Quantitative experiment research and analysis method for action potential refractory period phenomenon of neural stem
By adopting the "single-double-single" electrical stimulation series experiments and data processing methods in the study of refractory period phenomena of neural stem AP, the problem that traditional research methods cannot conduct quantitative analysis is solved, and accurate quantitative research on refractory period phenomena and accurate determination of critical value ranges are achieved.
Patent Information
- Application Number
- CN202411792490.3
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-12-07
- Publication Date
- 2025-03-18
AI Technical Summary
The traditional research method for the refractory phenomenon of neural stem action potential cannot conduct accurate and systematic quantitative research, and can only conduct qualitative observations, and cannot accurately measure the amplitude of the AP waveform, resulting in the inaccurate analysis of refractory phenomena.
A quantitative experimental research and analysis method for the refractory period of neural stem AP was used. By selecting multiple different double-pulse wave interval time values, a series of "single-double-single" electrical stimulation experiments were carried out, and the AP waveform data induced by single pulse and double pulse were recorded respectively. Through superposition and subtraction processing, the AP waveforms induced by the first pulse and the second pulse were split and accurately measured, the maximum amplitude ratio was calculated, and the coordinate graph was drawn to determine the critical value range of the refractory period.
The accurate quantitative study of the refractory period of neural stem AP has been achieved, and the critical value range of absolute refractory period, relative refractory period and non-refractory period can be accurately determined, which improves the accuracy and unity of the research.
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Abstract
Description
Technical Field
[0001] The invention belongs to the technical field of body function experimental research, and in particular relates to a quantitative experimental research and analysis method for the refractory period phenomenon of nerve stem action potential. Background Art
[0002] The outer layer of the nerve trunk is the epineurium, which contains many nerve fibers. Nerve fibers achieve rapid long-distance transmission of information by generating and conducting action potentials. Nerve trunk action potential (abbreviated as nerve trunk AP, where AP is the abbreviation of action potentials) is a comprehensive electrical signal recorded when many nerve fibers contained in the nerve trunk generate action potentials due to excitement and are transmitted to the surface of the nerve trunk at the recording electrode.
[0003] In a short period of time after a nerve trunk experiences a suprathreshold electrical stimulation, the sodium ion channel is temporarily not revived. If a new suprathreshold electrical stimulation is given during this period, its response will decrease or even disappear. This period is called the refractory period. The traditional research method of nerve trunk AP refractory period phenomenon generally adopts the following double pulse electrical stimulation method: multiple rounds of suprathreshold double pulse electrical stimulation are applied to the nerve trunk, and the parameters of the first pulse and the second pulse in each double pulse electrical stimulation (such as stimulation amplitude, stimulation wave width, etc.) are the same, inducing a nerve trunk AP mixed waveform induced by double pulse electrical stimulation (which can be represented by APall, including the nerve trunk AP waveform AP1 induced by the first pulse electrical stimulation and the nerve trunk AP waveform AP2 induced by the second pulse electrical stimulation); in each round of electrical stimulation, the time interval between the two pulses (also called the wave interval, represented by t) gradually decreases from large to small. By analyzing the waveforms obtained from the above-mentioned double-pulse electrical stimulation, it can be found that: when the wave interval t between the double-pulse electrical stimulation is large enough (such as t = 30 ms), two nerve trunk AP waveforms with almost identical sizes and shapes can be recorded one after another, such as Figure 1 As shown in A, there is no refractory period phenomenon in this process (for the convenience of description, the stage in which the wave interval t is large enough and the refractory period phenomenon does not occur is called the "non-refractory period"); when the wave interval t of the double pulse electrical stimulation is reduced to a certain range (such as t = 3.6 ms), it can be seen that the amplitude of the AP waveform of the second nerve trunk is significantly smaller than that of the first one. This is the "relative refractory period" phenomenon, as shown in Figure 1 As shown in Figure B; when the wave interval t of the double pulse electrical stimulation continues to decrease to a sufficiently small range (such as t = 0.8 ms), it can be seen with the naked eye that the AP waveform of the second nerve trunk has completely disappeared. This is the "absolute refractory period" phenomenon. Figure 1 As shown in C.
