From the Queensland Brain Institute (M.T., O.H.Z., B.D.K., S.K., V.A., B.v.S.) School of Biomedical Sciences (O.H.Z.) Clem Jones Centre for Ageing Dementia Research (V.A.), The University of Queensland, Brisbane, Queensland, Australia School of Medical Science and Griffith Health Institute, Griffith University Gold Coast Campus, Queensland, Australia (S.K.).
Anesthesiology. 2019 Sep;131(3):555-568. doi: 10.1097/ALN.0000000000002850.
Mutations in the presynaptic protein syntaxin1A modulate general anesthetic effects in vitro and in vivo. Coexpression of a truncated syntaxin1A protein confers resistance to volatile and intravenous anesthetics, suggesting a target mechanism distinct from postsynaptic inhibitory receptor processes. Hypothesizing that recovery from anesthesia may involve a presynaptic component, the authors tested whether syntaxin1A mutations facilitated recovery from isoflurane anesthesia in Drosophila melanogaster.
A truncated syntaxin1A construct was expressed in Drosophila neurons. The authors compared effects on isoflurane induction versus recovery in syntaxin1A mutant animals by probing behavioral responses to mechanical stimuli. The authors also measured synaptic responses from the larval neuromuscular junction using sharp intracellular recordings, and performed Western blots to determine whether the truncated syntaxin1A is associated with presynaptic core complexes.
Drosophila expressing a truncated syntaxin1A (syx, n = 40) were resistant to isoflurane induction for a behavioral responsiveness endpoint (ED50 0.30 ± 0.01% isoflurane, P < 0.001) compared with control (0.240 ± 0.002% isoflurane, n = 40). Recovery from isoflurane anesthesia was also faster, with syx-expressing flies showing greater levels of responsiveness earlier in recovery (reaction proportion 0.66 ± 0.48, P < 0.001, n = 68) than controls (0.22 ± 0.42, n = 68 and 0.33 ± 0.48, n = 66). Measuring excitatory junction potentials of larvae coexpressing the truncated syntaxin1A protein showed a greater recovery of synaptic function, compared with controls (17.39 ± 3.19 mV and 10.29 ± 4.88 mV, P = 0.014, n = 8 for both). The resistance-promoting truncated syntaxin1A was not associated with presynaptic core complexes, in the presence or absence of isoflurane anesthesia.
The same neomorphic syntaxin1A mutation that confers isoflurane resistance in cell culture and nematodes also produces isoflurane resistance in Drosophila. Resistance in Drosophila is, however, most evident at the level of recovery from anesthesia, suggesting that the syntaxin1A target affects anesthesia maintenance and recovery processes rather than induction. The absence of truncated syntaxin1A from the presynaptic complex suggests that the resistance-promoting effect of this molecule occurs before core complex formation.
突触蛋白 1A 的突变可调节体外和体内全身麻醉的作用。截短的突触蛋白 1A 蛋白的共表达赋予对挥发性和静脉内麻醉的抗性，表明与突触后抑制性受体过程不同的靶机制。假设麻醉恢复可能涉及一个突触前成分，作者测试了是否突变的突触蛋白 1A 有助于果蝇中的异氟烷麻醉恢复。
在果蝇神经元中表达截短的突触蛋白 1A 构建体。作者通过探测对机械刺激的行为反应来比较突变动物中异氟烷诱导与恢复的影响。作者还使用尖锐的细胞内记录测量幼虫神经肌肉接头的突触反应，并进行 Western blot 以确定截短的突触蛋白 1A 是否与突触前核心复合物相关。
表达截短的突触蛋白 1A 的果蝇（syx，n = 40）对行为反应终点（ED50 0.30 ± 0.01%异氟烷，P < 0.001）的异氟烷诱导具有抗性，而对照（0.240 ± 0.002%异氟烷，n = 40）。异氟烷麻醉的恢复也更快，syx 表达的苍蝇在恢复过程中更早地显示出更高水平的反应性（反应比例 0.66 ± 0.48，P < 0.001，n = 68）比对照（0.22 ± 0.42，n = 68 和 0.33 ± 0.48，n = 66）。测量共表达截短的突触蛋白 1A 蛋白的幼虫的兴奋性突触后电位显示突触功能的恢复更大，与对照相比（17.39 ± 3.19 mV 和 10.29 ± 4.88 mV，P = 0.014，n = 8）。在存在或不存在异氟烷麻醉的情况下，促进抗性的同种神经突变的突触蛋白 1A 与突触前核心复合物无关。
在细胞培养和线虫中赋予异氟烷抗性的相同新形态的突触蛋白 1A 突变也在果蝇中产生异氟烷抗性。然而，在麻醉恢复水平上，果蝇的抗性最为明显，这表明突触蛋白 1A 的靶标影响麻醉维持和恢复过程而不是诱导。截短的突触蛋白 1A 从突触前复合物缺失表明该分子的抗性促进作用发生在核心复合物形成之前。
