Acoustic escape reaction detection device and method for detecting functions of fish hair cells

Abstract

The invention belongs to the technical field of biology, and particularly relates to an acoustic escape reaction detection device and a method for detecting zebra fish hair cell functions. The acoustic escape reaction detection device provided by the invention comprises the optical detection module, the illumination module, the culture module, the sound stimulation module and the transmission module, and special parameter design is carried out for the illumination module, so that a new method for detecting a to-be-detected sample by using the acoustic escape reaction is provided; the acoustic escape reaction detection device provided by the invention can more sensitively reflect the condition of functional damage of the sample, and is more convenient and faster to operate.

Description

Technical Field

[0001] This application belongs to the field of biotechnology, and specifically relates to an acoustic escape response detection device and a method for detecting the function of fish hair cells. Background Art

[0002] Zebrafish (Danio rerio) is one of the commonly used model animals. Due to its many advantages such as small size, convenient cultivation, short life cycle, many offspring, transparent embryos throughout the body, and high similarity to the human genome, it is widely used in scientific research, environmental pollutant, cosmetic, and drug screening fields. Among them, zebrafish have two types of hair cells. One type is located in the inner ear, which is similar to human cochlear hair cells and plays a key role in the formation of the vestibule and hearing. The other type is located in the neuromasts of the lateral line system on both sides of the body. The neuromast hair cells are similar in structure and function to the inner ear hair cells, and also have functions such as sensing and maintaining body balance. They can sense signals such as water temperature, water flow, and sound, and transmit these signals to the central nervous system.

[0003] Based on the above characteristics, zebrafish have also become an important model animal for studying the function of hair cells, regeneration mechanism, and deafness mechanism. At present, many clinical drugs, small molecule inhibitors, environmental pollutants, etc. have ototoxicity, which can cause irreversible damage to hair cells, and then lead to hearing loss or loss; this seriously affects the quality of life of patients. Therefore, how to efficiently and accurately screen out ototoxic reagents, drugs, and otoprotective agents, and how to formulate the optimal concentration of drug use to minimize ototoxicity as much as possible has become a hot topic of concern for many biomedical researchers.

[0004] Currently, in traditional technology, the detection of hair cell function is mainly evaluated by DASPEI staining. This evaluation method has the following problems: it is only applicable to model animals, usually requires staining, photographing one by one, and then counting the number of cells and fluorescence intensity, with low efficiency, large workload, and long process. In addition, it is also unable to evaluate minor damage or early damage with no significant damage to the morphology and number of hair cells, that is, it is only applicable to organic damage and unable to evaluate functional damage. Summary of the Invention

[0005] Based on this, an embodiment of this application provides an acoustic escape response detection device and a method for detecting the function of fish hair cells.

[0006] This application provides an acoustic escape response detection device on the one hand, including an optical detection module, an illumination module, a culture module, a sound stimulation module, and a transmission module;

[0007] The culture module is used to load the test substance; the optical detection module is used to record the motion data of the test substance in the culture module;

[0008] The sound stimulation module is used to stimulate the object to be tested in the culture module by generating sound;

[0009] The transmission module is used to transmit the motion information detected by the optical detection module;

[0010] The illumination module includes an infrared light source.

[0011] In one embodiment, the outer diameter of the infrared light source is 110 mm to 130 mm.

[0012] In one embodiment, the inner diameter of the infrared light source is 70 mm to 90 mm.

[0013] In one embodiment, the wavelength band of the infrared light source is 840 nm to 860 nm.

[0014] In one embodiment, the infrared light source can deflect towards the central area of the culture module at an angle of 25° to 35°.

[0015] In one embodiment, the distance between the infrared light source and the culture module is 3.5 cm to 5.5 cm.

[0016] In one embodiment, the optical detection module includes a motion camera.

[0017] In one embodiment, the distance between the motion camera and the culture module is 8 cm to 9 cm.

[0018] In one embodiment, the focal length of the lens of the motion camera is 2.8 mm to 12 mm.

[0019] In one embodiment, the resolution of the motion camera is 2.8 MP to 3.2 MP.

[0020] In one embodiment, the recording frequency of the motion camera is 0.1 ms / frame to 33 ms / frame.

[0021] In one embodiment, the frequency of the sound stimulation module is 50 Hz to 100 Hz.

[0022] In one embodiment, the loudness is 28 dB to 38 dB.

[0023] In one embodiment, the interval between stimulations is 2 min to 4 min, and the number of stimulations is 8 to 12 times.

[0024] In one embodiment, the sound stimulation module includes a micro vibration device.

[0025] In one embodiment, the micro vibration device is an audio signal board.

