Yang C M, Farley J M, Dwyer T M
Department of Pharmacology, University of Mississippi Medical Center, Jackson 39216-4505.
J Appl Physiol (1985). 1988 Jan;64(1):200-9. doi: 10.1152/jappl.1988.64.1.200.
The properties of muscarinic acetylcholine receptors (mAChR) on tracheal explants and isolated submucosal gland cells were determined using [3H]quinuclidinyl benzilate ([3H]QNB) and N-[3H]methylscopolamine ([3H]NMS) as ligands. Analysis of competitive displacement of ([3H]NMS binding by pirenzepine demonstrated the presence of M1- (27 +/- 2%) and M2G- (73 +/- 2%) receptors on isolated tracheal submucosal gland cells (TSGC's) in control. Daily administration of diisopropylfluorophosphate (DFP) inhibited cholinesterase activity by greater than 95%. After 7 days of DFP treatment, [3H]QNB binding to intact TSGC's decreased from 14.2 +/- 0.6 to 6.3 +/- 0.8 fmol/10(6) cells; similarly, [3H]NMS binding fell from 8.1 +/- 1.9 to 2.0 +/- 0.8 fmol/10(6) cells. The loss of mAChR's was predominantly of the M2G subtype with the relative proportion dropping to 33%. In addition, 90% of the receptors assumed the high-affinity state for carbachol displacement of [3H]NMS. Mucus secretion was quantitated by measuring the release of 3H-labeled mucus macromolecules from explants of tracheal submucosal glands and isolated cells. Acetylcholine (ACh), 2 X 10(-5) M, stimulated mucus secretion by 2.5 and 2.3 times the basal rate, respectively. Elimination of acetylcholinesterase (AChe) by DFP increased the ACh sensitivity by 18- and 5-fold. Tracheal explants or TSGC's obtained 2 h after an in vivo DFP treatment showed a 6- and 3-fold ACh stimulation. This ACh sensitivity decreased during the continued daily dosing with DFP such that only a 1.3- and 1.1-fold ACh stimulation was apparent after 7 days of treatment.(ABSTRACT TRUNCATED AT 250 WORDS)
利用[3H]喹核醇基苯甲酸酯([3H]QNB)和N-[3H]甲基东莨菪碱([3H]NMS)作为配体，测定气管外植体和分离的黏膜下腺细胞上毒蕈碱型乙酰胆碱受体(mAChR)的特性。用哌仑西平对([3H]NMS)结合的竞争性置换分析表明，在对照条件下，分离的气管黏膜下腺细胞(TSGC's)上存在M1-(27±2%)和M2G-(73±2%)受体。每日给予二异丙基氟磷酸酯(DFP)可抑制胆碱酯酶活性超过95%。DFP处理7天后，[3H]QNB与完整TSGC's的结合从14.2±0.6降至6.3±0.8 fmol/10(6)细胞；同样，[3H]NMS结合从8.1±1.9降至2.0±0.8 fmol/10(6)细胞。mAChR的丢失主要是M2G亚型，其相对比例降至33%。此外，90%的受体对卡巴胆碱置换[3H]NMS呈现高亲和力状态。通过测量气管黏膜下腺外植体和分离细胞中3H标记的黏液大分子的释放来定量黏液分泌。2×10(-5)M的乙酰胆碱(ACh)分别刺激黏液分泌至基础速率的2.5倍和2.3倍。DFP消除乙酰胆碱酯酶(AChe)使ACh敏感性提高了18倍和5倍。体内DFP处理2小时后获得的气管外植体或TSGC's对ACh的刺激分别为6倍和3倍。在持续每日给予DFP期间，这种ACh敏感性降低，以至于处理7天后，ACh刺激仅为1.3倍和1.1倍。(摘要截短至250字)
