Characterization of opioid receptors in intestinal muscle cells by selective radioligands and receptor protection.
Characterization of opioid receptors in intestinal muscle cells by selective radioligands and receptor protection.
复制标题
通过选择性放射性配体和受体保护来表征肠肌细胞中的阿片受体。
DOI：
10.1152/ajpgi.1992.263.2.g269
复制
发表时间：
1992
期刊：
影响因子：
--
通讯作者：
Makhlouf,GM
中科院分区：
文献类型：
--
作者：
Kuemmerle,JF;Makhlouf,GM
阿片受体的特点分别从兔肠的环形和纵向肌肉层分离的肌肉细胞。将κ-（[3 H]U69，593）、δ-（[3 H][D-Pen 2，5]脑啡肽，DPDPE）和μ-（[3 H][D-Ala 2，N-Me-Phe 4，Gly 5-ol]脑啡肽，DAGO）阿片受体的选择性放射性配体与受体保护技术结合使用，该技术旨在富集具有特定受体类型的细胞。仅在环形肌肉层的细胞中观察到结合。结合是快速的（在2分钟内达到峰值），具有温度依赖性和浓度依赖性。从饱和曲线得出的高亲和力结合位点的解离常数（Kd）（U69，593为1.1 +/- 0.3 nM，DPDPE为0.39 +/- 0.04 nM，DAGO为1.9 +/- 0.3 nM）与从竞争曲线得出的Kd值相似。在竞争性研究中，阿片配体抑制结合的效力顺序取决于所用的放射性配体：U69，593（Kd 1.5 +/- 0.2 nM）优先抑制[3 H]U69，593、DPDPE（Kd 0.72 +/- 0.16 nM）、[3 H]DPDPE和DAGO（Kd 1.2 +/- 0.3 nM）的结合以及[3 H]DAGO的结合。在每种情况下，其他两种配体的效力都要低400- 12，000倍。在富含一种受体类型的细胞中，仅观察到与相应的选择性配体的结合和收缩。配体的效力略微增强，而其他两种配体的效力进一步降低（大于10，000倍）。我们的结论是，不同的κ-，δ-和μ-阿片受体存在于肌肉细胞的循环，但不纵向肌肉层的肠。
Opioid receptors were characterized on muscle cells isolated separately from the circular and longitudinal muscle layers of rabbit intestine. Selective radioligands for kappa- ([3H]U69,593), delta- ([3H][D-Pen2,5]enkephalin, DPDPE), and mu- ([3H][D-Ala2,N-Me-Phe4,Gly5-ol]enkephalin, DAGO) opioid receptors were used in conjunction with a technique of receptor protection designed to enrich cells with a specific receptor type. Binding was observed only in cells from the circular muscle layer. Binding was rapid (peak within 2 min), temperature dependent, and concentration dependent. Dissociation constants (Kd) for high-affinity binding sites derived from saturation curves (1.1 +/- 0.3 nM for U69,593, 0.39 +/- 0.04 nM for DPDPE, and 1.9 +/- 0.3 nM for DAGO) were similar to Kd values derived from competition curves. In competition studies, the order of potency with which opioid ligands inhibited binding depended on the radioligand used: U69,593 (Kd 1.5 +/- 0.2 nM) inhibited preferentially the binding of [3H]U69,593, DPDPE (Kd 0.72 +/- 0.16 nM) the binding of [3H]DPDPE and DAGO (Kd 1.2 +/- 0.3 nM) the binding of [3H]DAGO. In each instance the other two ligands were 400-12,000 times less potent. In cells enriched with one receptor type, binding and contraction were observed only with the corresponding selective ligand. The potency of the ligand was slightly enhanced, whereas the potencies of the other two ligands were further reduced (greater than 10,000-fold). We conclude that distinct kappa-, delta-, and mu-opioid receptors are present on muscle cells of the circular but not longitudinal muscle layer of the intestine.