[0026] In one embodiment, the culture module includes a culture container, and the diameter of the culture container is 6 cm to 10 cm.

[0027] In one embodiment, a metal disk with a thickness of 5 mm to 2.5 mm is further provided at the bottom of the culture container.

[0028] On the other hand, the present application provides a method for detecting the function of fish hair cells, including:

[0029] Using the above-mentioned acoustic escape response detection device to detect the function of the target zebrafish hair cells.

[0030] In one embodiment, it includes the following steps:

[0031] Controlling the operation of the optical detection module and the lighting module;

[0032] Placing the fish in the culture module, controlling the operation of the sound stimulation module to perform sound stimulation on the zebrafish; controlling the optical detection module to detect the movement information of the fish; and,

[0033] Controlling the transmission module to output data, and analyzing the function of fish hair cells according to the data analysis.

[0034] In one embodiment, the function of fish hair cells is analyzed according to the relative value of the movement information of the fish to be tested and the control fish.

[0035] In one embodiment, the fish includes zebrafish.

[0036] In one embodiment, the movement information includes one or more of the number of fish that have a reaction, the escape distance of each fish, and the C-type reaction.

[0037] On the other hand, the present application provides the application of the above-mentioned acoustic escape response detection device in drug screening.

[0038] In one embodiment, the drug includes one or two of otoprotective drugs and ototoxic drugs.

[0039] Details of one or more embodiments of the present application are set forth in the following description, and other features, objects, and advantages of the present application will become apparent from the specification and its claims. Description of the Drawings

[0040] To more clearly illustrate the technical solutions in the embodiments of the present application and more comprehensively understand the present application and its beneficial effects, the following will briefly introduce the attached drawings required for the description of the embodiments. Obviously, the attached drawings in the following description are only some embodiments of the present application. For those skilled in the art, without creative efforts, other attached drawings can be obtained based on these drawings.

[0041] Figure 1 Structural schematic diagram of the acoustic escape response detection device of the present application;

[0042] Figure 2 Signal reception schematic diagram of the transmission module of the acoustic escape response detection device of the present application;

[0043] Figure 3 Data result diagram of the zebrafish DASPEI staining method;

[0044] Figure 4 Data result diagram detected by the acoustic escape response detection device provided by the present application;

[0045] Figure 5 Detection results of various reagents by the staining method and the acoustic escape response detection device;

[0046] Figure 6 Detection results of various drugs by the acoustic escape response detection device provided by the present application.

[0047] Host: Host; wireless: Radio; receiver: Receiver; camera recording: Camera recording; soundcard: Sound card; Infrared indicator: Infrared indicator; acoustic escape response: Acoustic escape response; velocity max relative to control: Maximum velocity relative to the control; distancerelative to control; Distance relative to the control; concentration of cisplatin: Cisplatin concentration; number of labelled hair cells: Number of labeled hair cells.

[0048] Explanation of the reference numerals in the attached drawings: 10. Acoustic escape response detection device; 100. Optical detection module; 200. Lighting module; 300. Culture module; 400. Sound stimulation module; 500. Transmission module. Detailed implementation manners

[0049] The present application will be further described in detail below in conjunction with embodiments and examples. It should be understood that these embodiments and examples are only used to illustrate the present application and not to limit the scope of the present application. The purpose of providing these embodiments and examples is to make the disclosure of the present application more thoroughly and comprehensively understood. It should also be understood that the present application can be implemented in many different forms and is not limited to the embodiments and examples described herein. Those skilled in the art can make various changes or modifications without departing from the connotation of the present application, and the equivalent forms obtained also fall within the protection scope of the present application. In addition, in the following description, a large number of specific details are given to provide a more thorough understanding of the present application. It should be understood that the present application can be implemented without one or more of these details.

[0050] Unless otherwise defined, all technical and scientific terms used in the present application have the same meaning as commonly understood by those skilled in the technical field to which the present application belongs.

[0051] The term

[0052] Unless otherwise stated or there is a contradiction, the terms or phrases used herein have the following meanings:

[0053] The term "and / or", "or / and", "and / or" used herein includes any one of two or more related listed items, and also includes any and all combinations of the related listed items. The said any and all combinations include any two related listed items, any more related listed items, or the combination of all related listed items. It should be noted that when at least three items are connected by at least two conjunctions selected from "and / or", "or / and", "and / or", it should be understood that in the present application, this technical solution undoubtedly includes the technical solution connected by "logical AND", and also undoubtedly includes the technical solution connected by "logical OR". For example, "A and / or B" includes three parallel solutions: A, B, and A + B. Another example, the technical solution of "A, and / or, B, and / or, C, and / or, D" includes any one of A, B, C, D (that is, the technical solution connected by "logical OR"), and also includes any and all combinations of A, B, C, D, that is, it includes the combination of any two or any three of A, B, C, D, and also includes the combination of the four items of A, B, C, D (that is, the technical solution connected by "logical AND").

