Xenbase Image ID: 82992

Fig. 5. Itpkb is required for Ca2+ wave propagation following wounding. (A) Knock down of itpkb impairs Ca2+ wave propagation after wounding. Embryos were coinjected with 2.5 ng mRNA encoding GEM GECO and 40 ng of itpkb-mo or moc. Examples of Ca2+ ratio high/low over time in a wounded moc embryo (Top), wounded itpkb-mo embryo (Middle), and wounded itpkb-mo embryo treated with 0.5 μM aIP4 (Bottom). (Bars, 20 μm.) Asterisks indicate wound site. (B) 3D spatiotemporal quantification of Ca2+ ratio high/low on moc embryos after wounding, in the first cell or wounded cell (solid black line), second cell (dark gray line), and third cell (light gray line). For first cell n = 11, second cell n = 17, and third cell n = 10. (C�E) Temporal quantification of Ca2+ ratio high/low after wounding in embryos treated with moc (black line), itpkb-mo (red line), and itpkb-mo + 0.5 μM aIP4 (green line), in the first cell line or wounded cell (C); second cell line (D); and third cell line (E). For moc first cell n = 11, second cell = 17, third cell = 10; for itpkb-mo first cell n = 5, second cell = 24, third cell = 23; for itpkb-mo + aIP4 first cell n = 5, second cell = 13, third cell = 19. (F�M) Temporal quantification of Ca2+ ratio high/low in second cell line after wounding embryos. (F and G) Knock down of itpkb impairs intracellular Ca2+ wave propagation after wounding. Embryos wounded in medium without (F) or with calcium (G). Gray, moc embryos; red, itpkb-mo embryos. (H and I) XeC delays Ca2+ wave propagation after wounding in moc embryos and has an additive inhibitory effect on itpkb-mo�injected embryos. (H) Moc-injected embryos. (I) itpkb-mo�injected embryos. Green, DMSO-treated embryos; pink, 4 μM XeC-treated embryos. (J and K) InsP3 rescues the delayed Ca2+ wave propagation on itpkb-mo�injected wounded embryos. (J) Moc-injected embryos. (K) Itpkb-mo�injected embryos. Fuchsia, water-treated embryos; purple, 0.5 μM inositol-1,4,5-triphosphate (IP3)-treated embryos. (L and M) Ry delays Ca2+ wave propagation after wounding in moc embryos, but does not have an additive inhibitory effect on itpkb-mo embryos. (L) Moc-injected embryos. (M) Itpkb-mo�injected embryos. Blue, ethanol-treated embryos; orange, 30 μM Ry-treated embryos. N is as in Fig. S6. Each graph was analyzed by two-way ANOVA. Results are shown as means � SEM. **P < 0.01, ***P < 0.001. NS, not significant. Image published in: Soto X et al. (2013) Copyright © 2013. Image reproduced with permission of the publisher and the copyright holder. This is an Open Access article distributed under the terms of the Creative Commons Attribution License. Larger Image Printer Friendly View

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