ã€Abstract】 Objective To treat Alzheimer's disease model rats by acupuncture and observe the improvement of learning and memory. Methods A rat model of Alzheimer's disease was induced by administering D-galactose to rats. The normal group, the model group, the drug group, the acupuncture group and the electroacupuncture group were respectively established. In the normal group and the model group, only the same dose of normal saline was administered, and the drug group was treated with the Huperzine A tablet (Haberin) suspension. The acupuncture group used acupuncture Baihui, Shenting, Shenshu, too The method of brook and Zusanli was used for treatment. The electroacupuncture group was treated with acupuncture at Baihui, Shenting, Shenshu, Taixi and Zusanli, and connected with an electroacupuncture instrument for treatment. Take 10 days as a course of treatment for a total of 3 courses. The rat's learning and memory ability was measured by a rat platform test, and the tau protein content in the hippocampal CA1 region of the rat brain was observed. Results After modeling, the memory escape latency of the normal group was significantly lower than that of the model group (P<0.05). After the treatment, the escape latency of the drug group, acupuncture group and electroacupuncture group was shorter than that of the model group (P<0.05). ). The efficacy of the electroacupuncture group was better than that of the acupuncture group and the drug group (P<0.05). After treatment, the P-tau protein expression group in the hippocampal CA1 area was lower than the model group, and the electroacupuncture group was superior to the drug group and the acupuncture group (P<0.05). Conclusion Electroacupuncture can improve the memory ability of Alzheimer's disease model rats and has a certain therapeutic effect on Alzheimer's disease.
ã€Key words】 Acupuncture; Alzheimer's disease; Memory ability; Rat
With the increase of age, the incidence of dementia in the elderly is gradually increasing, among which Alzheimer's disease (AD) is the most common. This is a demyelinating degeneration of the central nervous system (brain) nerve cells. Clinically, the main manifestations are memory, judgment, learning, mobility, and self-care ability, which have adversely affected the quality of life and family life. At present, there are no special effects drugs and treatment methods available, and the acupuncture treatment of Chinese medicine has received relatively obvious improvement effects in clinical practice. The mechanism of action has not been revealed. This article has observed from the aspects of neurobehavioral and histochemistry, and hopes to provide evidence for the mechanism of acupuncture treatment of the disease. 1 Materials and methods
1.1 Materials
1.1.1 Main equipment
The Morris water maze produced by Shanghai Xinsoft Information Technology Co., Ltd. is an improved water maze, and its platform can appear regularly. A B50 type balance produced by Japan Co., Ltd., an in situ hybridization kit for testing AD P-tau protein supplied by Sigma Biotechnology Co., Ltd., USA. The "Great Wall" electroacupuncture instrument produced by Jiangsu Huasheng Medical Equipment Factory in China.
1.1.2 Main application reagents
The D-galactose used in the induced AD model was produced by Beijing Berry and Kang Biotechnology Co., Ltd. The drug used in the drug group was a suspension of Huperzine A (Habbin) produced by Henan Zhulin Zhongsheng Pharmaceutical Co., Ltd.
1.2 Animal screening
All Wistar rats purchased from the Animal Experimental Center of Heilongjiang University of Traditional Chinese Medicine were placed in a water maze to conduct adaptive swimming training, record their swimming distance, learn to escape the incubation period and the number of learning errors, and train for 1 week. Rats with impaired swimming ability and learning and memory ability were eliminated. Finally, 50 rats that met the experimental conditions were selected for the final experimental procedure.
1.3 Model preparation method
Place the rats ready for modeling in the same environment for adaptive feeding for 1 week, keeping plenty of light, moisture and food. Then, D-galactose was injected into the navel and abdomen at a dose of 200 mg per kilogram of body weight per rat, once a day. It lasted for 6 weeks. Then all the model rats were numbered and divided into 10 model groups, 10 drug groups, 10 acupuncture groups, and 10 electroacupuncture groups according to the random principle. Ten normal rats were taken as the normal group.
1.4 treatment
After the start of treatment, the normal group and the model group were given daily bundling for 30 min, and the same amount of physiological saline was administered to the drug group; the drug group was administered with 2 mL of the Huperzine A suspension ( 17 mg); acupuncture group used acupuncture needles of Huashi brand 0.30×13 mm to acupuncture rats Baihui, Shenting, Shenshu, Taixi and Housanli points, and then acupuncture for 2 min, leaving needle for 30 min; The needle set is connected to the "Great Wall" electroacupuncture instrument on the basis of the acupuncture group, using a sparse wave for 30 min. Baihui Point is at the center of the skull in the midline of the head; Shenting is at the front of the midline of the head; Shenshu is on the posterior midline, 1 inch below the spine of the second lumbar spine; Taixi is at the lateral rear of the hindquarters The depression is in the posterior and third in the lateral anterior and posterior aspect of the hind limb of the rat. All rats were treated for 6 weeks and treated once a day.
1.5 Test and material observation methods
Platform test. The Morris water maze has a rubber platform in the swimming pool, 1 cm from the water. The rats were placed in the pool to start recording, and when the rats were swimming to find the platform under water, they were recorded as learning escape latency. The above test was repeated after 24 h, and the second time required was the memory escape latency. This is used as an indicator of the learning and memory ability of rats. The rats were tested for escape latency and memory escape latency at the first week before the model establishment, the seventh week before the treatment, and the third week after the treatment.