[0054] In the present application, terms such as "multiple", "multiple types", "multiple times", "multiple elements", etc., unless otherwise specified, refer to a quantity greater than 2 or equal to 2. For example, "one or more" means one or greater than or equal to two.

[0055] As used herein, "its combination", "any combination thereof", "any combination mode thereof", etc. include all suitable combination modes of any two or more of the listed items.

[0056] In this application, the "suitable" in "suitable combination mode", "suitable mode", "any suitable mode", etc. is subject to being able to implement the technical solution of this application, solve the technical problems of this application, and achieve the expected technical effects of this application.

[0057] In this application, "further", "even further", "especially", etc. are used for descriptive purposes, indicating differences in content, but should not be construed as limiting the protection scope of this application.

[0058] In this application, "optionally", "optional", "option", mean that it can be either present or absent, that is, it refers to any one of the two alternative schemes of "present" or "absent". If "optional" appears multiple times in a technical solution, without special instructions and without contradictions or mutual restrictions, each "optional" is independent.

[0059] In this application, for the technical features described in an open-ended manner, it includes a closed technical solution composed of the listed features, and also includes an open-ended technical solution containing the listed features.

[0060] In this application, regarding a numerical interval (i.e., a numerical range), without special instructions, the optional numerical values are considered continuous within the above numerical interval, and include the two numerical endpoints (i.e., the minimum value and the maximum value) of this numerical range, as well as each numerical value between these two numerical endpoints. Without special instructions, when the numerical interval only refers to the integers within this numerical interval, it includes the two endpoint integers of this numerical range, as well as each integer between the two endpoints. In this article, it is equivalent to directly listing each integer. For example, if t is an integer selected from 1 to 10, it means that t is any integer selected from the integer group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10. In addition, when providing multiple range descriptions for features or characteristics, these ranges can be combined. In other words, unless otherwise specified, the ranges disclosed herein should be understood to include any and all sub-ranges subsumed therein.

[0061] The temperature parameter in this application, without special limitations, allows both constant temperature treatment and also allows fluctuations within a certain temperature range. It should be understood that the so-called constant temperature treatment allows the temperature to fluctuate within the accuracy range controlled by the instrument. Fluctuations within a range such as ±5°C, ±4°C, ±3°C, ±2°C, ±1°C are allowed.

[0062] In this application, both %(w / w) and wt% represent weight percentage, %(v / v) refers to volume percentage, and %(w / v) refers to mass-volume percentage.

[0063] All documents mentioned in this application are incorporated herein by reference as if each document were individually incorporated by reference. Unless it conflicts with the inventive purpose and / or technical solution of this application, the cited documents involved in this application are cited for their entire content and entire purpose. When this application involves cited documents, the definitions of relevant technical features, terms, nouns, phrases, etc. in the cited documents are also incorporated by reference. When this application involves cited documents, examples and preferred methods of the relevant technical features cited can also be incorporated into this application as references, but only to the extent that this application can be implemented. It should be understood that when the cited content conflicts with the description in this application, this application shall prevail or be amended adaptively according to the description of this application.

[0064] The term "acoustic escape response" The acoustic escape response is a rapid evasive behavior exhibited by organisms when subjected to acoustic stimuli. This response is prevalent in many animals and is an evolutionarily conserved behavioral mechanism for coping with potential threats or dangers.

[0065] The term "hair cell" Hair cells are a type of specialized sensory epithelial cells mainly present in the cochlea and vestibular system of vertebrates, responsible for converting mechanical waves (such as sound waves or vibrations generated by head movements) into neural signals, thus enabling auditory and balance functions.

[0066] The term: "dpf" In zebrafish research, dpf (Days Post Fertilization) refers to the number of days after fertilization and is used to describe the developmental stage of zebrafish embryos or larvae. For example: 1 dpf represents 1 day after fertilization; 5 dpf represents 5 days after fertilization.

[0067] It can be understood that the principle of the staining method is to evaluate the function and quantity of hair cells based on the principle that hair cells can uptake drugs or small molecule reagents through their cilia. When the cilia of hair cells are damaged or malfunction, they are unable to uptake drugs, dyes, etc., resulting in fewer stained hair cells and lighter staining; however, this method has requirements for the concentration control of dyes, staining time, shooting conditions, and shooting time. If any link is not properly handled or the conditions are not well controlled, serious errors or mistakes may occur.

