CN116492022A - Method for preparing cerebral hemorrhage animal model and method for screening cerebral hemorrhage treating medicine - Google Patents
Method for preparing cerebral hemorrhage animal model and method for screening cerebral hemorrhage treating medicine Download PDFInfo
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Abstract
The invention relates to the technical field of animal model preparation, and discloses a method for preparing a cerebral hemorrhage animal model and a method for screening a medicine for treating cerebral hemorrhage, which are characterized in that: it comprises the following steps: 1) Preparing a molding animal, and preprocessing the head; 2) Marking a target site after pretreatment, and fixing an animal on an objective table of the acoustic intervention instrument to enable the target site of the head of the animal to be aligned with an ultrasonic focusing point of the acoustic intervention instrument; 3) Intravenous infusion of a microbubble contrast agent; 4) And after the concentration of the microbubble contrast agent in the animal blood tends to be stable, continuing to inject the microbubble contrast agent, and simultaneously starting an acoustic intervention instrument to implement ultrasonic intervention. The invention realizes accurate positioning of focused ultrasound by an acoustic intervention instrument, and is a method for establishing a cerebral hemorrhage model with non-invasiveness, high reproducibility and high controllability by utilizing high-intensity focused ultrasound.
Description
Technical Field
The invention relates to the technical field of animal model preparation, in particular to a method for preparing a cerebral hemorrhage animal model and a method for screening medicines for treating cerebral hemorrhage.
Background
Cerebral hemorrhage (Intracerebral hemorrhage, ICH) is a common cerebrovascular disease in neurology, most of which are caused by vascular rupture of hypertension arteriosclerotic blood vessels, and has the characteristics of higher morbidity, disability rate and mortality rate, gradually rising morbidity and trend to younger morbidity age, and no mature diagnosis and treatment method exists clinically. Improving diagnosis and treatment level of cerebral hemorrhage, and preventing research of cerebral hemorrhage animal model. Therefore, the animal model of cerebral hemorrhage with high repeatability, good stability and high similarity to human can objectively and accurately reflect a series of changes of cerebral hemorrhage, and has important effects on researching pathogenesis, mechanism, pathophysiological change and the like of cerebral hemorrhage.
At present, common modeling methods for the rat cerebral hemorrhage animal model include an autologous blood injection method, a collagenase induction method and a microsphere inflation method.
Autologous blood infusion refers to taking rat femoral artery blood using an insulin needle and injecting it into the striatal region of the brain in a stereotactic manner, where hematoma forms. The autologous blood injection method has the disadvantages of complex operation, indeterminate form and size of hematoma, uncertain pressure of hematoma, easy flow of the spalling brain tissue into the ventricle, and hematoma not formed by cerebral vascular rupture.
The collagenase induction method is to use a microinjector to stereoscopically inject collagenase into the striatum region of the brain, and the blood brain barrier is composed of continuous capillary endothelium of the brain and close connection between cells, complete basal membrane, pericytes and glial membrane surrounded by astrocyte foot plates, and the collagenase can hydrolyze collagen on the intercellular matrix and the basal membrane of the blood vessel so as to induce cerebral tissue hemorrhage. The collagenase induction method has the defects that collagenase has obvious inflammatory reaction, damages brain tissues, blood vessels and blood brain barriers, and forms hematoma not in a real sense but diffusely seeps blood, so that the bleeding amount is difficult to control and the repeatability is relatively poor.
The balloon inflation method is a purely mechanical brain hemorrhage model making method, and has the defects that the model is different from true brain hemorrhage, has no blood component, cannot simulate the important functions of the components of blood per se in the formation and development of brain injury and cerebral edema after cerebral hemorrhage, and cannot observe the cytotoxicity reaction caused by cerebral hemorrhage, so that the brain hemorrhage model is rarely adopted in recent years.
