Construction method and application of chronic epilepsy co-disease depression mouse model

Abstract

The invention provides a construction method and application of a chronic epilepsy co-disease depression mouse model. The method comprises the following steps: firstly, sequentially carrying out operations of intraperitoneal injection of a lithium chloride solution, intraperitoneal injection of an atropine solution and intraperitoneal injection of a pirolocarb solution on mice in an experimental group to induce generation of the epileptic status; recording the epileptic seizure behavior of the mice in the experimental group, and evaluating the epileptic seizure grade of the mice in the experimental group by referring to a Racine standard to obtain a lithium chloride-pirocapene induced chronic epilepsy mouse model; evaluating the mouse depression-like behavior degree by combining a depression-like behavior experiment, screening the lithium chloride-pirocapene induced chronic epilepsy mouse model so as to determine a modeling standard and a modeling time point of the chronic epilepsy co-disease depression mouse model, and obtaining a chronic epilepsy co-disease depression mouse and a chronic epilepsy non-depression mouse; and performing depression-like behavioral comparison on the chronic epilepsy co-disease depression mouse and the chronic epilepsy non-depression mouse to obtain the chronic epilepsy co-disease depression mouse model.

Description

Technical Field

[0001] The present invention relates to the technical field of animal model construction, and in particular to a construction method and application of a chronic epilepsy comorbid depression mouse model. Background Art

[0002] The latest report on epilepsy published by the World Health Organization (WHO) in 2016 shows that there are about 2.4 million new epilepsy patients each year worldwide. As of 2016, there are about 50 million diagnosed epilepsy patients, of which 80% are from developing countries. Depression is the most common comorbidity of epilepsy. According to statistics, the incidence of comorbid or concomitant depression in epilepsy is 30-50%, which is significantly higher than the incidence of depressive disorders in the general population (5-17%). Studies have found that epileptic seizures in patients with comorbid or concomitant depression are not ideally controlled, and depressive disorders significantly aggravate the degree and duration of epileptic seizures in epilepsy patients, affecting the efficacy of epilepsy. It can be seen that depressive disorders are one of the important factors affecting epilepsy control. At present, epilepsy comorbid depression has gradually received attention, but prevention and treatment measures are limited. Many antidepressants claim in the drug instructions that they should be used with caution or prohibited for use in patients with epilepsy, such as paroxetine, fluoxetine, sertraline, escitalopram, venlafaxine, etc.

[0003] Therefore, in order to further study the pathophysiological mechanisms of co-morbidity of epilepsy and depression, it is very necessary to establish a more stable and reproducible mouse model of epilepsy and depression, and develop new intervention methods based on this, in order to provide new ideas and methods for the treatment of co-morbidity of epilepsy and depression. Summary of the invention

[0004] In order to solve the technical defects of unstable success rate of epilepsy-comorbid depression mouse model, uncertain modeling cycle and inconsistent modeling method, the present invention discloses a method for constructing and applying a chronic epilepsy-comorbid depression mouse model, which no longer requires external factors to stimulate mice to achieve depressive-like behavior. Instead, a clear modeling experiment cycle is established and behavioral test methods are used to screen the chronic epilepsy-comorbid depression mouse model, thereby reducing the interference and uncertainty of external stimuli.

[0005] The chronic epilepsy-comorbid depression mouse established by the present invention can be applied to research fields such as neurobiology, traditional Chinese and Western medicine research, and pharmacology. The model can more accurately simulate the pathophysiological mechanism of human chronic epilepsy-comorbid depression, and provides a powerful tool for studying the pathogenesis, drug screening, and treatment methods of the disease.