[0068] The principle of the acoustic escape response provided in this application is evaluated through the ability of hair cells to sense sound signals. When a certain sound stimulus is given, the hair cells sense the sound signal and randomly convert it into an electrical signal that is transmitted to the central nervous system, and the zebrafish will respond to the corresponding sound signal, such as quickly escaping or turning around to form a C shape (commonly known as the C-type response); therefore, it can more intuitively detect whether the main functions of hair cells are affected.

[0069] As shown Figure 1 in the figure, the present application provides an acoustic escape reaction detection device 10, and the detection device includes: an optical detection module 100, an illumination module 200, a culture module 300, a sound stimulation module 400, and a transmission module 500; wherein, the culture module 300 is used to load the object to be tested; the optical detection module 100 is used to record the motion data of the object to be tested in the culture module 300; the optical detection module 100 is used to record the motion data of the object to be tested in the culture module 300; the sound stimulation module 400 is used to stimulate the object to be tested in the culture module 300; the transmission module 500 is used to transmit the detection information detected by the optical detection module 100; and the illumination module 200 includes an infrared light source.

[0070] The acoustic escape reaction detection device provided by the present application includes an optical detection module, an illumination module, a culture module, a sound stimulation module, and a transmission module, and special parameter design is carried out for the illumination module, so as to provide a new method for detecting a sample to be tested by using the acoustic escape reaction. Compared with the ordinary staining method, the acoustic escape reaction detection device provided by the present application can more sensitively reflect the functional damage of the sample, and the operation is more convenient and fast.

[0071] In some embodiments, the outer diameter of the infrared light source is 110 mm to 130 mm. For example, the diameter of the infrared light source is 110 mm, 111 mm, 112 mm, 113 mm, 114 mm, 115 mm, 116 mm, 117 mm, 118 mm, 119 mm, 120 mm, 121 mm, 122 mm, 123 mm, 124 mm, 125 mm, 126 mm, 127 mm, 128 mm, 129 mm or 130 mm, and any value in between.

[0072] In some embodiments, the inner diameter of the infrared light source is 70 mm to 90 mm. For example, the inner diameter of the infrared light source is 70 mm, 71 mm, 72 mm, 73 mm, 74 mm, 75 mm, 76 mm, 77 mm, 78 mm, 79 mm, 80 mm, 81 mm, 82 mm, 83 mm, 84 mm, 85 mm, 86 mm, 87 mm, 88 mm, 89 mm or 90 mm, and any value in between.

[0073] In some of these embodiments, the wavelength band of the infrared light source is 840 nm to 860 nm. For example, the wavelength band of the infrared light source is 840 nm, 841 nm, 842 nm, 843 nm, 844 nm, 845 nm, 846 nm, 847 nm, 848 nm, 849 nm, 850 nm, 851 nm, 852 nm, 853 nm, 854 nm, 855 nm, 856 nm, 857 nm, 858 nm, 859 nm, or 860 nm, as well as any value in between.

[0074] In some of these embodiments, the infrared light source deflects towards the central region of the culture module 300 at an angle of 25° to 35°. For example, the angle of the infrared light source is 25°, 26°, 27°, 28°, 29°, 30°, 31°, 32°, 33°, 34°, or 35°, as well as any value in between. This setting can provide uniform infrared illumination to the entire culture module 300, reduce the visual interference of light on the zebrafish in the petri dish with each other, and ensure the quality of the captured images.

[0075] In some of these embodiments, the distance between the infrared light source and the culture module 300 is 3.5 cm to 5.5 cm. For example, the distance is 3.5 cm, 3.6 cm, 3.7 cm, 3.8 cm, 3.9 cm, 4.0 cm, 4.1 cm, 4.2 cm, 4.3 cm, 4.4 cm, 4.5 cm, 4.6 cm, 4.7 cm, 4.8 cm, 4.9 cm, 5.0 cm, 5.1 cm, 5.2 cm, 5.3 cm, 5.4 cm, or 5.5 cm, as well as any value in between.

[0076] In some of these embodiments, the optical detection module 100 includes a motion camera. Among them, the model of the motion camera is not particularly limited. For example, the Hikvision S6 motion camera is used.

[0077] In some of these embodiments, the distance between the motion camera and the culture module 300 is 8 cm to 9 cm. For example, the distance is 8.0 cm, 8.1 cm, 8.2 cm, 8.3 cm, 8.4 cm, 8.5 cm, 8.6 cm, 8.7 cm, 8.8 cm, 8.9 cm, or 9.0 cm, as well as any value in between.

