Fig. 3: Cholinergic stimulation triggers contractile motion of Acti-stained eccrine glands.
a Strategy to detect cell and nuclear displacement by color aberration. The top scheme shows the original video images of two merged channels. The middle scheme shows pseudocolored images of the actin channel at the starting point (T0, green) and at other time points (T1 and T2, magenta). When the images in magenta are merged with the initial image in green, cell displacement can be detected by the appearance of color aberrations. Cells that did not move are observed as entirely white due to the complete overlap of green and magenta. The bottom scheme shows pseudocolored images of the nuclei, showing color aberration. b Representative sequential time-lapse images of an eccrine gland after the application of PBS (control). The arrow indicates the time point at which PBS was added (t = 0’00”). The upper panels are overlaid fluorescence images of Acti-stain (yellow) and Hoechst 33342 (light blue). The lower panels show the color aberration images for actin (middle) and nuclei (bottom). The merged images remained white, indicating that actin filaments and nuclei did not move. c Representative sequential time-lapse images after 10 mM pilocarpine treatment. The panels are aligned in the same order as in b. The merged images yielded color aberration, indicative of displacement of actin filaments (arrows) and nuclei (arrowheads) after pilocarpine stimulation. d Hoechst 33342-stained eccrine gland. Representative sequential color-aberration images without Acti-stain (red box in the left image). The blue arrow indicates the time point of pilocarpine addition (t = 0’00”). e The maximum displacement of nuclei with (blue, n = 9) or without (red, n = 7) costaining with Acti-stain. **P < 0.001, unpaired t test. Panels (b), (c), and (d) have a temporal resolution of 30 seconds.
