[125I]博尔顿-亨特[Sar9,Met(O2)11]P物质的结合特性，一种新型NK1受体选择性放射性配体。

Binding characteristics of [125I]Bolton-Hunter [Sar9,Met(O2)11]substance P, a new selective radioligand for the NK1 receptor.

作者信息

Lew R, Geraghty D P, Drapeau G, Regoli D, Burcher E

机构信息

Department of Biological Sciences, Deakin University, Geelong, Victoria, Canada.

出版信息

Eur J Pharmacol. 1990 Aug 2;184(1):97-108. doi: 10.1016/0014-2999(90)90670-2.

DOI:10.1016/0014-2999(90)90670-2

PMID:1698645

Abstract

The selective tachykinin agonist [Sar9,Met(O2)11]substance P (Sar-SP) was radioiodinated with [125I]Bolton-Hunter reagent and the product [125I]Bolton-Hunter-[Sar9,Met(O)2)11]SP (BHSar-SP) purified using reverse phase HPLC. Autoradiographic studies showed dense specific binding of BHSar-SP over the rat submandibular gland and over several regions in rat brain, with very low nonspecific binding, identical with the pattern of binding sites seen in a parallel study with [125I]Bolton-Hunter SP (BHSP). In homogenate binding experiments, BHSar-SP bound with high affinity to a single site in membranes from rat brain (KD 261 pM) and rat submandibular gland (KD 105 pM). Comparative values for BHSP were 495 and 456 pM, i.e. of two and four fold lower affinity than BHSar-SP. Association of BHSar-SP to membranes from brain (k+1 3.7 x 10(9) M-1 min-1) was faster than to membranes from salivary gland (k+1 5.6 x 10(8) M-1 min-1). In competition studies, BHSar-SP was displaced from salivary gland membranes by substance P (SP) approximately physalaemin greater than or equal to Sar-SP approximately SP-(3-11) greater than SP-(5-11) much greater than neurokinin A (NKA) approximately eledoisin = kassinin = SP-methyl ester greater than or equal to neurokinin B (NKB) much greater than [Nle10]NKA-(4-10) greater than [MePhe7]NKB-(4-10). In brain membranes, the rank potency order was SP greater than Sar-SP greater than or equal to physalaemin greater than SP-(3-11) greater than SP-(5-11) greater than NKA greater than or equal to eledoisin much greater than NKB greater than kassinin greater than SP-methyl ester: however [MePhe7]NKB-(4-10) and [Nle10]NKA-(4-10) were ineffective competitors at concentrations up to 1 microM. Both binding patterns are consistent with BHSar-SP binding to an NK1 site. With the exception of SP, Sar-SP, SP-(3-11) and physalaemin, all competitors were 5 to 54 times less potent at BHSar-SP binding sites in brain than in salivary gland. These data reveal some differences in characteristics of NK1 binding sites in brain and submandibular gland. Although of higher affinity, BHSar-SP does not appear greatly more selective than BHSP in its ability to define NK1 binding sites.

摘要

将选择性速激肽激动剂[Sar9,Met(O2)11]P物质（Sar-SP）用[125I]博尔顿-亨特试剂进行放射性碘化，产物[125I]博尔顿-亨特-[Sar9,Met(O)2)11]SP（BHSar-SP）通过反相高效液相色谱法进行纯化。放射自显影研究显示，BHSar-SP在大鼠颌下腺及大鼠脑内多个区域有密集的特异性结合，非特异性结合极低，这与在一项使用[125I]博尔顿-亨特P物质（BHSP）的平行研究中观察到的结合位点模式相同。在匀浆结合实验中，BHSar-SP以高亲和力与大鼠脑（KD 261 pM）和大鼠颌下腺（KD 105 pM）膜上的单一位点结合。BHSP的相应值分别为495和456 pM，即亲和力比BHSar-SP低两倍和四倍。BHSar-SP与脑膜的结合（k+1 3.7×10(9) M-1 min-1）比与唾液腺膜的结合（k+1 5.6×10(8) M-1 min-1）更快。在竞争研究中，P物质（SP）、physalaemin、Sar-SP、SP-(3-11)、SP-(5-11)、神经激肽A（NKA）、eledoisin、kassinin、SP-甲酯、神经激肽B（NKB）、[Nle10]NKA-(4-10)、[MePhe7]NKB-(4-10)从唾液腺膜上置换BHSar-SP的能力顺序为：physalaemin≥Sar-SP≥SP-(3-11)≥SP-(5-11)＞NKA≥eledoisin = kassinin = SP-甲酯≥NKB＞[Nle10]NKA-(4-10)＞[MePhe7]NKB-(4-10)。在脑膜中，效价顺序为：SP＞Sar-SP≥physalaemin＞SP-(3-11)＞SP-(5-11)＞NKA≥eledoisin＞NKB＞kassinin＞SP-甲酯；然而，[MePhe7]NKB-(4-10)和[Nle10]NKA-(4-10)在浓度高达1 microM时是无效的竞争者。两种结合模式均与BHSar-SP结合至NK1位点一致。除SP、Sar-SP和physalaemin外，所有竞争者在脑膜中BHSar-SP结合位点的效力比在唾液腺中低5至54倍。这些数据揭示了脑和颌下腺中NK1结合位点特征的一些差异。尽管BHSar-SP具有更高的亲和力，但其在定义NK1结合位点的能力方面似乎并不比BHSP更具选择性。