Sharif N A, Burt D R
Regul Pept. 1983 Dec;7(4):399-411. doi: 10.1016/0167-0115(83)90111-8.
Optimal conditions for measuring receptor binding for thyrotropin-releasing hormone (TRH) in the rat central nervous system (CNS) have been determined using 3H-labelled [3-Me-His2]TRH [( 3H]MeTRH). Binding assays conducted at 0 degree C for 5-6 h using sodium phosphate- and/or Hepes-buffered tissue resuspensions, with subsequent filtration through Whatman GF/B filters, yielded the best results. Association and dissociation of [3H]MeTRH binding to amygdala membranes were time and temperature dependent. Dissociation kinetics appeared biphasic. Progressive reduction in receptor affinity and capacity and increased radioligand breakdown were observed at elevated temperatures. Bacitracin (25-1000 microM) prevented peptide degradation but inhibited receptor binding (8-37%). Detailed competition experiments using MeTRH and other drugs yielded a pharmacological profile similar to that observed previously in other tissues indicating TRH receptor identification. Highest density of TRH receptors was observed in the retina and numerous limbic areas. Monovalent and divalent cations modulated [3H]MeTRH binding by reducing apparent receptor number.
利用3H标记的[3-甲基-组氨酸2]促甲状腺激素释放激素（TRH）（[3H]MeTRH），已确定了大鼠中枢神经系统（CNS）中测量促甲状腺激素释放激素（TRH）受体结合的最佳条件。使用磷酸钠和/或Hepes缓冲的组织重悬液，在0℃下进行5-6小时的结合测定，随后通过Whatman GF/B滤纸过滤，可获得最佳结果。[3H]MeTRH与杏仁核膜的结合和解离具有时间和温度依赖性。解离动力学呈双相性。在升高的温度下，观察到受体亲和力和容量逐渐降低以及放射性配体分解增加。杆菌肽（25-1000微摩尔）可防止肽降解，但会抑制受体结合（8-37%）。使用MeTRH和其他药物进行的详细竞争实验产生了与先前在其他组织中观察到的相似的药理学特征，表明TRH受体得到了鉴定。在视网膜和许多边缘区域观察到TRH受体的最高密度。单价和二价阳离子通过减少表观受体数量来调节[3H]MeTRH的结合。
