Menthol-induced sedation and hemodynamic safety limits in Nile tilapia: defining a therapeutic window

Menthol, a natural monoterpene extracted from Mentha species, has gained attention as an alternative sedative and anesthetic in aquaculture. However, its physiological safety range and potential sublethal toxicity remain insufficiently defined. This study investigated the cardiac and behavioral effects of menthol sedation in Oreochromis niloticus , aiming to identify a therapeutic window and elucidate its underlying mechanisms of action. Juvenile fish were exposed to graded concentrations of menthol (100–250 mg L⁻¹) to evaluate induction and recovery times, cardiac rhythm stability, and electrocardiographic parameters, including heart rate, QRS amplitude, and PQ and QT intervals. Additional assays with the GABAAA​ receptor antagonist flumazenil (1 mg kg⁻¹, i.p.) were performed to determine neurophysiological mediation. Menthol induced a concentration-dependent decrease in heart rate and prolongation of ventricular conduction intervals, characterizing sinus bradycardia without arrhythmia. Concentrations between 100 and 200 mg L⁻¹ provided effective sedation and rapid recovery while maintaining hemodynamic stability, defining a safe therapeutic window. Above 200 mg L⁻¹, pronounced bradycardia and delayed recovery were observed, indicating sublethal cardiac depression. Flumazenil significantly prolonged anesthesia induction, confirming GABAAA​-mediated mechanisms in menthol sedation. These findings establish the physiological safety limits for menthol use in Nile tilapia and highlight its ecotoxicological significance, as excessive or environmental exposure may impair cardiac homeostasis in aquatic organisms. The results contribute to defining safe anesthetic practices and to understanding the toxicodynamic implications of natural sedatives in aquatic ecosystems. Graphical Abstract [Insert Fig. 1 about here]