Discussion on the model of electroacupuncture anti-depression research

Discussion on the model of electroacupuncture anti-depression research

【Abstract】 Objective: To explore the selection of suitable models for electroacupuncture antidepressive research.

METHODS: The drug experiment method was used to observe the mortality of norepinephrine toxicity test mice and the number of taro counts in serotonin test mice. The method of changing the environmental conditions was used to observe the immobility time of the tail of the tail-suspended mice and the activity of the rats in the wild-type experiment by the chronic stress model and the intake of the syrup in the syrup test.

RESULTS: Compared with the blank control group, electroacupuncture could not reduce the mortality of norepinephrine-toxic mice, and could not reduce the number of steamed mice in serotonin test mice; The immobile time of the tail; in the chronic stress model, compared with the model group and the model restraint group, the electroacupuncture group and Baiyoujie group can significantly increase the number of vertical movements and the intake of sugar water in the open field experiment.

Conclusion: The experimental study of acupuncture on depression should choose animal models of depression induced by environmental conditions. Drug-induced depression models may be inappropriate.

【Key words】 Depression, chronic stress, electroacupuncture, animal model

Depression is a group of mood disorders or affective disorders caused by various causes, with depression as the main symptom. In recent years, the incidence rate has been rising, which seriously affects people's physical and mental health. At the same time, depressed patients are accompanied by serious suicidal tendencies. . The disease has taken the second place in China's disease economic burden ranking, 6.2%. The current treatment of depression is mainly medical treatment. In the past, monoamine oxidase inhibitors and tricyclic antidepressants were the mainstays. In recent years, serotonin antidepressants represented by Baiyoujie (fluoxetine hydrochloride) have been developed. . Because depression is a multi-system disease, drug treatment is mostly directed to a single target, the long-term application of the effect is reduced, and there are serious toxic side effects and excessive risk. Acupuncture treatment of depression has achieved certain curative effect in clinical practice, and has the advantage of overall regulation, and is non-toxic and side effects.

There are two main types of animal models for depression research, drug-induced animal models of depression and animal models of depression induced by environmental conditions [1]. Many scholars have evaluated these two types of models from the perspective of drug research. However, the mechanism of action of acupuncture is different from that of drugs, so the study of acupuncture and depression should first select a depression model that conforms to the principle of acupuncture. We selected the norepinephrine toxicity test and the 5-hydroxytryptamine taro test in the drug-induced model and the mouse tail suspension test and chronic stress model in the animal model of depression induced by environmental conditions.

Materials and Methods

1.1 Materials

Norepinephrine (NE), 5-hydroxytryptophan (5-HTP), hydrochlo-ric pargylin, positive control drug Baiyou solution. Electro-acupuncture instrument. Male Kunming mice and SD rats, clean grade.

1.2 Electroacupuncture method

According to the animal selection map, the Yintang and Baihui points, which are equivalent to the human anatomy, are selected, which are common points for acupuncture treatment of depression. The needle insertion needle, the electroacupuncture instrument, the electroacupuncture frequency of the rat and the mouse are both 2 Hz, the depth of the needle of the mouse is 0.2 to 0.5 cm, the current intensity is 0.2 to 0.3 mA, and the depth of the needle of the rat is 0.5. ~1cm, the current intensity is 0.6 mA, and it is better to use head tremor. Because the "Baihui" and "Yintang" points are close to each other, care should be taken to avoid short circuits in the electro-acupuncture operation.

1.3 Norepinephrine (NE) toxicity test [2] Male Kunming mice (18 ~ 22 g) were randomly divided into 3 groups according to body weight, blank group, Baiyou solution group, electroacupuncture group, 10 in each group. The dosage of Baiyoujie group was 2.6 mg/kg, and it was intragastrically administered at 0.2 mL/10 g once a day for 1 week. The electroacupuncture group was electroacupuncture once a day for 20 minutes for 1 week. Each group was injected subcutaneously with NE (12 mg / kg) [2] on the 7th electroacupuncture and 1h after administration. The drug concentration was 1.2 mg / mL. The mice were placed in plastic cages and allowed to drink water and eat freely. After 48 hours, the number of deaths in each group of mice was observed.

1.4 The serotoninic acid (5-HTP) taro experiment [2] animal and group, Baiyou solution administration and electroacupuncture method is the same as 1.3. On the 7th day, Paglin hydrochloride 100 mg / kg was intraperitoneally injected. After 30 min, the electroacupuncture group was given acupuncture for 20 min. After 1 h, Baiyoujie group was intragastrically administered. After 30 min, the tail vein was injected with 5-HTP 10 mg / kg [3]. ], and immediately placed in a 24 cm × 15 cm × 15 cm mouse box, the number of taro in mice every 5 min, a total of 25 min.

