JP2017096838A - Method of screening oral control agents for skin sympathetic nerve activity or skin microcirculation - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a method of screening oral control agents for skin sympathetic nerve activity or skin microcirculation.SOLUTION: A method of screening oral control agents for skin sympathetic nerve activity or skin microcirculation involves measuring TRPV4 activity control capacity under the presence of oral control agent candidate substances for skin sympathetic nerve activity or skin microcirculation, and selecting a candidate subject capable of controlling TRPV4 activity as an oral control agent for skin sympathetic nerve activity or skin microcirculation.SELECTED DRAWING: None

Description

The present invention relates to a screening method for an oral regulator of skin sympathetic nerve activity or skin microcirculation.
The present invention also relates to a non-therapeutic method of controlling skin sympathetic nerve activity or skin microcirculation.
The present invention further relates to an oral sympathetic nerve activity control agent or an oral skin microcirculation control agent.

  In the skin, a vascular network composed of three layers of vascular beds parallel to the skin surface is formed. The most important function of the skin microcirculation is the regulation of body temperature. When a cold stimulus or a thermal stimulus is applied, the skin blood vessels contract or expand to change the blood flow of the skin microcirculation, thereby suppressing or promoting body heat dissipation. Maintain body temperature. In addition, skin microcirculation plays an important role in regulating the distribution of blood flow throughout the body, and is caused by information and exercise from pressure or chemoreceptors in addition to information from central and peripheral thermal receptors. Also affected by vasomotor reflexes.

The skin microcirculation is subject to dual regulation of a mechanism through the sympathetic nervous system, which is one of the autonomic nervous systems, and a local mechanism by local regulators. One local mechanism is axonal reflex. This is because impulses generated in the sensory nerves of the skin by stimulation are transmitted to other branches in the middle of conduction to the center, and substance P (SP) and calcitonin gene-related peptide (CGRP) are transmitted from the nerve endings. And release neuropeptides. As a result, the skin blood vessels dilate. On the other hand, the sympathetic nervous system dominates arteriole smooth muscles in the blood vessels of the skin, and the increase in the blood flow of the fingers during heat is caused by a decrease in the skin sympathetic nerve activity. On the other hand, a decrease in blood flow during cold is caused by an increase in skin sympathetic nerve activity.
Therefore, skin sympathetic nerve activity plays an important role in skin microcirculation control.

It is known that the skin microcirculation is affected by various effects such as aging, stress, and lifestyle habits. Normal control of the skin microcirculation is important for maintaining health, and when the skin blood vessels are too dilated and the skin blood flow is too high, symptoms such as redness (capillary telangiectasia) are caused. Conversely, when the skin blood vessels contract too much and the skin blood flow is low, symptoms such as poor circulation and coldness are caused.
Therefore, if the skin sympathetic nerve activity that controls the skin microcirculation can be controlled, it is considered that the above symptoms can be effectively improved.

  As components for controlling conventional skin microcirculation, collagen peptide (Patent Document 1), Sendangsa plant (Patent Document 2), γ-linolenic acid alone or γ-linolenic acid and a fat-soluble antioxidant (Patent Document 3), Diraceb or other addition salts (Patent Document 4), hydroxymethylfurfural derivatives (Patent Document 5), bilobalide (Patent Document 6), which is a component of ginkgo biloba, are known as blood circulation promoters. However, these are not always satisfactory techniques in terms of effect and specificity.

  Therefore, more effective skin microcirculation and skin sympathetic nerve control agents are desired. However, none of these screening methods are satisfactory, and establishment of an efficient screening method for agents for controlling skin microcirculation and skin sympathetic nerves is desired.

On the other hand, TRPV4 (Transient receptor potential cation channel subfamily V member 4) is a member of the transient receptor potential (TRP) superfamily of cation channels, and has low osmotic pressure, heat (27 ° C <), arachidonic acid metabolite. It is known that it is activated by the above (Non-patent Documents 1 and 2).
TRPV4 is expressed in a wide range of tissues such as kidney, lung, bladder, heart, skin, brain, gastrointestinal tract, and is thought to play a wide variety of different physiological roles. It is particularly strongly expressed in the distal tubular epithelial cells of the kidney, and it is suggested that urine osmotic pressure and flow rate are sensed (Non-patent Document 3). It has also been reported that when a TRPV4 activator is administered intravenously, arterial blood vessels are dilated and blood pressure is reduced (Non-patent Document 4).
However, there is no known effect on the skin sympathetic nerve activity and skin microcirculation when a TRPV4 activity regulator is orally administered.