[0006] Specifically, the present invention adopts the following technical solutions:

[0007] The first aspect of the present invention provides a method for constructing a chronic epilepsy comorbid depression mouse model, which comprises: step 1, firstly, intraperitoneally injecting lithium chloride solution, intraperitoneally injecting atropine solution and intraperitoneally injecting pilocarpine solution into the experimental group mice in sequence to induce status epilepticus; recording the epileptic seizure behavior of the experimental group mice, and evaluating the epileptic seizure grade of the experimental group mice with reference to the Racine standard to obtain a lithium chloride-pilocarpine induced chronic epilepsy mouse model; step 2, combining a depressive-like behavior experiment to evaluate the degree of depressive-like behavior of mice to screen the lithium chloride-pilocarpine induced chronic epilepsy mouse model to determine the modeling standard and modeling time point of the chronic epilepsy comorbid depression mouse model, and obtaining chronic epilepsy comorbid depression mice and chronic epilepsy without depression mice; comparing the depressive-like behavior of the chronic epilepsy comorbid depression mice and the chronic epilepsy without depression mice to obtain a chronic epilepsy comorbid depression mouse model.

[0008] Furthermore, in step 1, the invention specifically comprises: intraperitoneally injecting the experimental group mice with a lithium chloride solution at a dose of 127 mg / kg, intraperitoneally injecting atropine solution at a dose of 1 mg / kg 20 hours later, and intraperitoneally injecting pilocarpine solution at a dose of 30 mg / kg 30 minutes later to induce status epilepticus.

[0009] Furthermore, the step 1 also includes: for mice that do not experience grade III to grade V epileptic seizures within 30 minutes after the first injection of pilocarpine, pilocarpine is injected again at a dose of 10 mg / kg every 10 minutes, and no more than 3 times; if no grade IV or above seizures occur after 3 consecutive injections, the mouse modeling is deemed to have failed and will not be included in subsequent further experiments; after the mouse has status epilepticus for 2 hours, diazepam is injected at a dose of 15 mg / kg to terminate the epileptic seizure, and the mouse is kept warm; from the mice that have successfully modeled in the acute phase to the next 72 hours, 5 mg / kg of diazepam is injected intraperitoneally once a day % glucose sodium chloride solution to supplement energy; wherein, the Racine scoring standard includes: grade 0, grade I, grade II, grade III, grade IV, grade V and grade VI; grade 0 represents no convulsive seizure; grade I represents facial clonus, including blinking, vibrissa movement, rhythmic chewing, etc.; grade II represents aggravation of grade I and facial clonus with rhythmic nodding; grade III represents aggravation of grade II and accompanied by forelimb myoclonus, but no hindlimb upright position; grade IV represents aggravation of grade III and bilateral forelimb clonus with standing; grade V represents continuous standing with falling; grade VI represents status epilepticus, tonic-clonic seizures with falling, jumping, and even death.

[0010] Furthermore, the step 1 specifically includes: the mice that enter status epilepticus enter the chronic phase of spontaneous epileptic seizures after a latent period of 72 hours to 2 weeks after modeling, and in the chronic phase, the presence or absence of spontaneous epileptic seizures, the number of seizures per day, the level of seizures, and the duration of each seizure are recorded for 12 hours every day for 4 weeks, and judged according to the Racine scoring standard to obtain a lithium chloride-pilocarpine-induced chronic epilepsy mouse model.

[0011] Furthermore, in step 2, the depressive-like behavioral experiment includes: a forced swimming test and a sucrose consumption test.

[0012] Furthermore, in step 2, depressive-like behavioral experiments are performed at two time points, 4 weeks after status epilepticus and 6 weeks after status epilepticus, respectively, for evaluation and comparison, and the modeling time point of the chronic epilepsy comorbid depression mouse model is determined to be 6 weeks after status epilepticus, and the screening inclusion criteria of the chronic epilepsy comorbid depression mouse model are determined, thereby obtaining chronic epilepsy comorbid depression mice and chronic epilepsy without depression mice; the screening inclusion criteria of the chronic epilepsy comorbid depression mouse model include: when evaluating the depressive-like behavioral experiments of sucrose consumption experiments and forced swimming experiments on chronic epilepsy mice 6 weeks after status epilepticus, in the sucrose consumption experiment, the sugar water preference rate needs to be between 48% and 65%; and in the forced swimming experiment, the immobility time is ≥105s.

[0013] Furthermore, the method for constructing the chronic epilepsy comorbid depression mouse model has an average modeling success rate of 30%.

[0014] The second aspect of the present invention provides a chronic epilepsy comorbid depression mouse model constructed by the above-mentioned method for constructing a chronic epilepsy comorbid depression mouse model.