The Chinese patent application CN 111528179A (invention name: method for establishing rat cerebral hemorrhage model by ultrasonic combined microbubbles) discloses a method for establishing rat cerebral hemorrhage model by ultrasonic combined microbubbles, which has the advantages of noninvasive, simple and convenient operation and easy popularization by acting on contrast agent through ultrasonic targeting and by means of the instant cavitation effect of microbubbles to generate destructive action on local blood vessels in target areas in the brain, so that the phenomenon of cerebral parenchyma internal hemorrhage occurs in rats, and the research requirements of spontaneous cerebral hemorrhage are met; however, when the inventor of the present application performs experiments using parameters of the ultrasonic device, it was found that parameters far greater than those disclosed in the above patent application are selected, and the parameters are compared with the following table 1, so that a cerebral hemorrhage model cannot be prepared, and a brain slice diagram after the experiments is shown in fig. 1, so that it can be deduced that a series of parameters in the above patent application may not be used for preparing a cerebral hemorrhage model.
TABLE 1 ultrasonic device parameter comparison
The spontaneous cerebral hemorrhage is cerebral vascular rupture hemorrhage, and the method can not simulate the pathological process and has the problems of high technical requirements, incapability of standardization, poor repeatability, poor stability and the like in the modeling process. Therefore, a method is needed to fundamentally solve the problems encountered in the current scheme of modeling cerebral hemorrhage.
Disclosure of Invention
In order to solve the defects in the prior art, the invention provides a method for preparing a cerebral hemorrhage animal model and a method for screening a cerebral hemorrhage treating drug, which can establish a cerebral hemorrhage model with non-invasiveness, high reproducibility and high controllability by utilizing high-intensity focused ultrasound and can be used for screening the cerebral hemorrhage treating drug.
In order to achieve the technical purpose, the invention adopts the following technical scheme:
a method of preparing an animal model of cerebral hemorrhage comprising the steps of:
1) Preparing a molding animal, and preprocessing the head;
2) Marking a target site after pretreatment, and fixing an animal on an objective table of the acoustic intervention instrument to enable the target site of the head of the animal to be aligned with an ultrasonic focusing point of the acoustic intervention instrument;
3) Intravenous infusion of a microbubble contrast agent;
4) And after the concentration of the microbubble contrast agent in the animal blood tends to be stable, continuing to inject the microbubble contrast agent, and simultaneously starting an acoustic intervention instrument to implement ultrasonic intervention.
Wherein the animals in step 1 comprise rats, mice, rabbits, pigs and monkeys.
The pretreatment method in the step 1) comprises the following steps: the anesthetized animals are removed from the head hair, the head skin of the head area is cut by the wale, the periosteum is cut and pushed to four sides, and the head skin is wiped by the disinfectant fluid.
The acoustic intervention instrument in the step 2) comprises an operation table, wherein the operation table comprises a frame, a transducer module assembly, an objective table and a laser collimator, and the frame comprises a supporting table, a vertical wall and an installation table from bottom to top in sequence; the transducer module assembly is movably arranged in the mounting table of the frame, and a transducer is embedded in the transducer module assembly; the objective table is positioned right below the transducer module assembly and is fixed on the mounting table through a telescopic rod; the laser collimator is fixed on the top wall of the mounting table through a connecting rod and is opposite to the center of the transducer module assembly.
Further, a water bag outer film is arranged below the transducer module assembly, and a through hole is formed in the center of the transducer module.
The specific method for aligning the target site of the head of the animal to the ultrasonic focusing point of the acoustic intervention instrument in the step 2) comprises the following steps: aiming the target site of the head of the animal at an imaging point of a laser light column emitted by a laser collimator on an objective table, and adjusting the height of the objective table to aim the target site at an ultrasonic focusing point of an ultrasonic module assembly.
Wherein, after fixing the animal in the step 2), water is injected into the water sac outer membrane, and an ultrasonic couplant is smeared between the skull of the animal and the water sac outer membrane, so that the water sac outer membrane is tightly connected with the skull.
Wherein, the injection speed of the microbubble contrast agent injected in the step 3) is 5ul/s, and the injection time is 50s.
Wherein, the parameters of the ultrasonic intervention in the step 4) are as follows: frequency: 0.1-10Mhz, sound pressure: 9-12 MPa, sound intensity: 5000-10000W/cm 2, duty ratio: 5-20%, and the action time is as follows: 30-120 s.
