Sundaram H, Turner J D, Strange P G
Research School of Biosciences, The University, Canterbury, England.
J Neurochem. 1995 Nov;65(5):1909-16. doi: 10.1046/j.1471-4159.1995.65051909.x.
Serotonin 5-HT1A receptors expressed stably in recombinant Chinese hamster ovary cells have been studied using radioligand binding with the radiolabelled agonist [3H]lisuride. Competition studies with a range of antagonists versus [3H]lisuride confirmed that all of the specific [3H]lisuride binding was to 5-HT1A receptors on the cells. Competition studies with the antagonist spiperone and several agonists gave data that fitted best to two-binding-site models. The affinities of these competing ligands at the two classes of sites were generally in agreement with their corresponding affinities determined in previous work with either 8-[3H]hydroxydipropylaminotetralin ([3H]8-OH-DPAT; labels receptor coupled to G protein) or [3H]spiperone (labels free receptor). Saturation analyses with [3H]lisuride showed that this radioligand labels a single class of binding sites, but the level of radioligand binding was approximately twice that seen when either [3H]8-OH-DPAT or [3H]spiperone was used. [3H]Lisuride binding was partially inhibited by addition of guanine nucleotides, and the extent of inhibition decreased as the [3H]lisuride concentration was increased. This inhibition was due to the effect of guanine nucleotide to decrease slightly the affinity of [3H]lisuride for binding to the 5-HT1A receptors on the cells. It is concluded that [3H]lisuride can label both the free receptor and the receptor coupled to G proteins but with slightly different affinities and that these two states of the receptor exist in roughly equal amounts in the cells. Agonists generally have a higher affinity for the receptor coupled to G protein, whereas antagonists, with the exception of spiperone (which has a higher affinity for the free receptor), have roughly equal affinities for the free receptor and the receptor coupled to G proteins.
利用放射性配体与放射性标记激动剂[3H]利舒脲结合，对稳定表达于重组中国仓鼠卵巢细胞中的5-羟色胺5-HT1A受体进行了研究。一系列拮抗剂与[3H]利舒脲的竞争研究证实，所有特异性[3H]利舒脲结合均针对细胞上的5-HT1A受体。拮抗剂螺哌隆和几种激动剂的竞争研究给出的数据最符合双结合位点模型。这些竞争配体在两类位点的亲和力通常与其在先前使用8-[3H]羟基二丙基氨基四氢萘（[3H]8-OH-DPAT；标记与G蛋白偶联的受体）或[3H]螺哌隆（标记游离受体）的研究中确定的相应亲和力一致。[3H]利舒脲的饱和分析表明，该放射性配体标记单一类结合位点，但放射性配体结合水平约为使用[3H]8-OH-DPAT或[3H]螺哌隆时的两倍。加入鸟嘌呤核苷酸可部分抑制[3H]利舒脲结合，且随着[3H]利舒脲浓度增加，抑制程度降低。这种抑制是由于鸟嘌呤核苷酸略微降低[3H]利舒脲与细胞上5-HT1A受体结合亲和力的作用。结论是，[3H]利舒脲可标记游离受体和与G蛋白偶联的受体，但亲和力略有不同，且这两种受体状态在细胞中大致等量存在。激动剂通常对与G蛋白偶联的受体具有更高亲和力，而拮抗剂（除螺哌隆对游离受体具有更高亲和力外）对游离受体和与G蛋白偶联的受体具有大致相等的亲和力。
