Operation method suitable for electrophysiological detection in cardiac cavity of rat / mouse

Abstract

The invention discloses an operation method suitable for electrophysiological detection in cardiac cavities of rats / mice, and relates to the technical field of experimental animal medical detection, and the operation method comprises the following steps: anesthesia of rats / mice, supine position fixation, and neck skin preparation and hair removal; separating the neck artery of the rat / mouse, ligating the artery blood vessel at the telecentric end, and clamping the artery blood vessel at the proximal end by using an artery clamp; the artery blood vessel in the middle section is temporarily retained after being punctured by an L-shaped guide needle, and the electrophysiology catheter is inserted along the notch; slowly moving the electrode catheter into the left ventricle, fixing the electrode catheter, and performing electrophysiological detection. According to the method, the detection electrode is inserted from the neck artery, the detection electrode is sent into the heart cavity of the rat / mouse, the integrity of the heart organ is not damaged, the risk of cardiac bleeding is avoided, accurate electrocardio data can be detected for a long time, meanwhile, after one carotid artery is cut off, the rat / mouse can maintain basic physiological activities through the other carotid artery, and therefore the detection effect is good. Continuous electrocardio detection can be carried out so as to complete the whole set of electrocardio recording.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of experimental animal medical detection, and particularly relates to an operation method suitable for intracardiac electrophysiological detection of mice. BACKGROUND

[0002] Cardiac arrhythmias are a major cause of morbidity and mortality worldwide. Although recent advances in cell-based models, including human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CM), have contributed to our understanding of electrophysiology and arrhythmia mechanisms, preclinical animal studies of cardiovascular diseases remain the mainstream. Over the past few decades, animal models of cardiovascular diseases have improved our understanding of pathological remodeling, arrhythmia mechanisms, and drug effects, and have led to major improvements in pacing and defibrillation therapy.

[0003] In conventional animal studies, the electrocardiogram of mice is usually detected on the body surface, that is, electrodes are connected to the limbs of mice for electrocardiogram detection. This method is simple and effective, but its accuracy is difficult to guarantee, and it is easily affected by the environment, and the recorded data is relatively single. In order to ensure the accuracy of electrophysiological detection and enrich the indicators of electrophysiological detection of model animals, intracardiac detection method is derived, that is, the heart is exposed by thoracotomy, the recording electrode is placed at the apex of the heart, and the electrophysiological activity of the heart is recorded by giving programmed electrical stimulation. However, the animal survival time is only a few minutes after thoracotomy, and the complete electrocardiogram recording cannot be completed. In addition, the heart is a blood storage organ, and there is a risk of bleeding. Moreover, the recorded parameters of model animals and normal animals differ greatly, and fixed recording parameters can cause fatal arrhythmias. Based on this, an operation method suitable for intracardiac electrophysiological measurement of mice is proposed. SUMMARY

[0004] The present application aims to provide an operation method suitable for intracardiac electrophysiological detection of mice, to ensure the accuracy of electrophysiological detection, enrich the indicators of electrophysiological detection of model animals, and solve the problem that complete electrocardiogram recording cannot be stably completed under thoracotomy.

[0005] To achieve the above-mentioned purpose, the present application provides the following technical scheme:

[0006] An operation method suitable for intracardiac electrophysiological detection of mice, comprising the following steps:

[0007] (1) Anesthetize the mouse, fix it in a supine position, and depilate and depilate the neck;

[0008] (2) Isolate the arterial blood vessels of the mouse, ligate the arterial blood vessels at the distal end of the heart, and use an arterial clamp to close the arterial blood vessels at the proximal end of the heart;

[0009] (3) using L-shaped guide needle to puncture the middle segment of the proximal end artery, temporarily leaving it, using physiological saline to soak the tip of the electrophysiological catheter to keep it lubricated, then inserting the electrophysiological catheter along the needle hole, withdrawing the L-shaped guide needle after the electrophysiological catheter is successfully inserted into the blood vessel, continuing to push the electrophysiological catheter inward, and then closing the artery opening after the electrode tip of the catheter is completely inserted into the blood vessel, and then releasing the proximal end artery clamp;

[0010] (4) slowly moving the electrode catheter to make the electrode tip enter the left ventricle along the artery, judging the entry of the electrode catheter into the left ventricle according to the intracardiac electrocardiogram, fixing the electrode catheter, and performing intracardiac electrocardiogram measurement through a program electric stimulation.

[0011] Further, in step (3), the L-shaped guide needle with a diameter of 0.5 mm is used for rats, and the L-shaped guide needle with a diameter of 0.3 mm is used for mice.

[0012] Further, in step (4), after the electrode reaches the left ventricle, the minimum diastolic period threshold, effective refractory period, ventricular fibrillation threshold and ventricular fibrillation induction rate of the rat / mouse are recorded in sequence, and the ventricular fibrillation duration and recovery sinus rate time are calculated.