JP 2002-121148 A JP 2002-205954 A JP 2000-302677 A JP-A-11-92382 JP-A-11-228561 JP 7-53371 A

Guler AD., Et al. Neurosci., 2002, vol. 22, p. 6408-6414 Vriens J., et al., Proc. Natl. Acad. Sci. USA, 2004, vol. 101, p.396-401 Tian W., et al., Am. J. Physiol. Renal. Physiol., 2004, vol. 287, p. F17-F24 Gao F., et al., Hypertension, 2010, vol. 55, p. 1018-1025

  Controlling skin sympathetic nerve activity and controlling skin microcirculation is important in maintaining health. However, there has been no skin microcirculation control technology using skin microcirculation control agents and skin sympathetic nerve control agents that have been satisfactory to date, and efficient methods for finding such skin microcirculation control agents and skin sympathetic nerve control agents. There is no simple screening method.

An object of the present invention is to provide a screening method for an oral regulator of skin sympathetic nerve activity or skin microcirculation using TRPV4 activity control as an index.
Another object of the present invention is to provide a method for controlling non-therapeutic skin sympathetic nerve activity or skin microcirculation by controlling TRPV4 activity by oral administration or ingestion.
Another object of the present invention is to provide an oral skin microcirculation inhibitor or an oral skin sympathetic nerve activity enhancer comprising a TRPV4 activity enhancer as an active ingredient for controlling skin microcirculation by controlling skin sympathetic nerve activity.
Furthermore, this invention makes it a subject to provide the oral skin microcirculation enhancer or oral skin sympathetic nerve activity inhibitor which uses TRPV4 activity inhibitor as an active ingredient.

In view of the above problems, the present inventors examined the effects of TRPV4 activity regulators on skin sympathetic nerve activity and skin microcirculation function. As a result, it was found that oral ingestion of a TRPV4 activity controlling agent has an action of enhancing the skin sympathetic nerve activity and suppressing the skin microcirculation function.
The present invention has been completed based on this finding.

In the present invention, the ability to control TRPV4 activity is measured in the presence of a candidate substance for an oral regulator of skin sympathetic nerve activity or skin microcirculation, and a candidate substance capable of controlling TRPV4 activity is determined as the target of skin sympathetic nerve activity or skin microcirculation. The present invention relates to a screening method for an oral regulator of skin sympathetic nerve activity or skin microcirculation selected as an oral regulator.
The present invention also relates to a non-therapeutic method of controlling skin sympathetic nerve activity or skin microcirculation by controlling gastrointestinal TRPV4 activity by oral administration or ingestion.
The present invention further relates to an oral skin microcirculation control agent or an oral skin sympathetic nerve activity control agent comprising a TRPV4 activity control agent as an active ingredient.

According to the screening method for an oral regulator of skin sympathetic nerve activity or skin microcirculation of the present invention using TRPV4 activity control as an index, an oral regulator of skin sympathetic nerve activity or skin microcirculation can be efficiently screened. .
Further, according to the method for controlling skin sympathetic nerve activity or skin microcirculation of the present invention by controlling TRPV4 activity by oral administration or ingestion, skin sympathetic nerve activity or skin microcirculation can be effectively controlled. .
Moreover, the oral skin microcirculation controlling agent or the oral skin sympathetic nerve activity controlling agent of the present invention can effectively control the skin sympathetic nerve activity and control the skin microcirculation.

It is a graph which shows a time-dependent change of the afferent vagus nerve activity at the time of orally administering a TRPV4 activity inhibitor and a TRPV4 activity enhancer to a rat. FIG. 2 (A) is a graph showing changes over time in skin sympathetic nerve activity when a TRPV4 activity enhancer is orally administered to a rat, and FIG. It is a graph which shows a time-dependent change of the skin sympathetic nerve activity at the time of administering orally. FIG. 3 (A) is a graph showing the time course of skin blood flow when a TRPV4 activity enhancer is orally administered to rats, and FIG. 3 (B) is a graph showing a TRPV4 activity enhancer applied to a rat with afferent vagus nerve excised. It is a graph which shows the time-dependent change of the skin blood flow volume at the time of oral administration.

