内脏高敏感模型大鼠结肠特异背根神经节神经元的形态学和兴奋性研究

摘要

目的探讨内脏高敏感模型大鼠结肠特异背根神经节神经元的形态学和兴奋性研究。方法取出生10 d的大鼠,结肠内给予0.5%乙酸溶液制备内脏高敏感大鼠模型(AA组),对照组(NS组)给予等量0.9%生理盐水。对成年后的大鼠进行结直肠扩张(CRD),用腹部回撤反射(AWR)值来评估内脏敏感性。在结肠壁上注射1,1’-二(十八烷基)-3,3,3’,3’-四甲基吲哚羰基花青高氯酸盐(DiI)逆行标记的背根神经节(DRG)神经元为结肠特异DRG神经元,酶解分离神经节成单细胞后,荧光显微镜下观察细胞形态并计数。采用全细胞膜片钳技术记录神经元的阈电位和动作电位频率。结果与NS组比较,AA组大鼠在相同结直肠扩张压力(5.332、7.998、10.664 kPa)下 AWR值明显增高(P <0.05)；AA组结肠特异DRG神经元细胞直径为35~45μm, NS组为25~35μm；AA组神经元阈电流低于 NS组(P <0.05),2倍阈电流刺激下,引发出的AA组动作电位频率高于NS组(P<0.05)。结论大鼠结肠特异背根神经节神经元在内脏高敏感后细胞增大且兴奋性增加,这有利于今后更深层次地研究内脏敏感性增高的神经机制。%Objective To investigate the morphology and excitability of the colon-specific dorsal root ganglion (DRG) neurons in a rat model of colonic visceral hypersensitivity (CVH). Methods CVH was induced by colonic injection of 0.5% acetic acid (AA) in 10-day-old rats while control(NS) induced with 0.9% normal saline. The abdominal withdrawal reflex (AWR), induced by colorectal distention, was used to quantify the level of colonic sensitivity in adult rats. Colon-specific DRG neurons were retrogradely labeled by injection of 1,1’-dioctadecyl-3, 3,3’,3’-tetramethylindocarbocyanine perchlorate(DiI) into the colon wall and observed by fluorescence microscope after separated into a single cell with digestive enzyme. Rheobase and the frequency of action potentials(APs) in response to a current stimulation twice the rheobase from neurons were measured using whole patch clamp tech-niques. Results Colonic sensitization with acetic acid increased the AWR to innocuous levels of colorectal disten-tion (5.332,7.998,10.664 kPa)(P<0.05). DiI labeled positive neurons distributed between 35 to 45μm in AA group while between 25 to 35 μm in control. The average rheobase of AA neurons was significantly lower than that of NS group (P<0.05). The frequency of APs in AA neurons in response to a 2írheobase stimulation was signifi-cantly higher than that of NS group (P<0.05). Conclusion These data suggest that colon-specific DRG neurons in a rat model increase in size and excitability after colonic visceral hypersensitivity which is conducive to future study in neural mechanisms in visceral hypersensitivity.