[0013] Further, the measurement of the minimum diastolic period threshold includes: performing S1S1 short array stimulation to determine the diastolic period threshold, and the stimulation mode includes: starting with a stimulation voltage of 3000 mV, increasing or decreasing by 500 mV each time, an interval of 90% of the RR interval ms, a pulse width of 2 ms, and a duration of 30 times, with an interval of 1 min;

[0014] The measurement of the effective refractory period includes: performing S1S2 program stimulation, adding 1 S2 after 8 S1, recording the effective refractory period, and the stimulation mode includes: S1 stimulation voltage = 1.5 times the minimum diastolic period threshold, interval = 90% of the RR interval ms, pulse width 2 ms, 8 times; S2 interval = S1 interval-5 ms, -5 ms reverse scan, pulse width 2 ms, 1 time; the triggering delay is 0 ms, and the repetition interval is 1000 ms;

[0015] The measurement of the ventricular fibrillation threshold includes: performing high-intensity S1S1 stimulation to determine the ventricular fibrillation threshold, and the stimulation mode includes: the stimulation voltage is the minimum diastolic period threshold, and increases by 1000 mV each time; the stimulation interval is 33.6 / 25.2 / 16.8 ms; the pulse width is 2 ms; the duration is 1 min, and the repetition interval is 1 min;

[0016] The measurement of the ventricular fibrillation induction rate includes: after determining the ventricular fibrillation threshold, performing high-intensity S1Sx stimulation to determine the ventricular fibrillation induction rate, and the stimulation mode includes: performing 10 burst stimulations, the stimulation parameters are the same as the ventricular fibrillation threshold parameters, recording and analyzing the ventricular fibrillation induction rate, and finally calculating the ventricular fibrillation duration and recovery sinus rate time.

[0017] The principle and beneficial effects of the technical solution include at least:

[0018] The application provides a method for inserting a detection electrode from a carotid artery to send the detection electrode into a heart cavity of a mouse to perform electrocardio detection.

[0019] 1. The method does not damage the integrity of the heart organ, avoids the risk of heart bleeding, and can accurately detect electrocardio activity.

[0020] 2. Meanwhile, since the carotid artery of a mouse has two roots, after one carotid artery is clamped, the mouse can maintain basic physiological activities through the other carotid artery, and can perform long-term continuous electrocardio detection to complete a complete electrocardio record. After the experiment is completed, the punctured carotid artery can be ligated to ensure the survival of the mouse, or the mouse can be directly taken for further research.

[0021] 3. The structure of the model animal heart changes, and the electrical physiological parameters differ greatly. The method can accurately obtain the electrocardio parameters of each animal by giving a low-to-high step stimulation, and can avoid the phenomenon that different groups of animals are not tolerant due to uniform and non-difference stimulation parameters, and even death due to excessive stimulation parameters. BRIEF DESCRIPTION OF DRAWINGS

[0022] Figure 1 It is an operation method flowchart suitable for electrical physiological detection in a mouse heart cavity.

[0023] Figure 2 It is a schematic view after a catheter is placed along the carotid artery of a mouse.

[0024] Figure 3 It is a typical view of determining the catheter position in the heart cavity of a mouse.

[0025] Figure 4 It is a typical view of giving a programmed electrical stimulation to determine the minimum diastolic period threshold after the catheter position of a mouse is determined.

[0026] Figure 5 It is a typical view of giving a programmed electrical stimulation to determine the minimum diastolic period threshold after the catheter position of a mouse is determined.

[0027] Figure 6 It is a typical view of determining the ventricular fibrillation threshold after the catheter position of a mouse is determined.

[0028] Figure 7 It is a typical view of determining the catheter position in the heart cavity of a mouse.

[0029] Figure 8 It is a typical view of giving a programmed electrical stimulation to determine the minimum diastolic period threshold after the catheter position of a mouse is determined.

[0030] Figure 9Typical diagram of effective refractory period after giving programmed electrical stimulation reverse sweep to mice after determining the minimum diastolic period threshold;

[0031] Figure 10 Typical diagram of fibrillation after determining the fibrillation threshold of mice; DETAILED DESCRIPTION

[0032] In order to make the objects, technical solutions and advantages of the present application clearer, the present application will be further described in detail below with reference to the embodiments and drawings. Herein, the illustrative embodiments of the present application and the descriptions thereof are used to explain the present application, but are not used as the limitation of the present application.

[0033] It should also be noted that, in order to avoid the present application being obscured by unnecessary details, only the structures and / or processing steps closely related to the solutions according to the present application are shown in the drawings, and other details not closely related to the present application are omitted.

[0034] It should be emphasized that the term "comprises / comprising" as used herein indicates the presence of the stated features, elements, steps or components, but does not exclude the presence or addition of one or more other features, elements, steps or components.

