右美托咪定通过PI3K/Akt通路改善睡眠剥夺大鼠异氟醚暴露后的认知损害

Dexmedetomidine improves cognitive impairment in sleep deprived rats after exposure to isoflurane via the phosphoinositide 3‑kinase/protein kinase B pathway

目的探索右美托咪定(Dex)作为麻醉前药物是否可以改善睡眠剥夺大鼠异氟醚暴露后的认知损害及其可能的机制。方法采用随机数字表法将42只雄性SD大鼠分为7组(每组6只):对照组(C组)、异氟醚组(Iso组)、睡眠剥夺组(SD组)、睡眠剥夺+异氟醚组(SD+Iso组)、Dex+睡眠剥夺+异氟醚组(D+SD+Iso组)、磷脂酰肌醇3‐激酶(PI3K)抑制剂(LY294002)+睡眠剥夺+异氟醚组(L+SD+Iso组)、Dex+LY294002+睡眠剥夺+异氟醚组(D+L+SD+Iso组)。C组不做处理,Iso组进行异氟醚吸入3 h处理,SD组进行48 h睡眠剥夺,SD+Iso组在睡眠剥夺48 h后吸入异氟醚3 h,D+SD+Iso组、L+SD+Iso组在睡眠剥夺48 h和吸入异氟醚3 h同时分别给予Dex(20μg/kg)、LY294002(25μg/kg),D+L+SD+Iso组在睡眠剥夺48 h和吸入异氟醚3 h同时给予Dex(20μg/kg)、LY294002(25μg/kg)。莫里斯水迷宫评估大鼠认知能力,免疫荧光检测海马自噬体数量变化,免疫印迹法(Western blot)检测PI3K/蛋白激酶B(Akt)信号通路和自噬相关蛋白[微管相关蛋白1轻链3B(LC3B)、P62和Beclin‐1]水平。结果与C组比较:SD组和SD+Iso组穿越平台次数较少,目标象限停留时间较短,海马内LC3B表达增加,磷酸化PI3K(p‐PI3K)、磷酸化Akt(p‐Akt)水平较低,LC3B、P62和Beclin‐1蛋白水平较高(均P<0.05)。与SD组比较:SD+Iso组穿越平台次数较少,目标象限停留时间较短,p‐PI3K、p‐Akt水平较低,LC3B、p62和Beclin‐1蛋白水平较高(均P<0.05)。与SD+Iso组比较:D+SD+Iso组穿越平台次数较多,目标象限停留时间较长,海马内LC3B表达减少,p‐PI3K、p‐Akt水平较高,LC3B、P62和Beclin‐1蛋白水平较低(均P<0.05)。与D+SD+Iso组比较:D+L+SD+Iso组穿越平台次数较少,目标象限停留时间较短,海马内LC3B表达增加,p‐PI3K、p‐Akt水平较低,LC3B、P62和Beclin‐1蛋白水平较高(均P<0.05)。结论睡眠剥夺大鼠异氟醚暴露后发生认知损害,Dex改善了睡眠剥夺大鼠的认知能力,与抑制过度的自噬以及激活PI3K/Akt信号通路有关。

Objective To explore the potential role of dexmedetomidine(Dex)as an anesthetic pro‑drug to improve cognitive impairment in sleep deprived rats after exposure to isoflurane and the possible mechanisms.Methods According to the random number table method,42 male SD rats were divided into the seven groups(n=6):a control group(group C),an isoflurane group(group Iso),a sleep deprivation group(group SD),a sleep deprivation+isoflurane group(group SD+Iso),a Dex+sleep deprivation+isoflurane group(group D+SD+Iso),a phosphatidy linositol 3‑kinase(PI3K)inhibitor(LY294002)+sleep deprivation+isoflurane group(group L+SD+Iso),and a Dex+LY294002+sleep deprivation+isoflurane group(group D+L+SD+Iso).No treatment was given to mice in group C,while those in group Iso was given isoflurane by inhalation for 3 h.Group SD was deprived of sleep for 48 h.Group SD+Iso was given isoflurane by inhalation for 3 h after sleep deprivation for 48 h.Group D+SD+Iso and group L+SD+Iso were given 20μg/kg Dex and 25μg/kg LY294002,respectively,after sleep deprivation for 48 h and inhalation of isoflurane for 3 h.Group D+L+SD+Iso received both 20μg/kg Dex and 25μg/kg LY294002,after sleep deprivation for 48 h and inhalation of isoflurane for 3 h.The cognitive abilities of the rats were evaluated by Morris water maze.The changes in the number of hippocampal autophagosomes were detected by immunofluorescence.The levels of the PI3K/protein kinase B(Akt)signaling pathway and autophagy‑related proteins[microtubule associated protein 1 light chain 3B(LC3B),P62 and Beclin‑1]were measured by Western blot.Results Compared with group C,group SD and group SD+Iso showed decreases in the number of crossing the platform and target quadrant residence time,as well as increased expression of hippocampal LC3B,reduced levels of phosphorylated phosphatidy linositol 3‑kinase(p‑PI3K)and phosphorylated protein kinase B(p‑Akt),and increased amounts of LC3B,P62 and Beclin‑1(all P<0.05).Compared with group SD,group SD+Iso presented decreases in the number of crossing the platform and target quadrant residence times,as well as decreased levels of p‑PI3K and p‑Akt,and increased amounts of LC3B,p62 and Beclin‑1 proteins(all P<0.05).Compared with group SD+Iso,group D+SD+Iso showed increases in the number of crossing the platform and target quadrant residence times,as well as reduced expression of hippocampal LC3B,increased levels of p‑PI3K and p‑Akt,and decreased amounts of LC3B,p62 and Beclin‑1 proteins(all P<0.05).Compared with group D+SD+Iso,group D+L+SD+Iso showed decreases in the number of crossing the platform and target quadrant residence times,as well as increased expression of hippocampal LC3B,decreased levels of p‑PI3K and p‑Akt,and increased amounts of LC3B,p62 and Beclin‑1 proteins(all P<0.05).Conclusions Exposure to isoflurane cause cognitive impairment in sleep‑deprived rats.Dex can improve the cognitive ability of sleep‑deprived rats,which is associated with inhibiting excessive autophagy and activating the PI3K/Akt signaling pathway.