Study Of Acupuncture Treatment On Functional Dyspepsia Rats With Syndrome Of Liver Stagnation And Spleen Deficiency Through The Brain Gut Axis Way

Objective:To observe the influence of electro acupuncture(EA)treatment on functional dyspepsia(FD) rats with synd-rome of liver stagnation and spleen deficiency in general evaluation, daily behavioral changes, gastrointestinal dynamic changes, both central and peripheral related brain gut peptide :VIP CGRP and its receptor VPAC1 RAMP1 expre-ssion.To investigate the role of brain gut axis and explore the mechanism of EA treatment on functional dyspepsia rats with syndrome of liver stagnation and spleen deficiency from the brain-gut axis way. Methods:72 SPF Sprague-Dawley rats(weight 200±20g)with female and male each half were randomly divided into three groups(n = 24 each): a blank group, a model group, and an EA group. Except for the blank group(far away from modeling environment during the modeling period and avoid the interference),the other two groups underwent modeling totally 14 days by tail clamped stimulation(2 times/d), giving an irregular diet(fastingevery other day, with free access to water), and gavage of ice physiological saline(-4℃ 0.9%Na Cl injection 2 m L, 2 times/d).During the modeling period, general situations of rats were observed and recorded, including the mental state, hair color,ear color, activity, diet and drink, defecation frequency and stool quality, weight and so on.As FD was successfully induced, EA treatment started(28d, once a day) and acupuncture at Zusanli(ST36)and Taichong(LR3). After 28 days, the rats were killed to take tissue samples. The rates of gastric emptying and small intestinal transit were determined; gastricantrum and jejunum tissue were observed by HE staining method; Serum VIP CGRP content were determinated by ELISA; VIP CGRP expression levels of each rat in gastric, intestinal tissue were determinated by PCR; the expression levels of VIP CGRP and receptors VPAC1 RAMP1 in the hypothalamus, gastric and jejunum were determinated respectively by Western blotting; the expression levels of VIP CGRP and their receptors VPAC1 RAMP1 in jejunum,hypoth-alamus and hippocampus were determinated respectively by IHC.Results: 1.Observingthe states before and after treatment:Before treatment, the blank group rats in the mental state and activity are good, fur shiny and neat, stool was dry brown globosity. The model group and the EA group rats fur withered knot, activity decreased significantly, arrest unresponsive, stool was watery yellow. Compared with the blank group, the model and EA group rats in general score, body weight growth, growth of food and water intake per day were significantly decreased(P<0.01); After treatment, EA group rats fur to restore luster and tidy, good activity, food intake increased significantly, response for catching quick, stool turned to dry brown globosity, general score, body weight growth, growth of food and water intake per day significantly increased(P<0.01); the model group rats did not change significantly than before treatment(P>0.05).2. The histomorphology :light microscope(×400) observed in the gastric and jejunum tissue of rats,no organic change was found,gastric mucosa and jejunal wall layers of epithetlial structure integrity,cells arranged in neat rows, submucosa and muscle layers is clear and consecutive, no inflammatory infiltration and glandular epithelial lesions pathological manifestations.3. GI dynamics change: Compared with the blank group, gastric residual rate significantly increased(P<0.01), small intestinal transit rate significantly decreased(P<0.01) in the model group; compared with the model group,the gastric residual rate decreased significantly(P<0.01) and small intestinal propulsion rate increased significantly(P<0.01)in the EA group.Suggest that acupuncture can significantly promote gastrointestinal function, increase gastrointestinal motility.4. Effect of the expression of VIP CGRP :1Serum VIP, CGRP content were determinated by ELISA: Compared with the blank group, the model group rats serum VIP, CGRP content were significantly increased(P<0.01);compared with the model group, the EA group rats serum VIP, CGRP content signify-cantly reduced(P<0.01).2The relative expression levels of VIPm RNA,CGRPm RNA in gastric, intestine and hypothalamus were determinated by PCR: Compared with the blank group, the model group relative expression levels of VIPm RNA CGRPm RNA both in gastric, intestine and hypothala-mus was significantly increased(P<0.05);compared with the model group, the results were significantly decreased in the EA group(P<0.05).5. Effect of the expression of VIP CGRP and corresponding receptor RAMP1, VPAC1:1The expression of VIP CGRP RAMP1 and VPAC1 in gastric, jejunum, hypothalamic determinated by WB:Compared with the blank group, the model group relative expression levels of VIP, CGRP, VPAC1 and RAMP1 in gastric, jejunum and hypothalamus significantly increased(P<0.05); compared with the model group, the results were significantly decreased in the EA group(P<0.05).2The expression of VIP CGRP RAMP1 and VPAC1 in jejunum, hypothalamic and hippocampus was determinated by IHC: Compared with the blank group, the model group expression of VIP, CGRP, VPAC1 and RAMP1 in jejunum,hypothalamic and hippocampus significantly increased(P<0.05); compared with the model group, the results were significantly decreased in the EA group(P<0.05).Conclusion: 1. Our experiment successfully reproduced the closest to the clinical onset and symptoms of liver stagnation and spleen deficiency type FD rat model, combining tissue morphology, suggesting the model established by the multifactorial intervention scheme in this study was in line with the diagnostic criteria of functional dyspepsia.2. EA treatment has significant therapeutic effect on FD rats, can significantly improve the FD symptoms, and effectively enhance the gastric emptying and small intesti-nal propulsion, reduce intestinal sensitivity.It indicates that the effect of EA on FD is the improvement of gastro-intestinal motility disorders and sensory abnormalities.3. In the model group, the expression of VIP CGRPin both peripheral and central was significantly increased, which indicated that the abnormal brain gut peptide secretion and brain gut interaction was closely related to the pathogenesis of FD.4. EA treatment can significantly decrease peripheral and central VIP CGRP and receptors VPAC1 RAMP1 expression.It indicates that EA can regulate the secretion of brain gut peptides through the brain gut axis, correct the abnormal expression of brain gut peptides, thereby improving the gastrointestinal motility disorders and sensory abnormal-ities,This is the mechanism of EA effective treatment on FD.Therefore, we believe that:the abnormal brain gut peptide secretion and brain gut interaction which leading to gastrointestinal motility disorder and sensory abnormal-lities was the pathophysiological basis of FD.EA can significantly improve the syndrome of FD with stagnation of liver and spleen deficiency, furthermore down regulated VIP CGRP VPAC1 RAMP1 expression in both peripheral and central. Owing to the regulation of different levels and multiple targets through the brain gut axis, EA can correct the abnormal secretion of brain gut peptides, so as to achieve the purpose of treatment on FD.