[0035] Hereinafter, the embodiments of the present application will be described with reference to the drawings. In the drawings, the same reference signs represent the same or similar parts or the same or similar steps.

[0036] It should be emphasized here that the step labels mentioned hereinafter are not the limitation of the order of the steps, and it should be understood that the steps can be performed in the order mentioned in the embodiments, or in an order different from the embodiments, or several steps are performed simultaneously.

[0037] Embodiment One

[0038] Reference Figure 1 With Figure 2 An operation method suitable for intracardiac electrophysiological detection of large / small mice, comprising the following steps:

[0039] 1. Anesthetize the rat, fix it in a supine position, and depilate and depilate the neck;

[0040] 2. Isolate the unilateral rat carotid artery, ligate the distal end of the arterial blood vessel with a fine thread, and close the proximal end of the arterial blood vessel with an arterial clamp;

[0041] 3. After the blood flow is blocked, a L-shaped guide needle with a sharp tip and a diameter of 0.5 mm (for rats) or 0.3 mm (for mice) is used to puncture the blood vessel and then temporarily left in place. The tip of the electrophysiological catheter is soaked in normal saline to keep it lubricated. Then the catheter is inserted along the needle hole. After the catheter is successfully inserted into the blood vessel, the L-shaped guide needle is withdrawn. The catheter is continuously moved inward. After the electrode tip of the catheter is completely inserted into the blood vessel, a thin wire is used to tie the arterial opening (No. 1 knot) to prevent bleeding. Then the proximal arterial clamp is released. It is worth noting that a 1 mm to 1.2 mm notch can also be cut on the arterial blood vessel. The electrophysiological catheter can be inserted along the notch to achieve the same effect of inserting the catheter.

[0042] 4. The distal end of the artery is clamped with an ophthalmic clamp, and the electrode catheter is slowly pushed and moved (keeping the catheter wall moist). The electrode tip is pushed into the left ventricle along the artery. Whether the catheter enters the left ventricle is determined according to the intracardiac electrocardiogram pattern. Reference is made to Figure 3 , which is a typical graph of the catheter position in the heart cavity. After No. 1 knot, the electrode catheter is fixed with a thin wire (No. 2 knot). Programmed electrical stimulation is performed through an electrical stimulator, and intracardiac electrocardiogram measurement is performed.

[0043] After the electrode reaches the left ventricle, the minimum diastolic threshold, effective refractory period, ventricular fibrillation threshold, and ventricular fibrillation induction rate of rats and mice are recorded in sequence. The ventricular fibrillation duration and the time to restore sinus rate are calculated.

[0044] Reference is made to Figure 4 , which is used to measure the minimum diastolic threshold of rats. S1S1 short array stimulation is performed to determine the diastolic threshold. The stimulation method includes: stimulation voltage 3000 mV starting (increasing / decreasing by 500 mV each time), interval 90% x RR interval ms, pulse width 2 ms, lasting for 30 times, and interval 1 min.

[0045] Reference is made to Figure 5 , which is used to measure the effective refractory period of rats. S1S2 program stimulation is performed, 8 S1s are followed by 1 S2, and the effective refractory period is recorded. The stimulation method includes: S1 stimulation voltage = 1.5 times the minimum diastolic threshold, interval = 90% x RR interval ms, pulse width 2 ms, 8 times; S2 interval = S1 interval - 5 ms, -5 ms reverse sweep (decrease by 5 ms each time), pulse width 2 ms, 1 time; the triggering delay is 0 ms, and the repeat interval is 1000 ms.

[0046] Reference is made to Figure 6 , which is used to measure the ventricular fibrillation threshold of rats. High-intensity S1S1 stimulation is performed to determine the ventricular fibrillation threshold. The stimulation method includes: stimulation voltage is the minimum diastolic threshold, increasing by 1000 mV each time; stimulation interval 33.6 / 25.2 / 16.8 ms (starting from 33.6 ms, increasing to 16.8 ms as needed); pulse width 2 ms; lasting for 1 min, repeat interval 1 min.

[0047] Measurement of the rate of ventricular fibrillation induction: After the determination of the threshold of ventricular fibrillation, high intensity S1Sx stimulation is performed to determine the rate of ventricular fibrillation induction, the stimulation mode includes: 10 times of burst stimulation (the stimulation parameters are the same as the parameters of the threshold of ventricular fibrillation) is performed, and the rate of ventricular fibrillation induction is recorded and analyzed. Finally, according to the recorded data, the duration of ventricular fibrillation and the time of recovery to sinus rate are further calculated.

[0048] Example 2

[0049] The difference from example 1 is that the experimental subject is a mouse.