[0004] Although the above traditional experimental research method can observe the refractory period phenomenon of nerve trunk AP, it cannot conduct accurate and systematic quantitative research on the refractory period phenomenon of nerve trunk AP, and can only conduct qualitative research, so it is generally called the observation of the refractory period phenomenon of nerve trunk AP. This is because the nerve trunk AP waveform APall obtained in the above double pulse electrical stimulation process is actually a mixed waveform formed by the superposition of two nerve trunk AP waveforms AP1 and AP2 induced by two electrical stimulation pulses at a certain interval, such as Figure 2 As shown ( Figure 2 (The mixed waveforms of AP in nerve trunks induced by various double-pulse electrical stimulations obtained under experimental conditions with a wave interval of 0.8 ms to 3.0 ms) People can generally roughly determine the manifestation of the refractory period by observing the changing characteristics of the mixed waveforms with the naked eye: if the naked eye feels that the second nerve trunk AP waveform has disappeared, it is judged to be in the "absolute refractory period"; if the naked eye feels that the second nerve trunk AP waveform is significantly smaller than the first, it is judged to be in the "relative refractory period"; if the naked eye feels that the size and shape of the two waveforms are almost exactly the same, it is judged to be in the "non-refractory period". In the above process, although the naked eye observation results of different people are roughly the same, there are individual differences in the specific judgment results. The method of judging by the naked eye alone will lead to inaccurate research and analysis of the AP refractory period phenomenon in the nerve trunk, the results are not standardized and unified, and the research is not in-depth and specific; even if it is not observed with the naked eye, but the waveform is measured with tools, it is impossible to accurately quantify. This is because when the wave interval of the double-pulse electrical stimulation is small, the two AP waveforms are actually partially superimposed on each other, which will cause the waveform to be deformed and distorted, especially the distortion of the AP waveform of the second nerve trunk is more serious, such as Figure 2 As shown; therefore, the traditional nerve trunk AP refractory period experimental research method is unable to accurately measure the two AP waveform amplitudes, especially the second nerve trunk AP waveform amplitude, and thus is unable to conduct a truly accurate quantitative study of the refractory period phenomenon. Summary of the invention
[0005] The purpose of the present invention is to overcome the above-mentioned shortcomings and provide an accurate quantitative experimental research and analysis method for the AP refractory period phenomenon of nerve trunks, which can obtain two complete and undistorted nerve trunk AP waveforms AP1 and AP2 induced by the first pulse and the second pulse respectively, and can accurately determine the critical value range of the absolute refractory period, the relative refractory period and the period without refractory phenomenon.
[0006] The purpose can be achieved according to the following scheme: A quantitative experimental research and analysis method for the refractory period phenomenon of nerve stem action potentials, comprising the following steps: Step 1: Select a plurality of different double pulse wave interval time values t as experimental conditions, and the wave interval time values t are from large to small, and the selected maximum wave interval time value is within the stage range of the non-refractory period, and the selected minimum wave interval time value is within the stage range of the absolute refractory period; for each wave interval time value t, a series of "single-double-single" electrical stimulation experiments are carried out, which sequentially include the following experimental sub-steps: ① Apply a single pulse electrical stimulation to the nerve trunk specimen, and record the nerve trunk AP waveform data AP1 induced by the single pulse electrical stimulation; ② Apply a wave interval to the nerve trunk specimen ③ Apply a double pulse electrical stimulation with a time value of t, and record the AP mixed waveform data APall of the nerve trunk induced by the double pulse electrical stimulation; ③ Apply a single pulse electrical stimulation to the nerve trunk specimen again, and record the AP waveform data AP1 of the nerve trunk induced by the single pulse electrical stimulation; The interval time between the above experimental sub-steps ① and ② is equal to the interval time between experimental sub-steps ② and ③, and the interval time is 1 to 10 seconds; Each pulse in the above experimental sub-steps ①, ② and ③ is a square wave, and the amplitude and wave width of all the square wave electrical stimulations are the same; Step 2: for each experimental result corresponding to the wave interval time value t obtained in step 1, the nerve trunk AP waveform data AP1 obtained in experimental sub-step ① and the nerve trunk AP waveform data AP1 obtained in experimental sub-step ③ are superimposed and divided by 2 to obtain the superimposed average waveform data of the nerve trunk AP induced by the two single-pulse electrical stimulations in experimental sub-steps ① and ③, and the nerve trunk AP superimposed average waveform data is used to characterize the nerve trunk AP waveform data AP1 induced by the first pulse electrical stimulation in the double-pulse electrical stimulation in experimental sub-step ②; Step 3: For each experimental result corresponding to the wave interval time value t obtained in step 1, the mixed waveform data APall obtained in experimental sub-step ② minus the nerve trunk AP waveform data AP1 induced by the first pulse electrical stimulation of experimental sub-step ② obtained in step 2 is obtained to obtain the nerve trunk AP waveform data AP2 induced by the second pulse of experimental sub-step ②; Step 4: for each result of step 2 and step 3 corresponding to each wave interval time value t, measure the maximum amplitude AAP1 of the AP waveform data of the nerve trunk induced by the first pulse of the experimental sub-step ② obtained in step 2, and measure the maximum amplitude AAP2 of the AP waveform data of the nerve trunk induced by the second pulse of the experimental sub-step ② obtained in step 3, wherein the maximum amplitude of the AP waveform data of the nerve trunk induced by the first pulse is referred to as the "maximum amplitude of the first pulse", and the maximum amplitude of the AP waveform data of the nerve trunk induced by the second pulse is referred to as the "maximum amplitude of the second pulse", and the ratio AAP2 / AAP1 of "maximum amplitude of the second pulse / maximum amplitude of the first pulse" corresponding to each double pulse is calculated, and the ratio of "maximum amplitude of the second pulse / maximum amplitude of the first pulse" is referred to as the "maximum amplitude ratio", thereby obtaining the corresponding "maximum amplitude ratio" under the condition that the double pulse wave interval time value is t; Step 5: Based on the result obtained in step 4, with the wave interval time value t as the horizontal coordinate and the "maximum amplitude ratio" as the vertical coordinate, the discrete points corresponding to each wave interval time value t are drawn in the plane rectangular coordinate system to form a coordinate diagram showing the relationship between the wave interval time value t and the "maximum amplitude ratio"; Step 6. Based on the coordinate graph obtained in step 5, determine the critical value range between the non-refractory period and the relative refractory period according to the following rules: along the direction of the wave interval time value t from small to large, the wave interval time value corresponding to the discrete point where the "maximum amplitude ratio" is not less than 100% for the first time is determined as the maximum value of the critical value range between the non-refractory period and the relative refractory period, and the wave interval time value corresponding to the adjacent discrete point whose wave interval time value is smaller than the discrete point is determined as the minimum value of the critical value range between the non-refractory period and the relative refractory period; Step 7. Based on the coordinate graph obtained in step 5, determine the critical value range between the absolute refractory period and the relative refractory period according to the following rules: along the direction of the wave interval time value t from large to small, the wave interval time value corresponding to the discrete point where the "maximum amplitude ratio" is not greater than 0% for the first time is determined as the minimum value of the critical value range between the absolute refractory period and the relative refractory period, and the wave interval time value corresponding to the adjacent discrete point whose wave interval time value is larger than that of the discrete point is determined as the maximum value of the critical value range between the absolute refractory period and the relative refractory period.
