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. 2018 Jan 12;8:625. doi: 10.1038/s41598-017-18611-4

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© The Author(s) 2018

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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Loss of buccal/opercular water by angiotensin II (AngII) and its induction of migration to water. (a) The effect of AngII on amount of drinking on land. The diagram shows the experimental design. After the intracerebroventricular injection of AngII or vehicle, water was completely drained out for 15 min. Mudskippers could hold water in the buccal/opercular cavity and drink it even in the empty tank. Two-way factorial ANOVA was used for statistical analysis and the interaction was significant. *P < 0.05, n = 6. (b) X-ray images showing the presence of water (deep gray) in the buccal and opercular cavity. Arrow heads indicate air in these cavities. A thick arrow in the high-contrast image shows water in the anterior part of the oesophagus before swallowing (see Supplementary Movie 1). Broken lines show the gastrointestinal tracts. (c) A picture showing a tube which pierced the operculum to artificially eliminate buccal/opercular water. (d) Amphibious behaviour of ‘pierced’ mudskippers for 30 min. *P < 0.05 with unpaired Wilcoxon signed-rank test, n = 6. (e) Effect of decreased buccal/opercular water by placing water-absorptive dry resin (‘dry resin’) on amphibious behaviour. Controls were treated with the water pre-absorbed resin (‘wet resin’). *P < 0.05 with paired Wilcoxon signed-rank test, n = 8. Data are shown as mean ± s.e.m.