[0015] The third aspect of the present invention provides the application of the above-mentioned chronic epilepsy comorbid depression mouse model in the fields of neurobiology, traditional Chinese and western medicine research, and pharmacology research.

[0016] The fourth aspect of the present invention provides the use of the above-mentioned chronic epilepsy comorbid depression mouse model in the field of studying the pathogenesis, drug screening and treatment methods of human chronic epilepsy comorbid depression.

[0017] Compared with the prior art, the present invention has at least the following beneficial effects:

[0018] 1. More accurately simulate the pathophysiological mechanism of human diseases: The present invention uses a method for constructing a chronic epilepsy-comorbid depression mouse model and uses behavioral methods to screen the degree of individual depressive-like behavior. There is no need to use external factors to stimulate mice to achieve depressive-like behavior, which can more accurately simulate the pathophysiological mechanism of human chronic epilepsy-comorbid depression.

[0019] 2. Easy operation and good repeatability: The preparation method of the present invention is easy to operate and only requires the establishment of a chronic epileptic mouse model and behavioral screening, and the experimental results are stable and have good repeatability.

[0020] 3. The model is stable and lasts for a long time: After the chronic epilepsy-comorbid depression mouse model of the present invention is established, the relationship between spontaneous seizures and depressive-like behaviors of mice can be continuously observed and studied. The depressive symptoms and behavioral manifestations can last for a long time, which is conducive to long-term research.

[0021] 4. Wide range of applications: The chronic epilepsy comorbid depression mouse model of the present invention can not only be used to study the pathogenesis of the disease, but also can be used for drug screening and development of treatment methods, and has broad application prospects.

[0022] 5. It helps to promote the research and treatment of epilepsy and depression: The chronic epilepsy and depression mouse model of the present invention provides a powerful tool for studying the interaction between the two diseases, and helps to promote the research and treatment of epilepsy and depression. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 Behavioral manifestations of lithium chloride-pilocarpine-induced epileptic seizures in mice in the examples of the present invention;

[0024] Figure 2 The lithium chloride-pilocarpine-induced chronic epilepsy mouse model with depressive-like behaviors at different stages and the chronic epilepsy comorbid depression mouse model in the embodiments of the present invention were established. DETAILED DESCRIPTION

[0025] As can be seen from the background technology, epilepsy is a common neurological disease, and depression is one of the most common comorbidities in patients with epilepsy. In order to study the relationship between epilepsy and depression and develop new treatments, researchers have established a variety of epilepsy comorbid depression mouse models. However, the current epilepsy comorbid depression mouse model still lacks a clear and quantitative method, and is mostly classified in groups rather than evaluated in individual form, so the modeling success rate is still a relatively vague number.

[0026] The inventors found that the most commonly used epilepsy comorbid depression mouse model currently uses chemical induction methods, such as intraperitoneal injection of kainic acid or lithium chloride-pilocarpine to induce epileptic seizures, and chronic unpredictable mild stress (CUMS) or social defeat stress (SDS) to induce depressive behavior. These models can simulate the depressive symptoms of co-morbidity in human epilepsy patients, such as lack of pleasure, loss of interest, sleep disorders, etc. Although the existing epilepsy comorbid depression mouse model has made some progress in studying the relationship between epilepsy and depression, there are still some defects: 1) Lack of complete representation of human diseases: It is difficult for mouse models to fully simulate all the symptoms and pathophysiological mechanisms of human epilepsy and depression. For example, the nervous system, behavior and social structure of mice are different from those of humans. 2) Repeatability and stability of the model: There may be differences in the epilepsy comorbid depression mouse model established using the same method in different laboratories, which may affect the reliability and repeatability of the research results. 3) Lack of understanding of the mechanism of comorbidity: Current research mainly focuses on the symptoms of epilepsy and depression, while the molecular and cellular mechanisms of comorbidity of epilepsy and depression are not yet well understood. 4) Lack of effective intervention methods: Due to insufficient understanding of the pathogenesis of comorbidity of epilepsy and depression, there is currently no specific treatment for this disease.

[0027] The present invention is further described in detail below through specific embodiments.

