Maintenance of ion homeostasis is essential for normal brain function. Inhalational anesthetics are known to act on various receptors but their effects on ion homeostatic systems, such as the Na+/K+-ATPase, remain largely unexplored. Based on reports demonstrating global network activity and wakefulness modulation by interstitial ions, we hypothesized that deep isoflurane anesthesia affects ion homeostasis and the key mechanism for clearing extracellular K+, the Na+/K+-ATPase.
Using ion-selective microelectrodes, we assessed isoflurane-induced extracellular ion dynamics in cortical slices of male and female Wistar rats in the absence of synaptic activity, the presence of two-pore-domain K+ channel antagonists, during seizures, and spreading depolarizations. We measured specific isoflurane effects on Na+/K+-ATPase function using a coupled enzyme assay and studied the relevance of our findings in vivo and in silico.
Isoflurane concentrations clinically relevant for burst suppression anesthesia increased baseline [K+]o (mean±SD: 3.0 ± 0.0 versus 3.9 ± 0.5mM, p<0.001, n=39) and lowered [Na+]o (153.4 ± 0.8 versus 145.2 ± 6.0mM, p<0.001, n=28). Similar changes in [K+]o, [Na+]o, and a substantial drop in [Ca2+]o (1.5 ± 0.0 versus 1.2 ± 0.1mM, p=0.001, n=16) during inhibition of synaptic activity and two-pore-domain K+ suggested a different underlying mechanism. Following seizure-like events and spreading depolarization, isoflurane greatly slowed [K+]o clearance (63.4 ± 18.2 versus 196.2 ± 82.4s, p<0.001, n=14). Na+/K+-ATPase activity was markedly reduced after isoflurane exposure (>25%), affecting specifically the α2/3 activity fraction. In vivo, isoflurane-induced burst suppression resulted in impaired [K+]o clearance and interstitial K+ accumulation. A computational biophysical model reproduced the observed effects on [K+]o and displayed intensified bursting when Na+/K+-ATPase activity was reduced by 35%. Finally, Na+/K+-ATPase inhibition with ouabain induced burst-like activity during light anesthesia in vivo.
Our results demonstrate cortical ion homeostasis perturbation and specific Na+/K+-ATPase impairment during deep isoflurane anesthesia. Slowed K+ clearance and extracellular accumulation might modulate cortical excitability during burst suppression generation while prolonged Na+/K+-ATPase impairment could contribute to neuronal dysfunction after deep anesthesia.