Atomoxetine hydrochloride impairs human sperm quality and function via disruption of calcium homeostasis and PKC signaling: A combined network pharmacology and in vitro study

Abstract

Atomoxetine Hydrochloride (ATX) is a widely prescribed first-line pharmacotherapy for adult attention deficit hyperactivity disorder (ADHD). Despite its chronic use among men of reproductive age, the potential impact of ATX on male fertility remains largely unknown. This study aimed to elucidate the toxicological effects of ATX on human sperm and to determine the underlying molecular mechanisms. In vitro exposure of human sperm to ATX (1–500 μM) revealed a significant concentration- and time-dependent inhibition of sperm motility. A critical mechanistic dissociation was observed: severe cytotoxicity, mitochondrial depolarization, and massive ROS accumulation (>6-fold increase) occurred exclusively at ≥ 100 μM. Conversely, 50 μM ATX potently reduced progressive motility (27.8% vs. 53.0% in controls at 120 min) without significantly compromising cell viability (68.9% vs. 74.4%) or mitochondrial function, indicating a specific signaling disruption. Network pharmacological analysis was employed to identify calcium signaling and Protein Kinase C (PKC) as key candidate pathways. Validation experiments using fluorescent assays confirmed that ATX disrupted physiological calcium homeostasis, specifically abolishing progesterone-induced calcium influx. Co-treatment with the PKC agonist PMA (100 nM) significantly reversed this calcium suppression (∼58%), alongside rescuing defects in motility, capacitation, and the acrosome reaction. These findings demonstrate that ATX compromises human sperm function primarily by disrupting PKC-dependent calcium channel regulation. These results highlight the potential reproductive risks, warranting further clinical assessment of patients undergoing chronic ATX therapy.