Biochemical characteristics of muscarinic cholinoreceptors in swine tracheal smooth muscle.
Biochemical characteristics of muscarinic cholinoreceptors in swine tracheal smooth muscle.
复制标题
猪气管平滑肌毒蕈碱胆碱受体的生化特征。
DOI：
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发表时间：
1986
期刊：
影响因子：
--
通讯作者：
T. Dwyer
中科院分区：
文献类型：
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作者：
C. Yang;J. Farley;T. Dwyer
用氚标记的毒蕈碱胆碱受体拮抗剂二苯甲酸奎宁环酯（[~ 3 H]QNB）对猪气管平滑肌匀浆膜上的毒蕈碱受体进行了表征。基于受体结合试验，匀浆具有特异性的，饱和的，高亲和力的[3 H]QNB受体。特异性结合具有时间和温度依赖性。在[3 H]QNB浓度为180 pM时，[3 H]QNB与毒蕈碱受体的结合在37 ℃比25 ℃更快地达到平衡（分别为30 min和2 h）。在1800 pM的[3 H]QNB浓度下，在5分钟内达到两个温度下的平衡。所有剩余实验均在37 ℃下进行。结合相对于[3 H]QNB和组织浓度是饱和的。结合等温线的分析产生51 +/-20 pM的表观平衡解离常数（KD）和2.17 +/-0.27 pmole/mg蛋白质的最大受体密度（Bmax）。[3 H]QNB结合的希尔系数为1.07 +/-0.16。结合速率常数（K1）和解离速率常数（K-1）分别为（5.51 ± 0.16）× 10（8）M-1 min-1和（1.41 ± 0.18）× 10（-2）min-1。根据K1和K-1的比值计算的KD为26.3 +/- 3.8 pM;该值接近根据结合等温线的Scatchard图计算的KD值。毒蕈碱受体结合位点的密度在气管平滑肌中比在气管上皮中大10倍（0.20 +/- 0.03 pmole/mg蛋白）。断奶仔猪和青年成年猪气管平滑肌M受体密度无差异。非选择性毒蕈碱拮抗剂阿托品、东莨菪碱和奎宁环二苯甲酸酯（QNB）竞争性抑制[~ 3 H]QNB与匀浆的结合，Hill系数为0.9-1.0，抑制常数（Ki）为纳摩尔范围。与选择性毒蕈碱拮抗剂哌仑西平和3-奎宁环基咕吨-9-羧酸酯（QNX）的竞争分别得到Ki值为0.26 M和0.78 nM，希尔系数约为1。对于所有测试的毒蕈碱拮抗剂来说，M2亚型的[3 H]QNB结合位点只有一个群体。竞争与选择性毒蕈碱激动剂毛果芸香碱和卡巴胆碱产生Ki值的微摩尔范围内，希尔系数小于1，并揭示了存在两个结合位点（P <0.01）。
The tritiated muscarinic cholinoreceptor antagonist quinuclidinyl benzilate, [3H]QNB, was used to characterize the muscarinic receptors associated with homogenized membrane of the smooth muscle from swine trachea. Based on receptor binding assays, the homogenate had specific, saturable, high-affinity receptors for [3H]QNB. Specific binding was time- and temperature-dependent. The association of [3H]QNB with the muscarinic receptor reached equilibrium much sooner at 37 degrees C than 25 degrees C at a [3H]QNB concentration of 180 pM (30 min and 2 h, respectively). Equilibrium at both temperatures was attained within 5 min at a [3H]QNB concentration of 1800 pM. All remaining experiments were performed at 37 degrees C. Binding was saturable with respect to [3H]QNB and tissue concentrations. Analysis of binding isotherms yielded an apparent equilibrium dissociation constant (KD) of 51 +/- 20 pM and a maximum receptor density (Bmax) of 2.17 +/- 0.27 pmole/mg protein. The Hill coefficient for [3H]QNB binding was 1.07 +/- 0.16. The association (K1) and dissociation (K-1) rate constants were determined to be (5.51 +/- 0.16) X 10(8) M-1 min-1 and (1.41 +/- 0.18) X 10(-2) min-1, respectively. KD calculated from the ratio of K1 and K-1 was 26.3 +/- 3.8 pM; this value is close to the value of KD calculated from Scatchard plots of binding isotherms. The density of muscarinic receptor binding sites was 10-fold greater in tracheal smooth muscle than in tracheal epithelium (0.20 +/- 0.03 pmole/mg protein). There is no difference between weanling and young adult swine in the density of muscarinic receptors in tracheal smooth muscle. The nonselective muscarinic antagonists atropine, scopolamine and quinuclidinyl benzilate (QNB) competitively inhibited [3H]QNB binding to the homogenate with Hill coefficients of 0.9-1.0 and inhibition constants (Ki) of nanomolar range. Competition with selective muscarinic antagonists pirenzepine and 3-quinuclidinyl xanthene-9-carboxylate (QNX) gave Ki values, 0.26 M and 0.78 nM, respectively, and Hill coefficients of approximately 1. There was a single population of [3H]QNB binding sites of the M2 subtype for all tested muscarinic antagonists. Competition with selective muscarinic agonists pilocarpine and carbachol yielded Ki values of micromolar range, Hill coefficients of less than 1, and revealed the existence of two binding sites (P less than 0.01).
气道平滑肌的毒蕈碱受体：其体外表征。
DOI：
10.1152/jappl.1982.52.4.1084
发表时间：
1982
期刊：
Journal of applied physiology: respiratory, environmental and exercise physiology
影响因子：
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作者：
Murlas,C;Nadel,JA;Roberts,JM
通讯作者：
Roberts,JM
药物-受体相互作用的定量分析：I.动力学和平衡特性的测定。
影响因子：
6.1
作者：
Weiland,GA;Molinoff,PB
通讯作者：
Molinoff,PB
胚胎和新生鸡心脏中毒蕈碱胆碱能受体的定量和定性差异。
DOI：
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发表时间：
1981
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Hosey,MM;Fields,JZ
通讯作者：
Fields,JZ