[0078] In some of these embodiments, the focal length of the lens of the action camera is 2.8 mm to 12 mm. For example, the focal length is 2.8 mm, 3.0 mm, 3.2 mm, 3.4 mm, 3.6 mm, 3.8 mm, 4.0 mm, 4.2 mm, 4.4 mm, 4.6 mm, 4.8 mm, 5.0 mm, 5.2 mm, 5.4 mm, 5.6 mm, 5.8 mm, 6.0 mm, 6.2 mm, 6.4 mm, 6.6 mm, 6.8 mm, 7.0 mm, 7.2 mm, 7.4 mm, 7.6 mm, 7.8 mm, 8.0 mm, 8.2 mm, 8.4 mm, 8.6 mm, 8.8 mm, 9.0 mm, 9.2 mm, 9.4 mm, 9.6 mm, 9.8 mm, 10.0 mm, 10.2 mm, 10.4 mm, 10.6 mm, 10.8 mm, 11.0 mm, 11.2 mm, 11.4 mm, 11.6 mm, 11.8 mm or 12.0 mm, as well as any value in between.

[0079] In some of these embodiments, the resolution of the action camera is 2.8 MP to 3.2 MP. For example, the resolution is 2.8 MP, 2.9 MP, 3.0 MP, 3.1 MP or 3.2 MP, as well as any value in between.

[0080] In some of these embodiments, the recording frequency of the action camera is 0.1 ms / frame to 33 ms / frame. For example, the frequency is 0.1 ms / frame, 0.2 ms / frame or 0.3 ms / frame, 1 ms / frame, 2 ms / frame, 3 ms / frame, 4 ms / frame, 5 ms / frame, 6 ms / frame, 7 ms / frame, 8 ms / frame, 9 ms / frame, 10 ms / frame, 11 ms / frame, 12 ms / frame, 13 ms / frame, 14 ms / frame, 15 ms / frame, 16 ms / frame, 17 ms / frame, 18 ms / frame, 19 ms / frame, 20 ms / frame, 21 ms / frame, 22 ms / frame, 23 ms / frame, 24 ms / frame, 25 ms / frame, 26 ms / frame, 27 ms / frame, 28 ms / frame, 29 ms / frame, 30 ms / frame, 31 ms / frame, 32 ms / frame or 33 ms / frame, as well as any value in between.

[0081] In some of these embodiments, the sound stimulation module is a micro vibration device.

[0082] In some of these embodiments, the micro vibration device is an audio signal board.

[0083] For example, the input signal of the micro vibrator is generated by an audio signal board (STM32F407VGT6). The signal board transmits data to the host through a wireless receiver. Previously, a USB cable was used in the original article; this greatly improves the transmission rate and ensures lossless audio quality. The schematic diagram of the receiving end is as Figure 2as shown

[0084] In some embodiments, the frequency of the sound stimulation module 400 is 50 Hz to 100 Hz. For example, the frequency is 50 Hz, 51 Hz, 52 Hz, 53 Hz, 54 Hz, 55 Hz, 56 Hz, 57 Hz, 58 Hz, 59 Hz, 60 Hz, 61 Hz, 62 Hz, 63 Hz, 64 Hz, 65 Hz, 66 Hz, 67 Hz, 68 Hz, 69 Hz, 70 Hz, 71 Hz, 72 Hz, 73 Hz, 74 Hz, 75 Hz, 76 Hz, 77 Hz, 78 Hz, 79 Hz, 80 Hz, 81 Hz, 82 Hz, 83 Hz, 84 Hz, 85 Hz, 86 Hz, 87 Hz, 88 Hz, 89 Hz, 90 Hz, 91 Hz, 92 Hz, 93 Hz, 94 Hz, 95 Hz, 96 Hz, 97 Hz, 98 Hz, 99 Hz or 100 Hz and any value in between.

[0085] In some embodiments, the loudness of the sound stimulation module 400 is 28 dB to 38 dB; for example, the loudness is 28 dB, 29 dB, 30 dB, 31 dB, 32 dB, 33 dB, 34 dB, 35 dB, 36 dB, 37 dB or 38 dB and any value in between.

[0086] In some embodiments, the interval of the stimulation of the sound stimulation module 400 is 2 min to 4 min, and the number of stimulations is 8 to 12 times. For example, the interval is 2 min, 3 min or 4 min, and the number of stimulations is 8 times, 9 times, 10 times, 11 times or 12 times.

[0087] In some embodiments, the culture module 300 includes a culture container, and the diameter of the culture container is 6 cm to 10 cm. It can be understood that the culture container is, for example, a cell culture dish, and its diameter is 6 cm, 7 cm, 8 cm, 9 cm or 10 cm and any value in between.

