Sense-of-smell stimulator for functional MRI imaging and sense-of-smell analysis method

Abstract

To provide a device capable of measuring a functional MRI when a sense of smell of an experimental animal is stimulated.SOLUTION: Provided are: a sense-of-smell stimulator for a functional MRI having a sense-of-smell stimulator for functional MRI imaging having a sense-of-smell stimulation substance supply part and a sense-of-smell stimulation substance suction part; and a sense-of-smell analysis method.SELECTED DRAWING: Figure 1

Description

Technical Field

[0001] The present invention relates to a functional MRI, for example, an olfactory stimulation device and an olfactory analysis method for functional MRI targeting experimental animals.

Background Art

[0002] Magnetic resonance imaging (MRI) is widely used as a means for measuring brain activity. For conventional awake functional MRI targeting small animals such as marmosets, mice, and rats, there are few examples of implementation without surgery using a small-bore MRI device with a bore diameter of less than 30 cm.

[0003] Among these, Patent Document 1 describes a bed for functional MRI and a bed system for functional MRI equipped with the same.

[0004] Except for some mammals, the olfactory system has a main pathway and an accessory pathway. The main pathway mainly senses the odor of substances. The time for olfactory nerves to respond to odor substances is relatively short, and when exposed to the same odor substance for a long time, the signal gradually attenuates (olfactory adaptation). The accessory pathway senses pheromones and is known to not only have an immediate response but also be less likely to undergo olfactory adaptation.

[0005] In recent years, the relationship between diseases and olfaction has attracted attention. For example, Parkinson's disease is a neurodegenerative disease associated with the degeneration of dopaminergic neurons in the substantia nigra, and the accumulation of alpha-synuclein in dopamine-producing cells is considered to be the cause. Since the accumulation of this abnormal protein starts from the olfactory bulb, recent studies have reported cases where olfactory impairment can occur before the appearance of cognitive impairment and motor symptoms associated with Parkinson's disease. Also, it has recently been found that Alzheimer's disease also has olfactory impairment before the onset of cognitive impairment. Therefore, if olfactory impairment can be evaluated, it can assist in the early diagnosis of Parkinson's disease and Alzheimer's disease. Similarly, it is known that sensory nerve disorders such as early olfactory impairment also occur in Lewy body dementia and COVID-19.

[0006] As a conventional olfactory meter, there is a reported example of a computer-controlled olfactory meter (Non-Patent Document 1). This is a device equipped with a nose piece for delivering odorant substances to the human nasal cavity. A check valve is adopted in the nose piece to prevent the leakage of odorant substances. However, this device is not made assuming an MRI device, assuming a distance of 2.5 m between the device and the nose, and not considering placing the device immediately in front of the nose (inside the MRI device), so there is no non-magnetic device. Also, in the case of a semi-closed space like a small-bore MRI device, there is no suction, resulting in the retention of odors.

[0007] Non-Patent Document 2 reports fixing a marmoset in a holder, exposing it to odorant substances, and measuring functional MRI. Since the holder was modified from one for rats, anesthesia was required during the animal preparation stage. Also, data collection was performed when the marmoset was exposed to sex pheromones for 7 minutes. Since sex pheromones mainly stimulate the accessory olfactory pathway, the reactivity to odorant substances for a short period was not sought in this experiment.

Prior Art Documents

Patent Documents

[0008]

Patent Document 1

Non-Patent Documents

[0009]

Non-Patent Document 1

Non-Patent Document 2

Summary of the Invention

Problems to be Solved by the Invention

[0010] The inventors of the present invention have worked on functional MRI imaging when exposing experimental animals to odorant substances using a bed for functional MRI. However, in the case of intravenous administration of odorant substances, residues of the odorant substances in the MRI space and exposure at non-uniform stimulation intensities occurred, and clear results could not be obtained. In particular, in functional MRI measurements in which marmosets are fixed with a holder, exposed to an odorant substance for 30 seconds, and then a cycle of providing a 5-minute interval is repeated, after the first cycle, the odorant substance has not sufficiently disappeared from around the nasal cavity of the marmoset during the 5-minute interval, and the results when exposed to the odorant substance in the second cycle were somewhat unclear. Also, due to non-uniform stimulation intensity, it was not possible to observe the state in which the brain activity rises with high temporal resolution.

