Dave J R, Rubinstein N, Eskay R L
Endocrinology. 1985 Oct;117(4):1389-96. doi: 10.1210/endo-117-4-1389.
With the use of [125I]acetyl human beta-endorphin (Ac-hBE), specific binding sites for beta-endorphin (BE) were identified in the liver, kidney, adrenal, spleen, and testis of adult male rats, whereas specific BE-binding sites were not present in the ventral prostate or pancreas. In those tissues containing specific BE-binding sites, microsomal membranes (15,000-100,000 X g pellet) exhibited higher BE-binding capacity than the crude homogenate (125-100,000 X g pellet). The binding of BE was saturable, and maximal, specific binding was achieved with a 60-min incubation at 22 C. Furthermore, optimal BE binding was dependent on the presence of magnesium chloride. Scatchard analysis of BE binding to hepatic membranes revealed the existence of two classes of binding sites. One class had an apparent Ka of 0.019 X 10(9) M-1 and a lower number of binding sites (9.1 pmol BE/mg protein), whereas the other class had a lower affinity (apparent Ka of 0.0006 X 10(9) M-1) and a higher number of binding sites (159 pmol/mg protein). Specific BE binding to hepatic membranes was inhibited (80-100%) by rat AcBE-(1-27) and -(1-31), nonacetylated rat BE-(1-31), and human beta-lipotropin. At substantially higher peptide concentrations (greater than 10(-5) M), gamma-endorphin, met-enkephalin, or leu-enkephalin inhibited BE binding by 20-40%. In addition, opiate receptor-binding drugs, such as morphine and naloxone, at 10(-5) M did not alter BE binding to hepatic membranes. Incubation of hepatic membranes with BE induced a dose-related increase in membrane adenylate cyclase activity, and 0.5 X 10(-10) M BE resulted in a maximal enhancement of adenylate cyclase activity to 148% above control values. Water-deprived or salt-loaded male rats with chronically lowered immunoreactive plasma BE exhibited substantially increased BE binding to adrenal and kidney tissue. Specific binding sites for BE occur in a variety of peripheral tissues, and alterations of circulating BE result in changes in the capacity of certain peripheral tissues to bind BE. Finally, occupancy of specific BE-binding sites in peripheral tissue stimulates the adenylate cyclase-cAMP system, which suggests that the peripheral actions of circulating BE may be mediated via this system.
利用[125I]乙酰化人β-内啡肽（Ac-hBE），在成年雄性大鼠的肝脏、肾脏、肾上腺、脾脏和睾丸中鉴定出β-内啡肽（BE）的特异性结合位点，而在腹侧前列腺或胰腺中不存在特异性BE结合位点。在那些含有特异性BE结合位点的组织中，微粒体膜（15,000 - 100,000×g沉淀）比粗匀浆（125 - 100,000×g沉淀）表现出更高的BE结合能力。BE的结合是可饱和的，在22℃孵育60分钟可实现最大特异性结合。此外，最佳的BE结合依赖于氯化镁的存在。对BE与肝细胞膜结合的Scatchard分析揭示了两类结合位点的存在。一类的表观解离常数Ka为0.019×10⁹ M⁻¹，结合位点数量较少（9.1 pmol BE/mg蛋白），而另一类亲和力较低（表观Ka为0.0006×10⁹ M⁻¹），结合位点数量较多（159 pmol/mg蛋白）。大鼠AcBE-(1 - 27)和-(1 - 31)、非乙酰化大鼠BE-(1 - 31)以及人β-促脂素可抑制BE与肝细胞膜的特异性结合（80 - 100%）。在肽浓度显著更高（大于10⁻⁵ M）时，γ-内啡肽、甲硫氨酸脑啡肽或亮氨酸脑啡肽可使BE结合减少20 - 40%。此外，10⁻⁵ M的阿片受体结合药物，如吗啡和纳洛酮，并不改变BE与肝细胞膜的结合。用BE孵育肝细胞膜会导致膜腺苷酸环化酶活性呈剂量相关增加，0.5×10⁻¹⁰ M的BE可使腺苷酸环化酶活性最大增强至比对照值高148%。长期免疫反应性血浆BE降低的缺水或盐负荷雄性大鼠，其肾上腺和肾脏组织对BE的结合显著增加。BE的特异性结合位点存在于多种外周组织中，循环BE的改变会导致某些外周组织结合BE的能力发生变化。最后，外周组织中特异性BE结合位点的占据会刺激腺苷酸环化酶 - cAMP系统，这表明循环BE的外周作用可能通过该系统介导。
