Experimental design. Fish in the stress group were exposed to an 8-day stress protocol (UCS), while fish in the control group remained under standard conditions. All fish were subjected to behavioral tests before and after the stress protocol. On day 10, caudal fins were amputated, and growth measurements were recorded on days 13, 16, 19, and 23.

Effects in behavioral parameters (open field test and light/dark preference test) in zebrafish after exposure to a chronic stress protocol at day 9. (A) Total distance traveled in the open field test. (B) Distance to the center of the arena to evaluate thigmotaxis. (C) Total number of transitions between white and black zones during light/dark preference test. (D) Total time spent in the white zone of the tank during light/dark preference test as an exploratory behavior. Boxplots in panels (A–D), are drawn such that the upper and lower whiskers limit interquartile range (Q1, Q3) and the line represents the median (Q2). (E) Diagrams illustrating variable being measured in each case. Top: track visualization used to obtain the total distance traveled of zebrafish. Middle: heatmap visualization and how distance to the center point of the arena was defined. Bottom: Illustration of the number of transitions between the white and the black zones, and time spend in the white zone only. All results are expressed as the mean ± standard error of mean (S.E.M). Statistical analysis corresponds to Welch´s t-test. *p < 0.05; **p < 0.01; ***p < 0.001.

Regeneration process after the stress protocol. The regeneration index is calculated as the length of the regenerated tissue/initial length of the caudal fin of each fish measured from the most proximal part of the lepidotrichia. (A) Images of caudal fin growth measurements after amputation for young control fish (top row) and young stressed fish (bottom row). (B) Images of caudal fin growth measurements after amputation for old control fish (top row) and old stressed fish (bottom row). Black arrows indicate the amputation site. An additional measurement was done at 9 dpa but the image was excluded for clarity; complete regeneration process shown in Supplementary Figure S2 for young and Supplementary Figure S3 for old fish. Note heterogeneous growth in old fish (dashed line arrow). Further results on these unusual growth pattern can be seen in Supplementary Figure S4. (C) Graph comparing the regeneration index of the control and stress young caudal fin at 3, 6, 9, and 13 dpa; two cohorts included in this experiment. (D) Graph comparing the regeneration index of the control and stress old caudal fin at 3, 6, 9, and 13 dpa; two cohorts included in this experiment. All results are expressed as the mean ± standard error of mean (S.E.M). *p < 0.05; **p < 0.01; ***p < 0.001.

Relative expression of (A)sam2 and (B)nlgn1 in the brain of old fish. Relative expression of sam2 and nlgn1 between control and stress groups, calculated as fold-change differences in expression to the average expression of two housekeeping genes All results are expressed as the mean ± standard error: *p < 0.05.