Method for establishing spontaneous low-cranial-pressure experimental animal model

Abstract

The invention discloses a spontaneous low-cranial-pressure experimental animal model building method, which comprises the following steps: building a dura mater crevasse by using a balloon on a certain spine segment of an experimental animal under the guidance of X rays so as to build a spontaneous low-cranial-pressure model, so that spontaneous low-cranial-pressure spinal cord and cerebrospinal fluid leakage is effectively simulated; the important clinical value is provided for the research and treatment of the disease.

Description

Technical Field

[0001] The invention relates to the medical field, and in particular to a method for establishing an experimental animal model of spontaneous intracranial hypotension. Background Art

[0002] Spontaneous intracranial hypotension (SIH) is a disabling disease caused by the loss of cerebrospinal fluid (CSF) in the spinal cord. The clinical manifestations are usually orthostatic headache and other associated symptoms. During the course of the disease, due to the occurrence of dural rupture and CSF leakage, the CSF pressure and volume decrease, which increases the compliance of the dural sac and the negative pressure in the cranial cavity. This is followed by dilation of the intracranial venous system, dilation of small blood vessels in the dura mater, enlargement of the pituitary gland, subdural effusion or hematoma, loss of perilymph in the inner ear, downward movement of brain tissue, and traction of pain-sensitive structures such as meningeal blood vessels and nerves, resulting in headache. This series of factors will reduce the patient's health-related quality of life, such as forcing the patient to stay in bed for a long time, and the occurrence of complications such as subdural hematoma, cerebral venous thrombosis, and coma.

[0003] Studies have shown that spontaneous intracranial hypotension can occur in all age groups, but is more common in young and middle-aged women. In spinal cord imaging, spontaneous intracranial hypotension spinal cerebrospinal fluid leakage is most common in the cervical thoracic segment, especially at the cervical thoracic junction. Characteristic abnormalities of cranial magnetic resonance imaging (MRI) include: subdural effusion / hematoma, dural enhancement, venous system dilation, pituitary congestion and enlargement, and brain ptosis.

[0004] Large mammals, including dogs, sheep, pigs, monkeys, and gorillas, are commonly used experimental animals. They have similar anatomical characteristics, etiology, and disease progression characteristics to humans, and have broad potential for application in the biomedical field. Since the main cause of spontaneous intracranial hypotension is dural rupture, rodents are small in size and have narrow spinal canals, making it difficult to damage the dura mater, so large mammals are needed to establish models.

[0005] Since there is currently no method for establishing an experimental animal model of spontaneous intracranial hypotension, developing a method for establishing an experimental animal model of spontaneous intracranial hypotension has important clinical value for the study and treatment of the disease. Summary of the invention

[0006] The technical problem to be solved by the present invention is to provide a method for establishing an experimental animal model of spontaneous intracranial hypotension. By using the "balloon method" to establish a dural rupture in the thoracic vertebral segment of the experimental animal to construct a spontaneous intracranial hypotension model, the cerebrospinal fluid leakage of spontaneous intracranial hypotension is effectively simulated, which provides important clinical value for the research and treatment of the disease.

[0007] The present invention is implemented in this way. The present invention provides a method for establishing an experimental animal model of spontaneous intracranial hypotension, characterized in that the method for establishing the experimental animal model mainly comprises the following steps:

[0008] Step S1: At the beginning of the experiment, select experimental animals in good health and prepare for surgery;

[0009] Step S2: The experimental animals were fasted and deprived of water before the operation;

[0010] Step S3: anesthetizing and preparing the skin of the experimental animal;

[0011] Step S4: performing spinal puncture: under X-ray guidance, a thoracic puncture is performed in the intervertebral space between the sixth and seventh thoracic vertebrae of the experimental animal, and the puncture needle enters the subarachnoid space, and cerebrospinal fluid can generally be seen flowing out;

[0012] Step S5: Using the balloon method to create a dural leak to construct a spontaneous intracranial hypotension model;

[0013] Step S6: performing a postoperative examination on the experimental animal;

[0014] Step S7: After the experiment is over, the experimental animals are humanely killed;

[0015] The step S5 comprises the following steps:

[0016] Step S51: inserting a guide wire into the subarachnoid space through the puncture needle, and keeping the position of the guide wire unchanged;

[0017] Step S52: inserting a balloon under the guidance of the guidewire: inserting the balloon along the guidewire into the subarachnoid space;

[0018] Step S53: After the balloon is pressurized and inflated, the balloon is pulled out in an inflated state, and the dura mater can be ruptured when the balloon is pulled out;

[0019] The step S6 comprises the following steps:

[0020] S61: 1 hour after the operation, the cerebrospinal fluid pressure of the experimental animal is measured;

[0021] S62: One week after the operation, the experimental animals were subjected to CT angiography to observe whether the thoracic vertebral segment where the operation was performed had a leak;

[0022] S63: Two weeks after the operation, the experimental animals were subjected to enhanced MRI scanning to observe whether there were dura mater enhancement, brain sagging and characteristic manifestations of spontaneous intracranial hypotension.