[0088] In some embodiments, a metal disk with a thickness of 1.5 mm to 2.5 mm is further provided at the bottom of the culture container. For example, the thickness is 1.5 mm, 1.6 mm, 1.7 mm, 1.8 mm, 1.9 mm, 2.0 mm, 2.1 mm, 2.2 mm, 2.3 mm, 2.4 mm or 2.5 mm and any value in between. This designed fish dish ensures uniform sound conduction and has also passed the laser Doppler test, and the vibration error between the center and the edge is within 10%.

[0089] In some embodiments, the sound stimulation module 400 includes a micro vibration device.

[0090] On the other hand, the present application provides a method for detecting the function of zebrafish hair cells, including: detecting the function of the target zebrafish hair cells by using the above-mentioned acoustic escape response detection device 10.

[0091] In some embodiments, the detection method includes:

[0092] Turn on the optical detection module 100 and the illumination module 200, place the zebrafish in the culture module 300, and perform sound stimulation on the zebrafish through the sound stimulation module 400; then detect the movement information of the zebrafish through the optical detection module 100; output data through the transmission module 500, and analyze the function of the zebrafish hair cells based on the data.

[0093] The present application adopts remote wireless control. The shooting of the camera and the WIFI transmission of data are controlled by a wireless relay. The host remotely controls the relay through wireless communication technology, and the transmission of the sound signal is transmitted from the host to the sound card through Bluetooth.

[0094] In some embodiments, the movement information includes: one or more of the number of zebrafish that react, the escape distance of each zebrafish, and the C-type response.

[0095] On the other hand, the present application provides the application of the above-mentioned acoustic escape response detection device 10 in drug screening.

[0096] In some embodiments, the drug includes one or both of otoprotective drugs and ototoxic drugs.

[0097] The gene mutation samples do not require additional treatment; for reagent and drug screening, only different concentration gradients of reagents / drugs need to be prepared, and the zebrafish are soaked in them for 5 min to 30 min, and then the detection can start. The sample preparation during detection is simple and does not require additional treatment or special staining. For example: after soaking 6 dpf zebrafish larvae in neomycin (a commonly used clinical chemotherapy drug) at different concentration gradients for 30 min, 20 drug-soaked and non-drug-soaked zebrafish at the same concentration gradient can be placed on the fish tray.

[0098] The following will describe the implementation schemes of the present application in detail with reference to the embodiments. It should be understood that these embodiments are only used to illustrate the present application and not to limit the scope of the present application. For the experimental methods without specific conditions in the following embodiments, the guidance given in the present application is preferably referred to, and it can also be in accordance with the experimental manuals or conventional conditions in the art, or in accordance with the conditions recommended by the manufacturer, or refer to the experimental methods known in the art.

[0099] In the following specific embodiments, for the measurement parameters of the raw material components, if there is no special description, there may be slight deviations within the weighing accuracy range. For the temperature and time parameters, acceptable deviations caused by instrument test accuracy or operation accuracy are allowed.

[0100] It should be understood that in various embodiments of the present application, the magnitudes of the serial numbers of the above processes do not imply the order of execution. The order of execution of each process should be determined by its function and internal logic, and should not constitute any limitation to the implementation process of the embodiments of the present application.

[0101] Embodiment 1

[0102] As Figure 1 shown, this embodiment provides a detection device, which includes: an optical detection module 100, an illumination module 200, a culture module 300, a sound stimulation module 400, and a transmission module 500; the optical detection module 100 is used to record the motion data of the object to be measured in the culture module 300; the sound stimulation module 400 is used to stimulate the object to be measured in the culture module 300; the transmission module 500 is used to transmit the detection information detected by the optical detection module 100; the illumination module 200 includes an infrared light source.

[0103] Among them, the optical detection module 100 is a Hikvision S6 motion camera. The camera lens selects a 3MP zoom lens with an adjustable focal length of 2.8mm. The Hikvision S6 motion camera is placed 8.5 cm above the culture dish to ensure the best lighting conditions and imaging range.

[0104] The illumination module 200 is a circular dense infrared LED light source with an outer diameter of 120 mm, an inner diameter of 80 mm, an infrared band of 850 nm, and the infrared lamp beads are deflected at an angle of 30 degrees towards the circle. The illumination module 200 is placed 4.5 cm above the culture dish to provide uniform infrared illumination to the entire culture dish, which is used to reduce the visual interference between zebrafish in the culture dish and ensure the quality of the captured images.

[0105] The culture module 300 is designed as a fish dish with a diameter of 8 cm, and a metal plate with a thickness of 2 mm is added below. The specially designed fish dish ensures uniform sound conduction and has also passed laser Doppler testing, with the vibration error between the center and the edge within 10%.

[0106] The sound stimulation module 400 uses an audio signal board to perform sound stimulation on the object to be measured in the culture module 300.