As used herein, “prevention” refers to preventing or delaying the onset of a disease or symptom in an individual, or reducing the risk of developing an individual's disease or symptom. In the present specification, “improvement” refers to improvement of disease, symptom or condition, prevention or delay of deterioration of disease, symptom or condition, or reversal, prevention or delay of progression of disease, symptom or condition.
Further, in this specification, “non-therapeutic” is a concept that does not include a medical act, that is, a treatment act on the human body for the purpose of treatment under the direction of a doctor or a doctor.
In the present specification, “skin sympathetic nerve activity” refers to the activity of sympathetic nerves that govern the arterioles of the skin.
In the present specification, “skin microcirculation” refers to the total blood flow of arterioles, capillaries, and venules in the skin.
Further, in this specification, “TRPV4 activity control” or “gastrointestinal TRPV4 activity control” refers to the function of TRPV4 channel or the gastrointestinal TRPV4 channel itself, that is, control of calcium ion influx into cells via TRPV4 channel. Say.

As shown in Examples below, there is a correlation between gastrointestinal TRPV4 activity and skin sympathetic nerve activity or skin microcirculation, and when a gastrointestinal TRPV4 activity enhancer is orally administered or ingested, skin sympathy Neural activity is increased and skin blood flow is reduced. Moreover, when a gastrointestinal TRPV4 activity inhibitor is orally administered or ingested, the skin sympathetic nerve activity is suppressed and the skin blood flow rate increases.
The method for screening an oral regulator of skin sympathetic nerve activity or skin microcirculation of the present invention is based on the correlation between TRPV4 activity and skin sympathetic nerve activity or skin microcirculation. Specifically, in the screening method of the present invention, the ability to regulate TRPV4 activity, preferably gastrointestinal TRPV4 activity, is measured in the presence of a candidate substance for a regulator of skin sympathetic nerve activity or skin microcirculation, and TRPV4 activity is controlled. A controllable candidate substance is selected as an oral regulator of skin sympathetic nerve activity or skin microcirculation.

The ability to control TRPV4 activity or the ability to control TRPV4 activity in the screening method of the present invention can be confirmed by the following method. However, the present invention is not limited to this.
For TRPV4 activity, a vector expressing TRPV4 is used to forcibly express TRPV4 in Hela cells, 293 cells, or the like. Then, using the cultured cells expressing TRPV4, intracellular Ca ²⁺ influx activity via TRPV4 in the presence of a candidate substance for an oral regulator of skin sympathetic nerve activity or skin microcirculation (hereinafter also referred to as “screening specimen”). It can be confirmed by measuring. In this case, the screening sample in which Ca ²⁺ inflow is observed into the cell has a TRPV4 activity enhancing action. In contrast, a screening specimen that suppresses Ca ²⁺ influx into cells has a TRPV4 activity inhibitory action.
The base sequence information and amino acid sequence information of human digestive tract TRPV4 are registered as NCBI Reference Sequence: NM — 021625.4 and NP — 067638.3, respectively. The base sequence information and amino acid sequence information of rat TRPV4 are registered as NCBI Reference Sequence: NM — 023970.1 and NP — 076460.1, respectively. Further, the base sequence information and amino acid sequence information of mouse TRPV4 are registered as NCBI Reference Sequence: NM_022017.3 and NP_071300.2, respectively. Such information is available from NCBI. These variants or homologues can also be the subject of the present invention.

In Examples described later, it was found that gastrointestinal TRPV4 activity control is involved in the control of skin sympathetic nerve activity and skin microcirculation activity via afferent vagus nerve activity. Therefore, in order to confirm the ability to control gastrointestinal TRPV4 activity, it is necessary to further affect the ability to regulate afferent vagus nerve activity from the gastrointestinal tract by allowing the screening specimen to act on the gastrointestinal tract of mice, rats, and the like. In other words, it is necessary to confirm the ability to control the digestive tract TRPV4 activity including mucosal permeability on the digestive tract, absorbability of the specimen, and the like.
Screening specimens having an action of enhancing gastrointestinal TRPV4 activity increase afferent vagus nerve activity. On the other hand, the screening specimen having the gastrointestinal TRPV4 activity inhibitory action decreases the afferent vagus nerve activity. Then, based on the above-described correlation between the gastrointestinal TRPV4 activity and the skin sympathetic nerve activity or the skin microcirculation, a screening sample having an action to enhance the gastrointestinal TRPV4 activity is used as the skin sympathy via the afferent vagus nerve activity. Select as a nerve activity enhancer or skin microcirculation inhibitor. On the other hand, a screening sample having a gastrointestinal TRPV4 activity inhibitory action is selected as a skin sympathetic nerve activity inhibitor or a skin microcirculation enhancer via afferent vagus nerve activity.

