Oshikawa Sayuri, Tanoue Akito, Koshimizu Taka-aki, Kitagawa Yoko, Tsujimoto Gozoh
Department of Molecular, Cell Pharmacology, National Research Institute for Child Health and Development, Tokyo, Japan.
Mol Pharmacol. 2004 Mar;65(3):623-9. doi: 10.1124/mol.65.3.623.
Vasopressin receptor subtype(s) responsible for stimulation of insulin release from pancreatic beta cells were investigated by using subtype-selective antagonists and mice that were genetically lacking either V1a or V1b receptors. Arginine vasopressin (AVP) increased insulin release from isolated mouse islet cells in a concentration-dependent manner, with a submaximal response at 100 nM. Reverse transcription-polymerase chain reaction (RT-PCR) analysis detected V1b and oxytocin, but not V1a or V2, receptor transcripts in mouse islet cells. We characterized the recently synthesized vasopressin receptor subtype antagonists (2S)1-[(2R 3S)-(5-chloro-3-(2-chlorophenyl)-1-(3,4-dimethoxybenzene-sulfonyl)-3-hydroxy-2,3-difydro-1H-indole-2-carbonyl)-pyrrolidine-2-carboxamide] (SR49059), 1-[1-[4-(3-acetylaminopropoxy)benzoyl]-4-piperidyl]-3,4-dihydro-2(1H)-quinolinone (OPC-21268), and (2S,4R)-1-[5-chloro-1-[(2,4-dimethoxyphenyl)sulfonyl]-3-(2-methoxy-phenyl)-2-oxo-2,3-dihydro-1H-indol-3-yl]-4-hydroxy-N,N-dimethyl-2-pyrrolidine carboxamide (SSR149415) using human embryonic kidney 293 cells stably expressing the three cloned mouse vasopressin receptors (V1a, V1b, and V2). A radioligand binding study showed that SR49059 and OPC-21268 potently inhibited [3H]AVP binding to the cloned mouse V1a receptor, with Ki values of 27 and 510 nM, respectively, whereas SSR149415 potently inhibited [3H]AVP binding to the cloned mouse V1b receptor with a Ki value of 110 nM. The inhibitory effects of vasopressin antagonists on AVP-induced insulin release correlate well with the rank order of potency to inhibit [3H]AVP binding to the V1b receptor; pancreatic islet cells were significantly inhibited by SSR149415 but not by SR49059 or OPC-21268. Furthermore, the AVP effect on insulin release was entirely lost in mice lacking the V1b receptor but was preserved in mice lacking the V1a receptor. Our study, which combined pharmacological and knockout approaches, clearly demonstrates that vasopressin-stimulated insulin release from islet cells is mediated via V1b receptors.
通过使用亚型选择性拮抗剂和基因敲除V1a或V1b受体的小鼠，研究了负责刺激胰腺β细胞释放胰岛素的血管加压素受体亚型。精氨酸血管加压素（AVP）以浓度依赖的方式增加了分离的小鼠胰岛细胞的胰岛素释放，在100 nM时出现亚最大反应。逆转录-聚合酶链反应（RT-PCR）分析在小鼠胰岛细胞中检测到V1b和催产素受体转录本，但未检测到V1a或V2受体转录本。我们使用稳定表达三种克隆的小鼠血管加压素受体（V1a、Vlb和V2）的人胚肾293细胞，对最近合成的血管加压素受体亚型拮抗剂（2S）1- [（2R，3S）-（5-氯-3-（2-氯苯基）-1-（3,4-二甲氧基苯磺酰基）-3-羟基-2,3-二氢-1H-吲哚-2-羰基）-吡咯烷-2-甲酰胺]（SR49059）、1- [1- [4-（3-乙酰氨基丙氧基）苯甲酰基]-4-哌啶基]-3,4-二氢-2（1H）-喹啉酮（OPC-21268）和（2S，4R）-1- [5-氯-1- [（2,4-二甲氧基苯基）磺酰基]-3-（2-甲氧基苯基）-2-氧代-2,3-二氢-1H-吲哚-3-基]-4-羟基-N，N-二甲基-2-吡咯烷甲酰胺（SSR149415）进行了表征。放射性配体结合研究表明，SR49059和OPC-21268强烈抑制[3H]AVP与克隆的小鼠V1a受体的结合，Ki值分别为27和510 nM，而SSR149415强烈抑制[3H]AVP与克隆的小鼠V1b受体的结合，Ki值为110 nM。血管加压素拮抗剂对AVP诱导的胰岛素释放的抑制作用与抑制[3H]AVP与V1b受体结合的效力顺序密切相关；胰腺胰岛细胞被SSR149415显著抑制，但未被SR49059或OPC-21268抑制。此外，在缺乏V1b受体的小鼠中，AVP对胰岛素释放的作用完全丧失，但在缺乏V1a受体的小鼠中得以保留。我们结合药理学和基因敲除方法的研究清楚地表明，血管加压素刺激胰岛细胞释放胰岛素是通过V1b受体介导的。
