Ca2+ indicator. Using this in situ indicator, changes in Ca2+ concentration were measured over several hours in each cell of a series of sections through the developing embryo. Transient increases in Ca2+ concentration that lasted 20-50 sec (Ca2+ spikes) were first triggered during the 32- to 128-cell stage in cells of the outer embryonic cell layer. These cells develop epithelial characteristics and specialize into the enveloping layer (EVL). No Ca2+ activity was observed during the earlier cleavage cycles or in deep blastomeres. Ca2+ spikes remained restricted to the EVL until the end of the blastula stage. Ca2+ spikes in neighboring EVL cells often occurred in the same short time interval, indicating that small groups of EVL cells can synchronize their activity. When averaged over several cell cycles, Ca2+ activity showed an even distribution in the EVL and did not indicate future polarities.