The method for controlling skin sympathetic nerve activity or skin microcirculation by oral administration or ingestion of the present invention, and the oral skin microcirculation regulator or oral skin sympathetic nerve activity control agent are also TRPV4 activity, preferably gastrointestinal TRPV4 activity, and afferent. This is based on the correlation between sympathetic nerve activity or skin microcirculation via vagal activity.
Specifically, the method for controlling skin sympathetic nerve activity or skin microcirculation by oral administration or ingestion of the present invention comprises a step of controlling TRPV4 activity, preferably gastrointestinal TRPV4 activity. In order to control TRPV4 activity or gastrointestinal TRPV4 activity, it is preferable to orally administer or ingest the oral skin microcirculation regulator or oral skin sympathetic nerve activity regulator of the present invention. The method for controlling skin sympathetic nerve activity or skin microcirculation by oral administration or ingestion of the present invention may be a therapeutic method (ie, a medical act) or a non-therapeutic method (non-medical act). May be.
Moreover, the oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent of the present invention comprises a TRPV4 activity control agent, preferably a gastrointestinal TRPV4 activity control agent as an active ingredient. The TRPV4 activity control agent or gastrointestinal TRPV4 activity control agent used in the present invention includes a TRPV4 activity enhancer or a gastrointestinal TRPV4 activity enhancer (hereinafter referred to as “the first embodiment of the present invention”). Mode regulator "or simply" the gastrointestinal TRPV4 activity enhancer of the present invention "), a TRPV4 activity inhibitor or gastrointestinal TRPV4 activity inhibitor (hereinafter referred to as" the present invention " 2 ”or simply“ the gastrointestinal TRPV4 suppression enhancer of the present invention ”). These control agents are used for skin sympathetic nerve activity or skin microcirculation activity control via afferent vagus nerve activity.

An oral skin microcirculation controller or an oral skin sympathetic nerve activity control agent (hereinafter referred to as “skin microcirculation”) comprising TRPV4 activity enhancer or gastrointestinal TRPV4 activity enhancer as an active ingredient as TRPV4 activity control agent or gastrointestinal TRPV4 activity control agent. The “suppressor or skin sympathetic nerve activity enhancer” is also described.
As the skin microcirculation inhibitor or the skin sympathetic nerve activity enhancer of the present invention, one or more of any TRPV4 activity enhancer or gastrointestinal TRPV4 activity enhancer can be used as an active ingredient.
Examples of the TRPV4 activity enhancer or gastrointestinal TRPV4 activity enhancer include 12,13-didecanoic acid 4α-phorbol (4α-Phorbol-12,13-Didecanoate, hereinafter also abbreviated as “4α-PDD”), 5,6-epoxy. Eicosatrienoic acid (5,6-epoxyeicosatrienoic acid, also known as: 5,6-EET), 1-benzyl-4- (4-chloro-2-nitrophyll) sulfonylbenzylpiperazine (1-benzyl-4- (4- chloro-2-nitrophenyl) sulfonylpiperazine, also known as: RN-1747), and N-[(2S) -1- [4-[(2S) 2-[(2,4-dichlorophenyl) sulfonylamino] -3-hydroxyprop Noyl] piperazin-1-yl] -4-methyl-1-oxopentan-2-yl] -1-benzothiophene-2-carboxamide (N-[(2S) -1- [4-[(2S) 2- [(2,4-dichlorophenyl) sulfonylamino] -3-hydroxypropanoyl] piperazin-1-yl] -4-methyl-1-oxopentan-2-yl] -1-benzothiophene-2-carboxamide, also known as: GSK1016790A) . 4α-PDD, 5,6-EET, RN-1747, and GSK1016790A each enhance TRPV4 activity by enhancing the function of TRPV4 itself.
As the TRPV4 activity enhancer or gastrointestinal TRPV4 activity enhancer that can be used in the present invention, those commercially available as reagents can be used.