[0011] Therefore, an object of the present invention is to provide a functional MRI imaging apparatus capable of measuring olfactory stimuli, or an olfactory stimulus apparatus for functional MRI imaging, which at least partially solves the above problems.

Means for Solving the Problems

[0012] The inventors of the present invention have conducted intensive research to solve the above problems. As an example, they have found that an olfactory stimulus can be measured by a functional MRI imaging apparatus having a storage part made of a non-magnetic material, a valve for releasing the odorant substance stored in the storage part, and a suction part, and have completed the present invention including this as one embodiment.

[0013] According to the present disclosure, an odorant substance is temporarily stored in a storage part made of a non-magnetic material, then the valve made of a non-magnetic material is opened to release the stored odorant substance and expose the experimental animal, and then the odorant substance released by the suction part is suctioned to quickly disappear from around the nasal cavity of the experimental animal, whereby functional MRI can be imaged when the sense of smell is stimulated.

[0014] The present invention includes the following embodiments. [1] An olfactory stimulation device for functional MRI imaging having an olfactory stimulant supply unit and an olfactory stimulant suction unit, The olfactory stimulant supply unit includes a supply tube, a tube for temporarily holding the olfactory stimulant, a valve, a valve opening / closing unit, an olfactory stimulant storage tank, and an air compressor. The supply tube is connected to the tube for temporarily holding the olfactory stimulant, a valve is connected to the tube for temporarily holding the olfactory stimulant, the tube for temporarily holding the olfactory stimulant is connected to the olfactory stimulant storage tank, the olfactory stimulant storage tank contains the olfactory stimulant inside, and the olfactory stimulant storage tank is connected to the air compressor. The olfactory stimulant suction unit includes a suction tube and a suction pump. The suction tube is connected to the suction pump. The valve is connected to the valve opening / closing unit. The air compressor can send air into the olfactory stimulant storage tank, and the olfactory stimulant storage tank can send the air containing the olfactory stimulant into the tube for temporarily holding the olfactory stimulant. When the valve is open, the air containing the olfactory stimulant can be sent to the supply tube, and the experimental animal can be exposed to the olfactory stimulant. When the valve is closed, the air containing the olfactory stimulant is held in the tube for temporarily holding the olfactory stimulant. The suction pump can suck air with or without the olfactory stimulant from the suction tube and exhaust it to the outside of the device. An olfactory stimulation device for functional MRI imaging. [2] The device according to Embodiment 1, wherein an exhaust storage tank is arranged between the suction tube and the suction pump. [3] The device according to Embodiment 1, wherein a filter is arranged between the olfactory stimulant storage tank and the air compressor and / or a filter is arranged between the suction tube and the suction pump. [4] The device according to Embodiment 1, which is for marmosets, mice, rats, macaques, guinea pigs, or humans. [5] The device according to Embodiment 1, wherein the supply tube, the tube for temporarily holding the olfactory stimulant, the valve, and the suction tube are made of a non-magnetic material. [6] A method for imaging functional MRI when an experimental animal is exposed to an olfactory stimulant using the device according to Embodiment 1. [7] The method according to Embodiment 6, wherein the valve is closed to hold the olfactory stimulant in the tube for temporarily holding the olfactory stimulant, then the valve is opened by the valve opening / closing unit to send the olfactory stimulant to the supply tube, the experimental animal is exposed to the olfactory stimulant, and functional MRI is imaged. [8] The method according to Embodiment 7, wherein air containing or not containing the olfactory stimulant is sucked from the suction tube by a suction pump, and functional MRI is imaged while exhausting the air to the outside of the device. [9] After the experimental animal is exposed to the olfactory stimulant for a certain period of time, the valve is closed by the valve opening / closing unit to hold the olfactory stimulant in the tube for temporarily holding the olfactory stimulant, the decrease in reactivity of the piriform cortex and the anterior olfactory cortex of the experimental animal is suppressed, and after a predetermined period, the experimental animal is exposed to the olfactory stimulant for a certain period of time and functional MRI is imaged. The method according to Embodiment 8.

