Objective: To observe the effect of acupuncture and moxibustion of the combined "Biao-Ben" acupoints on autonomic nervous function and related factors in rats with irritable bowel syndrome diarrhea (IBS-D).
Methods: Female SD rats were randomly divided into normal control, IBS-D model, "Biao-Ben" acupoint combination, and conventional acupoint combination groups, with 10 rats in each group. The IBS-D model was established by repeated chronic stress stimulation (water or food deprivation, painful tail pinching, exposure to a 43 ℃ environment, forced swimming in 4 ℃ water, day-night inversion and horizontal vibration) for 28 d, and followed by acute restraint stress (wrapping of shoulders, forelimbs and trunk) for 1 h and gavage of senna fluid, once daily for 28 d. For rats of the "Biao-Ben" acupoint combination group, acupuncture and moxibustion were applied to "Guanyuan" (CV4), bilateral "Zusanli" (ST36), and "Neiguan" (PC6), and for rats of the conventional acupoint combination group, acupuncture and moxibustion were applied to bilateral ST36, "Tianshu"(ST25), and "Shangjuxu"(ST37). Both acupuncture and suspension moxibustion treatment were conduced for 15 min, once daily for 21 days. The fecal water content was calculated, and the spontaneous activity behaviors (total distance of crossing and the number of squares crossed in 5 min) were evaluated by open-field tests. The abdominal withdrawal reflex (AWR) was detected. H.E. staining was used to observe the histopathological changes of colon tissue. The domains of heart rate variability (HRV) including low frequency (LF), high frequency (HF), and LF/HF were analyzed using an electrophysiological recorder. The contents of calcitonin gene-related peptide (CGRP) in serum and noradrenaline (NE) in plasma were detected using ELISA. The expression level and immunoactivity of 5-hydroxytryptamine (5-HT) in colon tissues were detected by Western blot and immunofluorescence histochemistry, separately.
Results: Compared with the normal group, the model group had a striking increase in fecal water content, AWR scores at 20, 40, 60, and 80 mm Hg, NE and CGRP contents, LF, LF/HF, and 5-HT protein expression and immunoactivity (P<0.01), and an obvious decrease in the total distance of crossing, the number of squares crossed, and HF of HRV (P<0.01). In comparison with the model group, the fecal water content, AWR scores at 20, 40, 60, and 80 mm Hg, NE and CGRP contents, LF, LF/HF, and 5-HT protein expression and immunoactivity were significantly decreased (P<0.01,P<0.05), while the total distance of cros-sing, number of squares crossed, HF of HRV were considerably increased (P<0.01, P<0.05) in both "Biao-Ben" acupoint combination and conventional acupoint combination groups. The effects of the "Biao-Ben" acupoint combination were apparently superior to those of conventional acupoint combination in down-regulating the fecal water content, AWR score at 20, 40, 60 and 80 mm Hg, NE and CGRP contents, LF, LF/HF, and 5-HT expression and immunoactivity, and in increasing the number of squares crossed, and HF of HRV (P<0.01, P<0.05). HE staining showed no pathological changes in colonic mucosa in all groups.
Conclusion: Acupuncture and moxibustion stimulation of combined "Biao-Ben" acupoints can effectively improve the symptoms (spontaneous activities, visceral hypersensitivity) of IBS-D model rats, which may be related to its functions in regulating autonomic nervous activities, and down-regulating blood NE and CGRP contents and colonic 5-HT protein expression and immunoactivity, and the effects of "Biao-Ben" acupoint combination are superior to those of conventional acupoint combination.
目的：观察“标本配穴”针灸对腹泻型肠易激综合征(IBS-D)大鼠自主神经功能及相关因子的影响。方法：雌性SD大鼠随机分为正常组、模型组、标本配穴组与常规配穴组，每组10只。采用急-慢性应激联合番泻叶灌胃法制备IBS-D大鼠模型。标本配穴组予“关元”、双侧“足三里”“内关”针灸治疗，常规配穴组予双侧“足三里”“天枢”“上巨虚”针灸治疗，均毫针直刺，留针15 min，并将艾条置于穴位上方2～3 cm，每穴悬灸15 min，1次/d，连续21 d。记录各组大鼠粪便含水率，旷场实验观察大鼠活动的总路程及方格间穿行次数，腹壁回缩反射(AWR)评分评价内脏高敏性，HE染色观察结肠组织病理形态，电生理记录仪分析大鼠心率变异性(HRV)的低频(LF)成分、高频(HF)成分及LF/HF值，ELISA法检测血浆去甲肾上腺素(NE)和血清降钙素基因相关肽(CGRP)的含量，Western blot法和免疫荧光染色法检测结肠组织中5-羟色胺(5-HT)的表达水平及荧光强度。结果：与正常组比较，造模大鼠粪便含水率显著升高(P<0.01)，模型组旷场内活动总路程及方格间穿行次数显著减少(P<0.01)，AWR评分、HRV的LF成分、LF/HF比值升高(P<0.01)，HF成分降低(P<0.01)，血浆NE含量、血清CGRP含量及结肠组织5-HT的表达水平及荧光强度显著升高(P<0.01)。与模型组比较，两针灸组治疗后粪便含水率、AWR评分明显降低(P<0.01)，旷场内活动总路程及方格间穿行次数增加(P<0.01)，HRV的LF值、LF/HF比值降低(P<0.05)，HF值升高(P<0.05)，血浆NE含量、血清CGRP含量及结肠组织5-HT的表达水平及荧光强度均显著降低(P<0.01)。与常规配穴组比较，标本配穴组治疗后粪便含水率、AWR评分降低(P<0.01，P<0.05)，旷场内方格间穿行次数增加(P<0.05)，HRV的LF值、LF/HF值降低(P<0.05)，HF值升高(P<0.05)，血浆NE含量、血清CGRP含量及结肠组织5-HT的表达水平及荧光强度降低(P<0.01，P<0.05)。HE染色显示，各组大鼠结肠黏膜均未见病理改变，与IBS-D的结肠组织形态学特征相吻合。结论：“标本配穴”针灸能有效调节IBS-D模型大鼠的HRV各项频域指标及血浆中NE含量，改善其自主神经功能，并对血清CGRP及结肠中5-HT等相关因子的表达有良性调整作用，进而改善其内脏高敏感性。.
Keywords: Acupuncture and moxibustion; Autonomic nervous function; Predominant irritable bowel diarrhea; Viscera hypersensitivity; “Biao-ben” acupoint combination.