Oral skin microcirculation controller or oral skin sympathetic nerve activity control agent (hereinafter referred to as “skin microcirculation”) comprising TRPV4 activity inhibitor or digestive tract TRPV4 activity inhibitor as an active ingredient as TRPV4 activity controller or digestive tract TRPV4 activity controller. An enhancer or a skin sympathetic nerve activity inhibitor ”) will be described.
As the skin microcirculation enhancer or the skin sympathetic nerve activity inhibitor of the present invention, one or more arbitrary TRPV4 activity inhibitors or gastrointestinal TRPV4 activity inhibitors can be used as active ingredients.
Examples of TRPV4 activity inhibitors or gastrointestinal TRPV4 activity inhibitors include 2,4-dichloro-N-propan-2-yl-N- [2- (propan-2-ylamino) ethyl] benzenesulfonamide (2,4- dichloro-N-propan-2-yl-N- [2- (propan-2-ylamino) ethyl] benzenesulfonamide, aka RN1734), N- [4- [2- [benzyl (methyl) amino] ethyl] phenyl] -5-Pyridin-3-yl-1,3-thiazol-2-amine; Hydrobromide (N- [4- [2- [benzyl (methyl) amino] ethyl] phenyl] -5-pyridin-3-yl- 1,3-thiazol-2-amine; hydrobromide, also known as GSK205) and 2-methyl-1- (3-morpholin-4-ylpropyl) -5-phenyl-N- [3- (trifluoromethyl) phenyl] And pyrrole-3-carboxamide (2-methyl-1- (3-morpholin-4-ylpropyl) -5-phenyl-N- [3- (trifluoromethyl) phenyl] pyrrole-3-carboxamide, also known as HC067047). RN1734, GSK205, and HC067047 each suppress the gastrointestinal TRPV4 activity by suppressing the function of TRPV4 itself.
As the TRPV4 activity inhibitor or the digestive tract TRPV4 activity inhibitor that can be used in the present invention, those commercially available as reagents can be used.

As also shown in the examples described later, the TRPV4 activity enhancer enhances afferent vagus nerve activity from the gastrointestinal tract, enhances skin sympathetic nerve activity, and suppresses skin microcirculation by enhancing TRPV4 activity in the gastrointestinal tract. Moreover, TRPV4 activity suppression can suppress afferent vagus nerve activity from the gastrointestinal tract, suppress skin sympathetic nerve activity, and enhance skin microcirculation.
Moreover, according to the screening method of the present invention using TRPV4 activity control or gastrointestinal TRPV4 activity control as an index based on the relationship shown above, an oral regulator of skin sympathetic nerve activity or skin microcirculation is efficiently screened. be able to.

The skin sympathetic nerve activity enhancer, the skin microcirculation inhibitor, the skin sympathetic nerve activity inhibitor and the skin microcirculation enhancer are one of the specific embodiments of the oral skin microcirculation regulator or the oral skin sympathetic nerve activity regulator. It can be applied to both therapeutic uses (medical uses) and non-therapeutic uses (non-medical uses). Specifically, the active ingredient alone may be used as the skin sympathetic nerve activity enhancer, skin microcirculation inhibitor, skin sympathetic nerve activity inhibitor or skin microcirculation enhancer of the present invention. Alternatively, the skin sympathetic nerve activity enhancer, skin microcirculation inhibitor, skin sympathetic nerve activity inhibitor or skin microcirculation enhancer of the present invention comprising the active ingredient and a pharmaceutically acceptable carrier, It can be used as pharmaceuticals, quasi drugs, etc. Alternatively, the skin sympathetic nerve activity enhancer, skin microcirculation inhibitor, skin sympathetic nerve activity inhibitor or skin microcirculation enhancer of the present invention may be contained in various foods, foods, feeds, pet foods and the like.
Here, the skin sympathetic nerve activity enhancer, the skin microcirculation inhibitor, the skin sympathetic nerve activity inhibitor or the skin microcirculation enhancer of the present invention includes the above-described substance alone as an active ingredient, and the substance as an active ingredient. Any of the embodiments of the composition containing a pharmaceutically acceptable carrier and various additives are included.
Hereinafter, an embodiment of a composition containing the substance as an active ingredient and containing a pharmaceutically acceptable carrier and various additives will be described.