[10] After the experimental animal is exposed to the first olfactory stimulant for a certain period of time, the valve is closed by the valve opening / closing unit, the first olfactory stimulant is sucked by a suction pump, and the second olfactory stimulant is held in the tube for temporarily holding the olfactory stimulant with the valve closed. After a predetermined period, the experimental animal is exposed to the second olfactory stimulant for a certain period of time and functional MRI is imaged. The method according to Embodiment 9. [Effect of the Invention]

[0015] According to the present disclosure, functional MRI can be imaged when the sense of smell of an animal is stimulated. [Brief Description of the Drawings]

[0016]

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Embodiments for Carrying Out the Invention

[0017] Hereinafter, the present invention will be described in detail with reference to the drawings.

[0018] In one embodiment, the present invention provides an olfactory stimulation device for functional MRI imaging. An example of the olfactory stimulation device for functional MRI imaging of the present invention is shown in FIG. 1. In one embodiment, the olfactory stimulation device for functional MRI imaging of the present invention has, in addition to a normal experimental animal holder, a part for supplying an olfactory stimulant (also referred to as an olfactory stimulant supply part) and a part for sucking an olfactory stimulant (also referred to as an olfactory stimulant suction part).

[0019] In one embodiment, the olfactory stimulant supply part has a supply tube 5, a tube 10, a valve 11, a valve opening / closing part, an olfactory stimulant storage tank 16, and an air compressor 15. The tube 10 may be referred to as a tube for temporarily holding an olfactory stimulant in this specification. In one embodiment, the tube 10 and the valve 11 are made of a non-magnetic material. The non-magnetic material tube 10 can be, for example, a silicone tube, but is not limited thereto. Examples of the non-magnetic material valve 11 include, but are not limited to, a balloon valve. In one embodiment, the balloon can be made of rubber. The part for supplying an olfactory stimulant may optionally have a filter 14 (on the supply side).

[0020] The olfactory stimulant attracting section may include a suction tube 6 and a suction pump 18. The olfactory stimulant attracting section may further include an exhaust storage tank 17 and / or a filter 14 (on the suction side). The suction tube 6 may be made of a non-magnetic material. The filter 14, the olfactory stimulant storage tank 16, and the exhaust storage tank 17 may also be made of non-magnetic materials.

[0021] The valve opening / closing section may be any type as long as it can close the valve 11. In one embodiment, the valve opening / closing section may include a syringe 13 and a tube 22. The tube 22 connects the syringe 13 and the tube 10. In one embodiment, the valve 11 may be connected to a syringe 13 that sends air 12 through the tube 22. The air 12 may be a gas, such as an inert gas like nitrogen gas or helium. The syringe 13 may be made of a non-magnetic material. The tube 22 that sends air 12 from the syringe 13 to the balloon valve may be made of a non-magnetic material.

[0022] The olfactory stimulant is stored in the olfactory stimulant storage tank 16. The air compressor 15 is connected to the filter 14, and the filter 14 is connected to the olfactory stimulant storage tank 16. The olfactory stimulant storage tank 16 is connected to the tube 10. The tube 10 is connected to a supply tube 5 for supplying the olfactory stimulant. The suction tube 6 is connected to the exhaust storage tank 17, the exhaust storage tank 17 is connected to the filter 14, and the filter 14 is connected to the suction pump 18. The air compressor 15 sends out air, and the filtered air passes through the filter 14 and is sent into the olfactory stimulant storage tank 16. Then, the air containing the olfactory stimulant is sent from the olfactory stimulant storage tank 16 into the tube 10. If the valve 11 is open, the air containing the olfactory stimulant is sent to the supply tube 5, and the experimental animal is exposed to the olfactory stimulant. Then, the air containing the olfactory stimulant is sucked from the suction tube 6 by driving the suction pump 18, passes through the exhaust storage tank 17, passes through the filter 14, and is exhausted to the outside of the device by the suction pump 18.

[0023] With this configuration, the olfactory stimulus given to the experimental animal can be controlled. The valve 11 can be closed by the valve opening / closing part. For example, when air 12 is sent from the syringe 13, the valve 11 expands (closes), the inside of the tube 10 is blocked, and thereby the olfactory stimulant does not pass through the tube 10. When the valve 11 is opened by the valve opening / closing part, for example, when the balloon valve is deflated, the inside of the tube 10 becomes passable, and thereby the olfactory stimulant can pass through the tube 10. For example, before giving the olfactory stimulant to the experimental animal, the balloon valve can be inflated to store the olfactory stimulant inside the tube 10. Next, the balloon valve is deflated to open the valve, and the olfactory stimulant that has passed through the tube 10 is supplied to the supply tube 5, and the olfactory stimulant can be given to the experimental animal from the supply tube 5 for a predetermined time, for example, 30 seconds. Then, the olfactory stimulant can be sucked and removed through the suction tube. At this time, the balloon valve may be inflated to close the valve 11, or may remain in the open state. For example, after a 5-minute interval, the olfactory stimulant can be given again.