1.5 Mouse tail suspension experiment

Animals and groupings, Baiyou solution administration and electroacupuncture methods are the same as 1.3. After the acupuncture and administration for 1 h on the 7th day, the mouse tail was adhered to the lower end of one side of the suspending line at a distance of 2 cm from the tip of the tail. The head of the mouse was 30 cm from the ground and the weight was hung on the other side. Adjust the balance on both sides. When recording, the animal end suspension wire of the stent passes through a 50g tension transducer, the animal end is weighted by 5g, and the balance is re-adjusted (as a preload), and the transducer is connected to the Rm6240 B/C physiological experiment. On the computer of the system, record 5 min [4] after 1 min according to the indication of the system, and convert the number of cells (s) included in the immovable time period into time (s). The time of suspension of the mice in each group was recorded.

1.6 chronic stress model

Modeling method: according to the literature method improvement [5], taking 50 SD rats, weighing 160-180g, through the open field experiment, the number of horizontal movements plus vertical movements less than 30 times or more than 120 times of the rats were removed, and then Randomly grouped according to body weight, divided into blank control group, model group, Baiyou solution group, model binding group (due to the selection of chronic stress model, electroacupuncture will also exert restraint stress stimulation on rats, in order to avoid this For the error, we added the model binding group, and the model was given to the rats in the same way as the acupuncture and the stimulation time, and the electroacupuncture group, 10 in each group. The model animals were all orphaned and received 7 different stress stimuli for 21 days, including cold water swimming (10 ° C, 5 min), tail (3 min), water ban (24 h), fasting (24 h), electric shock ( The voltage was 30V, electric shock was 5s, intermittent 5s for 2 min), day and night reversal (24 h), incubator (40 ° C, 5 min), and the average stimulation was 3 times. Rats in the normal group were housed in 5 cages without any stimulation. Simultaneous administration and acupuncture treatment (method same as above). On the 22nd day, the field experiment and the sugar water experiment were carried out.

Open field test method: According to the reference [6], the open box used in this experiment is cuboid, 40 cm high, 80 cm long and wide, the perimeter wall and the bottom surface are black, and the bottom surface is composed of 25 pieces of equal area. Line division.

Place the rat in the center square of the bottom of the open box, and use the number of cells across the bottom of the floor (the squares in which all four claws enter can be counted) as the number of horizontal movements, and the number of subsequent limbs upright (two forepaws vacating or climbing) Wall) is the number of vertical movements. Each animal was measured once, each time was measured for 180 s, and the number of horizontal movements and the number of vertical movements were recorded.

Sugar water test method: After the rats were banned from water for 24 h, 1% sucrose water was added, and the weight of the bottle was 250 g. After 24 h, the weight of the sucrose water bottle was weighed, and the sucrose water consumption of the rats was calculated within 24 h.

1.7 Statistical methods

Data were expressed as x±s, and one-way analysis of variance was performed using statistical software SPSS 11. 0 with <0.05 as the level of significance of difference. The NE toxicity test is expressed as a percentage and subjected to an X ² test.

result

2.1 Effect of electroacupuncture on mortality of mice with NE toxicity test

Table 1 Comparison of mortality of each group of NE toxicity test mice (%)

Group n mortality (%)

Blank group 10 20%

Baiyou Solution Group 10 70%*

Electro-acupuncture group 10 40%

Comparison with blank group: *P < 0.05.

The results in Table 1 showed that there was no significant difference in mortality between the sublethal dose of NE and the electroacupuncture group and the blank group, while the mortality of the Baiyoujie group was significantly higher than that of the blank group.

2.2 Effects of electroacupuncture on 5-HTP-induced mouse taro experiments

After the administration of 5-HTP, the mice quickly showed a "taro" action, and the frequency of the steamed bread reached a peak between 10 and 25 minutes. It can be seen from Table 2 that there is no significant difference between the number of steamed mice in the electroacupuncture group and the blank group, and the Baiyoujie group can significantly increase the 5-HTP-induced mouse head movement.

Table 2 Comparison of the number of 5-HTP-induced taro in each group (s±s)

Group n number of hoes

Blank group 10 68.3 ± 24.7

Baiyou Solution Group 10 94.8 ± 33.7*

Electroacupuncture group 10 70.2 ± 28.7

Comparison with blank group: *P < 0.05.

2.3 Effect of electroacupuncture on the immobility time of mouse tail suspension experiment

Table 3 Comparison of the time of suspension of mice in each group (x±s)

Group n no time (s)

Blank group 10 125.02 ± 25.42

Baiyou Solution Group 10 88.23 ± 21.78*

Electric needle set 10 36.49 ± 13.48* ^★

Compared with the blank group: *P < 0.05; compared with the Baiyou solution group: ^★ P < 0.05.

It can be seen from Table 3 that in the 5 min of the mouse tail suspension recording, the immobility time of the electroacupuncture group was significantly less than that of the blank group, and the Baiyoujie group also shortened the immobility time of the mouse tail, and at the same time, the electroacupuncture group There are also significant differences compared with the Baiyou solution group.