  When preparing a pharmaceutical composition as a skin sympathetic nerve activity enhancer, a skin microcirculation inhibitor, a skin sympathetic nerve activity inhibitor or a skin microcirculation enhancer of the present invention, usually the active ingredient and preferably pharmaceutically acceptable It is prepared as a preparation containing the carrier. A pharmaceutically acceptable carrier generally refers to an inert, non-toxic, solid or liquid, bulking agent, diluent or encapsulating material that does not react with the active ingredient, such as water. , Ethanol, polyols, suitable mixtures thereof, solvents or dispersion media such as vegetable oils, and the like.

  The pharmaceutical composition is administered or ingested orally. Examples of the preparation for oral administration include tablets, coated tablets, capsules, granules, powders, syrups, enteric solvents, troches, and drinks.

  The skin sympathetic nerve activity enhancer, skin microcirculation inhibitor, skin sympathetic nerve activity inhibitor and skin microcirculation enhancer of the present invention may contain other additives as long as they do not affect the effect. Good. Other additives include, for example, excipients, liquid carriers, oily carriers, stabilizers, wetting agents, emulsifiers, binders, tonicity agents, disintegrating agents, lubricants, bulking agents, surfactants, Dispersant, suspending agent, diluent, osmotic pressure adjusting agent, pH adjusting agent, preservative, antioxidant, colorant, UV absorber, moisturizer, thickener, brightener, buffer, preservative, candy Commonly used additives that are pharmaceutically acceptable, such as a flavoring agent, a fragrance, a film agent, a flavoring agent, and a bacteria inhibitor, can be used.

  When applying the skin sympathetic nerve activity enhancer, skin microcirculation inhibitor, skin sympathetic nerve activity inhibitor and skin microcirculation enhancer of the present invention to pharmaceuticals and quasi-drugs, it is necessary to contain an effective amount of the active ingredient, Can be prepared into various dosage forms by blending additives. What is necessary is just to manufacture in accordance with a conventional method using an additive, in preparing in various dosage forms.

  The skin sympathetic nerve activity enhancer, skin microcirculation inhibitor, skin sympathetic nerve activity inhibitor and skin microcirculation enhancer of the present invention should be added to foods, beverages, feeds, pet foods, etc. or mixed with these. Can do. Alternatively, it can be used as it is as a beverage or food. Alternatively, foods and beverages that indicate the concept of increased skin sympathetic nerve activity, suppression of skin microcirculation, suppression of skin sympathetic nerve activity, or increase of skin microcirculation, that is, health food, functional indication food, disease It can be used by being added to or blended with foods for consumers, health foods and foods for specified health use. In addition, treatment or prevention of a disease or condition that contains the active ingredient and can be treated, prevented, or improved by enhancing skin sympathetic nerve activity, inhibiting skin microcirculation, inhibiting skin sympathetic nerve activity, or enhancing skin microcirculation. Or it can be used by adding to or blending with foods and drinks that indicate that with the concept of improvement or the like, that is, health foods, functional labeling foods, sick foods, health foods and foods for specified health. The above-mentioned health foods, functional indication foods, foods for the sick, health foods and foods for specified health use are in various forms suitable for food or drink, for example, various preparations such as granules, granules, tablets, capsules, pastes, etc. It can be provided in the form. A food in the form of a preparation can be produced in the same manner as a pharmaceutical preparation, and the active ingredient is mixed with a food-acceptable carrier, such as an appropriate excipient, and then produced using conventional means. Can do. In addition, soups, juices, fruit juice drinks, milk, milk drinks, whey drinks, lactic acid bacteria drinks, tea drinks, alcoholic drinks, coffee drinks, carbonated drinks, soft drinks, water drinks, cocoa drinks, jelly drinks, sports Addition or control agents for skin sympathetic nerve activity or skin microcirculation of the present invention to liquid food compositions such as beverages and diet beverages, semi-solid food compositions such as pudding and yogurt, noodles, confectionery, spreads, etc. And a food composition can be produced. Examples of the feed include small animal feed used for rabbits, rats, mice and the like, pet food used for dogs, cats, small birds, squirrels, and the like.

  The food composition includes various food additives such as antioxidants, pigments, fragrances, seasonings, sweeteners, acidulants, pH adjusters, quality stabilizers, preservatives, etc. These additives may be used alone or in combination.