[0024] The valve 11 can be of any shape or form as long as it can send or cut off the gas in the tube 10 by opening and closing. Also, the valve 11 may be arranged outside the tube 10 instead of inside the tube 10. In certain embodiments, the valve 11 is arranged outside the tube 10. In certain embodiments, the valve 11 can be a donut-shaped external balloon. In certain embodiments, the donut-shaped external balloon can be installed in a form wrapped around the outer periphery of the tube 10. In this case, when the donut-shaped external balloon is inflated, the inflated donut-shaped external balloon compresses and deforms the tube 10, closing its inner cavity. Thereby, the donut-shaped external balloon can function as a valve. If it is a donut-shaped external balloon, there is no need to install anything inside the tube 10 for air supply, and it has the advantage of not obstructing the flow of odorant gas. Also, there is no need to drill a hole in the tube 10 to insert the balloon, and the risk of unintended gas leakage from the tube 10 can be avoided. Even when the valve 11 is a donut-shaped external balloon, the valve 11 can be closed by the valve opening / closing part. For example, when air 12 is sent in from the syringe 13, the valve 11 inflates (closes), and the inside of the tube 10 is blocked, whereby the olfactory stimulant does not pass through the tube 10. When the valve 11 is opened by the valve opening / closing part, for example, when the donut-shaped external balloon is deflated, the blockage of the tube 10 is eliminated, and its inside becomes passable, whereby the olfactory stimulant can pass through the tube 10. Both the balloon valve and the donut-shaped external balloon can be made of non-magnetic materials. When strict control of odorant substances is required, etc., a combination of a balloon valve arranged inside the tube 10 and a donut-shaped external balloon arranged outside may be used.

[0025] The olfactory stimulant is not particularly limited and includes any substance that can stimulate the olfaction of experimental animals. Examples of the olfactory stimulant include, but are not limited to, ester compounds and disulfide compounds such as thiamine propyl disulfide. In certain embodiments, the olfactory stimulation device for functional MRI imaging may have a plurality of olfactory stimulant storage tanks 16. In certain embodiments, each olfactory stimulant storage tank 16 may contain a different olfactory stimulant. For example, after exposure to the first olfactory stimulant contained in the first olfactory stimulant storage tank 16, the olfactory stimulant may be aspirated, and then the experimental animal may be exposed to the second olfactory stimulant contained in the second olfactory stimulant storage tank 16. In another embodiment, the olfactory stimulation device for functional MRI imaging may have a single olfactory stimulant storage tank 16. After exposure to the first olfactory stimulant contained in the olfactory stimulant storage tank 16, the olfactory stimulant may be aspirated, and then the olfactory stimulant in the olfactory stimulant storage tank 16 may be exchanged, and the experimental animal may be exposed to the second olfactory stimulant.

[0026] In certain embodiments, the present invention provides a method for stimulating the olfaction of experimental animals using an olfactory stimulation device for functional MRI imaging. The time for exposing the experimental animal to the olfactory stimulant is not particularly limited and may be, for example, 1 second, 2 seconds, 3 seconds, 4 seconds, 5 seconds, 10 seconds, 20 seconds, 30 seconds, 1 minute, 2 minutes, 3 minutes, 5 minutes, 10 minutes, 15 minutes, 30 minutes, for example, 60 minutes. After the experimental animal is exposed to the olfactory stimulant, a period during which the experimental animal is not exposed to the olfactory stimulant, that is, an interval period, can be provided. The interval is not particularly limited and may be, for example, 10 seconds, 20 seconds, 30 seconds, 1 minute, 2 minutes, 3 minutes, 5 minutes, 10 minutes, 15 minutes, 30 minutes, for example, 60 minutes.

