The impact of Electroacupuncture on chronic stress which induced depression rats’ hippocampal struc.doc

1、1The impact of Electroacupuncture on chronic stress which induced depression rats hippocampal strucAbstract. ObjectiveThe antidepressant mechanism of EA in repairing damaged areas of astrocytes.Methods66 adult female SD rats were randomly divided into four groups. The normal control group only for n

2、ormal feeding, the other three groups adopt long-term chronic unpredictable mild stimulation singly housed jointly, establishing model of depression. Blank group, after modeling successfully with no treatment; electro-acupuncture group, in the successful modeling begins the day after electro-acupunc

3、ture treatment four passes points, selecting a continuous wave electro-acupuncture device, alternating left and right points, continuous 21 days . Drug group, on the day after the successful modeling gavage given riluzole treatment, continuous treatment for 3 weeks.Rats were sacrificed by dislocatio

4、n, taking hippocampal tissue under light microscope. ResultsThe modeling light microscope rat hippocampus astrocytes significant change in the number and morphology, and the normal control group, the difference was statistically significant (P 0.05); electroacupuncture group 2number CA3 region astro

5、cytes no significant increase, with the drug group, the difference was statistically significant (P = 0.032), in the control group and the normal control group, the difference was not statistically significant (P 0.05). ConclusionEA can fix the depression caused by chronic stress hippocampal astrocy

6、tes injury, and similar efficacy with riluzole, which may be an important way for the antidepressant of EA. Key words: Rat; EA; light microscopy; hippocampus AST; astrocytes Introduction Depression, is known as major depressive disorder, with significant and persistent low mood as the main clinical

7、feature. Currently the clinical treatment of major depressive disorder divided into drug therapy and non-drug therapy. However, due to the slow onset of Western presence, recurrence rate, toxicity, and many other big problems, leading to the use of drug medicine is limited and poor patient complianc

8、e 1-3. Non-drug therapy like acupuncture is now representative, in recent years domestic and international research results gradually valued by clinicians and patients 4. Astrocytes 3(AST) is the largest number of the central nervous system, and the most widely distributed glial cells, neurons suppo

9、rt, nutrition, rehabilitation and protection. More and more evidence improve that AST morphology and function abnormalities involved in the development of depression 5. Domestic and international research results find AST “triple synaptic structures“ 6-7 and glutamate uptake through the circulatory

10、system to regulate the role of glutamate, AST in the brain to reduce the number and function, and the glutamate acid decreased uptake and accumulation in the synapses, causing excitatory neurotoxicity onset depressive disorder is considered a key link in a new research progress. In this study, rats

11、hippocampal AST as a target to study the morphology and function in the pathophysiology of depression and EA changes after antidepressant treatment. It is a new field in acupuncture treatment of depression mechanism, elucidating that acupuncture antidepressant mechanism of action and clinical treatm

12、ent of acupuncture has important guiding significance. Materials and methods Animals and Grouping 7-8 weeks old, weighing 220-250g, 66 adult female SD rats were randomly divided into four groups: normal control group (n 4= 12), control group (n = 18), EA group (n = 18) and drug group (n = 18). All r

13、ats were caged alone. The normal control group has no treatment, and the remaining three groups use chronic unpredictable mild stimulation singly housed jointly establish rat model of depression. Blank group, after modeling successfully have no treatment; electro-acupuncture group, in the successful

14、 modeling begins the day after electro-acupuncture treatment four customs points, electro-acupuncture device continuous wave, frequency 15-30Hz, every 15-20 minutes, alternating left and right. First week, treatment is everyday. From next week, the treatment is once every other day for three weeks;

15、drug group, on the day after the successful modeling gavage riluzole treatment given every 12 hours (4mg / kg), continuous treatment for 3 weeks. Reagents and Instruments Select G-6805electric acupuncture apparatus, XSP-2C light microscope by the Shanghai Optical Instrument Factory production; needl

16、e: Medical Devices Co., Ltd. Wujiang City Yunlong producing disposable cosmetic acupuncture needles, size 0.16 7mm (half an inch). Riluzole (Sanofi), Na2HPO4 ? 12H2O (Shanghai Sinopharm), NaH2PO4 2H2O (Shanghai Sinopharm), NaOH tablets (Shanghai Sinopharm), paraformaldehyde (Tianjin 5Chemical Co., L

17、td.), hematoxylin (Sigma H3136), aluminum potassium sulfate dodecahydrate (Guangdong Guanghua Chemical Reagent Factory), sodium iodide (Guangdong Guanghua Chemical Reagent Factory), xylene (Guangdong Guanghua Chemical Reagent Factory), water-soluble eosin (Guangdong Guanghua reagent Factory), ethano

18、l (Guangdong Guanghua reagent plant), 10% formalin (Guangdong Guanghua Chemical Reagent Factory). points positioning Hegu(LI4),Taichong(Liv3) acupoints in the pharmaceutical industry based on the national higher education “Twelfth Five-Year Plan“ materials (Ninth Edition) “experimental acupuncture“

19、(Guo Yi, Fang Jian Qiao editor of Chinese Medicine Press, 2012), and with reference to comparative anatomy learn and simulate the human body Meridian positioning point positioning method in rats. Hegu: between first and second metacarpal of the forelimb , so each one hole; Taichong: hind dorsal firs

20、t and second metatarsal depression, left and right side of a hole. Preparation of the model In this study, chronic unpredictable mild stimulation singly housed jointly modeling, in which the normal control rats per cage feeds three, without stimulation. Each model group rats were caged in 7 days for

21、 a period of a day to accept a stimulus.7 kinds of different 6stimuli, the cycle, a total of five cycles, with a total length of 35 days. Using water deprivation, fasting, Akira tail, tail clamp, ice swimming, day and night reversed, foot shock, over a variety of stimuli in order to follow the imple