[0007] Preferably, in the "single-double-single" electrical stimulation series experiments corresponding to two adjacent wave interval time values t, the experimental sub-step ③ corresponding to the previous wave interval time value t is used as the experimental sub-step ① corresponding to the next wave interval time value t.
[0008] It should be noted that, in the prior art, although it is impossible to accurately determine the critical value range of the "absolute refractory period", "relative refractory period" and "period without refractory phenomenon", it can still be roughly estimated; therefore, as long as the wave interval time value t selected in step one is long enough, it can be ensured that the selected maximum wave interval time value is within the stage range of the period without refractory phenomenon; as long as the wave interval time value t selected in step one is short enough, it can be ensured that the selected minimum wave interval time value is within the stage range of the absolute refractory period.
[0009] The specific amplitude and specific wave width of each of the above square waves can be determined according to the biological properties of the nerve trunk specimen so as to be able to induce a stable and typical nerve trunk AP waveform. This belongs to conventional technical content.
[0010] The reason why the above series of experiments for each wave interval time value t is named "single-double-single" electrical stimulation experiment is that it first performs a single pulse electrical stimulation, then a double pulse electrical stimulation, and then a single pulse electrical stimulation again, so the term "single-double-single" is used in the naming.
[0011] The present invention has the following advantages and effects: 1. The nerve trunk specimen is an in vitro biological specimen, and its biological properties may gradually change as the in vitro time gradually increases, and may also gradually change as the number of electrical stimulations gradually increases. In the present invention, the time when the first pulse of the double pulse in experimental sub-step ② occurs is basically equivalent to the midpoint between the time when experimental sub-step ① occurs and the time when experimental sub-step ③ occurs, and the number of electrical stimulations experienced when the first pulse of the double pulse in experimental sub-step ② occurs is also roughly equivalent to the midpoint between the number of electrical stimulations experienced when experimental sub-step ① occurs and the number of electrical stimulations experienced when experimental sub-step ③ occurs. Therefore, the superimposed average waveform data of the nerve trunk AP induced by the two single pulse electrical stimulations in experimental sub-steps ① and ③ is theoretically basically equal to the average waveform data of the double pulse in experimental sub-steps ① and ③. The nerve trunk AP waveform data induced by the first pulse of the double pulse in ② can be more accurately characterized by the superimposed average waveform data of the nerve trunk AP induced by two single-pulse electrical stimulations in experimental sub-steps ① and ③. Furthermore, the nerve trunk AP waveform data induced by the first pulse of the double pulse in experimental sub-step ② can be subtracted from the mixed waveform data obtained in experimental sub-step ② to obtain the nerve trunk AP waveform data induced by the second pulse of the double pulse electrical stimulation. In this way, the two nerve trunk AP waveforms induced by the first pulse and the second pulse in the double pulse are completely separated, thereby accurately measuring the relevant parameters of the two waveforms respectively, creating conditions for further precise and quantitative analysis and research.
[0012] Second, the present invention establishes the key index (i.e., "maximum amplitude ratio") for determining "absolute refractory period", "relative refractory period" and "period without refractory phenomenon", and draws the coordinate relationship diagram between "maximum amplitude ratio" and wave interval time value t. The characteristics of the coordinate relationship diagram can be used to accurately determine the critical range values of "absolute refractory period", "relative refractory period" and "period without refractory phenomenon". This method is clearer and more intuitive than the traditional judgment method, and can simplify the complex, make the judgment elements single and unique, and turn the uncertain subjective judgment into a certain objective result, so as to achieve true quantitative research and analysis.
[0013] 3. In the "single-double-single" electrical stimulation series experiments corresponding to the time values t of the intervals between two adjacent waves, the experimental sub-step ③ corresponding to the previous wave interval time value t is used as the experimental sub-step ① corresponding to the next wave interval time value t. This can reduce the number of experimental sub-steps, shorten the overall experimental time, reduce the number of electrical stimulations of the nerve trunk specimen, and try to keep the biological properties of the nerve trunk specimen stable, which is conducive to accurate experimental results and makes the experiment more convenient. BRIEF DESCRIPTION OF THE DRAWINGS
[0014] Figure 1 A is a schematic diagram of mixed waveform data with a wave interval time value of 30ms and no refractory period phenomenon.
[0015] Figure 1 B is a schematic diagram of mixed waveform data showing a relative refractory period phenomenon when the wave interval time value is 3.6ms.
[0016] Figure 1 C is a schematic diagram of mixed waveform data showing an absolute refractory period with a wave interval value of 0.8 ms.
