Biessen E A, Norder J A, Horn A S, Robillard G T
Department of Medicinal Chemistry, Subfaculty of Pharmacy, University of Groningen, The Netherlands.
Biochem Pharmacol. 1988 Oct 15;37(20):3959-66. doi: 10.1016/0006-2952(88)90080-9.
Chemical modification procedures have been used to study the interaction of tricyclic and non-tricyclic 5HT-reuptake inhibitors with the [3H]imipramine binding site (IBS). N-Ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) induced a pronounced loss in [3H]imipramine binding due to a reduction in Bmax. Preincubation with reuptake inhibitors and subsequent inactivation by EEDQ revealed that imipramine and 5HT prevented the EEDQ-induced inhibition, but citalopram and fluoxetine did not. Thiol modification studies demonstrated that reduction by dithiothreitol (DTT) enhanced the binding of [3H]imipramine by increasing the Bmax. The thioselective reagents 1,1-diazobis- (N,N-dimethylformamide) (diamide), phenyl-arsineoxide (PAO) and N-ethylmaleimide (NEM) attenuated the binding capacity by lowering the Bmax. PAO, a reversible thiol reagent, prevented NEM alkylation indicating that dithiols are involved in the NEM-induced inactivation. Binding of tricyclics or non-tricyclics prior to PAO inactivation revealed that tricyclics provide complete protection against thiol modification, while the non-tricyclics do not. The results support the hypothesis that the 5HT-reuptake system of human platelets possesses at least two distinguishable binding sites.
化学修饰程序已被用于研究三环和非三环5-羟色胺再摄取抑制剂与[3H]丙咪嗪结合位点（IBS）的相互作用。N-乙氧羰基-2-乙氧基-1,2-二氢喹啉（EEDQ）由于Bmax降低导致[3H]丙咪嗪结合显著丧失。与再摄取抑制剂预孵育并随后被EEDQ失活表明，丙咪嗪和5-羟色胺可防止EEDQ诱导的抑制作用，但西酞普兰和氟西汀则不能。硫醇修饰研究表明，二硫苏糖醇（DTT）还原通过增加Bmax增强了[3H]丙咪嗪的结合。硫醇选择性试剂1,1-重氮双（N,N-二甲基甲酰胺）（二酰胺）、苯胂氧化物（PAO）和N-乙基马来酰亚胺（NEM）通过降低Bmax减弱了结合能力。PAO是一种可逆的硫醇试剂，可防止NEM烷基化，表明二硫醇参与了NEM诱导的失活。在PAO失活之前结合三环或非三环化合物表明，三环化合物可提供完全的抗硫醇修饰保护，而非三环化合物则不能。结果支持这样的假设，即人血小板的5-羟色胺再摄取系统至少有两个可区分的结合位点。