[0023] Furthermore, the experimental animals are large mammals with anatomical features and the causes and disease progression characteristics of spontaneous intracranial hypotension similar to those of humans.

[0024] Furthermore, the diameter of the balloon is a different diameter selected according to experimental requirements.

[0025] Furthermore, the puncture needle is an epidural puncture needle.

[0026] Furthermore, the puncture site includes the entire spinal segment including the cervical vertebra, thoracic vertebra, and lumbar vertebra.

[0027] Compared with the prior art, the beneficial effect of the present invention lies in: the present invention provides a method for establishing an experimental animal model of spontaneous intracranial hypotension, which constructs a spontaneous intracranial hypotension model by using an X-ray-guided balloon method to establish a dural rupture in a certain spinal segment of the experimental animal, thereby effectively simulating spontaneous intracranial hypotension cerebrospinal fluid leakage, and providing important clinical value for the research and treatment of the disease. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the drawings required for use in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0029] Figure 1 It is a schematic diagram of the arachnoid membrane, dura mater, subarachnoid space, and subdural space involved in a method for establishing an experimental animal model of spontaneous intracranial hypotension provided in an embodiment of the present invention.

[0030] Figure 2 The present invention provides a flowchart of a method for establishing an experimental animal model of spontaneous intracranial hypotension.

[0031] Figure 3 It is a schematic diagram of an experimental animal preparation surgery for a method for establishing an experimental animal model of spontaneous intracranial hypotension provided by an embodiment of the present invention.

[0032] Figure 4 It is a schematic diagram of an experimental animal operation in a method for establishing an experimental animal model of spontaneous intracranial hypotension provided by an embodiment of the present invention.

[0033] Figure 5 It is a schematic diagram of measuring the cerebrospinal fluid pressure of experimental animals in a method for establishing an experimental animal model of spontaneous intracranial hypotension provided by an embodiment of the present invention.

[0034] Figure 6This is one of the schematic diagrams of a puncture needle and a balloon used in a method for establishing an experimental animal model of spontaneous intracranial hypotension provided in an embodiment of the present invention.

[0035] Figure 7 This is the second schematic diagram of a puncture needle and a balloon used in a method for establishing an experimental animal model of spontaneous intracranial hypotension provided in an embodiment of the present invention.

[0036] Figure 8 It is a schematic diagram of a method for establishing an experimental animal model of spontaneous intracranial hypotension provided by an embodiment of the present invention, in which a balloon is inserted into the subarachnoid space along a guide wire.

[0037] Fig. 9 It is a schematic diagram of a puncture needle penetrating into the subarachnoid space in a method for establishing an experimental animal model of spontaneous intracranial hypotension provided by an embodiment of the present invention.

[0038] Fig.10 It is a schematic diagram of a puncture needle insertion guide wire in a method for establishing an experimental animal model of spontaneous intracranial hypotension provided by an embodiment of the present invention.

[0039] Fig.11 It is a schematic diagram of the balloon position of a method for establishing an experimental animal model of spontaneous intracranial hypotension provided by an embodiment of the present invention.

[0040] Fig.12 It is a schematic diagram of the preparation of CT myelography of an experimental animal in a method for establishing an experimental animal model of spontaneous intracranial hypotension provided by an embodiment of the present invention.

[0041] Fig.13 This is one of the experimental animal CT myelography results of a method for establishing an experimental animal model of spontaneous intracranial hypotension provided by an embodiment of the present invention.

[0042] Fig.14 This is the second intention of the experimental animal CT myelography result of a method for establishing an experimental animal model of spontaneous intracranial hypotension provided by an embodiment of the present invention. DETAILED DESCRIPTION

[0043] In order to make the purpose, technical solution and advantages of the present invention more clearly understood, the present invention is further described in detail below in conjunction with the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not intended to limit the present invention.

[0044] The same or similar numbers in the drawings of this embodiment correspond to the same or similar parts; in the description of the present invention, it should be noted that when an element is referred to as "fixed to" another element, it can be directly on the other element or there can also be a central element. When an element is considered to be "connected" to another element, it can be directly connected to the other element or there can be a central element at the same time. It should be understood that if there are terms such as "upper", "lower", "left", "right", etc. indicating an orientation or positional relationship based on the orientation or positional relationship shown in the drawings, it is only for the convenience of describing the present invention and simplifying the description, rather than indicating or implying that the device or element referred to must have a specific orientation, be constructed and operated in a specific orientation. Therefore, the terms describing the positional relationship in the drawings are only used for exemplary descriptions and cannot be understood as limitations on this patent. For ordinary technicians in this field, the specific meanings of the above terms can be understood according to specific circumstances.