[0017] Figure 2 It is a schematic diagram of the mixed waveform data change trend with the wave interval time value ranging from 3.0ms to 0.8ms.
[0018] Figure 3 A is a schematic diagram of nerve trunk AP waveform data AP1_001 obtained in experimental sub-step 1 of a specific embodiment.
[0019] Figure 3 B is a schematic diagram of nerve trunk AP waveform data APall_002 obtained in experimental sub-step 2 of a specific embodiment.
[0020] Figure 3 C is a schematic diagram of nerve trunk AP waveform data AP1_003 obtained in experimental sub-step 3 of a specific embodiment.
[0021] Figure 3 D is a schematic diagram of nerve trunk AP waveform data APall_004 obtained in experimental sub-step 4 of the specific embodiment.
[0022] Figure 3 E is a schematic diagram of nerve trunk AP waveform data AP1_005 obtained in experimental sub-step 5 of a specific embodiment.
[0023] Figure 3 F is a schematic diagram of nerve trunk AP waveform data APall_006 obtained in experimental sub-step 6 of a specific embodiment.
[0024] Figure 3 G is a schematic diagram of nerve trunk AP waveform data AP1_129 obtained in the experimental sub-step 129 of the specific embodiment.
[0025] Figure 4 It is a schematic diagram of the process of obtaining the second pulse waveform data AP2_100 corresponding to the wave interval time value of 3.6ms.
[0026] Figure 5 It is a schematic diagram of the process of obtaining the second pulse waveform data AP2 corresponding to multiple wave interval time values.
[0027] Figure 6 It is a coordinate relationship diagram of the "maximum amplitude ratio" obtained in step five of the specific embodiment and the wave interval time value t. DETAILED DESCRIPTION
[0028] A quantitative experimental research and analysis method for the AP refractory period phenomenon of the sciatic nerve trunk of a bullfrog comprises the following steps: Step 1: Select 64 different double pulse wave interval time values t as experimental conditions, and the wave interval time values t are from large to small, as shown in Table 1 below, and also as shown in the third column of Table 2 below;
[0029] The maximum wave interval time value of 30ms selected above is within the range of the non-refractory period of the bullfrog's sciatic nerve trunk, and the minimum wave interval time value of 0.8ms selected is within the range of the absolute refractory period of the bullfrog's sciatic nerve trunk; for each wave interval time value t, a series of "single-double-single" electrical stimulation experiments are carried out, and the "single-double-single" corresponding to each wave interval time value includes the following experimental sub-steps in sequence: ① ① Apply single-pulse electrical stimulation to the nerve trunk specimen, and record the nerve trunk AP waveform data AP1 induced by the single-pulse electrical stimulation; ② Apply double-pulse electrical stimulation with a wave interval time value t to the nerve trunk specimen, and record the nerve trunk AP mixed waveform data APall induced by the double-pulse electrical stimulation; ③ Apply single-pulse electrical stimulation to the nerve trunk specimen again, and record the nerve trunk AP waveform data AP1 induced by the single-pulse electrical stimulation; the interval time between the above experimental sub-steps ① and ② is equal to the interval time between experimental sub-steps ② and ③, and the interval time is 2 seconds; each pulse in the above experimental sub-steps ①, ② and ③ is a square wave, and the amplitude and wave width of all pulse square wave electrical stimulations are the same, the amplitude is 2V, and the wave width of the square wave is 0.05ms; and in the "single-double-single" electrical stimulation series experiments corresponding to two adjacent wave interval time values t, use the experimental sub-step ③ corresponding to the previous wave interval time value t as the experimental sub-step ① corresponding to the next wave interval time value t.
[0030] Since step 1 has a total of 64 different double pulse wave interval time values t as experimental conditions, a total of the following 2×64+1=129 experimental sub-steps are required; step 1 is specifically described as follows: Experimental sub-step 1. Apply a single pulse electrical stimulation to the nerve trunk specimen, and obtain the nerve trunk AP waveform data AP1_001 induced by the single pulse electrical stimulation, such as Figure 3 As shown in A; Experimental sub-step 2. Apply double pulse electrical stimulation to the nerve trunk specimen, with the wave interval time t of the two pulses being 30 milliseconds (i.e., t1), and record the nerve trunk AP mixed waveform data APall_002 induced by the double pulse electrical stimulation, such as Figure 3 As shown in B; Experimental sub-step 3. Apply a single pulse electrical stimulation to the nerve trunk specimen, and obtain the nerve trunk AP waveform data AP1_003 induced by the single pulse electrical stimulation, such as Figure 3 As shown in C; Experimental sub-step 4. Apply double pulse electrical stimulation to the nerve trunk specimen, with the wave interval time t of the two pulses being 29 milliseconds (ie, t2), and record the nerve trunk AP mixed waveform data APall_004 induced by the double pulse electrical stimulation, such as Figure 3 As shown in D; Experimental sub-step 5. Apply a single pulse electrical stimulation to the nerve trunk specimen, and obtain the nerve trunk AP waveform data AP1_005 induced by the single pulse electrical stimulation, such as Figure 3 As shown in E; Experimental sub-step 6. Apply double pulse electrical stimulation to the nerve trunk specimen, with the wave interval time t of the two pulses being 28 milliseconds (ie, t3), and obtain the nerve trunk AP mixed waveform data APall_006 induced by the double pulse electrical stimulation, as shown in FIG. Figure 3 As shown in F; Similarly, experimental sub-step (2n-1). Apply a single pulse electrical stimulation to the nerve trunk specimen, and obtain the nerve trunk AP waveform data AP1_(2n-1) induced by the single pulse electrical stimulation; where n is the serial number of the wave interval time value t in Table 1, and the maximum value is 064; Experimental sub-step 2n. Apply double pulse electrical stimulation to the nerve trunk specimen, the wave interval time t of the two pulses is the corresponding tn in Table 1, and obtain the nerve trunk AP mixed waveform data APall_2n induced by the double pulse electrical stimulation; wherein n is the sequence number of the wave interval time value t in Table 1, and the maximum value is 064; Experimental sub-step 129. Apply a single pulse electrical stimulation to the nerve trunk specimen, and obtain the nerve trunk AP waveform data AP1_129 induced by the single pulse electrical stimulation, such as Figure 3 As shown in G.
