Sevoflurane induces the inattention, cognitive impairment, and motor dysfunction by hypermethylation of retinoic acid receptor α in the medial prefrontal cortex of neonatal mice

Abstract Background : External stimuli during neurodevelopment can alter DNA methylation, affecting gene expression that disrupt brain development. The impact of multiple non‐consecutive sevoflurane (MNS) exposures on DNA methylation patterns and key methylated genes involved in neurodevelopmental toxicity remains unclear. Methods : Neonatal mice were non‐consecutively exposed to 3% sevoflurane for 2 h on postnatal days 6, 8, and 10. On postnatal day 30, cognitive function, attention level, and motor skills were assessed. Whole‐genome bisulfite sequencing, RNA‐sequencing, and bisulfite target sequencing were performed to assess DNA methylation patterns, transcriptomic changes, and hypermethylation sites of retinoic acid receptor α (RARα) in the medial prefrontal cortex (mPFC), respectively. RARα expression was measured using RT‐qPCR and immunofluorescence staining. Adeno‐associated virus mediated the down‐regulation or overexpression of RARα in the mPFC of mice. Results : Neonatal mice MNS exposures led to inattention, cognitive impairments, and motor dysfunction. MNS exposures caused 941 significantly differentially methylated promoter genes and 316 differentially expressed genes in the mPFC. Notably, the correlation between RARα promoter hypermethylation and its reduced mRNA expression was the most statistically significant ( r = ‒0.89; p < .05). RARα expression was significantly reduced (1.00 ± 0.01 vs. 0.63 ± 0.03; p < .001). Knockdown RARα in the mPFC induced inattention, cognitive impairments, motor dysfunction, and dendrites morphology alterations. Restoration of RARα expression in the mPFC mitigated the neurotoxicity caused by neonatal MNS exposures. Conclusion : Neonatal MNS exposures induce inattention, cognitive impairment, and motor dysfunction by down‐regulating RARα through promoter hypermethylation. These findings suggest RARα as a promising therapeutic targeted for interventions in pediatric anesthesia to reduce long‐term neurodevelopmental toxicity.