The invention also provides a method for screening the cerebral hemorrhage model prepared by the method, which comprises the following steps:
a. taking a drug to be detected;
b. applying the medicine to be detected to the prepared cerebral hemorrhage animal model;
c. evaluating and judging the efficacy of the medicine to be detected.
Compared with the prior art, the invention has the beneficial effects that:
in the method for preparing the cerebral hemorrhage animal model, the focused ultrasound is accurately positioned through the acoustic intervention instrument, and ultrasound with different intensities acts on the microbubble contrast agent to cause the microbubble contrast agent to be instantaneously cavitated, and the mechanical force generated by the instantaneous cavitation causes the blood vessel in the brain to be ruptured, thereby causing cerebral hemorrhage; the method is a method for establishing a non-invasive, high-reproducibility and high-controllability cerebral hemorrhage model by utilizing high-intensity focused ultrasound, and the effect of simulating micro-vascular rupture hemorrhage in brain parenchyma caused by primary non-trauma of human is superior to that of the existing several cerebral hemorrhage models, so that the method can be used for researching pathophysiological mechanism of cerebral hemorrhage and evaluating preclinical research of new drugs for treating cerebral hemorrhage related diseases.
Drawings
In order to more clearly illustrate the technical solutions of the embodiments of the present application, the drawings that are needed in the embodiments will be briefly described below, it being understood that the following drawings only illustrate some embodiments of the present application and therefore should not be considered limiting the scope, and that other related drawings may be obtained according to these drawings without inventive effort for a person skilled in the art.
FIG. 1 is a graph of brain slices of the model prepared according to the invention under the parameters of Table 1;
FIG. 2 is a schematic diagram of the architecture of an acoustic interventional instrument in an embodiment of the invention;
FIG. 3 is a schematic diagram of a disassembled configuration of a transducer module assembly of the acoustic interventional instrument of FIG. 2;
FIG. 4 is a schematic cross-sectional structural view of the transducer module assembly shown in FIG. 3;
FIG. 5 (A) is a representation of fluorescence imaging of each group of brain slices of a cerebral hemorrhage model prepared in accordance with the present invention;
FIG. 5 (B) shows fluorescence intensity values of each group of brain sheets of the cerebral hemorrhage model prepared in the present invention;
FIG. 5 (C) is a general histological observation of each group of brain sheets of the cerebral hemorrhage model prepared in the present invention;
FIG. 5 (D) is a TTC-stained photograph of each group of brain sheets of the cerebral hemorrhage model prepared by the present invention;
FIG. 5 (E) is a graph showing the necrotic volume of brain tissue for each group of brain hemorrhage models prepared according to the present invention;
FIG. 6 (A) is a diagram of general histological observation of brain sheets of ICH-A group and ICH-A+Edaravone group of cerebral hemorrhage model prepared in the present invention and a TTC staining picture;
FIG. 6 (B) is the brain tissue necrosis volumes of ICH-A and ICH-A+Edaravone groups, which were the brain hemorrhage models prepared according to the present invention.
Reference numerals: 1-operation table, 2-control table, 10-rack, 101-support table, 102-vertical wall, 103-installation table, 104-scale mark marking ruler, 11-transducer module assembly, 111-transducer module, 112-transducer cap, 113-water bag outer membrane, 1111-through hole, 1112-concave surface, 1121-connecting hole, 1122-convex ridge, 12-water bag bracket, 13-laser collimator, 14-objective table, 21-controller, 22-display screen, 3-cable.
Detailed Description
For the purposes of making the objects, technical solutions and advantages of the embodiments of the present application more clear, the technical solutions of the embodiments of the present application will be clearly and completely described below with reference to the drawings in the embodiments of the present application, and it is apparent that the described embodiments are some embodiments of the present application, but not all embodiments. The components of the embodiments of the present application, which are generally described and illustrated in the figures herein, may be arranged and designed in a wide variety of different configurations. Thus, the following detailed description of the embodiments of the present application, as provided in the accompanying drawings, is not intended to limit the scope of the application, as claimed, but is merely representative of selected embodiments of the application. All other embodiments, which can be made by one of ordinary skill in the art based on the embodiments herein without making any inventive effort, are intended to be within the scope of the present application.