[0028] Example

[0029] Step 1: First, a lithium chloride-pilocarpine chronic epilepsy mouse model was constructed: the mice were fed for one week to adapt to the environment, and the model group mice were intraperitoneally injected with lithium chloride (127 mg / kg) solution; 20 hours later, scopolamine methyl nitrate (1 mg / kg, ip) was injected to reduce the peripheral nervous system reactions (blood tears, salivation, etc.) induced by pilocarpine; 30 minutes later, the acetylcholine (Ach) receptor agonist pilocarpine (30 mg / kg, ip) was injected to induce status epilepticus (SE), and the Racine scoring standard was used to evaluate the seizure level of mice. The latency of epileptic seizure refers to the time from pilocarpine injection to grade III epileptic seizure. SE refers to an epileptic seizure greater than grade IV that lasts for at least 30 minutes.

[0030] For mice that did not develop grade IV or higher seizures 30 minutes after the first injection of pilocarpine, pilocarpine (10 mg / kg, ip) was injected again every 10 minutes for up to 3 consecutive injections. If there was still no grade IV or higher seizure after 3 injections, the modeling of the mice was deemed to have failed and they were no longer included in subsequent experiments.

[0031] After the SE attack lasted for 2 hours, diazepam (15 mg / kg, ip) solution was injected to terminate the SE attack of mice, and an electric blanket was used to keep the mice warm. For the next 3 days, a sodium chloride solution containing 5% glucose was injected intraperitoneally once a day to replenish energy and increase the survival rate. During the experiment, the mice were monitored for seizures 24 hours a day using infrared video, and the time, number and duration of epileptic seizures were recorded. The Racine scoring criteria are shown in Table 1.

[0032] Table 1 Racine scoring criteria

[0033]

[0034] Step 2: Record the seizures of mice that entered status epilepticus (SE) in step 1 and entered the chronic phase (4 weeks) after the incubation period (2 weeks). The incubation period refers to the incubation period of 72 hours to 2 weeks after the acute modeling. During this process, mice are rarely seen to have epileptic seizures; the chronic phase refers to the chronic phase of 2 weeks to 2 months and beyond after the incubation period. Mice may have spontaneous epileptic seizures and can stop on their own, and the experimental mice become irritable, and the irritability gradually decreases over time (SE is more obvious after 2 weeks, such as mice running around, jumping up and down, turning in circles, drinking water constantly, increased defecation, increased activity, etc., and the above conditions are significantly reduced after 3 weeks of SE, mostly manifested as aggregation, reduced activity, sluggishness, etc.).

[0035] Forced swimming test (FST) and sucrose consumption test were performed at 4 weeks of chronic phase and 6 weeks of chronic phase, respectively, to assess the degree of depressive-like behavior in mice. Forced swimming is used to test adaptive strategies in an inescapable stress environment (to avoid the potential influence of acute seizures on FST, FST was only performed when there was no seizure at least 6 hours before the test) and is an indicator of despair and depressive-like behavior. Mice (without seizures for 6 hours before the test) were placed in a glass cylinder (60 cm high, 30 cm diameter, 45 cm deep, water temperature 22-23°C), with the water level occupying about 4 / 5, and each glass cylinder was separated by paper to avoid influence from each other. The swimming behavior was videotaped and analyzed by the researcher in a blinded manner for 6 minutes. For analysis, the first 2 minutes were excluded because the mice were mobile and familiar with the environment during that time, so any immobility during this time could not be attributed to despair. The cumulative immobility time in the last 4 minutes was recorded. An increase in the "immobility" time during swimming indicates the presence of despair. In this test, the "immobility period" is defined as the period when the animal stops struggling for ≥1s. The sucrose consumption experiment is to detect depression-related anhedonia: the sucrose consumption experiment is divided into 3 stages, and the amount of sucrose water consumed by each group of mice within 24 hours is observed and recorded. 1% disaccharide on the first day, no water and no food on the second day, and direct measurement of sugar and water on the third day - the direction of the water bottle is changed every 12 hours. Index: calculated by the percentage of 24h sucrose water / (sucrose water + regular water) intake. The calculation method is: sugar water consumption = amount of sucrose water before measurement - amount of sucrose water after measurement. The less sucrose water consumption, the greater the anhedonia. The present invention combines forced swimming and sucrose consumption experiments for analysis by designing a cluster analysis algorithm. The results show that the success rate of the chronic epilepsy comorbid depression mouse model 6 weeks after SE is about 38%, which is very close to the incidence of epilepsy comorbidity or concomitant depression in the population of 30-50%, so the establishment of this time point is clear and consistent.