[0027] A schematic diagram of the marmoset brain is shown in Fig. 13. When odor substances are detected in the olfactory bulb, the olfactory stimulus propagates through each region of the marmoset brain. While monitoring the brain activity of the experimental animal by functional MRI, the experimental animal can be exposed to the olfactory stimulus substance. Thereby, the brain activity before and after exposing the experimental animal to the olfactory stimulus substance, for example, the brain activity in various brain regions, can be compared. Examples of brain regions include, but are not limited to, the olfactory bulb (OB), anterior olfactory nucleus (On), piriform cortex (Piri), entorhinal cortex (Eth), amygdala (Amy), dorsomedial thalamic nucleus (MD), and orbitofrontal cortex (Orbi). Before exposing the experimental animal to the olfactory stimulus substance, first, the experimental animal can be fixed by the fixing device 3. A predetermined acclimation period may be provided until the brain activity of the experimental animal fixed by the fixing device 3 stabilizes. Thereafter, the valve 11 is opened to expose the experimental animal to the olfactory stimulus substance, and the brain activity can be monitored by functional MRI.

[0028] The fixing device 3 may include a bed portion and a helmet portion. Further, the fixing device 3 may include a collar portion and a U-shaped device. The collar portion is for restraining the neck and shoulders of the animal. The U-shaped device is for restraining the head of the animal.

[0029] To perform functional MRI measurement, an animal fixed so that its head does not move with a functional MRI imaging device is inserted into the MRI device. The coil is arranged so that MRI measurement of the fixed animal is possible. The coil can be cylindrical and can have a dimension such that the entire functional MRI bed can be inserted inside the coil. Further, the receiving coil can be built into the inner helmet. The MRI device is controlled by a computer. Also, the measurement data from the MRI device is sent to the computer.

[0030] The fixing device 3 may be referred to as a functional MRI bed in this specification. In this specification, when referring to an olfactory stimulus device system for functional MRI imaging, this does not mean only the functional MRI bed included in the system, but rather the entire functional MRI bed system including the functional MRI bed and the main body of the MRI device.

[0031] In one embodiment, it is desirable that the size of the fixing device 3 for use in a small-diameter MRI with a bore diameter of less than 30 cm is 15 cm or less in diameter. This can be used, for example, for marmosets. The fixing device 3 may have a collar portion for fixing the shoulders and neck of an individual, a tunnel portion for fixing the waist, and a helmet portion for fixing the head. The helmet portion is for accommodating the head of the animal. Corresponding to the helmet portion, the bed portion may have a platform for placing the chin. The helmet side of the tunnel portion may be provided with a collar mounting portion for mounting a collar. The side of the tunnel portion opposite to the helmet side may be provided with a corset. The corset may be made of a flexible material such as sponge. Also, the corset may have a binding band mounting portion so that the lower body or hind limbs of the animal can be restrained by a binding band. In the corset, it may be fixed with an adhesive tape instead of a binding band. Further, the fixing device 3 may have a pocket portion for the front limbs to enter when the animal is in the Sphinx posture.

[0032] The helmet portion may have a U-shaped device mounting portion for mounting one to four U-shaped devices. After the animal is placed in the Sphinx posture, the fixing device 3 can fix the head with a U-shaped device. The height at which the U-shaped device is attached can be adjusted by a plurality of adjustment screws. The fixing device 3 can be adjusted so that the head of the marmoset does not move more than 0.5 mm (in the vertical, horizontal, and front-back directions).

[0033] Improved fixing device 3 The improved fixing device 3 does not use a U-shaped device. The improved fixing device 3 is provided with a helmet portion having an inner helmet 21 and an outer helmet 20. The dimensions and shape of the inner helmet 21 can be designed according to the dimensions and shape of the head of the target experimental animal. By using the helmet portion having the inner helmet 21 and the outer helmet 20, body movement in the Z-axis direction, that is, the longitudinal direction, can be suppressed. The inner helmet 21 can be further fixed with sponge or the like. Thereby, body movement in the X-axis direction and the Y-axis direction, that is, the left-right and up-down body movement of the animal, can be reduced.

[0034] Unless otherwise specified, the fixing device 3 is made of a non-magnetic material. For example, the material of a certain fixing device 3 can be plastic, acrylic, or resin. Also, the screws for fixing the U-shaped device can be made of a non-magnetic material including resin. The inner helmet 21 can be made of an ultraviolet curable resin. For example, the large frame of the inner helmet 21 can be produced by 3D printing, and then coils can be combined with it, and further ultraviolet curable resin can be overlaid. The coils can be arranged inside (on the animal side) or outside the inner helmet 21.