The invention relates to an animal experiment acoustic intervention instrument used in a method for preparing a cerebral hemorrhage animal model, which is shown in figures 2,3 and 4 and comprises an operation table 1, wherein the operation table 1 comprises a frame 10, a transducer module assembly 11, an objective table 14 and a laser collimator 13, the frame 10 comprises a supporting table 101, a vertical wall 102 and an installation table 103 from bottom to top in sequence, the transducer module assembly 11 is movably installed in the installation table 103 of the frame, and a transducer is embedded in the transducer module assembly 11; the objective table 14 is located right below the transducer module assembly 11, and preferably, the objective table 14 is fixed on the mounting table 101 through a telescopic rod and can move up and down in height; the laser collimator 13 is fixed on the top wall of the mounting table 103 through a connecting rod, and the laser collimator 13 faces the center of the transducer module assembly 11; the laser beam emitted by the laser collimator forms a light spot on the upper surface of the objective table, and the height of the objective table is adjusted to enable the light spot to be located at the ultrasonic focusing point of the transducer module assembly, so that the situations of focus point deviation and inaccurate depth position can be avoided.
The transducer is mainly composed of piezoelectric ceramics, and generates vibration through the driving of an electric signal to form an ultrasonic signal so as to emit a specific ultrasonic signal; since the ultrasonic signal is extremely attenuated in the air, other media are needed to conduct the ultrasonic signal, here we select water as the media, a water bag outer membrane 113 is arranged below the transducer module assembly 11, a through hole 1111 is formed in the center of the transducer module assembly 11, water is injected into the water bag outer membrane 113 through the through hole 1111 in the center of the transducer module assembly 11, and the diameter of the through hole 1111 is larger than 5 mm. Preferably, the transducer module assembly 11 includes a transducer module 111 and a transducer cap 112, the transducer module 111 is mounted in the transducer cap 112, and the transducer cap 112 is slidably connected to the mounting table 103. Preferably, the transducer cap 112 is square, two opposite side walls of the transducer cap 112 are provided with protruding ridges 1122, a through square notch is arranged in the mounting platform 103 of the stand 10, two opposite side walls of the notch are provided with sliding grooves, and the protruding ridges 1122 of the transducer cap are slidably connected in the sliding grooves of the mounting platform 103. In one embodiment, the transducer module 111 is cylindrical, the inner bottom surface of the transducer module 111 is a concave surface 1112, the transducer is embedded in the transducer module 111, the transducer is adapted to the concave surface of the transducer module 111, and the concave surface can enable the transducer to be focused on a point better; the outer surface of the transducer module 111 is provided with external threads, the center of the transducer cap 112 is provided with a circular connecting hole 1121, the connecting hole 1121 is internally provided with internal threads matched with the external threads of the transducer module 111, and the transducer module 111 is in threaded connection with the transducer cap 112; the water bag outer membrane 113 is tightly pressed on the abutting surface of the transducer module 111 and the transducer cap 112, after the experiment is finished, the transducer module 111 is rotated to separate the transducer module 111 from the transducer cap 112, the water bag outer membrane 113 can be separated along with the separation, and the replacement and installation of the water bag outer membrane 113 are simple and quick; the sliding connection pluggable transducer module assembly can effectively solve the problems of water injection of the water bag, replacement of transducers with different standards and the like, and simultaneously, a simpler solution is provided for replacement of the loss of the transducers.
The operation table is further provided with a water bag bracket 12, the water bag bracket 12 is fixed on the vertical wall 102 and is positioned right below the transducer module assembly 11, and the water bag bracket 12 is used for fixing the water bag outer film 113 above the objective table 14, so that the water bag outer film 113 is suspended right below the transducer module assembly 11.