[0036] 1.1 Construction of LiCl-Pilocarpine-Induced Chronic Epilepsy Mouse Model

[0037] The present invention successfully constructed a chronic epilepsy mouse model induced by lithium chloride-pilocarpine. First, the experimental group mice were intraperitoneally injected with lithium chloride solution (dose: 127 mg / kg), and 20 hours later, atropine solution (dose: 1 mg / kg) was intraperitoneally injected to reduce the side effects such as cholinergic reaction caused by pilocarpine. After 30 minutes, pilocarpine solution (dose: 30 mg / kg) was intraperitoneally injected to induce SE. The epileptic seizure grade of mice was evaluated according to the Racine standard, and the epileptic seizure behavior of mice was observed. Mice may show slow movement at first, followed by facial dullness, staring motionless, and stiff tail, followed by wet dog-like shaking, standing on both hind legs, stumbling or jumping up and down, head and facial muscle clonus, limb clonus and generalized tonic-clonic seizures, such as Figure 1 As shown. The above manifestations may appear separately or simultaneously. As the grade and degree of seizures increase, the experimental mice gradually enter a state of epilepsy and cannot be relieved on their own. For mice that do not reach grade III to grade V epileptic seizures within 30 minutes after the first injection of pilocarpine, pilocarpine (dose: 10 mg / kg) can be injected again every 10 minutes, and the maximum number of times cannot exceed 3 times. If there is still no grade IV or above seizure after 3 consecutive injections, the mouse model is deemed to have failed and will not be included in further experiments. After the mouse SE seizure lasts for 2 hours, diazepam (dose: 15 mg / kg) needs to be injected to terminate the epileptic seizure, and the modeled mice should be placed on an electric blanket to keep warm. From the time the acute phase model is successfully established to the next 72 hours, 5% glucose and sodium chloride solution is injected intraperitoneally once a day to replenish energy.

[0038] The SE mice entered the chronic stage of spontaneous epileptic seizures after a latent period of 72 hours to 2 weeks after modeling. During the chronic stage, the Xiaomi gimbal was used for real-time video monitoring for 12 hours a day (8:00-20:00) for a total of 1 month. The number of spontaneous seizures per day, the grade of the seizure, and the duration of each seizure were recorded and judged according to the Racine scoring standard.

[0039] Figure 1 A: After pilocarpine induction, mice began to develop grade III epileptic seizures: facial clonic twitches, rhythmic nodding, and forelimb clonus; Figure 1 B is a grade IV seizure that gradually develops after pilocarpine induction: facial twitching, rhythmic nodding, and bilateral forelimb clonus with standing; Figure 1 C is a grade V seizure induced by pilocarpine in mice: in addition to grade IV symptoms, continuous falls occur; Figure 1 D: The mice died after pilocarpine-induced grade VI seizure.

[0040] 1.2 Screening and establishment of lithium chloride-pilocarpine-induced chronic epilepsy and depression mouse model