After the end of the water maze test, the rats were anesthetized and then died by excessive brain perfusion. The rat skull was incised and the rat brain tissue was removed and stored in a 1% deep DEPC solution at a depth of 4%, and then fixed with paraformaldehyde for testing. The fixed brain tissue was equally divided into 5 parts from the frontal pole to the occipital pole, and the second brain tissue was taken out and placed in water at 24 ° C for gradient dehydration with ethanol. The order was 70% ethanol, 80% ethanol, 95% ethanol, 100% ethanol 1 degree, 100% ethanol 2 degrees, 100% ethanol 3 degrees, and then the sections were stained and the tissue was made transparent with xylene. Positive detection of tau hyperphosphorylation was performed using the AD P-tau protein in situ hybridization kit. The brown-yellow particles appearing in the cells are hyperphosphorylated tau protein expression. Finally, statistics on the positive expression of tau protein were performed.
1.6 Statistical methods
Edit images and tables with an electronic computer. All data was counted using the SPSS 13.0 software package. The means of measurement were expressed as mean ± standard deviation, and the mean comparison between groups was analyzed by variance, with P < 0.05 as a measure. Each slide was taken for 3 fields of view when the number of positive cells was observed, and the average number of particles was counted.
2.1 General condition of the rat
The rats in the normal group were full of energy, sensitive, quick-acting, shiny, and normal diet and bowel movements. Rats in the model group showed varying degrees of fatigue, slow response, dull action, poor physical strength, rough coat, poor diet, less bowel movements or thin stools. This situation begins after the model is built until the end of the experiment.
2.2 Rat behavioral indicators observation results
It can be seen from Table 1 that the learning escape latency of the rats in each group before the model establishment was approximately equal, and the difference was not statistically significant (P > 0.05). Before treatment (ie, after 6 months of modeling), each module was large. The learning latency of the rats was significantly prolonged, and the difference was statistically significant compared with the normal group (P <0.05), suggesting that the model was successful, and the learning ability of the rats decreased, with the preliminary characteristics of AD. After treatment, that is, at the 13th week, the learning escape latency of the rats in each treatment group was significantly shorter than that of the model group, and the difference was statistically significant (P <0.05), indicating that after acupuncture or drug treatment, the learning ability of AD rats was A certain recovery. The situation of the electroacupuncture group was more optimistic. Compared with other treatment groups, the shortening of learning escape latency was more obvious, and the difference was statistically significant (P <0.05). It can be seen from Table 2 that the memory escape latency of the rats in each group before the model establishment was approximately equal, and the difference was not statistically significant (P > 0.05); before the treatment (ie, after 6 months of modeling), each module was large. The memory escape latency was significantly prolonged, and the difference was statistically significant compared with the normal group (P <0.05), suggesting that the model was successful, and the memory ability of the rats was decreased, with preliminary characteristics of AD. After treatment, that is, at the 13th week, the memory escape latency of the rats in each treatment group was significantly shorter than that of the model group, and the difference was statistically significant (P <0.05), indicating that after acupuncture or drug treatment, the memory ability of AD rats was A certain recovery. The situation of the electroacupuncture group was more optimistic. Compared with other treatment groups, the memory escape latency shortened more significantly, and the difference was statistically significant (P <0.05).
3 Discussion
Many studies have fully confirmed that humans' ability to learn and remember spatial structure is more based on hippocampal neurons in brain tissue, while AD is a persistent neuronal degeneration of brain tissue in this region. And make space learning and memory ability. Learning and memory ability as an important test point of mental indicators of dementia patients, with the indication status of gold indicators, so in the experiment we used the learning and memory escape latency of rats as an indicator. The above results clearly show that the spatial learning and memory ability of AD model rats has been significantly degraded after intraperitoneal administration of D-galactose, indicating that D-galactose absorption can cause degeneration of rat brain cells. Tau
The production of negative substances such as proteins degrades brain function. However, after acupuncture, the performance of the rat in the water maze was gratifying. Although the brain cells of the rats were still degenerated during the 6 weeks of treatment, the latency did not drop very quickly, even the electroacupuncture group. There are also improvements, experiments have confirmed that acupuncture treatment can play a role in improving brain function like Western medicine. After treatment, the learning and memory ability of the rats was higher than before the treatment. Of course, since AD ​​is not only a functional change, but also a part of the instrumental changes, acupuncture can only play a role in the recovery of treatment within a certain range, and can not completely restore the memory function of AD rats, but the experiment also shows that Electroacupuncture is more effective than ordinary acupuncture, indicating that if the acupuncture method and method are improved, the effect can be further expanded, and the effect after treatment is infinitely close to normal. From the biochemical level, the excessive production of tau protein has a negative effect on brain function, and both the drug group and the acupuncture group achieve the purpose of inhibiting the production of the protein to varying degrees. Electroacupuncture can reduce the expression rate of tau protein more effectively. On the one hand, it shows that tau does cause disorder in rat memory system. On the other hand, it also suggests that if it can reduce the expression of tau protein, it can improve brain function to a certain extent. .
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