Laufer R, Gilon C, Chorev M, Selinger Z
J Biol Chem. 1986 Aug 5;261(22):10257-63.
We have recently characterized a tachykinin receptor subtype (SP-N) whose preferred ligand is the mammalian neuropeptide, neurokinin B (Laufer, R., Wormser, U., Friedman, Z. Y., Gilon, C., Chorev, M., and Selinger, Z. (1985) Proc. Natl. Acad. Sci. U.S.A. 82, 7444-7448). To investigate this novel tachykinin receptor, we have now prepared a radiolabeled peptide, N alpha-[( 125I]desamino-3-iodotyrosyl)-[Asp5,6, N-methyl-Phe8]substance P (5-11) heptapeptide (125I-BH-NH-Senktide), which selectively interacts with the SP-N receptor subtype. The binding of 125I-BH-NH-Senktide to rat cerebral cortex membranes was studied under conditions that minimized nonspecific binding. Unlike other tachykinin receptor probes, this radioligand is not degraded during the binding experiment. Binding of 125I-BH-NH-Senktide is reversible, saturable, and of high affinity (KD = 0.9 nM). The radioligand labels a single class of binding site (122 fmol binding sites/mg of protein), as indicated by a linear Scatchard plot and a Hill coefficient close to unity (nH = 1.05). The pharmacological specificity of this binding site corresponds to that of the neuronal SP-N receptor in guinea pig ileum myenteric plexus, which was determined by a functional bioassay. Among various rat brain regions, the highest binding was observed in the cerebral cortex, olfactory bulb, hypothalamus, and hippocampus. These results suggest the existence and specific distribution of a neurokinin B receptor site of the SP-N type in rat brain. 125I-BH-NH-Senktide is the first selective and potent probe for this receptor and is thus an important tool for further studies of its distribution, regulation, and functional role.
我们最近鉴定了一种速激肽受体亚型（SP-N），其首选配体是哺乳动物神经肽神经激肽B（Laufer，R.，Wormser，U.，Friedman，Z.Y.，Gilon，C.，Chorev，M.，和Selinger，Z.（1985年）美国国家科学院院刊82，7444 - 7448）。为了研究这种新型速激肽受体，我们现已制备了一种放射性标记肽，Nα-[(125I]去氨基-3-碘酪氨酸基)-[天冬氨酸5,6，N-甲基苯丙氨酸8]P物质（5 - 11）七肽（125I-BH-NH-Senktide），它能选择性地与SP-N受体亚型相互作用。在使非特异性结合最小化的条件下，研究了125I-BH-NH-Senktide与大鼠大脑皮层膜的结合。与其他速激肽受体探针不同，这种放射性配体在结合实验过程中不会降解。125I-BH-NH-Senktide的结合是可逆的、可饱和的且具有高亲和力（KD = 0.9 nM）。如线性Scatchard图和接近1的希尔系数（nH = 1.05）所示，放射性配体标记了一类单一的结合位点（122 fmol结合位点/毫克蛋白质）。该结合位点的药理学特异性与豚鼠回肠肌间神经丛中神经元SP-N受体的特异性相对应，这是通过功能生物测定法确定的。在大鼠的各个脑区中，在大脑皮层、嗅球、下丘脑和海马体中观察到最高的结合。这些结果表明大鼠脑中存在SP-N型神经激肽B受体位点并具有特定分布。125I-BH-NH-Senktide是该受体的首个选择性强效探针，因此是进一步研究其分布、调节和功能作用的重要工具。