[0045] The technical solution of the present invention is described in detail below in conjunction with the accompanying drawings and specific embodiments.

[0046] like Figure 1 As shown in the figure, the spinal cord membranes are the pia mater, arachnoid mater, and dura mater from the inside to the outside. The space between the arachnoid mater and the pia mater is the subarachnoid space (same as Figure 1 The subdural space is the space between the dura mater and the arachnoid membrane.

[0047] Reference Figure 2 The method for establishing an experimental animal model of spontaneous intracranial hypotension provided by the present invention mainly comprises the following steps:

[0048] Step S1: At the beginning of the experiment, select experimental animals in good health and prepare for surgery;

[0049] Step S2: The experimental animals were fasted and deprived of water before the operation;

[0050] Step S3: anesthetizing and skin preparation of experimental animals;

[0051] Step S4: performing spinal puncture: under X-ray guidance, a thoracic puncture is performed in the intervertebral space between the sixth and seventh thoracic vertebrae of the experimental animal, and the puncture needle enters the subarachnoid space, and cerebrospinal fluid can generally be seen flowing out;

[0052] Step S5: Using the balloon method to create a dural leak to construct a spontaneous intracranial hypotension model;

[0053] Step S6: Performing postoperative examination on the experimental animals;

[0054] Step S7: After the experiment is over, the experimental animals are humanely killed;

[0055] The step S5 comprises the following steps:

[0056] Step S51: inserting a guide wire into the subarachnoid space through the puncture needle, and keeping the position of the guide wire unchanged;

[0057] Step S52: inserting a balloon under the guidance of the guidewire: inserting the balloon along the guidewire into the subarachnoid space;

[0058] Step S53: After the balloon is pressurized and inflated, the balloon is pulled out in an inflated state, and the dura mater can be ruptured when the balloon is pulled out;

[0059] The step S6 comprises the following steps:

[0060] S61: One hour after the surgery, the cerebrospinal fluid pressure of the experimental animals was measured;

[0061] S62: One week after the surgery, the experimental animals were subjected to CT angiography to observe whether there was a leak in the thoracic vertebral segment where the surgery was performed;

[0062] S63: Two weeks after the operation, the experimental animals were subjected to enhanced MRI scans to observe whether there was dura mater enhancement and brain sagging with characteristic manifestations of spontaneous intracranial hypotension.

[0063] Preferably, the experimental animals are large mammals with anatomical features and the etiology and disease progression characteristics of spontaneous intracranial hypotension similar to those of humans.

[0064] Preferably, the diameter of the balloon is a different diameter selected according to experimental requirements.

[0065] Preferably, the puncture needle is an epidural puncture needle.

[0066] Preferably, the puncture site includes the entire spinal segment including the cervical vertebrae, thoracic vertebrae, and lumbar vertebrae.

[0067] The above technical solution provides a method for establishing an experimental animal model of spontaneous intracranial hypotension. By using a balloon to create a dura mater rupture in a certain spinal segment of the experimental animal under X-ray guidance to construct a spontaneous intracranial hypotension model, the spontaneous intracranial hypotension spinal cord cerebrospinal fluid leakage is effectively simulated, which provides important clinical value for the research and treatment of the disease.

[0068] Specifically, since beagles are commonly used experimental animals and have been widely used in the construction of cardiovascular and endocrine disease models, a female beagle weighing about 15 kg and in good health was selected as the experimental animal in this embodiment. Before the operation, the animal was fasted and deprived of water. Anesthesia was induced by intramuscular injection of Shutai 50 (5 mg / kg) and Sumenxin II (0.04 mL / kg). After successful induction, the trachea was intubated and the experimental animal was fixed on the operating table in a side-lying position. Respiratory anesthesia was maintained with isoflurane (dose: 1.5%), and the animal's respiration, electrocardiogram and other indicators were monitored. The hair on the back of the waist was cut off, the skin was disinfected with 5% iodine, the surgical area was covered with a sheet, and the puncture point was located between the sixth and seventh lumbar vertebrae, such as Figure 3 As shown;

[0069] The specifications of the selected lumbar puncture needle are: No. 9 lumbar puncture needle, 10 cm long, 0.9 mm in diameter;