[0107] The transmission module 500 is for remote wireless control. Among them, the shooting of the camera and the WIFI transmission of data are controlled by a wireless relay. The host remotely controls the relay through wireless communication technology, and the transmission of the sound signal is transmitted from the host to the sound card through Bluetooth.

[0108] Embodiment 2

[0109] This embodiment provides a method for detecting the function of zebrafish hair cells, including the following steps:

[0110] 1. Turn on the acoustic escape response detection device 10 provided in this application.

[0111] 2. Place the zebrafish in a culture container.

[0112] 3. Use an audio signal board (STM32F407VGT6) to give the zebrafish a sound stimulus of 75 Hz and 32 dB, stimulate once every 3 minutes, stimulate a total of 10 times, and take the average value of the 10 data for statistical calculation.

[0113] 4. Through the optical detection module 100, record the response of each zebrafish in the culture container within 100 ms (one frame every 0.2 ms) when the sound stimulus is emitted.

[0114] 5. Count the number of zebrafish that react in the culture container and the escape distance of each, and analyze and form statistical data.

[0115] Among them, the schematic diagram of the ordinary staining method is as Figure 3 shown, and the schematic diagram of the acoustic escape method of this application is as Figure 4 shown.

[0116] Example 3

[0117] This example provides a method for verifying whether a reagent has otoprotective properties, including the following steps:

[0118] 1. Turn on the acoustic escape response detection device 10 provided in this application.

[0119] 2. Prepare test reagents of 10 μM quinine, 10 μM d-tubocurarine (d-tu), 10 μM acetylsalicylic acid (asa), 10 μM acetylcholine (ach), 5 μM isradipine, and 10 μM xe991.

[0120] 3. Immerse the zebrafish in it for 30 minutes, and then place the zebrafish in a culture container.

[0121] 4. Use an audio signal board (STM32F407VGT6) to give the zebrafish a sound stimulus of 75 Hz and 32 dB, stimulate once every 3 minutes, stimulate a total of 10 times, and take the average value of the 10 data for statistics and calculation.

[0122] 5. Select a 3MP zoom lens with an adjustable focal length of 2.8 - 12 mm through the camera lens of the Hikvision S6 motion camera to record the response of each zebrafish in the culture container within 100 ms (one frame every 0.2 ms) when the sound stimulus is emitted.

[0123] 6. Count the number of reactions occurring in the culture vessel and the escape distance of each reaction, analyze and form statistical data, and the results are as Figure 5 shown, Figure 5 where A in Figure 5 is the data obtained by using the acoustic escape reaction detection device 10 of the present application, Figure 5 B in

[0124] Example 4

[0125] This example provides a method for verifying whether an ototoxic drug has otoprotective properties, including the following steps:

[0126] 1. Turn on the acoustic escape reaction detection device 10 provided by the present application.

[0127] 2. Provide the drug to be tested and divide it into the following groups:

[0128] A: Control, Cisplatin 1000 μM, Edaravon 750 μM + Cisplatin 1000 μM, Quercetin 100 μM + Cisplatin 1000 μM. Treatment duration: 4 hours.

[0129] B: Control, Cisplatin 500 μM, Berbamine 10 μM + Cisplatin 500 μM. Treatment duration: 6 hours.

[0130] C: Control, Cisplatin 5 μM, Kenpaullone 50 μM + Cisplatin 5 μM. Treatment duration: 20 hours.

[0131] D: Control, Cisplatin 100 μM, Benzamil 50 μM + Cisplatin 100 μM, Paroxetine 5 μM + Cisplatin 100 μM. Treatment duration: 24 hours.

[0132] E: Control, Neomycin 20 μM, Acetylcysteine 1200 μM + Neomycin 20 μM, L-Ascorbic Acid 100 μM + Neomycin 20 μM. Treatment duration: 1 hour.

[0133] F: Control, Neomycin 100 μM, Quercetin 100 μM + Neomycin 100 μM, Tannic Acid 10 μM + Neomycin 100 μM, Berbamine 25 μM + Neomycin 100 μM. Treatment duration: 1 hour.

[0134] G: Control, neomycin 100 μM, kenpaullone 50 μM + neomycin 100 μM. Treatment duration: 1 hour.

[0135] H: Control, neomycin 100 μM, tamoxifen 10 μM + neomycin 100 μM, paroxetine 10 μM + neomycin 100 μM, benzamil 50 μM + neomycin 100 μM, loperamide 10 μM + neomycin 100 μM. Treatment duration: 1 hour.

[0136] 3. After soaking the zebrafish for the corresponding treatment duration, place the zebrafish in a culture container.