The food composition can be provided by filling a normal packaging container such as a molded container (PET bottle) containing polyethylene terephthalate as a main component, a metal can, a paper container combined with a metal foil or a plastic film, or a bottle. .
In addition, food compositions are manufactured under the sterilization conditions stipulated in the applicable regulations (Food Sanitation Law in Japan) if they can be sterilized by heating after filling in containers such as metal cans. can do. For those that cannot be sterilized by retort, such as PET bottles and paper containers, the above sterilization conditions are preliminarily sterilized at high temperature and short time in a plate heat exchanger, etc. Can be adopted.

The content of the active ingredient in the skin sympathetic nerve activity enhancer, skin microcirculation inhibitor, skin sympathetic nerve activity inhibitor and skin microcirculation enhancer of the present invention containing the active ingredient is appropriately determined depending on the use form. be able to.
For example, in the case of oral solid preparations such as tablets, coated tablets, granules, powders, capsules, and oral liquid preparations such as oral liquids and syrups, the oral skin microcirculation regulator or oral skin sympathetic nerve of the present invention. In the total amount of the activity control agent, the content of the active ingredient is preferably 0.001% by mass or more, more preferably 0.01% by mass or more, and further preferably 0.1% by mass or more as solid content concentration (solid content conversion). The upper limit is preferably 10% by mass or less, more preferably 1% by mass or less, and further preferably 0.5% by mass or less. Furthermore, 0.001-10 mass% is preferable, as for the numerical range of content of the said active ingredient, 0.01-1 mass% is more preferable, and 0.1-0.5 mass% is further more preferable.
When the oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent of the present invention is blended in food and drink, pet food, etc., the total amount of the oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent of the present invention The content of the active ingredient is preferably 0.0001% by mass or more, more preferably 0.001% by mass or more, further preferably 0.01% by mass or more, and the upper limit is 1% by mass as the solid content concentration. The following is preferable, 0.1 mass% or less is more preferable, and 0.05 mass% or less is further more preferable. Furthermore, 0.0001-1 mass% is preferable, as for the numerical range of content of the said active ingredient, 0.001-0.1 mass% is more preferable, and 0.01-0.05 mass% is further more preferable.

  The subject of administration or ingestion of the oral skin microcirculation controlling agent or oral skin sympathetic nerve activity controlling agent of the present invention is preferably a warm-blooded vertebrate, more preferably a mammal. As used herein, mammals include, for example, humans and non-human mammals such as monkeys, mice, rats, rabbits, dogs, cats, cows, horses, and pigs. The oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent of the present invention is suitable for administration to humans.

The oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent of the present invention is a subject who desires control of skin microcirculation or skin sympathetic nerve activity, skin microcirculation control or skin sympathetic nerve activity control. The present invention can be preferably applied to a required subject or a desired subject. Alternatively, it can be preferably applied to a person who desires the prevention or improvement effect of telangiectasia, the prevention or improvement effect of poor circulation, or the prevention or improvement effect of cold syndrome, and a preliminary group of the above symptoms.
Moreover, the oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent of the present invention can be preferably applied under conditions where normal skin sympathetic nerve activity and skin microcirculation are obstructed.

  In the method for controlling skin sympathetic nerve activity or skin microcirculation using the oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent of the present invention, the active ingredient applied by administering or ingesting the control agent Administration or intake can be appropriately determined according to the individual's condition, body weight, sex, age, material activity, administration or intake route, administration or intake schedule, formulation form, or other factors. For example, the administration or intake of the active ingredient is preferably 0.001 mg / kg body weight / day or more, more preferably 0.01 mg / kg body weight / day or more, further preferably 0.1 mg / kg body weight / day or more, The upper limit is preferably 100 mg / kg body weight / day or less, more preferably 10 mg / kg body weight / day or less, and further preferably 1 mg / kg body weight / day or less. Further, the numerical range of administration or intake of the active ingredient is preferably 0.001 to 100 mg / kg body weight / day, more preferably 0.01 to 10 mg / kg body weight / day, and 0.1 to 1 mg / kg body weight. / Day is more preferred. The active ingredient can be divided into once to several times a day, or can be administered or taken at an arbitrary period and interval.

  In relation to the above-described embodiments, the present invention further describes in detail the following screening method, skin sympathetic nerve activity or skin microcirculation control method, oral skin microcirculation control agent or skin sympathetic nerve activity control agent, and production method. .

