Dubocovich M L, Takahashi J S
Proc Natl Acad Sci U S A. 1987 Jun;84(11):3916-20. doi: 10.1073/pnas.84.11.3916.
2-[125I]Iodomelatonin binds with high affinity to a site possessing the pharmacological characteristics of a melatonin receptor in chicken retinal membranes. The specific binding of 2-[125I]iodomelatonin is stable, saturable, and reversible. Saturation experiments indicated that 2-[125I]iodomelatonin labeled a single class of sites with an affinity constant (Kd) of 434 +/- 56 pM and a total number of binding sites (Bmax) of 74.0 +/- 13.6 fmol/mg of protein. The affinity constant obtained from kinetic analysis was in close agreement with that obtained in saturation experiments. Competition experiments showed a monophasic reduction of 2-[125I]iodomelatonin binding with a pharmacological order of indole amine affinities characteristic of a melatonin receptor: 2-iodomelatonin greater than 6-chloromelatonin greater than or equal to melatonin greater than or equal to 6,7-dichloro-2-methylmelatonin greater than 6-hydroxymelatonin greater than or equal to 6-methoxymelatonin much greater than N-acetyltryptamine greater than N-acetyl-5-hydroxytryptamine greater than 5-methoxytryptamine greater than 5-hydroxytryptamine (inactive). The affinities of these melatonin analogs in competing for 2-[125I]iodomelatonin binding sites were correlated closely with their potencies for inhibition of the calcium-dependent release of [3H]dopamine from chicken and rabbit retinas, indicating association of the binding site with a functional response regulated by melatonin. The results indicate that 2-[125I]iodomelatonin is a selective, high-affinity radioligand for the identification and characterization of melatonin receptor sites.
2-[¹²⁵I]碘褪黑素与鸡视网膜膜中具有褪黑素受体药理学特性的位点具有高亲和力结合。2-[¹²⁵I]碘褪黑素的特异性结合是稳定、可饱和且可逆的。饱和实验表明，2-[¹²⁵I]碘褪黑素标记了一类单一的位点，其亲和常数（Kd）为434±56 pM，结合位点总数（Bmax）为74.0±13.6 fmol/mg蛋白质。从动力学分析获得的亲和常数与饱和实验中获得的结果密切一致。竞争实验表明，2-[¹²⁵I]碘褪黑素结合呈单相减少，其吲哚胺亲和力的药理学顺序具有褪黑素受体的特征：2-碘褪黑素＞6-氯褪黑素≥褪黑素≥6,7-二氯-2-甲基褪黑素＞6-羟基褪黑素≥6-甲氧基褪黑素＞N-乙酰色胺＞N-乙酰-5-羟色胺＞5-甲氧基色胺＞5-羟色胺（无活性）。这些褪黑素类似物竞争2-[¹²⁵I]碘褪黑素结合位点的亲和力与其抑制鸡和兔视网膜中[³H]多巴胺钙依赖性释放的效力密切相关，表明该结合位点与受褪黑素调节的功能反应相关。结果表明，2-[¹²⁵I]碘褪黑素是一种用于鉴定和表征褪黑素受体位点的选择性、高亲和力放射性配体。