[0137] 4. Use an audio signal board (STM32F407VGT6) to give the zebrafish a sound stimulus of 75 Hz and 32 dB, stimulate once every 3 minutes, for a total of 10 times, and take the average of the 10 data for statistics and calculation.

[0138] 5. Select a 3MP zoom lens with an adjustable range of 2.8 - 12 mm through the lens of the Hikvision S6 motion camera to record the response of each zebrafish in the culture container within 100 ms (one frame every 0.2 ms) when the sound stimulus is issued.

[0139] 6. Count the number of zebrafish that respond in the culture container and the escape distance of each, analyze and form statistical data, and the results are as Figure 6 shown. The above-described embodiments only represent several implementation manners of the present application, which are convenient for understanding the technical solutions of the present application specifically and in detail, but should not be construed as limiting the scope of patent protection of the application. It should be noted that for those of ordinary skill in the art, without departing from the concept of the present application, several modifications and improvements can be made, and these all belong to the protection scope of the present application. In addition, it should be understood that after reading the above teachings of the present application, those skilled in the art can make various changes or modifications to the present application, and the equivalent forms obtained also fall within the protection scope of the present application. It should also be understood that the technical solutions obtained by those skilled in the art through logical analysis, reasoning or limited experiments based on the technical solutions provided by the present application are all within the protection scope of the appended claims of the present application. Therefore, the protection scope of the patent of the present application should be subject to the content of the appended claims, and the specification can be used to explain the content of the claims.

Claims

1. An acoustic escape response detection device, characterized in that, It includes an optical detection module, a lighting module, a culture module, a sound stimulation module, and a transmission module; The culture module is used to load the object to be tested; the optical detection module is used to record the movement data of the object to be tested in the culture module; The sound stimulation module is used to stimulate the object to be tested in the culture module by generating sound; The transmission module is used to transmit the movement information detected by the optical detection module; The lighting module includes an infrared light source, and the infrared light source satisfies one or more of the following conditions: (1) The outer diameter of the infrared light source is 110 mm to 130 mm; (2) The inner diameter of the infrared light source is 70 mm to 90 mm; and, (3) The wavelength band of the infrared light source is 840 nm to 860 nm.

2. The acoustic escape response detection device according to claim 1, wherein The infrared light source can deflect at an angle of 25° to 35° towards the central area of the culture module; and / or, The distance between the infrared light source and the culture module is 3.5 cm to 5.5 cm.

3. The acoustic escape response detection device according to claim 1, wherein The optical detection module includes a motion camera; Optionally, the motion camera satisfies one or more of the following conditions: (1) The distance between the motion camera and the culture module is 8 cm to 9 cm; (2) The focal length of the lens of the motion camera is 2.8 mm to 12 mm; (3) The resolution of the motion camera is 2.8 MP to 3.2 MP; and, (4) The recording frequency of the motion camera is 0.1 ms / frame to 33 ms / frame.

4. The acoustic escape response detection device according to claim 1, characterized in that, The parameters of the sound stimulation module satisfy one or more of the following conditions: (1) The frequency is 50 Hz to 100 Hz; (2) The loudness is 28 dB to 38 dB; and, (3) The interval of stimulation is 2 min to 4 min, and the number of stimulations is 8 to 12 times.

5. The acoustic escape response detection device according to claim 4, characterized in that, The sound stimulation module includes a micro vibration device; Optionally, the micro vibration device is an audio signal board.

6. The acoustic escape response detection device according to any one of claims 1 to 5, characterized in that, The culture module includes a culture container, and the diameter of the culture container is 6 cm to 10 cm; Optionally, a metal disc with a thickness of 1.5 mm to 2.5 mm is further provided at the bottom of the culture container.

7. A method for detecting the function of fish hair cells, characterized in that, It includes: Using the acoustic escape response detection device according to any one of claims 1 to 6 to detect the function of the hair cells of the target fish.

8. The method for detecting the function of fish hair cells according to claim 7, characterized in that, It includes the following steps: Controlling the optical detection module and the lighting module to work; Placing the fish in the culture module, controlling the sound stimulation module to work to perform sound stimulation on the fish; controlling the optical detection module to detect the movement information of the fish; and, Controlling the transmission module to output data, and analyzing the function of the fish hair cells according to the data.

9. The method for detecting the function of fish hair cells according to claim 8, characterized in that, Analyzing the function of the fish hair cells according to the relative value of the movement information of the fish to be tested and the control fish; Optionally, the fish includes zebrafish; Optionally, the movement information includes: One or more of the number of fish that have a reaction, the escape distance of each fish, and the C-type reaction.

10. The application of the acoustic escape response detection device according to any one of claims 1 to 6 in drug screening; Optionally, the drug includes one or both of otoprotective drugs and ototoxic drugs.

Images