[0031] Among the above 129 experimental sub-steps, each experimental sub-step 2n with an even number and its two adjacent experimental sub-steps with odd numbers [i.e., experimental sub-step (2n-1) and experimental sub-step (2n+1)] constitute three experimental sub-steps of the "single-double-single" electrical stimulation process for the wave interval time value numbered n and the wave interval time value tn, among which experimental sub-step (2n-1) is experimental sub-step ① of the "single-double-single" electrical stimulation process, experimental sub-step 2n is experimental sub-step ② of the "single-double-single" electrical stimulation process, and experimental sub-step (2n+1) is experimental sub-step ③ of the "single-double-single" electrical stimulation process.
[0032] After step 1 is completed, the names of the various waveform data are obtained as shown in the second column of Table 2 below.
[0033] Step 2: for the experimental results corresponding to each wave interval time value t obtained in step 1, the nerve trunk AP waveform data AP1 obtained in experimental sub-step ① and the nerve trunk AP waveform data AP1 obtained in experimental sub-step ③ are superimposed and divided by 2 to obtain the superimposed average waveform data of the nerve trunk AP induced by the two single-pulse electrical stimulations in experimental sub-steps ① and ③, and the nerve trunk AP superimposed average waveform data is used to characterize (represent) the nerve trunk AP waveform data AP1 induced by the first pulse electrical stimulation in the double-pulse electrical stimulation in experimental sub-step ②; Step 2 is as follows: The nerve trunk AP waveform data AP1_001 obtained in the above experimental sub-step 1 and the nerve trunk AP waveform data AP1_003 obtained in the experimental sub-step 3 are superimposed and divided by 2 to obtain the superimposed average waveform data of the nerve trunk AP induced by the two single-pulse electrical stimulations in the experimental sub-step 1 and the experimental sub-step 3; the superimposed average waveform data of the nerve trunk AP is used to represent the nerve trunk AP waveform data AP1_002 induced by the first pulse electrical stimulation in the double-pulse electrical stimulation in the experimental sub-step 2, AP1_002=(AP1_001+AP1_003) / 2; The nerve trunk AP waveform data AP1_003 obtained in the above experimental sub-step 3 and the nerve trunk AP waveform data AP1_005 obtained in the experimental sub-step 5 are superimposed and divided by 2 to obtain the superimposed average waveform data of the nerve trunk AP induced by the two single pulse electrical stimulations in the experimental sub-step 3 and the experimental sub-step 5; the superimposed average waveform data of the nerve trunk AP is used to represent the nerve trunk AP waveform data AP1_004 induced by the first pulse electrical stimulation in the double pulse electrical stimulation in the experimental sub-step 4; AP1_004=(AP1_003+AP1_005) / 2; The nerve trunk AP waveform data AP1_005 obtained in the above experimental sub-step 5 and the nerve trunk AP waveform data AP1_007 obtained in the experimental sub-step 7 are superimposed and divided by 2 to obtain the superimposed average waveform data of the nerve trunk AP induced by the two single-pulse electrical stimulations in the experimental sub-step 5 and the experimental sub-step 7; the superimposed average waveform data of the nerve trunk AP is used to represent the nerve trunk AP waveform data AP1_006 induced by the first pulse electrical stimulation in the double-pulse electrical stimulation in the experimental sub-step 6, AP1_006=(AP1_005+AP1_007) / 2; Similarly, the nerve trunk AP waveform data AP1_(2n-1) obtained in the above experimental sub-step (2n-1) and the nerve trunk AP waveform data AP1_(2n+1) obtained in the experimental sub-step (2n+1) are superimposed and divided by 2 to obtain the superimposed average waveform data of the nerve trunk AP induced by the two single-pulse electrical stimulations in the experimental sub-step (2n-1) and the experimental sub-step (2n+1); the superimposed average waveform data of the nerve trunk AP is used to represent the nerve trunk AP waveform data AP1_2n induced by the first pulse electrical stimulation in the double-pulse electrical stimulation in the experimental sub-step 2n, AP1_2n=[AP1_(2n-1) + AP1_(2n+1)] / 2; wherein n is the serial number of the wave interval time value t in Table 1, and the maximum value is 064; in order to make the serial numbers neat, the serial numbers in Table 1 are represented by three digits. If the serial number has only a unit digit, two zeros are added in front of it; if it has only a ten digit, one zero is added in front of it, and the same applies below.
