Ketamine Metabolites Promote Anxiolysis and Hydrocortisone Stress Buffering in Zebrafish

Anxiety and stress-related disorders affect over 280 million individuals worldwide and remain inadequately treated by current pharmacotherapies, which are often limited in efficacy and tolerability. Ketamine produces rapid therapeutic effects in mood and anxiety disorders, but its clinical use is constrained by dissociative side effects and abuse liability. Ketamine metabolites, particularly (2 R,6 R)-hydroxynorketamine (RANK) and (2S,6S)-hydroxynorketamine (SHANK), may retain therapeutic benefits while minimizing adverse effects, but their behavioral and stress-dependent actions remain incompletely understood. In the present study, we used zebrafish to examine stereospecific behavioral effects of hydroxynorketamine isomers under baseline and hydrocortisone-induced stress conditions. Behavior was assessed using the novel tank diving test alongside measures of locomotor activity and erratic movement. Under baseline conditions, RANK produced a robust anxiolytic-like profile, characterized by reduced bottom-dwelling and decreased erratic movements, whereas SHANK increased anxiety-like behavior, reflected by increased bottom-dwelling and prolonged latency to ascend. Hydrocortisone exposure induced a hyperarousal phenotype marked by increased locomotion and erratic movements without altering classical anxiety-like measures. Pretreatment with either RANK or SHANK attenuated hydrocortisone-induced hyperarousal, restoring behavior toward control levels; however, only RANK produced consistent anxiolytic-like effects across conditions. These findings demonstrate stereospecific and context-dependent behavioral effects of hydroxynorketamine isomers and highlight their differential modulation of baseline anxiety-like behavior and stress-induced hyperarousal.