[0035] The fixing device 3 can be provided with means for fixing the shoulders, neck, and waist of the animal. The fixing device 3 can be provided with a configuration for inserting the front limbs of the animal, that is, a pocket.

[0036] The whole or part of the fixing device 3 can be produced by 3D printing or injection molding. The inner helmet 21 can be produced by 3D printing or injection molding.

[0037] The olfactory stimulation device for functional MRI imaging of the present invention can be used for both small animals and large animals. Examples of experimental animals include small animals such as marmosets, mice, rats, guinea pigs, and macaques, pet animals such as dogs and cats, but are not limited thereto. In a specific embodiment, the experimental subject can be a human, for example, for human neonates.

[0038] Known or commercially available MRI devices can be used. In one embodiment, the MRI device can have a magnetic flux density of 7 to 14, 7 to 11.7, 7 to 9.4 tesla, for example, 7.0, 9.4, 11.7, 14 tesla, but is not limited thereto. In one embodiment, the MRI device can have a bore diameter of 15 cm or less, 14 cm or less, 13 cm or less, 12 cm or less, 11 cm or less, 10 cm or less, 9 cm or less, 8 cm or less, for example, 7 to 15 cm, 7.2 cm to 14 cm, 7.2 cm to 10 cm. The inner diameter refers to the diameter inside the bore.

[0039] (Example) An olfactory stimulation device for functional MRI imaging as shown in Fig. 1 was fabricated. The design of the fixing device 3 was improved from that described in Patent Document 1 using 3D CAD (Fusion 360: Autodesk, San Francisco, CA). The helmet has an outer helmet 20 and an inner helmet 21. The inner helmet 21 incorporates a receiving coil 7 for improving the signal-to-noise ratio (S / N ratio) and reduces the gap between the head and the coil (Takashima Seisakusho, Tokyo, Japan). The gap between the helmets was adjusted for each marmoset by inserting a sponge between the helmets. The marmoset was placed in a Sphinx posture. A schematic diagram of the olfactory stimulation circuit is shown in Fig. 13. Two holes were drilled in the fixing device 3, and the supply tube 5 and the suction tube 6 were passed through. The tip of each tube was designed to be slightly bent so that it could be placed near the nose of the marmoset (Figs. 2 and 3). One of the tubes was used as the supply tube 5, and the other was used as the suction tube 6. A balloon valve was used as the valve 11. The balloon valve was set at a position 50 mm from the tip of the supply tube 5. The supply tube 5 was connected to an olfactory inspection device (O'HARA & CO., Tokyo, Japan), and the olfactory stimulant was supplied from an air compressor 15.

[0040] Visualization of the airflow of the olfactory stimulant Method To evaluate the gas flow and residual degree in a physical environment equivalent to an MRI experiment, the airflow of olfactory stimulants was evaluated using a simulation circuit. Figures 4 and 5 show the equipment for the simulation environment. The simulation environment consists of an acrylic cylinder 4 with an inner diameter of 74 mm and a total length of 650 mm that mimics the bore of an MRI, a face mold 8 that takes the shape of a marmoset, a marmoset fixator 3, a receiving coil 7, a silicon supply tube 5 with an inner diameter of 2 mm, and a suction tube 6. These were arranged in the same way as in an actual MRI. The smoke from a tracer particle source (not shown) (manufactured by Gastec, smoke tube 501) was irradiated with a laser light source 1 (manufactured by STS, GML-7010) at an irradiation angle of 70 degrees to visualize the gas flow. The visualized flow was photographed at 30 fps and a resolution of 1280x720 using a video camera 2. A transparent acrylic cylinder that mimics the bore of an MRI was used to observe the behavior of tracer particles inside the cylinder. The influence of the presence or absence of suction on the gas flow was visually examined under the condition that the supply and suction air flow rates were fixed at 0.3 L / min.

[0041] Results Figures 7, 8, and 9 show the results when only air was supplied and no suction was performed. The period from Figure 7 to Figure 9 is a series of continuous images showing the gas flow for about 1 second. The particles that were not exhausted formed a swirling flow and remained around the face of the marmoset. Figures 10, 11, and 12 are a series of continuous images of the gas flow for 1 second with and without suction. The exhaled smoke passed in front of the marmoset's nose and was smoothly exhausted as a laminar flow. No particles were observed to remain in the air for a long time.

