Mussap C J, Burcher E
School of Physiology and Pharmacology, University of New South Wales, Sydney, Australia.
J Pharmacol Exp Ther. 1993 Aug;266(2):1043-53.
Tachykinin receptors in rat gastric fundus were characterized using radioligand binding, functional and autoradiographic techniques. In crude homogenates of fundus, the specific binding of 125I-iodohistidyl-neurokinin A (INKA), 125I-Bolton-Hunter eledoisin (BHELE) and 125I-Bolton-Hunter [Sar9,Met(O2)11]-SP (BHSar-SP) was reversible and saturable. INKA and, in particular, BHSar-SP showed high affinity binding (Kds, 2.2 and 0.6 nM, respectively), with lower affinity for BHELE (Kd, 17 nM). The binding capacity was higher for INKA and BHELE than for BHSar-SP. The superior potency of neurokinin (NK)-2-preferring agonists (neuropeptide gamma > or = [Lys5,MeLeu9,Nle10]-NKA(4-10) > or = neuropeptide K > neurokinin A [NKA] > [Sar9,Met(O2)11]-SP >> senktide) and antagonists (SR 48,968 > GR 94,800 > MDL 29,913 > L-659,877 > MEN 10,207) as competitors for INKA and BHELE binding suggests interaction at mainly NK-2 sites. Additional competition studies showed that BHSar-SP was binding to NK-1 sites. Autoradiographic studies revealed very dense INKA and BHELE specific binding over the circular muscle and muscularis mucosae, while BHSar-SP binding was observed only to the circular muscle. The weak specific binding for 125I-Bolton-Hunter scyliorhinin II localized to the muscularis mucosae may indicate NK-3 sites. This was consistent with functional studies showing concentration-dependent contractions of fundus strips by NK-2-preferring tachykinin agonists (potency, pD2s, 7.1 to 8.1) and [Sar9, Met(O2)11]-SP (pD2, 7.1). The NK-2 selective antagonist MDL 29,913 inhibited INKA binding (Kd, 14 nM) with more than tenfold greater affinity than did MEN 10,207. The antagonism by MDL 29,913 was noncompetitive, with a nonparallel rightward shift of the concentration-response curves to the agonists neuropeptide gamma, neuropeptide K, NKA and [Lys5,MeLeu9,Nle10]-NKA(4-10) (dose ratios at 400 nM MDL 29,913 were 230, 62, 40 and 23, respectively). These data indicate that classic NK-2 receptors predominate in the rat fundus and that NK-1 and perhaps NK-3 receptors also exist.
采用放射性配体结合、功能及放射自显影技术对大鼠胃底中的速激肽受体进行了特性研究。在胃底粗匀浆中，125I-碘组氨酰神经激肽A（INKA）、125I-博尔顿-亨特章鱼涎肽（BHELE）和125I-博尔顿-亨特[Sar9,Met(O2)11]-P物质（BHSar-SP）的特异性结合是可逆且可饱和的。INKA，尤其是BHSar-SP表现出高亲和力结合（解离常数Kds分别为2.2和0.6 nM），对BHELE的亲和力较低（Kd为17 nM）。INKA和BHELE的结合容量高于BHSar-SP。偏爱神经激肽（NK）-2的激动剂（神经肽γ≥[Lys5,MeLeu9,Nle10]-NKA(4-10)≥神经肽K＞神经激肽A [NKA]＞[Sar9,Met(O2)11]-SP＞＞速激肽）和拮抗剂（SR 48,968＞GR 94,800＞MDL 29,913＞L-659,877＞MEN 10,207）作为INKA和BHELE结合的竞争剂，表明主要在NK-2位点发生相互作用。额外的竞争研究表明BHSar-SP与NK-1位点结合。放射自显影研究显示，在环形肌和黏膜肌层上INKA和BHELE有非常密集的特异性结合，而仅在环形肌上观察到BHSar-SP的结合。125I-博尔顿-亨特鲨肌肽II的弱特异性结合定位于黏膜肌层，可能表明存在NK-3位点。这与功能研究结果一致，该研究显示偏爱NK-2的速激肽激动剂（效价，pD2s为7.1至8.1）和[Sar9,Met(O2)11]-SP（pD2为7.1）可引起胃底条带浓度依赖性收缩。NK-2选择性拮抗剂MDL 29,913抑制INKA结合（Kd为14 nM），其亲和力比MEN 10,207高十多倍。MDL 29,913的拮抗作用是非竞争性的，浓度-反应曲线对激动剂神经肽γ、神经肽K、NKA和[Lys5,MeLeu9,Nle10]-NKA(4-10)呈非平行右移（在400 nM MDL 29,913时的剂量比分别为230、62、40和23）。这些数据表明，经典的NK-2受体在大鼠胃底中占主导地位，并且NK-1受体甚至可能NK-3受体也存在。