[0034] The names of the various waveform data obtained after step 2 are shown in the fourth column of Table 2 below.
[0035] Step 3: For each experimental result corresponding to the wave interval time value t obtained in step 1, the mixed waveform data APall obtained in experimental sub-step ② minus the nerve trunk AP waveform data AP1 induced by the first pulse electrical stimulation of experimental sub-step ② obtained in step 2 is obtained to obtain the nerve trunk AP waveform data AP2 induced by the second pulse of experimental sub-step ②; Step 3 is as follows: The mixed waveform data APall_002 obtained in the above experimental sub-step 2 is subtracted from the nerve trunk AP waveform data AP1_002 induced by the first pulse electrical stimulation of the experimental sub-step 2 obtained in step 2, to obtain the nerve trunk AP waveform data AP2_002 induced by the second pulse of the experimental sub-step 2, AP2_002 = APall_002 - AP1_002; The mixed waveform data APall_004 obtained in the above experimental sub-step 4 is subtracted from the nerve trunk AP waveform data AP1_004 induced by the first pulse electrical stimulation of the experimental sub-step 4 obtained in step 2, to obtain the nerve trunk AP waveform data AP2_004 induced by the second pulse of the experimental sub-step 4, AP2_004 = APall_004 - AP1_004; The mixed waveform data APall_006 obtained in the above experimental sub-step 6 is subtracted from the nerve trunk AP waveform data AP1_006 induced by the first pulse electrical stimulation of the experimental sub-step 6 obtained in step 2, to obtain the nerve trunk AP waveform data AP2_006 induced by the second pulse of the experimental sub-step 6, AP2_006 = APall_006 - AP1_006; Similarly, the AP waveform data APall_2n of the nerve trunk induced by the first pulse electrical stimulation of the experimental sub-step 2n obtained in step 2 is subtracted from the AP waveform data AP1_2n of the nerve trunk induced by the first pulse electrical stimulation of the experimental sub-step 2n obtained in step 2 to obtain the AP waveform data AP2_2n of the nerve trunk induced by the second pulse of the experimental sub-step 2n, AP2_2n = APall_2n - AP1_2n; wherein n is the serial number of the wave interval time value t in Table 1, and the maximum value is 064; for example Figure 4 The AP waveform data APall_100 of the nerve trunk induced by the first pulse electrical stimulation of the experimental substep 100 obtained in step 2 is subtracted from the AP waveform data AP1_100 of the nerve trunk induced by the first pulse electrical stimulation of the experimental substep 100 obtained in step 2, and the AP waveform data AP2_100 of the nerve trunk induced by the second pulse of the experimental substep 100 is obtained, AP2_100=APall_100-AP1_100; Figure 5 Represents AP waveform data of nerve trunk obtained by double pulse stimulation with multiple different wave interval time conditions APall ( Figure 5 The red waveform data on the left) minus the AP waveform data AP1 induced by the first pulse of electrical stimulation ( Figure 5 The middle blue waveform data) obtains the AP waveform data AP2 of the nerve trunk induced by the second pulse ( Figure 5 Brown waveform data on the right).
[0036] The names of the waveform data obtained after step 3 are shown in the sixth column of Table 2 below; through steps 2 and 3, each mixed waveform data APall_2n is split into two complete nerve trunk action potential waveforms AP1_2n and AP2_2n respectively.
[0037] Step 4: for each result of step 2 and step 3 corresponding to the time value t of each wave interval, measure the maximum amplitude AAP1 of the AP waveform data of the nerve trunk induced by the first pulse of the experimental sub-step ② obtained in step 2, and measure the maximum amplitude AAP2 of the AP waveform data of the nerve trunk induced by the second pulse of the experimental sub-step ② obtained in step 3. The maximum amplitude of the AP waveform data of the nerve trunk induced by the first pulse is referred to as the "maximum amplitude of the first pulse", and the maximum amplitude of the AP waveform data of the nerve trunk induced by the second pulse is referred to as the "maximum amplitude of the second pulse". Calculate the ratio AAP2 / AAP1 of "maximum amplitude of the second pulse / maximum amplitude of the first pulse" corresponding to each double pulse, and the ratio of "maximum amplitude of the second pulse / maximum amplitude of the first pulse" is referred to as "maximum amplitude ratio", thereby obtaining the corresponding "maximum amplitude ratio" under the condition that the double pulse wave interval is t; Step 4 is as follows: Measure the maximum amplitude AAP1_002 of the AP waveform data of the nerve trunk induced by the first pulse of experimental sub-step 2 obtained in step 2, and measure the maximum amplitude AAP2_002 of the AP waveform data of the nerve trunk induced by the second pulse of experimental sub-step 2 obtained in step 3, and calculate the ratio of "maximum amplitude of the second pulse / maximum amplitude of the first pulse" corresponding to experimental sub-step 2, that is, the ratio of AAP2_002 / AAP1_002, thereby obtaining the corresponding "maximum amplitude ratio" under the condition that the double pulse wave interval is 30ms; Measure the maximum amplitude AAP1_004 of the AP waveform data of the nerve trunk induced by the first pulse of experimental sub-step 4 obtained in step 2, and measure the maximum amplitude AAP2_004 of the AP waveform data of the nerve trunk induced by the second pulse of experimental sub-step 4 obtained in step 3, and calculate the ratio of "maximum amplitude of the second pulse / maximum amplitude of the first pulse" corresponding to experimental sub-step 4, that is, the ratio of AAP2_004 / AAP1_004, thereby obtaining the corresponding "maximum amplitude ratio" under the condition that the double pulse wave interval is 29ms; Measure the maximum amplitude AAP1_006 of the AP waveform data of the nerve trunk induced by the first pulse of experimental sub-step 6 obtained in step 2, and measure the maximum amplitude AAP2_006 of the AP waveform data of the nerve trunk induced by the second