In one embodiment, the telescopic rod is connected by damping, the telescopic length can be manually adjusted, and in another preferred embodiment, the telescopic rod is an electric telescopic rod and is connected with a motor, and the telescopic length of the telescopic rod is controlled by the motor. Preferably, be provided with scale mark marking ruler 104 on the vertical wall 102 of frame, under the assistance of scale mark marking ruler, but the objective table of reciprocates has solved the different problem of different transducer module focus degree of depth, finely tunes the objective table simultaneously, also can make the focus carry out the fine setting of upper and lower direction, has guaranteed the equipment foundation for the animal experiment on the different degree of depth.
The acoustic intervention instrument for animal experiments further comprises a control console 2, wherein the control console 2 consists of a controller 21 and a display screen 22, the control console 2 is connected with the operation console 1 through a cable 3, and the cable 3 is used for transmitting signals; the controller 21 is used for setting the transmitted ultrasonic waveform and ultrasonic frequency and setting a certain duty cycle and period; the display 22 is used to display ultrasound in real time, giving the user a visual display to help the user make adjustments better. In addition to adjusting the ultrasonic properties, the controller 21 is also used to control the elevation of the stage 14 at the end of the console 1, and by setting the controller, the stage can be fixed on a set plane, so as to perform accurate positioning and focusing of the ultrasonic waves.
The method for preparing the cerebral hemorrhage animal model takes a rat as an example, and of course, the method is not limited to the rat, and can also be an experimental animal such as a mouse, a rabbit, a pig, a monkey and the like, and the method comprises the following steps:
1) SPF-class male SD rats are prepared, the weight of the SD rats is 230-280g, 4 groups are arranged, the 1 st group is a non-injected microbubble group (w/o microbubbles group), the 2 nd group is a cerebral hemorrhage model-A group (ICH-A group), the 3 rd group is a cerebral hemorrhage model-B group (ICH-B group), and the 4 th group is a cerebral hemorrhage model-A+edaravone group (ICH-A+Edaravone group); pretreating the head of a rat, removing cranium top hair after the rat is anesthetized, cutting the scalp of the cranium top area by wales, cutting periosteum and pushing the periosteum to four sides, and wiping the skull by using a hydrogen peroxide cotton swab.
2) Referring to the rat brain in-vitro positioning map, marking a target site (3 mm on the right side of the bregma, 0.2mm in front) by taking the bregma as an origin; fixing the rat on a stage of the acoustic intervention instrument, and aligning a target site of the head of the rat with an ultrasonic focusing point of the acoustic intervention instrument;
the specific method for aligning the target site of the rat part with the ultrasonic focusing point of the acoustic intervention instrument comprises the following steps: aligning a target site of the head of the rat with an imaging point of a laser light column emitted by a laser collimator on an objective table, and adjusting the height of the objective table to align the target site with an ultrasonic focusing point of an ultrasonic module assembly;
after the rat is fixed, water is injected into the water sac outer membrane 113 through the central through hole of the ultrasonic module assembly 11, the water sac outer membrane 113 after water injection is abutted against the skull of the rat, and an ultrasonic couplant is smeared between the water sac outer membrane and the skull, so that the water sac outer membrane is tightly connected with the skull.
3) The microbubble contrast agent was injected intravenously from the tail of the rat with the aid of a microinjection pump, except for the group of microbubbles, which in this example were sulfur hexafluoride microbubbles.
4) Microbubble contrast agent (concentration: 8 ul/ml) is 5ul/s, the total injection is 80s, 50s is injected firstly to enable the microbubble concentration in the blood to be stable, the microbubble concentration in the blood is about 0.8-1.0 ml/kg, an acoustic intervention instrument is started in the last 30s, ultrasonic intervention is implemented, ultrasonic intervention parameters of each group are shown in Table 2, so that the microbubble contrast agent reaches an instantaneous cavitation threshold value, and the mechanical force generated by the injection enables blood vessels in the brain to be ruptured, thereby leading to cerebral hemorrhage; immediately after the end of the ultrasonic intervention, the group 4 rats were injected with edaravone at a dose of 6mg/kg, a volume of 5ml/kg and a concentration of 1.2mg/ml.
