Studies on Tyrosine Hydroxylase System in Rat Brain Slices Using High‐Performance Liquid Chromatography with Electrochemical Detection
Studies on Tyrosine Hydroxylase System in Rat Brain Slices Using High‐Performance Liquid Chromatography with Electrochemical Detection
复制标题
高效液相色谱电化学检测大鼠脑切片酪氨酸羟化酶系统研究
作者：
Y. Hirata;A. Togari;T. Nagatsu
建立了一种测定脑片中酪氨酸羟化酶(TH；EC 1.14.16.2)活性的新方法。为了评价脑片中TH活性，包括四氢生物蝶呤、二氢蝶呤还原酶和TH本身，脑片与芳香族L氨基酸脱羧酶抑制剂NSD-1055孵育，用高效液相色谱-电泳法测定内源性酪氨酸生成的3，4-二羟基苯丙氨酸。纹状体脑片内源性酪氨酸羟化为DOPA在37°C持续90min内呈线性关系，加入20 mM K+去极化神经细胞后，内源性酪氨酸羟基化增强。此外，在所检测的所有脑区都可以检测到DOPA的形成，并且在该切片系统中的活性几乎与以酪氨酸和6-甲基四氢蝶呤为辅因子的饱和浓度的切片作为酶的匀浆的最大速度平行。该检测系统可用于研究TH在相对完整的组织制剂中的调节机制。
A new method for the measurement of tyrosine hydroxylase (TH; EC 1.14.16.2) activity in brain slices was developed by using high‐performance liquid chromatography (HPLC) with electrochemical detection (ED). To estimate TH activity in brain slices containing all of the components of the enzyme system, tetrahydrobiopterin, dihydropteridine reductase, and TH itself, slices were incubated with NSD‐1055, an inhibitor of aromatic l‐amino acid decarboxylase, and 3,4‐dihydroxyphenylalanine (DOPA) formed from endogenous tyrosine was measured using HPLC‐ED. Hydroxylation of endogenous tyrosine to DOPA in striatal slices was linear up to 90 min at 37°C, and increased by incubation with 20 mM K+ to depolarize the nerve cells. Furthermore, the formation of DOPA could be detected in all parts of brain regions examined, and the activity in this slice system was nearly parallel to the maximal velocity of the homogenate from the slices as enzyme in the presence of saturating concentrations of tyrosine and 6‐methyltetrahydropterin as cofactor. This assay system should be useful to study the regulatory mechanisms of TH in relatively intact tissue preparations.
视网膜细胞悬浮液中酪氨酸羟化酶活性的调节：钾和 8-溴环 AMP 的作用。
影响因子：
6.1
作者：
Iuvone,PM;Marshburn,PB
通讯作者：
Marshburn,PB