Figure 4

Isoflurane transactivates TrkB. (A) Pre-treatment with the AMPA receptor blocker NBQX (10 mg/kg, i.p.) does not prevent isoflurane (15 min) induced phosphorylations of TrkB (two-way ANOVA: F_3,20 = 4.379, p = 0.017; Tukey HSD post hoc test: CTRL vs. NBQX p = 0.972, CTRL vs. NBQX + ISO p = 0.075, CTRL vs. ISO p = 0.050, NBQX + ISO vs. ISO p = 0.996), CREB (two-way ANOVA: F_3,20 = 24.731, p < 0.001; Tukey HSD post hoc test: CTRL vs. NBQX p = 0.648, CTRL vs. NBQX + ISO p < 0.001, CTRL vs. ISO p < 0.001, NBQX + ISO vs. ISO p = 1.000) or p70S6K (two-way ANOVA: F_3,20 = 6.228, p = 0.0037; Tukey HSD post hoc test: CTRL vs. NBQX p = 0.859, CTRL vs. NBQX + ISO p = 0.041, CTRL vs. ISO p = 0.143, NBQX + ISO vs. ISO p = 0.916). pTrkB normalized to total TrkB, other phospho-proteins to GAPDH. n = 6/group. (B) The phosphorylation of eEF2 was not affected by isoflurane anesthesia (30 min) in the prefrontal cortex. n = 6. Student’s t test. (C) Total Bdnf mRNA (normalized to Gapdh mRNA) levels remain unaltered after isoflurane anesthesia (30 min) (n_Ctrl = 6; n_Iso = 5). (D) Mature BDNF protein levels remain unaltered after isoflurane anesthesia (30 min). Analysis done using ELISA and western blot. A representative western blot on right shows the mature BDNF band, which is absent in a sample obtained from conditional BDNF knockout (KO) mouse. (E) Isoflurane anesthesia activates TrkB in the hippocampus of conditional BDNF knockout mice (Two-way ANOVA treatment effect F_3,12 = 41,843, p < 0.001; Tukey HSD post hoc test WT CTRL vs. WT ISO p₌0.001, WT CTRL vs. cKO ISO p = 0.015). Representative western blots (A, B, D) have been cropped from complete immunoblots shown in Supplementary Information file. *p < 0.05, **p < 0.01, ***p < 0.001; Abbreviations: CTRL, control treatment; ISO, isoflurane; AMPA, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid; NBQX, 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulfonamide; CREB, cAMP response element binding protein; eEF2, Eukaryotic elongation factor 2.