[0041] In order to establish a chronic epilepsy with depression mouse model (Epilepsy with depression, EWD) and formulate a systematic evaluation standard, the present invention firstly established a chronic epilepsy mouse model by inducing lithium chloride-pilocarpine, and then performed depressive-like behavioral experiments (sucrose consumption experiment and forced swimming experiment) at different time points (i.e., 4 weeks after SE and 6 weeks after SE) for evaluation and comparison, and finally determined the modeling standard, screened and constructed a chronic epilepsy with depression mouse model ( Figure 2 A). The results showed that in the sucrose consumption experiment, the preference for sugar water in chronic epileptic mice was significantly reduced at 4 and 6 weeks after SE compared with the control group, and the difference was statistically significant ( Figure 2 B, C); Compared with the control group, in the forced swimming test, there was no significant difference in the immobility time of chronic epileptic mice 4 weeks after SE, while the immobility time of chronic epileptic mice 6 weeks after SE increased significantly, and compared with the chronic epileptic mice 4 weeks after SE, the immobility time of chronic epileptic mice 6 weeks after SE was longer, and the difference was statistically significant ( Figure 2 D, E). Therefore, the present invention determines that the modeling time point of the chronic epilepsy comorbid depression mouse model is 6 weeks after SE based on the results of the sucrose consumption experiment and the forced swimming experiment. Accordingly, the present invention has formulated a screening inclusion standard for a chronic epilepsy comorbid depression mouse model, which is as follows: Sucrose consumption experiment and forced swimming experiment depression-like behavior experiment evaluation are performed on chronic epilepsy mice 6 weeks after SE, and it must meet the following requirements at the same time: ① In the sucrose consumption experiment, the sugar water preference rate needs to be between 48% and 65%, indicating that the mouse is in a "hedonia state"; ② In the forced swimming experiment, the immobility time ≥105s indicates that the mouse is in a "desperate state" before it can be included in the chronic epilepsy comorbid depression mouse model.

[0042] Subsequently, the present invention screened out chronic epilepsy-comorbid depression mice and chronic epilepsy without depression mice (Epilepsy with non-depression, EWND) according to this standard, and compared the depressive-like behaviors. The results showed that in the sucrose consumption experiment, the preference for sugar water in chronic epilepsy-comorbid depression mice was significantly reduced compared with the control group and chronic epilepsy without depression mice, showing a "hedonia state", and the difference was statistically significant. Compared with the control group, there was no significant difference in the preference for sugar water in chronic epilepsy without depression mice ( Figure 2 F); In the forced swimming test, the immobility time of the chronic epilepsy comorbid depression mice was significantly longer than that of the control group and the chronic epilepsy without depression mice, showing a "desperate state", and the difference was statistically significant. In addition, there was no significant difference in the immobility time of the chronic epilepsy without depression mice compared with the control group ( Figure 2G). It is worth noting that the present invention has established a chronic epilepsy comorbid depression mouse model after ≥3 batches of screening, and the average modeling success rate is 30%, which is basically consistent with the clinical incidence of epilepsy comorbid depression ( Figure 2 H). The above indicates that the successful establishment of a lithium chloride-pilocarpine-induced chronic epilepsy comorbid depression mouse model in the present invention can be used for further follow-up experiments to explore the related mechanisms of chronic epilepsy comorbid depression.

[0043] Those skilled in the art will appreciate that the above embodiments are specific examples of the present invention, and in practical applications, various changes may be made to the embodiments in form and detail without departing from the spirit and scope of the present invention. Any person skilled in the art may make changes and modifications without departing from the spirit and scope of the present invention, and therefore the scope of protection of the present invention shall be subject to the scope defined in the claims.

Claims

1. A method for constructing a mouse model of chronic epilepsy and comorbid depression, characterized in that: The method includes: Step 1. First, the mice in the experimental group were intraperitoneally injected with lithium chloride solution, atropine solution, and pilocarpine solution in sequence to induce status epilepticus; the epileptic seizure behavior of the mice in the experimental group was recorded, and the epileptic seizure grade of the mice in the experimental group was evaluated according to the Racine standard to obtain a lithium chloride-pilocarpine-induced chronic epilepsy mouse model; Step 2: The lithium chloride-pilocarpine induced chronic epilepsy mouse model is screened by evaluating the degree of depressive-like behavior of mice in combination with a depressive-like behavior experiment to determine the modeling standard and modeling time point of the chronic epilepsy comorbid depression mouse model, and obtain chronic epilepsy comorbid depression mice and chronic epilepsy without depression mice; the depressive-like behavior of the chronic epilepsy comorbid depression mice and the chronic epilepsy without depression mice is compared to obtain the chronic epilepsy comorbid depression mouse model.

