胃动素受体激动剂对肠肌间神经元内钙的影响

目的探讨胃动素受体激动剂对体外培养大鼠肠肌间神经元内钙的影响。方法采用免疫荧光标记技术鉴定原代培养的肠肌间神经元;应用钙离子(Ca2 )指示剂Fluo-3/AM作为细胞内Ca2 的荧光探针,对负载培养的神经元,应用激光共聚焦显微镜技术检测胃动素及红霉素引起的单个神经元内Ca2 荧光强度(FI)的变化。结果胃动素和红霉素引起的神经元内Ca2 浓度变化为:(1)在Hank's液中,胃动素浓度为10-8、10-7、10-6mol/L时,神经元内Ca2 浓度逐渐升高,其峰高(峰值减去静息值)分别为10.6±2.1、15.9±1.2、30.6±3.7,相邻两组间存在显著性差异(P<0.05)。FI相对变化百分比分别为(40.1±6.3)%、(63.0±11.2)%、(100.8±18.4)%,相邻两组间存在显著性差异(P<0.05),且呈明显剂量依赖性。(2)在Hank's液中,10μg/ml红霉素可引起神经元内Ca2 浓度升高,其峰高为23.2±5.6,FI相对变化百分数为(82.8±13.0)%。(3)当用胃动素受体的抗体孵育后,再加入10μg/ml红霉素,其胞内Ca2 浓度的升高几乎完全被抑制。结论胃动素和红霉素均可引起体外培养肠肌间神经元的功能应答。胃动素可明显升高肠肌间神经元内Ca2 浓度,且呈剂量依赖性;红霉素可作用于肠肌间神经元胃动素受体引起胞内Ca2 浓度的升高。

Effects of motilin agonists on intracellular calcium mobilization in cultured rat myenteric neurons

OBJECTIVE: To investigate the effects of motilin agonists on intracellular Ca(2+) mobilization in primary cultured rat myenteric neurons. METHODS: Motilin-induced and erythromycin-induced intracellular Ca(2+) signaling was studied in primary cultures of rat myenteric neurons using the radiometric Ca(2+) indicator Furo3/AM with a laser confocal microscope. RESULTS: In Hank's solution, 10(-8), 10(-7), and 10(-6) mol/L motilin could elevate intracellular Ca(2+) concentration ([Ca(2+)]i) to the peak levels of 10.6-/+2.1, 15.9-/+1.2, and 30.6-/+3.7 respectively with their relative percentage change in fluorescent intensity of (40.1-/+6.3)%, (63.0-/+11.2)%, and (100.8-/+18.4)% respectively, indicating the dose-dependent effect of motilin on [Ca(2+)]i. In Hank's solution, 10 microg/ml erythromycin could induce the elevation of [Ca(2+)]i to the average peak of 23.2-/+5.6 with the relative percentage change in fluorescent intensity of (82.8-/+13.0)%. When pretreated with the antibody against motilin receptor in Hank's solution, the effect of 10 microg/ml erythromycin was almost inhibited completely. CONCLUSION: Motilin can increase [Ca(2+)]i, and erythromycin also has this effect by binding to motilin receptor.