Repeated Sevoflurane Exposure Acutely Does Not Disrupt the Gut Microbiome in Young Adult Female Rats

Abstract

Surgery and general anesthesia (GA) may accelerate cognitive decline, referred to as perioperative neurocognitive disorder (PND), and potentially contribute to neurodegenerative conditions such as Alzheimer’s disease. However, the underlying mechanisms, vulnerable populations, and prevention strategies remain unclear. Previous studies have shown that repeated exposure to the general anesthetic sevoflurane (Sevo) disrupts the brain-gut-microbiota (BGM) axis and induces neurocognitive deficits in male rats and their offspring. The present study investigates whether Sevo exerts similar disruptions in the gut microbiota in female rats and whether inhibition of the Na⁺-K⁺-2Cl⁻ (NKCC1) chloride importer with bumetanide provides protection comparable to that observed in Sevo exposed male rats, thereby elucidating potential sex-dependent mechanisms underlying Sevo-induced neurocognitive effects. Female Sprague-Dawley rats were exposed to 2.1% Sevo for 3 hours on postnatal days 56 (P56), P58, and P60 (Sevo group). Rats in the BS group received bumetanide (1.84 mg/kg, intraperitoneally) before each Sevo exposure. Rats in the Control group were placed in a novel cage and housed singly for an equivalent time on P56, P58, and P60. To control for nonspecific effects of bumetanide injections, rats in the Control and Sevo groups were injected with an equivalent volume of saline. Gut tissue samples were collected on P60, and bacterial diversity was assessed using 16S rRNA gene sequencing and bioinformatics analyses. Compared to Control rats, Sevo exposure produced no significant changes in the gut microbiome. Alpha and beta diversity metrics remained consistent across groups.  Differential abundance analysis revealed modest changes in gut microbial amplicon sequencing variants (ASVs), with more taxa depleted than enriched in both the Sevo and BS groups compared to the Control group. The genera Lactobacillus and Bacteroides showed increased abundance following treatment, while the overall community structure across all taxonomic levels remained similar among groups. The Firmicutes/Bacteroidetes ratios and Statistical Analysis of Metagenomic Profiles (STAMP) results further confirmed minimal compositional differences, consistent with the overlapping clustering patterns observed. Predictive functional profiling indicated a reduction in stress-tolerant bacterial phenotypes following Sevo exposure, which was restored by bumetanide. These findings suggest that, acutely, repeated Sevo exposure does not measurably alter the gut microbiome of young adult female rats and therefore is unlikely to contribute to their PND pathophysiology. Further studies are needed to investigate the long-term consequences of repeated Sevo exposure on the BGM axis in young adult female rats.