Chen N H, Ding J H, Wang Y L
Department of Pharmacology, Nanjing Medical University, China.
Zhongguo Yao Li Xue Bao. 1997 Mar;18(2):115-20.
To characterize the binding of [125I]3 beta-(4-iodophenyl) tropane-2 beta-carboxylic acid isopropyl ester (RTI-121) to the dopamine transporter (DAT) under physiologically relevant conditions.
[125I]RTI-121 was used to label the DAT on fresh rat striatum synaptosomal membranes in artificial cerebrospinal fluid (ACSF) at 37 degrees C.
[125I]RTI-121 binding reached equilibrium within 3 min and remained at its plateau value for at least 9 min. The data from kinetic, saturation, and competition studies supported a one-site model for the binding of [125I]RTI-121 to the DAT. Various DAT blockers (oocaine, GBR12935, and BTCP) and substrates (dopamine and d-amphetamine) competitively inhibited the binding of [125I]RTI-121. Compared with NaPhos-KCl-NaCl assay buffer, ACSF containing Ca2+ and Mg2+ markedly increased the IC50 of DAT blockers for inhibiting [125I]RTI-121 binding with less effect on that of substrates. Various D2 receptor ligands (pergolide, quinirole, sulpiride, and l-stepholidine) had no direct effect on the binding of [125I]RTI-121.
[125I]RTI-121 binding under physiologically relevant conditions fulfills the basic criteria for DAT binding assay.
在生理相关条件下，表征[125I]3β-(4-碘苯基)托烷-2β-羧酸异丙酯（RTI-121）与多巴胺转运体（DAT）的结合情况。
在37℃下，使用[125I]RTI-121标记人工脑脊液（ACSF）中新鲜大鼠纹状体突触体膜上的DAT。
[125I]RTI-121结合在3分钟内达到平衡，并在至少9分钟内保持在平台值。动力学、饱和和竞争研究的数据支持[125I]RTI-121与DAT结合的单点模型。各种DAT阻滞剂（育亨宾、GBR12935和BTCP）和底物（多巴胺和d-苯丙胺）竞争性抑制[125I]RTI-121的结合。与NaPhos-KCl-NaCl测定缓冲液相比，含有Ca2+和Mg2+的ACSF显著增加了DAT阻滞剂抑制[125I]RTI-121结合的IC50，而对底物的影响较小。各种D2受体配体（培高利特、喹尼洛尔、舒必利和左旋千金藤啶碱）对[125I]RTI-121的结合没有直接影响。
在生理相关条件下，[125I]RTI-121结合满足DAT结合测定的基本标准。