[0050] Reference Figure 7 Typical diagram of determining the position of the catheter in the heart cavity of a mouse;

[0051] Reference Figure 8 Typical diagram of determining the minimum diastolic period threshold of a mouse after the position of the catheter is determined and programmed electrical stimulation is given;

[0052] Reference Figure 9 Typical diagram of determining the minimum diastolic period threshold of a mouse after the position of the catheter is determined and programmed electrical stimulation is given;

[0053] Reference Figure 10 Typical diagram of determining the threshold of ventricular fibrillation of a mouse after the threshold of ventricular fibrillation is determined.

[0054] It should be noted that the present application is not limited to the specific configurations and processes described above and shown in the drawings. For the sake of brevity, detailed descriptions of well-known methods are omitted here. In the above examples, several specific steps are described and shown as examples. However, the method process of the present application is not limited to the specific steps described and shown, and those skilled in the art can make various changes, modifications and additions, or change the order between steps, after understanding the spirit of the present application.

[0055] In the present application, the features described and / or exemplified for one embodiment can be used in the same way or in a similar way in one or more other embodiments, and / or in combination with or instead of the features of other embodiments.

[0056] The above description is only the preferred embodiment of the present application and is not intended to limit the present application. For those skilled in the art, the embodiments of the present application can be variously changed and modified. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present application shall be included in the protection scope of the present application.

Claims

1. A method for conducting intracardiac electrophysiological testing in mice and rats, characterized in that, Includes the following steps: (1) Anesthetize the mouse, fix it in a supine position, and prepare the skin and remove the hair from its neck; (2) Separate the rat's carotid artery, ligate the distal end of the artery, and clamp the proximal end of the artery with an arterial clamp. (3) After puncturing the middle segment of the proximal artery with an L-shaped guide needle, leave it temporarily, moisten the tip of the electrophysiological catheter with normal saline to keep it lubricated, and then insert the electrophysiological catheter along the needle hole. After the electrophysiological catheter is successfully inserted into the blood vessel, withdraw the L-shaped guide needle and continue to push the electrophysiological catheter inward. After the electrode tip of the catheter is fully inserted into the blood vessel, close the arterial opening and then release the proximal arterial clamp. (4) Slowly move the electrode catheter so that the electrode tip enters the left ventricle along the artery. After the electrode enters the left ventricle according to the morphology of the intracardiac electrocardiogram, fix the electrode catheter and deliver programmed electrical stimulation through the electrical stimulator to perform intracardiac electrocardiogram measurement.

2. The operating method for intracardiac electrophysiological detection in mice / large mice according to claim 1, characterized in that, In step (3), a 0.5 mm diameter L-shaped guide needle was used for rats and a 0.3 mm diameter L-shaped guide needle was used for mice.

3. The operating method for intracardiac electrophysiological detection in mice / large mice according to claim 1, characterized in that, In step (4), after the electrode reaches the left ventricle, the minimum diastolic threshold, effective refractory period, ventricular fibrillation threshold and ventricular fibrillation induction rate of mice / greater mice are recorded in sequence, and the duration of ventricular fibrillation and the time to recovery of sinus rate are calculated.

4. The operating method for intracardiac electrophysiological detection in mice / large mice according to claim 4, characterized in that, The measurement of the minimum diastolic threshold includes: performing short S1S1 stimulation to determine the diastolic threshold. The stimulation method includes: starting with a stimulation voltage of 3000mV, increasing or decreasing by 500mV each time, with an interval of 90% × RR interval ms, a pulse width of 2ms, lasting for 30 cycles, and interrupting for 1min. The effective refractory period was measured by performing S1S2 programmed stimulation, firing 8 S1 pulses followed by 1 S2 pulse, and recording the effective refractory period. The stimulation method included: S1 stimulation voltage = 1.5 times the minimum diastolic threshold, interval = 90% × RR interval ms, pulse width 2 ms, 8 times; S2 interval = S1 interval - 5 ms, -5 ms reverse scan, pulse width 2 ms, 1 time; priming delay was 0 ms, and the repetition interval was 1000 ms. Measuring the ventricular fibrillation threshold involves performing high-intensity S1S1 stimulation to determine the ventricular fibrillation threshold. The stimulation method includes: stimulation voltage equal to the minimum diastolic threshold, increasing by 1000mV each time; stimulation intervals of 33.6 / 25.2 / 16.8ms; pulse width of 2ms; duration of 1min, repeated at 1min intervals. The measurement of ventricular fibrillation induction rate includes: after determining the ventricular fibrillation threshold, high-intensity S1Sx stimulation is performed to determine the ventricular fibrillation induction rate. The stimulation method includes: performing 10 burst stimulations, with each stimulation parameter being the same as the ventricular fibrillation threshold parameter. The ventricular fibrillation induction rate is recorded and analyzed. Finally, the duration of ventricular fibrillation and the time to recovery of sinus rate are calculated.

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