七氟醚麻醉通过c-fos和神经振荡影响小鼠苏醒期恐惧记忆

目的 探讨c-fos和神经振荡在七氟醚麻醉小鼠苏醒期恐惧记忆中的作用。
方法 选择SPF级雄性成年C57BL/6小鼠270只,8～10周龄,体重18～24 g。将小鼠随机分为三组：A组吸入2.5%七氟醚麻醉2 h;B组吸入2.5%七氟醚麻醉6 h;C组吸入2.5%七氟醚麻醉2 h,连续3 d,每组90只。所有小鼠先进行恐惧条件实验训练,训练后1 d进行相应的七氟醚麻醉处理,记录麻醉停止到翻正反射出现（苏醒）的时间。于麻醉前30 min（T₀）、翻正反射出现后5 min（T₁）、30 min（T₂）和60 min（T₃）对小鼠进行恐惧条件实验测试,记录冻结时间百分比。于T₀、T₁时进行旷场试验,记录小鼠运动总路程和在中心区域停留时间。A组随机另取6只小鼠于麻醉前1周在海马组织植入微电极,后将小鼠放回原来的笼子恢复1周,于T₀—T₃时记录Delta、Theta、Alpha、Beta和Gamma波相对能量值。于T₀—T₃时采用免疫荧光法检测海马组织c-fos阳性细胞数,采用Western blot法记录海马组织c-fos蛋白相对含量。
结果 与T₀时比较,T₁时A组和B组冻结时间百分比明显升高（P＜0.05）,B组中心区域停留时间明显缩短（P＜0.05）。B组停止麻醉到翻正反射出现的时间明显长于A组（P＜0.05）,T₁时冻结时间百分比明显低于A组（P＜0.05）,T₁时中心区域停留时间明显短于A组（P＜0.05）。C组停止麻醉到翻正反射出现的时间明显短于B组（P＜0.05）,T₁时冻结时间百分比明显低于A组和B组（P＜0.05）,T₁时中心区域停留时间明显长于B组（P＜0.05）。与T₀时比较,A组T₁时Theta波和T₁、T₂时Gamma波相对能量值明显升高（P＜0.05）,T₁时海马CA1区c-fos阳性细胞数明显增多（P＜0.05）,T₁时海马组织c-fos蛋白相对含量明显升高（P＜0.05）。与T₁时比较,A组T₃时Theta波和T₂、T₃时Gamma波相对能量值明显降低（P＜0.05）,T₂、T₃时海马CA1区c-fos阳性细胞数明显减少（P＜0.05）。
结论c-fos表达上调和神经振荡增强参与单次短时间吸入七氟醚增强小鼠苏醒短时间内恐惧记忆的过程,但这一效应随着苏醒后时间的延长而逐渐消失。

Sevoflurane anesthesia affects fear memory of mice in recovery period through c-fos and neural oscillation

Ojective To investigate the effect of c-fos and neural oscillation on fear memory in sevoflurane-anesthetized mice in recovery period.
Methods A total of 270 SPF male adult C57BL/6 mice were selected, aged 8-10 weeks, weighing 18-24 g. Mice were randomly divided into three groups: group A inhaled 2.5% sevoflurane for 2 hours, group B inhaled 2.5% sevoflurane for 6 hours, group C inhaled 2.5% sevoflurane for 3 days, 2 hours each day, 90 mice in each group. All mice were trained by fear conditioning test, and sevoflurane anesthesia was performed 1 day after training, and the time from the end of anesthesia to the positive righting reflex (awakening) was recorded. Fear conditioning test was performed 30 minutes before anesthesia (T₀), 5 minutes (T₁), 30 minutes (T₂), 60 minutes (T₃) after positive righting reflex, and the percentage of freezing time was recorded. Open field test was carried out at T₀ and T₁, and the total distance of movement and time in the central area were recorded. Another 6 mice in group A were implanted with microelectrodes in the hippocampus one week before anesthesia, and then put back to the original cage for one week to recovery. The relative powers of delta, theta, alpha, beta, and gamma waves were recorded at T₀-T₃. The number of c-fos positive cells in hippocampus was detected by immunofluorescence method at T₀-T₃. Western blot was used to record the relative expression of c-fos protein in hippocampus at T₀-T₃.
Results Compared with T₀, the percentage of freezing time in groups A and B were significantly increased at T₁ (P<0.05), time in the central area was significantly shortened in group B (P<0.05). Compared with group A, the time from the end of anesthesia to the positive righting reflex was significantly prolonged in group B (P<0.05), the percentage of freezing time at T₁ was significantly decreased in group B (P<0.05), time in the central area at T₁ was significantly shortened in group B (P<0.05). The time from the end of anesthesia to the positive righting reflex in group C was significantly shorter than that in group B (P<0.05), and the percentage of freezing time at T₁ in group C was significantly lower than that in groups A and B (P< 0.05), and time in the central area at T₁ in group C was significantly longer than that in group B (P<0.05). Compared with T₀, the relative powers of Theta wave at T₁ and the relative powers of Gamma wave at T₁ and T₂ in group A were significantly increased (P<0.05), the number of c-fos positive cells in hippocampus CA1 was area at T₁ significantly increased in group A (P<0.05), the relative expression of c-fos protein in hippocampus at T₁ was significantly increased in group A (P<0.05). Compared with T₁, the relative powers of Theta wave at T₃ and the relative powers of Gamma wave at T₂ and T₃ in group A were significantly decreased (P<0.05), the number of c-fos positive cells in hippocampus CA1 area at T₂ and T₃ significantly were decreased in group A (P<0.05).
Conclusion The up-regulation of c-fos expression and the enhancement of neural oscillation may be involved in the the process of a single short-duration inhalation of sevoflurane in strengthening fear memory in anesthesia recovery period within a short period of time, and this effect disappeared gradually with the extension of time after awakening.