Marcusson J O, Bäckström I T, Ross S B
Mol Pharmacol. 1986 Aug;30(2):121-8.
Recently, a high affinity [3H]imipramine-binding site of protein nature that appeared to be related to the 5-hydroxytryptamine (5-HT, serotonin) uptake mechanism was demonstrated. This binding site was only part of desipramine-displaceable [3H]imipramine binding, which contained a significant amount of additional binding not related to 5-HT uptake. The present study further investigates the [3H]imipramine-binding site of protein nature in the rat brain. Displacement by 5-HT and 6-methoxytetrahydro-beta-carboline (6-MeO-TH beta C) revealed monophasic displacement patterns with 60% displaceable binding. This binding fraction was abolished by protease treatment of the brain tissue prior to binding assay. Saturation studies of [3H]imipramine binding (1-30 nM) in rat cortex showed that the binding displaced by 30 microM 5-HT [Bmax 322 +/- 16 fmol/mg of protein, Kd 4.17 +/- 1.07 nM (means +/- SE)] was not different from the binding displaced by 1.0 microM norzimeldine (Bmax 349 +/- 15 fmol/mg of protein, Kd 4.47 +/- 1.07 nM) or 30 microM 6-MeO-TH beta C (Bmax 439 +/- 28 fmol/mg of protein, Kd 5.49 +/- 1.09 nM). When 100 microM desipramine was used in saturation studies, the binding was different from that displaced by 5-HT with Bmax 608 +/- 42 fmol/mg of protein and Kd 6.68 +/- 1.09 nM. Both displacement and saturation studies in which two displacing agents were combined indicated that most of the binding competed by 5-HT (30 microM) and norzimeldine (1.0 microM) is identical. Similarly, the binding displaced by 5-HT or norzimeldine is subsumed within 6-MeO-TH beta C (30 microM)-displaceable binding. Lesion studies with parachloroamphetamine, a selective toxin for 5-HT terminals, which resulted in a 83% reduction of [3H] 5-HT uptake ( [3H]noradrenaline uptake unaffected), abolished cortical [3H]imipramine binding displaced by 30 microM 5-HT or 1.0 microM norzimeldine. (greater than 80% reduction). However, with 100 microM desipramine as displacer, 40% of the binding remained in lesioned animals. The [3H]imipramine binding displaced by 30 microM 5-HT or 1.0 microM norzimeldine was sodium dependent, and an increase in NaCl concentration from 0 to 120 mM resulted in a 10-fold increase in affinity without effect on Bmax, whereas no change in binding was observed with increasing concentrations of LiCl.(ABSTRACT TRUNCATED AT 400 WORDS)
最近，人们证实了一种具有蛋白质性质的高亲和力[³H]丙咪嗪结合位点，该位点似乎与5-羟色胺（5-HT，血清素）摄取机制有关。这个结合位点只是地昔帕明可置换的[³H]丙咪嗪结合的一部分，后者还包含大量与5-HT摄取无关的额外结合。本研究进一步探究了大鼠脑中具有蛋白质性质的[³H]丙咪嗪结合位点。5-HT和6-甲氧基四氢-β-咔啉（6-MeO-THβC）的置换显示出单相置换模式，其中60%的结合可被置换。在结合测定前用蛋白酶处理脑组织可消除这一结合部分。大鼠皮层中[³H]丙咪嗪结合（1 - 30 nM）的饱和研究表明，被30μM 5-HT置换的结合（Bmax为322±16 fmol/mg蛋白质，Kd为4.17±1.07 nM（均值±标准误））与被1.0μM去甲替林置换的结合（Bmax为349±15 fmol/mg蛋白质，Kd为4.47±1.07 nM）或30μM 6-MeO-THβC置换的结合（Bmax为439±28 fmol/mg蛋白质，Kd为5.49±1.09 nM）没有差异。当在饱和研究中使用100μM地昔帕明时，其结合与被5-HT置换的结合不同，Bmax为608±42 fmol/mg蛋白质，Kd为6.68±1.09 nM。用两种置换剂联合进行的置换和饱和研究均表明，大部分被5-HT（30μM）和去甲替林（1.0μM）竞争的结合是相同的。同样，被5-HT或去甲替林置换的结合包含在6-MeO-THβC（30μM）可置换的结合范围内。用对氯苯丙胺（一种5-HT终末的选择性毒素）进行的损伤研究导致[³H] 5-HT摄取减少83%（[³H]去甲肾上腺素摄取未受影响），消除了被30μM 5-HT或1.0μM去甲替林置换的皮层[³H]丙咪嗪结合（减少大于80%）。然而，如果用100μM地昔帕明作为置换剂，损伤动物中仍有40%的结合存在。被30μM 5-HT或1.0μM去甲替林置换的[³H]丙咪嗪结合是钠依赖性的，将NaCl浓度从0增加到120 mM会使亲和力增加10倍，而对Bmax没有影响，而随着LiCl浓度增加，结合未观察到变化。（摘要截于400字）