[0042] Discussion When the olfactory stimulant was suctioned by the suction pump 18, the olfactory stimulant from the supply tube 5 passed through the tip of the marmoset's nose, was suctioned from the suction tube 6, and was discharged as a laminar flow (Figures 10, 11, and 12). When there was no suction and the olfactory stimulant was simply released into the marmoset's nose, the olfactory stimulant remained in front of the marmoset unintentionally for a long time (Figures 7, 8, and 9), and the odor remained, causing olfactory adaptation. Therefore, this result demonstrated that suction of the olfactory stimulant is useful in observing the olfactory stimulation of experimental animals.

[0043] MRI experiment Method For the experiment, an animal MRI device, the 7.0T Biospec 70 / 16 scanner system, was used (Bruker biospin MRI GmbH; Ettlingen, Germany). The receiving coil 7 was a 4-channel phased array surface coil, and the transmitting coil was a 72-mm orthogonal transmitting RF coil (Bruker BioSpin). Functional MRI was performed on two trained marmosets. Imaging was performed for 60 seconds per trial. For the first 30 seconds, there was no exposure to the olfactory stimulant (off), and for the next 30 seconds, exposure to the olfactory stimulant was carried out (on). The olfactory stimulant was thiamine propyl disulfide (Alinamin (registered trademark) injection solution 10 mg, Teva Pharmaceutical Industries, Ltd., Tokyo, Japan), which is used in human intravenous olfactory tests. The imaging interval was set at 5 minutes. During imaging and the interval, the olfactory stimulant was always aspirated from the suction tube 6 by the suction pump 18. This trial was repeated 5 sets. The flow rate was 3 L / min for both air supply and suction.

[0044] Results In the olfactory bulb, anterior olfactory nucleus, piriform cortex (p < 0.05), entorhinal field, and olfactory cortex (p < 0.01), a significant difference in AUC (Area Under the Curve) was observed before and after stimulation. No significant difference was found in the three olfactory-related regions, the amygdala, medial thalamus, and prefrontal orbital cortex. The olfactory pathway and the responses of each site are shown in FIGS. 14 to 20.

[0045] Discussion The experimental results demonstrated the advantages of the olfactory stimulation device for functional MRI imaging of the present disclosure. After olfactory stimulation, an increase in signals was observed in five brain regions related to the olfactory pathway. On the other hand, no significant brain activity was seen in regions not related to olfactory function, such as the somatosensory cortex and gustatory cortex. Therefore, it was demonstrated that the olfactory stimulation device for functional MRI imaging of the present disclosure can specifically evaluate olfactory function. Furthermore, the MRI signal showed a significant difference within 30 seconds after stimulation with an olfactory stimulant, enabling evaluation with a high temporal resolution compared to previous studies. In this example, fMRI was performed 5 times at 5-minute intervals, and since the reaction in the first half of the olfactory pathway from the olfactory bulb to the piriform cortex was observed in each execution, it is considered that the ventilation of this device is sufficient. It is known that when olfactory stimulation is continuously applied for a long period, the reactivity of the piriform cortex and the anterior olfactory cortex decreases. The olfactory stimulation device for functional MRI imaging of the present disclosure can avoid continuous long-term exposure of experimental animals to olfactory stimulants, thereby suppressing the decrease in the reactivity of the piriform cortex and the anterior olfactory cortex. Also, being able to conduct experiments without being affected by unnecessary excitement or body movement due to visual stimulation is also a feature of the olfactory stimulation device for functional MRI imaging of the present disclosure. In addition, when using the olfactory stimulation device for functional MRI imaging of the present disclosure, when changing the olfactory stimulation presented to experimental animals, it is not necessary to change the odor source in front of the animals' eyes as in conventional devices. Therefore, it is possible to avoid problems such as other sensory stimuli and excitement caused by the operation of changing the odor source.

[0046] In this specification, a number of documents including the manufacturer's manuals are cited. The disclosures of these documents are not considered to be relevant to the patentability of the present invention, but the entire contents thereof are hereby incorporated by reference into this specification. More specifically, all the reference documents are incorporated by reference into this specification in the same manner as if each individual document were specifically and individually indicated as being incorporated by reference.

