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. 2019 Aug 23;10:3830. doi: 10.1038/s41467-019-11739-z

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 4 — Glial cells are highly active and strongly synchronized already during preictal period. a Confocal image of a transgenic zebrafish larvae expressing GCaMP6s in GFAP positive glial cells (green) and jRCaMP1a in elavl3 positive neurons (magenta), dorsal view. b Glial calcium signals measured by two-photon microscopy in Tg(GFAP:Gal4)nw7;Tg(UAS:GCaMP6s) transgenic zebrafish larva. White dashed line indicates the start of 20 mM pentylenetetrazole (PTZ) perfusion. c Simultaneous patch-clamp recording of glial membrane potential and epifluorescence calcium recordings of Tg(GFAP:Gal4)nw7;Tg(UAS:GCaMP6s) transgenic zebrafish, during PTZ-induced epileptic activity. d Correlation of electrical activity and calcium signals from individual glial cells, n = 6 cells/zebrafish. e Percentage of active glial cells (>3std_baseline) during baseline (black), preictal (gray), and seizure (red) periods. f Average activity of the active glial cells, defined by the area under the curve of the ΔF/F trace. g Average pairwise Pearson’s correlation between glial cells. h Relation between pairwise correlation of glial activity and the distance between each glial pair. Dotted lines represent the results when glial locations are shuffled. i Histogram representing the distribution of the correlation coefficients between glial cells during baseline (black), preictal (gray) and seizure periods (red). j Confocal image showing GFAP positive radial glia expressing GCaMP6s (green) and neurobiotin coupling between glial cells (magenta) after filling a single glia with neurobiotin by patch-clamp electrode. Location of the patch-clamped glial cell is indicated with *. A, anterior; P, posterior; L, left; R, right. White bars reflect 100 µm (a, j). ***p = < 0.001, *p = < 0.05, ns = not significant, Wilcoxon signed-rank test. Error bars and shaded regions represent the s.d. of n = 6 cells (d) or s.e.m. of n = 11 fish (e–i)