Motilin release in perifused isolated canine duodenal epithelial cells.

尽管许多研究致力于胃动素对胃肠平滑肌的运动刺激作用，但对胃动素产生细胞释放胃动素的机制知之甚少。本研究旨在1)建立犬十二指肠上皮细胞分散的灌流系统，2)评价乙酰胆碱(ACh)、生长抑素和蛙皮素对胃动素释放的影响，3)确定胃动素产生细胞上的M受体亚型。胶原酶分散和Percoll分离获得分离的上皮细胞。将分散的细胞与溶胀前的Bio-Gel P2混合，等量成柱(1ml体积)。灌流液(HEPES缓冲的HANK‘s液)保持在37ﾟC，并用100%O2连续鼓泡。流速0.3ml/m in，分数…每隔3min采集S标本，用双抗体放射免疫法测定胃动素浓度。乙酰胆碱(Ach)激动剂卡巴胆碱(CCH)以剂量依赖的方式刺激胃动素的释放，其最大反应出现在10^~(-4)M时，这种反应可被阿托品(10^&lt；-4&gt；M)阻断，而生长抑素(10^&lt；-4&gt；M)对ACh刺激的胃动素释放无影响。蛙皮素在实验浓度范围内(10^~10~10)对胃动素的释放也没有影响。M1受体激动剂吡伦西平和M2受体激动剂AF-DX116BS对CCH刺激的胃动素释放无明显影响，而M3受体激动剂4-DAMP可完全阻断CCH刺激的胃动素释放。结论：1)建立了一个能监测CCH刺激胃动素快速释放的灌流系统。2)Ach通过细胞上的M3受体直接刺激胃动素的释放，提示细胞胃动素的释放受可碱能神经刺激的调节。较少

In spite of numerous invesigations devoted to the motor stimulatory effect of motilin on gastrointestinal smooth muscle, little is known regarding the mechanisms controlling the release of motilin from its producing cells. The present study was designed to 1) develop a perifusion system with dispersed canine duodenal epithelial cells, and 2) assess the effect of acetylcholine (ACh), somatostatin and bombesin on motilin release, and 3) determine the muscarinic receptor subtypes on the motilin producing cells.Segments of the duodenum were dissected out from anesthetized dogs, and the mucosal layr was removed from the muscle layr with fine scissors. Isolated epithelial cells were obtained by collagenase dispersion and percoll separation. The dispersed cells were mixed with preswollen Bio-Gel P2 and aliquoted into columns (1 ml volume). The perifusion medium (HEPES buffered Hank's solution) was kept at 37ﾟC and continuously bubbled with a 100% O2. The flow rate was 0.3 ml/min, the fraction … More s were collected at 3 min intervals, and the motilin concentration was determined by double-antibody radio-immunoassay. Carbachol (Cch), Ach agonist, (10^<-10>M to 10^<-1>M) stimulated release of motilin from the isolated cell column in a dose-dependent manner with the maximum response at 10^<-4>M.This response to Cch was abolished by atropine (10^<-4>M), but somatostatin (10^<-4>M) had no effect on the ACh-stimulated motilin release. Bombesin also had no effect on motilin release in the concentration range tested (10^<-10>M to 10^<-6>M). Pirenzepine (M1 receptor agonist) and AF-DX116BS (M2 receptor agonist) had no effect on Cch (10^<-4>M) Stimulated motilin release at 10^<-6>M,but 4-DAMP (M3 receptor agonist) (10^<-6>M) completely abolished Cch-stimulated motilin release.Conclusion : 1) A perifusion system which can monitor rapid motilin release was established. 2) Ach directly stimulates motilin release though M3 receptors on the cell.These results indicate that release of motilin from the cells are regulated by the colinergic neural stimulation. Less