Explanation of Signs

[0047] 1 Laser light source 2 Video camera 3 Restrainer 4 Cylinder 5 Supply tube 6 Suction tube 7 Receiver coil 8 Facial mold of marmoset 9 Simulated marmoset nose tip 10 Tube 11 Valve 12 Air 13 Syringe 14 Filter 15 Air compressor 16 Olfactory stimulant reservoir 17 Exhaust reservoir 18 Suction pump 19 Laboratory animal 20 Outer helmet 21 Inner helmet 22 Tube 30 Olfactory bulb 31 Olfactory tract 32 Olfactory nucleus 33 Piriform cortex 34 Tonsil 35 Olfactory entorhinal area 36 Dorsomedial thalamic nucleus 37 Orbitofrontal cortex 100 Olfactory stimulation device for functional MRI imaging

Claims

1. An olfactory stimulation device for functional MRI imaging having an olfactory stimulant supply unit and an olfactory stimulant suction unit, the olfactory stimulant supply unit includes a supply tube, a tube for temporarily holding the olfactory stimulant, a valve, a valve opening / closing unit, an olfactory stimulant storage tank, and an air compressor; the supply tube is connected to a tube for temporarily storing the olfactory stimulant, a valve is connected to the tube for temporarily storing the olfactory stimulant, the tube for temporarily storing the olfactory stimulant is connected to an olfactory stimulant storage tank, the olfactory stimulant storage tank contains the olfactory stimulant therein, and the olfactory stimulant storage tank is connected to an air compressor; The olfactory stimulant suction unit has a suction tube and a suction pump, The suction tube is connected to a suction pump. The valve is connected to the valve opening and closing portion, The air compressor can send air to an olfactory stimulant storage tank, and the olfactory stimulant storage tank can send the air containing the olfactory stimulant to a tube for temporarily holding the olfactory stimulant; When the valve is open, air containing an olfactory stimulus can be sent to the supply tube, and the experimental animal can be exposed to the olfactory stimulus, and when the valve is closed, the air containing the olfactory stimulus is retained in the tube for temporarily retaining the olfactory stimulus, The suction pump can suck air containing or not containing olfactory stimulants through the suction tube and exhaust it outside the device. Olfactory stimulation device for functional MRI imaging.

2. 10. The apparatus of claim 1, further comprising an exhaust reservoir disposed between the suction tube and the suction pump.

3. 2. The device of claim 1, wherein a filter is disposed between the olfactory stimulant reservoir and the air compressor, and / or a filter is disposed between the suction tube and the suction pump.

4. 10. The device of claim 1, for use in a marmoset, a mouse, a rat, a macaque, a guinea pig, or a human.

5. 10. The device of claim 1, wherein the supply tube, the tube for temporarily holding the olfactory stimulant, the valve, and the suction tube are constructed of non-magnetic materials.

6. 13. A method for imaging functional MRI of laboratory animals upon exposure to olfactory stimuli using the device of claim 1.

7. The method according to claim 6, wherein the valve is closed to hold the olfactory stimulant in a tube for temporarily holding the olfactory stimulant, and then the valve is opened by the valve opening / closing section to send the olfactory stimulant to the supply tube, exposing the experimental animal to the olfactory stimulant, and performing functional MRI imaging.

8. The method according to claim 7, wherein air containing or not containing an olfactory stimulant is sucked through a suction tube by a suction pump and exhausted to the outside of the device while functional MRI is performed.

9. The method according to claim 8, wherein the experimental animal is exposed to the olfactory stimulant for a certain period of time, and then the valve is closed by the valve opening / closing unit to retain the olfactory stimulant in a tube for temporarily retaining the olfactory stimulant, thereby suppressing a decrease in reactivity of the piriform cortex and anterior olfactory cortex of the experimental animal, and after a predetermined period of time, the experimental animal is exposed to the olfactory stimulant for a certain period of time and functional MRI is taken.

10. The method according to claim 9, wherein after exposing an experimental animal to a first olfactory stimulant for a certain period of time, the valve is closed using a valve opening / closing unit, the first olfactory stimulant is sucked in using a suction pump, and while the valve is closed, the second olfactory stimulant is held in a tube for temporarily holding olfactory stimulants, and after a predetermined period of time, the experimental animal is exposed to the second olfactory stimulant for a certain period of time and functional MRI is taken.

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