2. The method for constructing a mouse model of chronic epilepsy and comorbid depression according to claim 1, characterized in that: The step 1 specifically includes: intraperitoneally injecting a lithium chloride solution at a dose of 127 mg / kg into the experimental group mice, intraperitoneally injecting atropine solution at a dose of 1 mg / kg 20 hours later, and intraperitoneally injecting pilocarpine solution at a dose of 30 mg / kg 30 minutes later to induce status epilepticus.

3. The method for constructing a mouse model of chronic epilepsy and comorbid depression according to claim 2, characterized in that: The step 1 also includes: if the mouse does not develop grade III to grade V epileptic seizures within 30 minutes after the first injection of pilocarpine, pilocarpine is injected again at a dose of 10 mg / kg every 10 minutes, for no more than 3 times; if no grade IV or higher seizures occur after 3 consecutive injections, the mouse model is deemed to have failed and will not be included in subsequent further experiments; After the mice had status epilepticus for 2 hours, diazepam was injected at a dose of 15 mg / kg to terminate the epileptic seizure, and the mice were kept warm. From the acute phase of the successful modeling mice to the next 72 hours, 5% glucose and sodium chloride solution was injected intraperitoneally once a day to replenish energy. Among them, the Racine scoring standard includes: grade 0, grade I, grade II, grade III, grade IV, grade V and grade VI; grade 0 represents no convulsive seizure; grade I represents facial clonus, including blinking, vibrissa movement, rhythmic chewing, etc.; grade II represents aggravation of grade I and facial clonus with rhythmic nodding; grade III represents aggravation of grade II and accompanied by forelimb myoclonus, but no hindlimb upright position; grade IV represents aggravation of grade III and bilateral forelimb clonus with standing; grade V represents continuous standing with falls; grade VI represents status epilepticus, tonic-clonic seizures with falls, jumping, and even death.

4. The method for constructing a mouse model of chronic epilepsy and comorbid depression according to claim 3, characterized in that: The step 1 specifically includes: the mice that enter status epilepticus enter the chronic phase of spontaneous epileptic seizures after a latent period of 72 hours to 2 weeks after modeling, and in the chronic phase, the presence or absence of spontaneous epileptic seizures, the number of seizures per day, the severity of the seizures, and the duration of each seizure are recorded for 12 hours every day for 4 weeks, and the results are evaluated according to the Racine scoring standard to obtain a lithium chloride-pilocarpine-induced chronic epilepsy mouse model.

5. The method for constructing a mouse model of chronic epilepsy and comorbid depression according to claim 1, characterized in that: In step 2, the depressive-like behavioral experiment includes: a forced swimming test and a sucrose consumption test.

6. The method for constructing a mouse model of chronic epilepsy and comorbid depression according to claim 5, characterized in that: In the step 2, depressive-like behavioral experiments were performed at two time points, 4 weeks after status epilepticus and 6 weeks after status epilepticus, respectively, to evaluate and compare, determine that the modeling time point of the chronic epilepsy comorbid depression mouse model is 6 weeks after status epilepticus, and determine the screening inclusion criteria for the chronic epilepsy comorbid depression mouse model, thereby obtaining chronic epilepsy comorbid depression mice and chronic epilepsy without depression mice; The screening inclusion criteria for the chronic epilepsy comorbid depression mouse model include: when evaluating the depressive-like behavior experiments of chronic epilepsy mice 6 weeks after status epilepticus by sucrose consumption experiment and forced swimming experiment, the preference rate for sugar water needs to be between 48% and 65% in the sucrose consumption experiment; and the immobility time in the forced swimming experiment is ≥105s.

7. The method for constructing a mouse model of chronic epilepsy and comorbid depression according to claim 1, characterized in that: The modeling success rate of the method for constructing the chronic epilepsy comorbid depression mouse model is 30% on average.

8. The chronic epilepsy comorbid depression mouse model constructed by the method for constructing a chronic epilepsy comorbid depression mouse model according to any one of claims 1 to 7.

9. Application of the chronic epilepsy comorbid depression mouse model as claimed in claim 8 in the fields of neurobiology, traditional Chinese and western medicine research, and pharmacology research.

10. Use of the chronic epilepsy comorbid depression mouse model as claimed in claim 8 in the study of the pathogenesis, drug screening and treatment methods of human chronic epilepsy comorbid depression.

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