Regulation of signal transduction and GTP binding proteins for mucin release from submandibular glands

已普遍认为颌下腺的粘蛋白释放是由β-肾上腺素能受体-腺苷环化酶系统介导的，而α-肾上腺素能和毒鼠碱胆碱能系统不参与粘蛋白的释放。然而，我们先前的研究表明，狗颌下腺的粘蛋白释放主要是由M受体(MAChR)介导的。本研究探讨了犬颌下腺细胞胞吐作用的信号转导系统和mAChR偶联的GTP结合蛋白。ACh、α肾上腺素能激动剂、离子霉素和thapsigardin以钙依赖的方式刺激粘蛋白的释放。ACh可增加通透性细胞内游离钙离子浓度([Ca^lt；2+>t；i)、IP_3和IP_4。IP_3、兰尼定、环化ADP-核糖而不是咖啡因可刺激通透性细胞内钙离子释放。通透性细胞中的GTPGammaS和TPA可促进粘蛋白的释放。GTPGammaS刺激DTT不敏感的胆碱磷酸转移酶合成血小板激活因子(PAF)。PAF抑制细胞膜Na~+，K~(++)-ATPase活性，使通透性细胞内Ca~(2+)~(2+)~(gt；)活化。提示，mAChR刺激通过IP_3介导细胞内Ca~(2+)~(2+)~(gt；)的动员和质膜上Ca~(2+)~(2+)~(gt；)的释放机制和激活Ca~(^&lt；2+)~(gt；)进入机制，从而引起[Ca~(^&lt；2+&gt；]i的释放。在犬颌下腺中，PKC和GTP结合蛋白可能参与粘蛋白的释放和PAF的合成，并作为信号转导系统。

It has been generally accepted that mucin release from submandibular gland is mediated by beta adrenergic receptor-adenylate cyclase system, and alpha adrenergic and muscarinic cholinergic systems are not involved in mucin release. However, we have previously shown that mucin release from dog submandibular gland is mainly mediated by muscarinic ACh receptor(mAChR)stimulation. The present study investigated signal transduction systems and GTP binding proteins coupled to mAChR for exocytosis in dog submandibular gland cells.Mucin release was stimulated by ACh, alpha adrenergic agonists, ionomycin and thapsigardin in Ca^<2+>-dependent manner. ACh increased intracellular free Ca^<2+> concentration ( [Ca^<2+>] i), IP_3 and IP_4. IP_3, ryanodine, cyclic ADP-ribose but not caffeine stimulated Ca^<2+> release in permeabilized cells. GTPgammaS in permeabilized cells and TPA increase mucin release. GTPgammaS stimlated DTT-insensitive choline-phosphotransferase activity for platelet-activating factor (PAF) synthesis. PAF inhibited membrane Na^+, K^+-ATPase activity and mobilized Ca^<2+> in permeabilized cells.These results suggest that mAChR stimulation causes mucin release by increasing [Ca^<2+>] i through IP_3-mediated mobilization of Ca^<2+> from intracellular stores, stimulation of Ca^<2+> induced Ca^<2+> release mechanism and activation of Ca^<2+> entry mechanism through plasma membrane. PKC and GTP binding proteins may be involved in mucin release and PAF synthesis, and serve as signal transduction systems in dog submandibular glands.