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. 2017 Oct 11;7:12970. doi: 10.1038/s41598-017-13438-5

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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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Circulation of the cell membrane during cell migration. Photobleaching experiments in fast-migrating cells. Aggregation-competent cells were stained with Polaric, and the small vertical rectangle regions in the ventral (A and C) and dorsal cell membrane (B and D) were photobleached using a confocal microscope. Middle (A and B) and posterior (C and D) regions of the cells were bleached, respectively. In each of panel A–D, phase-contrast (left) and fluorescence (right) images before photobleaching, immediately after photobleaching, and after fluorescence recovery are shown. The directions of cell migration are indicated by black arrows. The lower figure in each panel shows a kymograph of the white rectangle shown in each top figure. The white dotted lines show the fluorescence recovery of bleached regions, and the solid lines show the advance of both ends (anterior and posterior) of the migrating cells, respectively. Graphs E and F show summaries of forward displacements of the bleached zones and tail edges of multiple cells (n = 18 each) in the ventral (E) and dorsal (F) for 4.8 sec, respectively. Note that the bleached regions did not move relative to the substratum, whereas the cell advanced. Data are presented as mean ± SD and analyzed by unpaired two-tailed Student’s t-test. ***P ≤ 0.0001. Bar, 10 µm.