大鼠丘脑酪氨酸羟化酶调控丙泊酚诱导的镇静作用  

Tyrosine hydroxylase regulates propofol-induced sedation in rat thalamus

作　　者：张一佳 傅风华[1] 曹燕卿 邵帅 谭博 苏瑞斌 ZHANG Yi-jia;FU Feng-hua;CAO Yan-qing;SHAO Shuai;TAN Bo;SU Rui-bin(School of Pharmacy,Yantai University,Yantai 264005,China;State Key Laboratory of Toxicology and Medical Countermeasures,Institute of Pharmacology and Toxicology,Academy of Military Medical Sciences,Academy of Military Sciences,Beijing 100850,China)

机构地区：[1]烟台大学药学院,山东烟台264005 [2]军事科学院军事医学研究院毒物药物研究所,抗毒药物与毒理学国家重点实验室,北京100850

出　　处：《中国药理学与毒理学杂志》2020年第2期95-103,共9页Chinese Journal of Pharmacology and Toxicology

摘　　要：目的通过定量磷酸化蛋白质组学研究丙泊酚诱导大鼠丘脑镇静作用的新型调节机制。方法①采用同位素标记相对和绝对定量(iTRAQ)技术对丙泊酚(100 mg·kg-1,ip)诱导镇静模型SD大鼠丘脑差异磷酸化蛋白质进行标记和定量分析;基因本体(GO)富集分析差异磷酸化蛋白质的生物进程、细胞组分及分子功能;Western印迹法检测大鼠丘脑酪氨酸羟化酶(TH)第19位丝氨酸磷酸化表达水平。②利用TH抑制剂甲基酪氨酸(20 mg·kg-1,提前30 min ip给药),考察其对丙泊酚诱导C57BL/6J小鼠翻正反射消失的诱导时间、持续时间和翻正反射消失率的影响。结果质谱结果显示,与正常对照组相比,丙泊酚组SD大鼠丘脑中共92个磷酸化蛋白质表达水平发生改变,其中TH第19位丝氨酸磷酸化水平显著下调(P<0.01);与Western印迹法结果一致(P<0.05)。丙泊酚(70~120 mg·kg-1)剂量依赖性地增加小鼠的翻正反射消失率,TH抑制剂甲基酪氨酸预处理可使丙泊酚诱导小鼠翻正反射消失率的半数有效剂量(ED50)由92.32 mg·kg-1(95%CI:90.60~94.08,R2=0.9969)降至85.38 mg·kg-1(95%CI:81.30~89.67,R2=0.9768),量效曲线左移。与单用丙泊酚组比较,甲基酪氨酸预处理组可显著延长丙泊酚(90,100和120 mg·kg-1)诱导的C57BL/6J小鼠翻正反射消失持续时间(P<0.05)。结论本研究筛选出大鼠丘脑中多种磷酸化蛋白质参与调控丙泊酚诱导的镇静效应,抑制TH的活性可显著增强丙泊酚的镇静作用。OBJECTIVE To identify a novel regulatory mechanism of propofol-induced sedation in the rat thalamus by quantitative phosphoproteomics.METHODS①Isobaric tags for relative and absolute quantification(iTRAQ)technology was used to label and quantify differentially expressed phosphorylated proteins in the thalamus of a propofol-induced(100 mg·kg-1,ip)sedation model in SD rats.Gene ontology(GO)analyses were conducted to predict and annotate the biological processes,cellular components and molecular functions of these phosphorylated proteins.Western blotting was used to detect the expression of phosphorylated tyrosine hydroxylase(TH)at Ser19〔p-TH(Ser19)〕in the thalamus of rats.②Metyrosine(20 mg·kg-1,ip pretreated for 30 min),a TH inhibitor,was to evaluated in terms of the latency,duration and ratio of loss of righting reflex(LORR)induced by propofol(70,80,90,100 and 120 mg·kg-1,ip)in C57 BL/6 J mice.RESULTS A total of 92 phosphorylated proteins were found to be differentially expressed in the propofol-induced rat sedation group compared with the expressions of the normal control.Among them,p-TH(Ser19)in the rat thalamus was significantly down-regulated(P<0.01)and verified by Western blotting(P<0.05).Propofol(70~120 mg·kg-1)dose-dependently increased the ratio of LORR in mice,with a median effective dose(ED50)of 92.32 mg·kg-1(95%CI:90.60~94.08,R2=0.9969).Pretreatment of metyrosine reduced the ED50 of propofol to 85.38 mg·kg-1(95%CI:81.30~89.67,R2=0.9768)and the dose-effect curve shifted to the left.In addition,the duration of propofol(90,100 and 120 mg·kg-1)-induced LORR was significantly prolonged under metyrosine pretreatment(P<0.05).CONCLUSION Multiple phosphorylated proteins in the rat thalamus have been screened out to be candidates that participate in the regulation of propofol-induced sedation.Moreover,propofol-induced sedation could be significantly enhanced by TH inhibition.

关 键 词：丙泊酚 翻正反射 丘脑 磷酸化蛋白质组学 酪氨酸羟化酶 

分 类 号：R964[医药卫生—药理学] R971.2[医药卫生—药学]