Fig. 5. Lymph node drainage exhibits circadian variation.

a Experimental outline. Lymph nodes in purple, black dashed square: area imaged. White dashed lines: regions of interest (ROI) around lymph nodes. b Mean intensity (arbitrary units, A.U.) of lymph node fluorescence during day (orange, for b–e: n = 12 mice) and night (blue, for b–e: n = 15 mice). Time 0 on all graphs indicate start of imaging, on average 8 min after tracer infusion start. Thick line: mean, shading: SEM. c Boxplot of lymph node intensity at minute 50. Two-sided t test, p = 0.0335. d Boxplot of the rate of lymph node filling calculated from the first 20 min recording. Mann–Whitney test, p = 0.0321. e Representative images from in vivo lymph node imaging across 50 min for day (orange) and night (blue). White scale bar: 2 mm. f Experimental outline. g Average fluorescence over time (thick line: mean, SEM: shading). h Boxplot of fluorescence in lymph nodes at 50 min, during active (green, for f–j: n = 6 mice) and rest (yellow, for f–j n = 8 mice) phases. Two-sided t test, p = 0.0231. i Boxplot of rate of lymph node filling. j Representative time lapse images for animals housed in constant light for 10 days. White scale bar: 2 mm. k Diagram of experiment for lymph node contraction rate after FITC-dextran cheek injection, black dashed square: imaging area (left) with representative lymph vessel and trace of contractions from two regions of interest over 2 min (right). White scale bar: 1 mm. White dotted lines: area of measurement for traces. l Boxplot of contractions per minute of lymph vessels during the day (n = 5 mice) and night (n = 7 mice) in LD, and active (n = 7 animals) and rest (n = 7 animals) after 10 days in constant light. m Summary diagram of circadian regulation of CSF distribution. All boxplots minima: minimum value, maxima: maximum value, center: median, box and whiskers: quartiles, individual animals: colored dots. Asterisk indicates p < 0.05; ns: not significant. Source data are provided as a Source Data file.