22、mentation of the principles from weak to strong, in order to avoid stimulating interaction. Daily use one kind of stimulation, which is adopted average five times each, the same stimulation did not consecutive, so that animals can not anticipate the occurrence of stimulating 8-9. Interventions Norma

23、l control rats are feeding without special treatment. After modeling, the control group are without any intervention. Acupuncture group were successful in modeling after acupuncture treatment, selecting G-6805 type electro-acupuncture device density wave, frequency 215 Hz, amplitude of 3V, acupunctu

24、re 1 times/d, 15 min/times, acupuncture the Hegu, Taichong, alternating left and right, 1 times/d, were treated for 21 d. Drugs in rats after a successful modeling for drug riluzole orally at a dose of 4 mg / kg, 1 times/d, treated for 21days. F. Specimen collection and testing Rats were anesthetize

25、d with 10% chloral hydrate (40mg/kg) intraperitoneally and thoracotomy, the ascending aorta was 7intubated, being cut right atrium, the first is to be perfused 0.1M PBS 250ml, 150 ml infusion followed by light microscopy specimens were fixed solution (4% paraformaldehyde +0.1 MPBS mixed) last 30min;

26、 decapitated, brains were removed, blunt dissection hippocampus, immersed in the same fixative fixed after 612h; hippocampus were then immersed in 10%, 20% and 30% sucrose PBS, until the tissue in the end sinking conducted after tissue dehydration, sectioned and stained. Image Analysis With the Leic

27、a electron microscope image acquisition, each rat select relatively intact hippocampus HE staining of five, each HE staining under 200 times the light microscope observation side of hippocampal CA1, CA3 region, DG random region the five non-overlapping same area (0.1mm2) vision, using image-Pro Plus

28、 6.0 image analysis system (Media Cybernetics Inc., USA), taking into account the number of astrocytes. Section analysis are in the same strength, carried out under the same magnification, averaged for statistical analysis. Statistical Using PASW statistics18.0 statistical software for statistical a

29、nalysis of the experimental data, the experimental measurement data if normally distributed computing, groups were 8compared using analysis of variance, not normal or heterogeneity of variance, using the non-parametric test; between groups LSD test was used to compare, test level = 0.05. Results The

30、 impact of CUMS singly housed joint modeling on hippocampal districts astrocyte morphology Through Figure one we can see, Blank group compared with the control group, CA1 region: pyramidal cell layer thinning, fewer number of cells (20-30%). Some astrocytes karyopyknosis (5-10%); CA3 region: part py

31、ramidal cell degeneration, cytoplasmic condensation, stain (60-70%), partly astrocytes karyopyknosis (5-10 %); DG region: some granular cytoplasmic vacuoles (10-20%), partly astrocytes karyopyknosis (1-5%). These results suggest that, CUMS singly housed after modeling combined hippocampal astrocyte

32、morphology changes, mainly as pyknotic nuclei. While some brain regions (CA1 region, CA3 region) pyramidal cell degeneration occurs, reducing the number of cases, some granular cytoplasmic vacuoles in DG area. The impact of EA and riluzole gavage in depression rat hippocampus districts astrocyte mor

33、phology Through Figure two we can see, CA1 region:pyramidal cells 9in the EA group were normal, some astrocytes karyopyknosis (5-10%); drug group were normal pyramidal cells, some astrocytes karyopyknosis (20-30%); CA3 area:EA group astrocytes reduce, some astrocytes s karyopyknosis (1-5%); drug gro

34、up pyramidal cells and astrocytes showed no abnormalities. DG area:EA group granule cells and astrocytes showed no abnormalities; drug group were normal granulosa cells, some astrocytes karyopyknosis (5-10%). The experimental results showed that: compared with rats in Figure one under the light micr

35、oscopy, After EA and riluzole, electro-acupuncture group and medication groups CA1 pyramidal cells were normal, astrocytes karyopyknosis situation is not see significant improvement. CA3 pyramidal cells in both groups returned to normal, the drug group returned to normal astrocytes, glial in EA grou

36、p karyopyknosis partially restored. DG granule cells and astrocytes in EA group returned to normal, DG granule cells in drug group return to normal, astrocytes recovery is not obvious. Tip: EA is effective to CA3 and DG regions astrocyte morphology, but on the CA1 region the repair is not obvious; r

37、iluzole on astrocyte morphology CA3 region has a good repair ,but the CA1 and DG regions is not obvious. EA and riluzole 10impact on astrocytes, and on hippocampal pyramidal cells, granule cells have a good repair. Discussion Depression attributed to Chinese medicine, “ Melancholia “ category , the

38、cause is emotional injuries , the main lesions is the stagnation , both blood stasis, phlegm knot of fire and so on , liver, spleen and related organs , especially the liver are closely related, “liver depress “is the key to disease pathogenesis. Four passes points due to the Hegu, Taichong two pair

39、s of points in the clinical use of mutual compatibility of the name. Hegu is hand Yangming large intestine by the original hole , Yangming as much gas and more blood through the hole with qi and blood. Taichong is Jueyin Liver Meridian of Foot point, the original hole, the less gas as much blood thr

40、ough . Liver is the main catharsis. Taichong have qi and blood, liver and qi , have the effect of calm the liver to stop the wind. Hegu match with Taichong transfer gas can enhance the role of liver. In recent years, our group consider “ Soothing the liver and regulating the God “ as the treatment method , selecting the four passes points based, with Baihui , Yintang, to carry out acupuncture treatment of depression in clinical studies , the results showed that acupuncture and a new generation of glutamyl acid antagonist riluzole quite effective , has good