pulse of experimental sub-step 6 obtained in step 3, and calculate the ratio of "maximum amplitude of the second pulse / maximum amplitude of the first pulse" corresponding to experimental sub-step 6, that is, the ratio of AAP2_006 / AAP1_006, thereby obtaining the corresponding "maximum amplitude ratio" under the condition that the double pulse wave interval is 28ms; Similarly, the maximum amplitude AAP1_2n of the AP waveform data of the nerve trunk induced by the first pulse of the experimental sub-step 2n obtained in step 2 is measured, and the maximum amplitude AAP2_2n of the AP waveform data of the nerve trunk induced by the second pulse of the experimental sub-step 2n obtained in step 3 is measured, and the ratio of "the maximum amplitude of the second pulse / the maximum amplitude of the first pulse" corresponding to the experimental sub-step 2n is calculated, that is, the ratio of AAP2_2n / AAP1_2n, thereby obtaining the double pulse wave interval tn The corresponding "maximum amplitude ratio" under the condition of; Where n is the sorting number of the wave interval time value t in Table 1, and the maximum value is 064; After completing step 4, the data in columns 5, 7, and 8 of Table 2 below are obtained:
[0038] Step 5. Based on the results obtained in step 4, with the wave interval time value t (the duration value in the third column, third sub-column of Table 2) as the horizontal coordinate and the "maximum amplitude ratio" (the value in the eighth column of Table 2) as the vertical coordinate, the discrete points corresponding to each wave interval time value t are plotted in the plane rectangular coordinate system to form a coordinate diagram representing the relationship between the wave interval time value t and the "maximum amplitude ratio", as shown below: Figure 6 shown.
[0039] Step 6. Based on the coordinate graph obtained in step 5, determine the critical value range between the non-refractory period and the relative refractory period according to the following rules: along the direction of the wave interval time value t from small to large, the wave interval time value corresponding to the discrete point where the "maximum amplitude ratio" is not less than 100% for the first time is determined as the maximum value of the critical value range between the non-refractory period and the relative refractory period, and the wave interval time value corresponding to the adjacent discrete point whose wave interval time value is smaller than the discrete point is determined as the minimum value of the critical value range between the non-refractory period and the relative refractory period; Specifically, in Figure 6 In the direction from small to large wave interval time value t (i.e. Figure 6 In the direction from left to right in the graph), the discrete point whose "maximum amplitude ratio" is not less than 100% for the first time is the 32nd discrete point from left to right, and the wave interval time value corresponding to the 32nd discrete point is 11.5ms, and the adjacent discrete point whose wave interval time value is smaller than the 32nd discrete point is the 31st discrete point from left to right, and the wave interval time value corresponding to the 31st discrete point from left to right is 11.0ms. Based on this, 11.5ms is defined as the maximum value of the critical value range between the two time periods of the non-refractory period and the relative refractory period, and 11.0ms is defined as the minimum value of the critical value range between the two time periods of the non-refractory period and the relative refractory period, that is, the critical value range between the two time periods of the non-refractory period and the relative refractory period is 11.0ms to 11.5ms. Figure 6 As indicated by arrow b.
[0040] Step 7. Based on the coordinate graph obtained in step 5, determine the critical value range between the absolute refractory period and the relative refractory period according to the following rules: along the direction of the wave interval time value t from large to small, the wave interval time value corresponding to the discrete point where the "maximum amplitude ratio" is not greater than 0% for the first time is determined as the minimum value of the critical value range between the absolute refractory period and the relative refractory period, and the wave interval time value corresponding to the adjacent discrete point whose wave interval time value is larger than the discrete point is determined as the maximum value of the critical value range between the absolute refractory period and the relative refractory period; Specifically, in Figure 6 In the direction from large to small wave interval time value t (i.e. Figure 6In the direction from right to left in the figure), the discrete point where the "maximum amplitude ratio" is not greater than 0% for the first time is the 62nd discrete point from right to left, and the wave interval time value corresponding to the 62nd discrete point from right to left is 1.0ms, and the adjacent discrete point whose wave interval time value is larger than the 62nd discrete point from right to left is the 61st discrete point from right to left, and the wave interval time value corresponding to the 61st discrete point from right to left is 1.2ms. Based on this, 1.0ms is defined as the minimum value of the critical value range between the absolute refractory period and the relative refractory period, and 1.2ms is defined as the maximum value of the critical value range between the absolute refractory period and the relative refractory period, that is, the critical value range between the absolute refractory period and the relative refractory period is 1.0ms to 1.2ms. Figure 6 As indicated by arrow a.
[0041] In the above embodiment, as long as the points of the double pulse wave interval time value t selected in step 1 are dense enough, the critical value range obtained in step 6 and step 7 is small enough, and the judgment result is accurate enough. The density of the points of the double pulse wave interval time value t selected in step 1 is determined according to the accuracy requirement of the experimental results.
