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. 2017 Aug 4;7:7351. doi: 10.1038/s41598-017-07820-6

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© The Author(s) 2017

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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In vivo imaging of cerebral cortical astrocytes using Pink Flamindo. (a) Image showing the experimental setup of a mouse under anaesthesia, positioned under a two-photon microscope. (b) Schematic of the cranial window for in vivo two-photon imaging. (c) Sequential images of cortical astrocytes expressing Pink Flamindo during exposure to 50 μM Fsk and 500 μM IBMX. Scale bar represents 100 μm. (d) Enlarged image of c (24 min) to indicate the regions of interest. Analysed cells are marked in red. Yellow dashed line indicates the coverslip border (see b). Scale bar represents 50 μm. (e) Population mean trace of fluorescence intensity for Pink Flamindo from the images shown in c. The topical application of Fsk, IBMX and compound washout (indicated by black horizontal bars) was repeated twice. The data represent the means ± standard deviation (n = 9 cells).