A drop in interstitial osmotic pressure triggers ATP release in wounded tail fins. (a) Representative fluorescence/luminescence images from a zebrafish larva at the indicated times after tail fin amputation in isotonic mounting agarose. After 20 min of isotonic incubation (left), the fish was overlaid with a bolus of hypotonic bathing solution (right). Red, SYTOX orange. Green, luminescence. Orientation of the tissue relative to the wound margin (yellow broken line) is indicated. Bars, 100 µm. (b) Representative kymograph (n = 6 for 20 min isotonic preincubation, n = 4 for 10 min isotonic preincubation) of luminescence emissions acquired by line scans of the wound margin (yellow broken line in Fig. 5 a). Asterisks mark the representative time points (shown in a) on the kymograph. White/yellow, high luminescence emission. Blue/green, low luminescence emission. Bar, 100 µm. (c) Representative intensity plots of the time-lapse data shown in panel a displaying either luciferase luminescence (left) or SYTOX orange fluorescence (right); plots correspond to intensity measurements taken within a 100-µm-diameter region of interest around the center of each ATP luminescent flash. Measurements start one frame before, and end 1–2 frames after each luminescence peak (40 s per frame). Color-coded traces correspond to matching measurements of respective luminescence and fluorescence. The intensity measurements were performed on one representative sample from the dataset depicted in a.