|Budget Amount *help
¥2,100,000 (Direct Cost : ¥2,100,000)
Fiscal Year 1995 : ¥700,000 (Direct Cost : ¥700,000)
Fiscal Year 1994 : ¥1,400,000 (Direct Cost : ¥1,400,000)
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
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