5) Post-processing, observing, recording and analyzing data are carried out on the rat; specific:
5.1 30min after the ultrasonic intervention, 10% sodium fluorescein (NaFL) is injected into the abdominal cavity, and the administration volume is 4ml/kg;
5.2 24h after ultrasonic intervention, dissecting and removing brain tissue, and observing the general histology;
5.3 Cutting brain tissue into 6 pieces of coronal brain slices with the thickness of about 2mm, observing bleeding sites, and using a living animal imager (. About.)Lumia series iii)) to detect blood brain barrier (Blood brain barrier, BBB) permeability, with Total Radiant Efficiency (TRE) representing fluorescence intensity;
5.4 Brain pieces were then stained with 2,3, 5-triphenyltetrazolium chloride (TTC), and the necrotic foci size of each group of brain tissue was counted and analyzed.
TABLE 2 ultrasonic device parameters for preparing a rat cerebral hemorrhage model
As can be seen from fig. 5 (a) to 5 (E), the fluorescence intensity of brain sheets of the ICH-a group is significantly higher than that of (w/o) Microbubbles group (p=0.0010), which indicates that, under the same ultrasonic equipment parameters, the injection of Microbubbles into rats can increase BBB permeability and can rupture the walls of cerebral blood vessels by using mechanical force generated at the moment of rupture of the Microbubbles to cause cerebral hemorrhage, but the ICH-a group increases cavitation effect by the Microbubbles and also increases thermal effect, resulting in a brain tissue necrosis factor significantly greater than that of (w/o) Microbubbles (p=0.0169). To reduce necrotic lesions, further optimization of device parameters, the BBB permeability of the ICH-B group was found to increase to a lesser extent than that of the ICH-a group (p=0.0282), which had macroscopic lesions, and brain tissue necrosis volumes significantly lower than that of the ICH-a group (p=0.0027), but lesions were smaller than that of the ICH-a group. In summary, the ultrasonic equipment parameters of ICH-A group and ICH-B group can prepare the cerebral hemorrhage models of rats with different hemorrhagic foci sizes.
Observing the general histology of ICH-A and ICH-A+Edaravone brain chips, see FIG. 6 (A), with macroscopic lesions, and with lesions larger than ICH-A+Edaravone; observing TTC staining patterns of two groups of brain slices, wherein the red part is normal tissue, and the white part is necrotic foci; analysis of the necrotic volume size with Image J software, as shown in fig. 6 (B), the necrotic volume of the ICH-a+edaravone group was significantly reduced (p= 0.0208) compared to the ICH-a group, indicating that Edaravone has therapeutic effect on the model, and thus it can be deduced that the model can be used for preclinical study of new drugs for treating cerebral hemorrhage-related diseases.
The ICH-A model is suitable for researching pathogenesis, pathological damage and symptom manifestations of the middle and severe cerebral hemorrhage of rats. The pathophysiological process of spontaneous cerebral hemorrhage and the morphological structure change characteristics of brain tissue provide basis for clinical treatment; can be used for researching inflammation and edema after cerebral hemorrhage; research on pathogenesis of acute cerebrovascular disease induced multiple organ dysfunction syndrome; preclinical studies of new drugs for the treatment of diseases associated with moderate and severe cerebral hemorrhage.
The ICH-B model is suitable for researching the pathogenesis of the mild cerebral hemorrhage of rats, pathological damage and symptom manifestation. The pathophysiological process of spontaneous cerebral hemorrhage and the morphological structure change characteristics of brain tissue provide basis for clinical treatment; can be used for researching inflammation and edema after cerebral hemorrhage; research on pathogenesis of acute cerebrovascular disease induced multiple organ dysfunction syndrome; preclinical studies of new drugs for the treatment of diseases associated with mild cerebral hemorrhage.
The invention utilizes high-intensity focused ultrasound to build a non-invasive, high-reproducibility and high-controllability cerebral hemorrhage model, and the effect of the model for simulating micro-vascular rupture hemorrhage in brain parenchyma caused by primary non-trauma of human is superior to that of the existing several cerebral hemorrhage models, so that the model can be used for researching pathophysiological mechanism of cerebral hemorrhage and evaluating preclinical research of new drugs for treating cerebral hemorrhage related diseases.
