Pre-Activation of Mitophagy Protects Against Hyperbaric Oxygen-Induced Central Nervous System Oxygen Toxicity

Central nervous system oxygen toxicity (CNS-OT) is a major complication of hyperbaric oxygen (HBO) characterized by seizures and neuronal damage, yet the underlying mechanisms remain incompletely understood. Using male Sprague Dawley rats (n = 6 per group) and HT22 neurons exposed to either HBO (6 ATA, 100% O₂) or hyperbaric normoxia (HNO), our results demonstrate that HBO, but not HNO, caused mitochondrial structural damage and loss of mitochondrial membrane potential (ΔΨm). Transcriptomic analysis revealed enrichment of apoptosis and mitogen-activated protein kinase (MAPK) signaling pathways. Using HeLa cells stably overexpressing Parkin and Mito-Keima, a pH-sensitive mitochondrial probe system for monitoring mitophagy, we observed that mitophagic flux was initiated but proceeded too slowly to clear damaged mitochondria in a timely manner in HBO-exposed neurons. Pharmacological preconditioning to activate mitophagy enabled the prompt elimination of dysfunctional mitochondria and rescued HBO-induced mitochondrial dysfunction and cell death. In vivo, everolimus treatment promoted timely mitophagic clearance, prolonged seizure latency, attenuated ΔΨm loss, and suppressed p-p38 activation. These findings demonstrate that HBO exposure disrupts mitochondrial homeostasis and activates pro-apoptotic MAPK signaling. Meanwhile, endogenous mitophagy is initiated but fails to clear damaged mitochondria in a timely manner. Pre-activation of mitophagy by everolimus enables the timely clearance of damaged mitochondria, protecting against CNS-OT and highlighting a promising therapeutic strategy.