Boja J W, Carroll F I, Vaughan R A, Kopajtic T, Kuhar M J
N.E. Ohio Universities College of Medicine, Dept. of Pharmacology, Rootstown 44266, USA.
Synapse. 1998 Sep;30(1):9-17. doi: 10.1002/(SICI)1098-2396(199809)30:1<9::AID-SYN2>3.0.CO;2-7.
In an effort to identify novel binding sites for cocaine and its analogs, we carried out binding studies with the high-affinity and selective ligand [125I]RTI-121 in rat frontal cortical tissue. Very low densities of binding sites were found. Saturation analysis revealed that the binding was to both high- and low-affinity sites. Pharmacological competition studies were carried out with inhibitors of the dopamine, norepinephrine, and serotonin transporters. The various transporter inhibitors inhibited the binding of 15 pM [125I]RTI-121 in a biphasic fashion following a two-site binding model. The resultant data were complex and did not suggest a simple association with any single transporter. Correlational analysis supported the following hypothesis: [125I] RTI-121 binds to known transporters and not to novel sites; these include dopamine, norepinephrine, and serotonin transporters. Immunoprecipitation of transporters photoaffinity labeled with [125]RTI-82 and subsequent analysis of SDS-page gels revealed the presence of authentic dopamine transporters in these samples; displacement of the photoaffinity label occurred with a typical dopamine transporter pharmacology. These data are compatible with the binding properties of RTI-121 and the presence of several known transporters in the tissue studied.
为了确定可卡因及其类似物的新结合位点，我们用高亲和力和选择性配体[125I]RTI-121在大鼠额叶皮质组织中进行了结合研究。发现结合位点的密度非常低。饱和分析表明，结合是针对高亲和力和低亲和力位点的。用多巴胺、去甲肾上腺素和5-羟色胺转运体抑制剂进行了药理学竞争研究。各种转运体抑制剂按照双位点结合模型以双相方式抑制15 pM [125I]RTI-121的结合。所得数据很复杂，并未表明与任何单一转运体有简单关联。相关分析支持以下假设：[125I]RTI-121与已知转运体结合，而非与新位点结合；这些转运体包括多巴胺、去甲肾上腺素和5-羟色胺转运体。对用[125]RTI-82进行光亲和标记的转运体进行免疫沉淀，并随后对SDS-聚丙烯酰胺凝胶进行分析，结果显示这些样品中存在真正的多巴胺转运体；光亲和标记的置换呈现典型的多巴胺转运体药理学特征。这些数据与RTI-121的结合特性以及所研究组织中几种已知转运体的存在情况相符。