Of course, the present invention is capable of other various embodiments and its several details are capable of modification and variation in light of the present invention by one skilled in the art without departing from the spirit and scope of the invention as defined in the appended claims.
Claims (10)
1. A method for preparing an animal model of cerebral hemorrhage, comprising: it comprises the following steps:
1) Preparing a molding animal, and preprocessing the head;
2) Marking a target site after pretreatment, and fixing an animal on an objective table of the acoustic intervention instrument to enable the target site of the head of the animal to be aligned with an ultrasonic focusing point of the acoustic intervention instrument;
3) Intravenous infusion of a microbubble contrast agent;
4) And after the concentration of the microbubble contrast agent in the animal blood tends to be stable, continuing to inject the microbubble contrast agent, and simultaneously starting an acoustic intervention instrument to implement ultrasonic intervention.
2. The method of preparing an animal model of cerebral hemorrhage according to claim 1, wherein: the animals in step 1 include rats, mice, rabbits, pigs and monkeys.
3. The method of preparing an animal model of cerebral hemorrhage according to claim 1, wherein: the pretreatment method of the step 1) comprises the following steps: the anesthetized animals are removed from the head hair, the head skin of the head area is cut by the wale, the periosteum is cut and pushed to four sides, and the head skin is wiped by the disinfectant fluid.
4. The method of preparing an animal model of cerebral hemorrhage according to claim 1, wherein: step 2) the acoustic intervention instrument comprises an operation table (1), wherein the operation table (1) comprises a frame (10), a transducer module assembly (11), an objective table (14) and a laser collimator (13), and the frame (10) comprises a supporting table (101), a vertical wall (102) and an installation table (103) from bottom to top in sequence; the transducer module assembly (11) is movably arranged in the mounting table (103) of the frame, and the transducer is embedded in the transducer module assembly (11); the objective table (14) is positioned right below the transducer module assembly (11) and is fixed on the mounting table (101) through a telescopic rod; the laser collimator (13) is fixed on the top wall of the mounting table (103) through a connecting rod, and the laser collimator (13) is opposite to the center of the transducer module assembly (11).
5. The method of preparing an animal model of cerebral hemorrhage according to claim 4, wherein: a water bag outer film (113) is arranged below the transducer module assembly (11), and a through hole (1111) is formed in the center of the transducer module assembly (11).
6. The method of preparing an animal model of cerebral hemorrhage according to claim 5, wherein: the specific method for aligning the target site of the head of the animal with the ultrasonic focusing point of the acoustic intervention instrument in the step 2) is as follows: aiming the target site of the head of the animal at an imaging point of a laser light column emitted by a laser collimator on an objective table, and adjusting the height of the objective table to aim the target site at an ultrasonic focusing point of an ultrasonic module assembly.
7. The method of preparing an animal model of cerebral hemorrhage according to claim 6, wherein: after the animal is fixed in the step 2), water is injected into the water sac outer membrane, and an ultrasonic couplant is smeared between the animal skull and the water sac outer membrane, so that the water sac outer membrane is tightly connected with the skull.
8. The method of preparing an animal model of cerebral hemorrhage according to claim 1, wherein: the injection speed of the microbubble contrast agent injected in the step 3) is 5ul/s, and the injection time is 50s.
9. The method of preparing an animal model of cerebral hemorrhage according to claim 1, wherein: the parameters of the ultrasonic intervention in the step 4) are as follows: frequency: 0.1-10Mhz, sound pressure: 9-12 MPa, sound intensity: 5000-10000W/cm 2, duty ratio: 5-20%, and the action time is as follows: 30-120 s.
10. A method for screening a medicament for treating cerebral hemorrhage, which is characterized by comprising the following steps: the method comprises the following steps:
a. taking a drug to be detected;
b. applying a drug to be tested to a cerebral hemorrhage animal model prepared according to the method for preparing a cerebral hemorrhage animal model of any one of claims 1 to 9;
c. evaluating and judging the efficacy of the medicine to be detected.
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