[0070] The specifications of the selected hard anesthesia needles are: AN-E 1.6×80, total length 10cm, outer diameter 1.6mm, inner diameter 1.2mm;

[0071] The intervertebral space between the sixth and seventh thoracic vertebrae was selected for subarachnoid puncture, such as Figure 3 As shown, there is a breakthrough feeling during puncture, and clear cerebrospinal fluid can be seen flowing out after the puncture needle is pulled out;

[0072] like Figure 4 As shown, after the puncture needle is pulled out, the pressure measuring tube is connected to measure the cerebrospinal fluid pressure of the dog;

[0073] The needle and balloon Figure 5 As shown, the blue arrow indicates the epidural puncture needle, and the red arrow indicates the balloon in the uninflated state;

[0074] The balloon is inflated. Figure 6 As shown, the white arrow indicates the balloon in the inflated state, and the diameter of the inflated balloon selected in this embodiment is 3.5 mm;

[0075] The balloon is placed along the guide wire into the subarachnoid space. Figure 7 As shown;

[0076] Schematic diagram of the puncture needle entering the subarachnoid space Figure 8 As shown, the yellow dotted line in the figure shows the spinal cavity, indicating that the puncture needle has penetrated into the subarachnoid space at this time;

[0077] Insert the guide wire through the puncture needle and then keep the guide wire in place. Fig. 9 As shown, the white arrow in the figure indicates the guide wire;

[0078] The balloon is implanted under the guidance of a guidewire, and the balloon is pressurized and inflated before being removed in the inflated state. Fig.10As shown, the white arrow in the figure indicates the position of the balloon;

[0079] One hour after surgery, the cerebrospinal fluid pressure was measured from 80 mmH 2 O becomes 60mmH 2 O;

[0080] One week after surgery, the experimental animals were prepared for CT myelography, as Fig.11 As shown, after lumbar puncture, contrast medium was injected, and the dog was scanned in a prone position, with the lower limbs and back slightly elevated to allow the contrast medium to fill the subarachnoid space;

[0081] CT myelography results Fig.12 and Fig.13 As shown, the white arrow in the figure indicates cerebrospinal fluid leakage, indicating that there is a leakage point in the dog's dura mater.

[0082] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. A method for establishing an experimental animal model of spontaneous intracranial hypotension, characterized in that: The experimental animal model establishment method mainly includes the following steps: Step S1: At the beginning of the experiment, select experimental animals in good health and prepare for surgery; Step S2: The experimental animals are fasted and deprived of water before the operation; Step S3: anesthetizing and preparing the skin of the experimental animal; Step S4: performing spinal puncture: under X-ray guidance, a thoracic puncture is performed in the intervertebral space between the sixth and seventh thoracic vertebrae of the experimental animal, and the puncture needle enters the subarachnoid space, and cerebrospinal fluid can generally be seen flowing out; Step S5: Using the balloon method to create a dural leak to construct a spontaneous intracranial hypotension model; Step S6: performing a postoperative examination on the experimental animal; Step S7: After the experiment is over, the experimental animals are humanely killed; The step S5 comprises the following steps: Step S51: inserting a guide wire into the subarachnoid space through the puncture needle, and keeping the position of the guide wire unchanged; Step S52: inserting a balloon under the guidance of the guidewire: inserting the balloon along the guidewire into the subarachnoid space; Step S53: After the balloon is pressurized and inflated, the balloon is pulled out in an inflated state, and the dura mater can be ruptured when the balloon is pulled out; The step S6 comprises the following steps: S61: 1 hour after the operation, the cerebrospinal fluid pressure of the experimental animal is measured; S62: One week after the operation, the experimental animals were subjected to CT angiography to observe whether the thoracic vertebral segment where the operation was performed had a leak; S63: Two weeks after the operation, the experimental animals were subjected to enhanced MRI scanning to observe whether there were dura mater enhancement, brain sagging and characteristic manifestations of spontaneous intracranial hypotension.

2. The method for establishing an experimental animal model of spontaneous intracranial hypotension according to claim 1, characterized in that: The experimental animals are large mammals with similar anatomical features and the causes and development characteristics of spontaneous intracranial hypotension as those of humans.

3. The method for establishing an experimental animal model of spontaneous intracranial hypotension according to claim 1, characterized in that: The diameter of the balloon is different and is selected according to experimental requirements.

4. The method for establishing an experimental animal model of spontaneous intracranial hypotension according to claim 1, characterized in that: The puncture needle is an epidural puncture needle.

5. The method for establishing an experimental animal model of spontaneous intracranial hypotension according to claim 1, characterized in that: The puncture sites include the entire spinal segment including the cervical, thoracic and lumbar vertebrae.

Images