Chronic Administration of Calendula officinalis Ethanolic Extract Mitigates Anxiety-like Behavior and Cognitive Impairment Induced by Acute Scopolamine Exposure in Zebrafish

<b>Background/Objectives:</b> Scopolamine (SCO) is widely employed as a pharmacological model of anxiety and amnesia in both rodents and zebrafish, the latter representing a valuable translational model in neuropsychopharmacology. The present study aimed to evaluate the neuroprotective and antioxidant potential of chronic administration of an ethanolic extract from <i>Calendula officinalis</i> flowers (CEE). <b>Methods:</b> Adult zebrafish (n = 10/group, both sexes) were exposed to CEE at concentrations of 1, 3, and 10 mg/L, administered daily for 22 consecutive days. After the initial 7-day pretreatment period, fish were challenged with SCO (100 μM, immersion for 30 min) followed by behavioral testing, including the Novel Tank Diving Test, Light/Dark Test, Novel Approach Test, Y-Maze, and Novel Object Recognition. Subsequently, brain homogenates were analyzed for acetylcholinesterase (AChE) activity, antioxidant enzymes (superoxide dismutase-SOD, catalase-CAT, glutathione peroxidase-GPx), reduced glutathione (GSH), protein carbonyls, and malondialdehyde (MDA). <b>Results:</b> Chronic CEE administration significantly attenuated scopolamine-induced anxiety-like behaviors and improved spatial memory (Y-maze) and recognition memory (NOR), as well as reduced anxiety-like behavior in the SCO-induced zebrafish model. Biochemical analyses revealed that CEE restored AChE activity, enhanced the activity of SOD, CAT, and GPx, and increased GSH levels, while concomitantly reducing protein oxidation and lipid peroxidation. The most pronounced effects were observed at 3 mg/L, which nearly normalized both behavioral and biochemical parameters. <b>Conclusions:</b> The CEE exerted anxiolytic and procognitive effects in zebrafish through combined cholinergic and antioxidant mechanisms. These findings highlight its translational potential as a promising candidate for the prevention and treatment of anxiety-related and cognitive disorders.