Isoflurane activates the type 1 ryanodine receptor to induce anesthesia in mice
Fig 4
Resistance to isoflurane in RyR1(M4003F) KI mice.
(A) Amino acid sequence around M4000 (rabbit RyR1) and the generation of RyR1(M4003F) KI mice. (B) The genomic sequences of the KI site for each genotype. (C) Dose–response curves for isoflurane-induced loss of righting reflex (LORR). Data were fitted with logistic functions (equation 5 in Materials and methods). N = 11 (WT), 14 (heterozygous [Hetero] KI), and 13 (homozygous [Homo] KI). The shaded regions represent 95% CIs. (D) Comparison of the concentrations at which the mice reached LORR (Mean ± SD with individual data points). N = 11 (WT), 14 (Hetero KI), and 13 (Homo KI). The adjusted P-values (Adj. P) by the Dunnett’s test are shown. (E) Representative transition of EEG signals, the spectrograms, and EMG signals during isoflurane treatment. At the baseline, mice were kept awake by gentle handling. (F) Normalized delta power (0.4–4 Hz) on EEG during isoflurane treatment for each animal. The mean values and individual data are shown. N = 8 (WT), 10 (Hetero KI), and 9 (Homo KI). (G) Changes in delta power over the baseline (0.0% isoflurane). Mean ± SD is shown. N = 8 (WT), 10 (Hetero KI), and 9 (Homo KI). Data were fitted using logistic functions (equation 6 in Materials and methods). RyR1, the type 1 ryanodine receptor; ssODN, single-stranded oligodeoxynucleotide; WT, wild-type; Hetero, heterozygous; Homo, homozygous; PAM, protospacer-adjacent motif; Iso., Isoflurane; LORR, loss of righting reflex; EEG, electroencephalogram; EMG, electromyogram; The mice illustration was modified from Openclipart (https://openclipart.org/).