2.4 Effects of electroacupuncture on the open field experiment of chronic stress model rats

Table 4 Changes in activity of rats in each group (x±s, 180s)

Group n horizontal motion vertical motion

Blank group 10 44.6±12.69 8.6±2.29

Model group 10 11.0 ± 6.09* 3.7 ± 2.21*

Baiyou Solution Group 10 15.7 ± 7.61 6.5 ± 2.40 ^#

Model binding group 10 6.7 ± 3.65 1.1 ± 0.85

Electro-acupuncture group 10 17.0 ± 8.14 ^★ 9.6 ± 3.43 ^{# ★}

Compared with the blank group: P < 0.05; compared with the model group: # P < 0.05;

Comparison with the model binding group: ^★ P < 0.05. (the same below)

It can be seen from Table 4 that both chronic stress and restraint stimulation on the basis of chronic stress have an effect on the behavior of rats. The number of horizontal movements and vertical movements of the model group and the model-binding group were significantly less than the blanks. The rats given the positive control drug Baiyou solution had significant difference in the number of vertical movements compared with the model group; the effect of the electroacupuncture group on the depression behavior of the stress model was expressed in both the number of horizontal movements and the number of vertical movements, that is, The model restraint group has significant differences in horizontal motion, and the number of vertical movements is significantly greater than that of the model group and the model binding group.

2.5 Effects of electroacupuncture on sucrose intake in rats with chronic stress

Table 5 Changes in sugar water intake of rats in each group (x±s)

Group n water intake (mL)

Blank group 10 126.0±29.1

Model group 10 86.4±23.4*

Baiyou Solution Group 10 109.7±18.3 ^#

Model binding group 10 76.9±18.4*

Electric needle set 10 101.5±22.3 ^★

It can be seen from Table 5 that the sucrose intake of the model group and the model restraint group is significantly lower than that of the blank group; the sucrose intake is significantly increased in the electroacupuncture group compared with the model restraint group; the Baiyoujie group is also compared with the model group rats. There are significant differences.

discuss

The drug-induced depression animal model appeared earlier, based on the monoamine hypothesis of depression, and was used to evaluate the pharmacological effects of antidepressants with a specific neurochemical effect. One of the pathogenesis of depression is thought to be due to a decrease in the content of monoamine transmitters in the body. Drugs with antidepressant effects may have an antidepressant effect by increasing the amount of monoamine transmitters in the body. The NE toxicity test was designed, and the mice were intraperitoneally injected with a sublethal dose of NE, and then the test drug was administered. If the test drug can increase the NE content in the mouse, a lethal dose or a lethal dose of NE can cause death in the mouse. The 5-HTP taro experiment is similar to the NE toxicity test. The difference is that the test gives pajrin hydrochloride and 5-HTP to cause the girl to move. If the test drug is administered to increase the 5-HT content in the body, The number of taro in mice over a certain period of time can also increase.

In recent years, studies have found that stressful life events are a significant trigger for depression, and that chronic, low-intensity, long-term daily stress is the main cause of depression. To this end, a depressive animal model that induces animal behavior abnormalities by changing the surrounding environment is designed. This kind of animal model tries to simulate the environmental incentives of depressed patients, and uses abnormal behavior as an observation index to study the etiology of depression. The mouse tail suspension experiment was designed. After the test drug was administered, the duration of the desperate psychology caused by the environmental change was judged by observing the immobility time of the mouse after the tail was no longer struggling, thereby observing the resistance of the test drug. Depressive efficacy. Simulating human chronic stress stimulation to design a chronic stress model, by giving long-term chronic stress stimulation to rats, causing depression in rats, adding intervention means at the same time of modeling, and judging intervention through behavioral observation after modeling The antidepressant effect of the means. The mechanism of action of acupuncture is different from that of drugs. Acupuncture has a two-way regulation effect on the body. Through the benign adjustment of the body, the body is in a state of dynamic equilibrium. In the NE toxicity test and the 5-HTP taro experiment, the effect of acupuncture should be reflected in a decrease in the elevated substance content in the body. However, the results of this NE toxicity test showed that electroacupuncture did not improve the toxicity of mice caused by sublethal NE. We believe that the NE toxicity test model makes the NE content in the body rise rapidly, and the electroacupuncture cannot quickly block the spread of NE in the body, thus not reducing the mortality of mice. Similarly, in the 5-HTP taro experiment, 5-HTP and pajilin hydrochloride were rapidly administered, and the two substances in the mice were rapidly increased. The electroacupuncture could not completely block the effects of these two substances in a short time. Behaviorally, it is not possible to rapidly slow down the behavior of steamed bread caused by 5-HTP and pajilin hydrochloride. It can be seen that the acute experiment induced by this drug does not conform to the mechanism of action of acupuncture. The drug-induced depression animal model is an acute experimental model, so the inference is not suitable for the study of acupuncture anti-depression. Although acupuncture has a two-way regulation effect, it has problems of strength of action and accumulation of time. In a model of environmentally induced depression, acupuncture can improve an animal's response to adverse stimuli. The electroacupuncture balances the function by adjusting the function, thereby suppressing the desperate behavior of the mouse caused by the tail suspension and shortening the immobility time of the mouse tail. The electroacupuncture can enhance the activity of the rat after chronic stress and enhance its inquiry behavior. Improve depression due to chronic stress stimuli [7].

In summary, it can be seen that the experimental study of acupuncture on depression should choose to change the animal model induced by environmental conditions, and the drug-induced depression model may not be suitable for acupuncture research.