Figure 1

Experimental timeline and design—sequence of events during all the experiment (n = 15/group). The procedures start with fish acclimatizing to the isolated aquarium for 7 days (Acclimatization Week). Followed this step, firstly, on Day 1, basal fish’s aggressiveness level (represented in the figure by the green “A” letter) and stress response (Ventilation rate [VR]) to a social stimulus (represented in the figure by the green “B” letter”) were accessed through the mirror test. Afterward, fish started to receive the treated diets containing different cannabidiol (CBD) doses—0 (control), 1 (CBD 1), 10 (CBD 10) and 20 mg/kg (CBD 20). During the first four experimental weeks (Day 1 to Day 28), the aggressive behavior and the stress responses to a social stimulus were accessed once a week. In the fifth experimental week, some stress responses to a non-social stimulus (confinement stress) were measured (represented in the figure by the yellow “C” letter): on Day 29, blood samples were collected to analyze fish’s cortisol baseline levels; and on Day 35, confinement stress was applied to fish, and the VR and cortisol stress-induced levels were collected. Lastly, fish were euthanized (1500 µl/l of clove oil), and their testis were collected for morpho-histological analysis (gonads’ size and the number of spermatozoa).