[0042] In the above embodiment, the interval time between experimental sub-step ① and experimental sub-step ② can be changed to 1 second, or to 10 seconds; the interval time between experimental sub-step ② and experimental sub-step ③ is changed accordingly.
Claims
1. A quantitative experimental research and analysis method for the refractory period phenomenon of nerve stem action potential, comprising the following steps: Step 1: Select multiple different double pulse wave interval time values t as experimental conditions, and the wave interval time values t are from large to small, and the selected maximum wave interval time value is within the stage range of no refractory period phenomenon, and the selected minimum wave interval time value is within the stage range of absolute refractory period; for each wave interval time value t, a "single-double-single" electrical stimulation series experiment is carried out, which includes the following experimental sub-steps in sequence: ① Apply a single pulse electrical stimulation to the nerve trunk specimen, and record the nerve trunk AP waveform data AP1 induced by the single pulse electrical stimulation; ② Apply a wave interval to the nerve trunk specimen ③ Apply a double pulse electrical stimulation with a time value of t, and record the AP mixed waveform data APall of the nerve trunk induced by the double pulse electrical stimulation; ③ Apply a single pulse electrical stimulation to the nerve trunk specimen again, and record the AP waveform data AP1 of the nerve trunk induced by the single pulse electrical stimulation; The interval time between the above experimental sub-steps ① and ② is equal to the interval time between experimental sub-steps ② and ③, and the interval time is 1 to 10 seconds; Each pulse in the above experimental sub-steps ①, ② and ③ is a square wave, and the amplitude and wave width of all the square wave electrical stimulations are the same; Step 2: for each experimental result corresponding to the wave interval time value t obtained in step 1, the nerve trunk AP waveform data AP1 obtained in experimental sub-step ① and the nerve trunk AP waveform data AP1 obtained in experimental sub-step ③ are superimposed and divided by 2 to obtain the superimposed average waveform data of the nerve trunk AP induced by the two single-pulse electrical stimulations in experimental sub-steps ① and ③, and the nerve trunk AP superimposed average waveform data is used to characterize the nerve trunk AP waveform data AP1 induced by the first pulse electrical stimulation in the double-pulse electrical stimulation in experimental sub-step ②; Step 3: For each experimental result corresponding to the wave interval time value t obtained in step 1, the mixed waveform data APall obtained in experimental sub-step ② minus the nerve trunk AP waveform data AP1 induced by the first pulse electrical stimulation of experimental sub-step ② obtained in step 2 is obtained to obtain the nerve trunk AP waveform data AP2 induced by the second pulse of experimental sub-step ②; Step 4: for each result of step 2 and step 3 corresponding to each wave interval time value t, measure the maximum amplitude AAP1 of the AP waveform data of the nerve trunk induced by the first pulse of the experimental sub-step ② obtained in step 2, and measure the maximum amplitude AAP2 of the AP waveform data of the nerve trunk induced by the second pulse of the experimental sub-step ② obtained in step 3, wherein the maximum amplitude of the AP waveform data of the nerve trunk induced by the first pulse is referred to as the "maximum amplitude of the first pulse", and the maximum amplitude of the AP waveform data of the nerve trunk induced by the second pulse is referred to as the "maximum amplitude of the second pulse", and the ratio AAP2 / AAP1 of "maximum amplitude of the second pulse / maximum amplitude of the first pulse" corresponding to each double pulse is calculated, and the ratio of "maximum amplitude of the second pulse / maximum amplitude of the first pulse" is referred to as the "maximum amplitude ratio", thereby obtaining the corresponding "maximum amplitude ratio" under the condition that the double pulse wave interval time value is t; Step 5: Based on the result obtained in step 4, with the wave interval time value t as the horizontal coordinate and the "maximum amplitude ratio" as the vertical coordinate, the discrete points corresponding to each wave interval time value t are drawn in the plane rectangular coordinate system to form a coordinate diagram showing the relationship between the wave interval time value t and the "maximum amplitude ratio"; Step 6. Based on the coordinate graph obtained in step 5, determine the critical value range between the non-refractory period and the relative refractory period according to the following rules: along the direction from small to large wave interval time value t, the wave interval time value corresponding to the discrete point where the "maximum amplitude ratio" is not less than 100% for the first time is determined as the maximum value of the critical value range between the non-refractory period and the relative refractory period, and the wave interval time value corresponding to the adjacent discrete point whose wave interval time value is smaller than the discrete point is determined as the minimum value of the critical value range between the non-refractory period and the relative refractory period; Step 7. Based on the coordinate graph obtained in step 5, determine the critical value range between the absolute refractory period and the relative refractory period according to the following rules: along the direction of the wave interval time value t from large to small, the wave interval time value corresponding to the discrete point where the "maximum amplitude ratio" is not greater than 0% for the first time is defined as the minimum value of the critical value range between the absolute refractory period and the relative refractory period, and the wave interval time value corresponding to the adjacent discrete point whose wave interval time value is larger than that of the discrete point is defined as the maximum value of the critical value range between the absolute refractory period and the relative refractory period.
2. The quantitative experimental research and analysis method of the AP refractory period phenomenon of a nerve trunk according to claim 1, characterized in that: In the "single-double-single" electrical stimulation series experiments corresponding to two adjacent wave interval time values t, the experimental sub-step ③ corresponding to the previous wave interval time value t is used as the experimental sub-step ① corresponding to the next wave interval time value t.