<1> In the presence of a candidate substance for an oral regulator of skin sympathetic nerve activity or skin microcirculation, the ability to control TRPV4 activity is measured, and the candidate substance capable of controlling TRPV4 activity is taken orally for skin sympathetic nerve activity or skin microcirculation. A screening method for an oral regulator of skin sympathetic nerve activity or skin microcirculation, which is selected as a regulator.
<2> The screening method according to <1>, wherein the TRPV4 activity is gastrointestinal TRPV4 activity.
<3> The screening method according to <1> or <2>, wherein the TRPV4 or gastrointestinal tract TRPV4 is human, rat or mouse-derived TRV4 or gastrointestinal TRPV4.
<4> The screening method according to any one of <1> to <3>, wherein TRPV4 or digestive tract TRPV4 expression cultured cells are used to measure TRPV4 or digestive tract TRPV4 activity.
<5> The screening method according to any one of <2> to <4>, wherein the control of the gastrointestinal TRPV4 activity controls skin sympathetic nerve activity or skin microcirculation via afferent vagus nerve activity.

<6> A method for controlling skin sympathetic nerve activity or skin microcirculation by controlling TRPV4 activity by oral administration or ingestion.
<7> The method according to <6>, wherein the TRPV4 activity is gastrointestinal TRPV4 activity.
<8> The method according to <7>, wherein the control of gastrointestinal TRPV4 activity controls skin sympathetic nerve activity or skin microcirculation via afferent vagus nerve activity.
<9> The method according to any one of <6> to <8>, wherein the TRPV4 activity enhancer or the digestive tract TRPV4 activity enhancer is orally administered or ingested.
<10> The TRPV4 activity enhancer or gastrointestinal TRPV4 activity enhancer is one or more selected from the group consisting of 4α-PDD, 5,6-EET, RN-1747, and GSK1016790A, Item 9>.
<11> The method according to any one of <6> to <8>, wherein the TRPV4 activity inhibitor or the digestive tract TRPV4 activity inhibitor is orally administered or ingested.
<12> The method according to <11>, wherein the TRPV4 activity inhibitor or the gastrointestinal TRPV4 activity inhibitor is one or more selected from the group consisting of RN1734, GSK205, and HC067047.
<13> TRPV4 activity regulator or gastrointestinal TRPV4 activity regulator is required to enhance or inhibit skin sympathetic nerve activity or skin microcirculation, human or animal desiring to enhance or inhibit skin sympathetic nerve activity or skin microcirculation <6> to <12> applied to humans or animals, humans who desire prevention or improvement effect of telangiectasia, prevention or improvement of poor circulation, or prevention or improvement effect of cold syndrome, or the preliminary group of the above symptoms The method of any one of these.
<14> The administration or intake of the active ingredient of the TRPV4 activity regulator or gastrointestinal TRPV4 activity regulator is 0.001 mg / kg body weight / day or more, preferably 0.01 mg / kg body weight / day or more, more preferably 0.1 mg / kg body weight / day or more, 100 mg / kg body weight / day or less, preferably 10 mg / kg body weight / day or less, more preferably 1 mg / kg body weight / day or less, <6> to <13> The method according to any one of the above.
<15> The method according to any one of <6> to <14>, wherein the method is non-therapeutic.

<16> An oral skin microcirculation control agent or an oral skin sympathetic nerve activity control agent comprising a TRPV4 activity control agent as an active ingredient.
<17> The oral skin microcirculation control agent or the oral skin sympathetic nerve activity control agent according to <16>, wherein the TRPV4 activity control agent is a gastrointestinal TRPV4 activity control agent.
<18> The oral skin microcirculation regulator or the oral skin sympathetic nerve according to <17>, wherein the gastrointestinal TRPV4 activity regulator controls skin sympathetic nerve activity or skin microcirculation via afferent vagus nerve activity Activity control agent.
<19> The above <16> to <16, wherein the TRPV4 activity regulator or gastrointestinal TRPV4 activity regulator is a TRPV4 activity enhancer, a gastrointestinal TRPV4 activity enhancer, or a TRPV4 activity inhibitor or a gastrointestinal TRPV4 activity inhibitor. The oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent according to any one of 18>.
<20> The TRPV4 activity enhancer or gastrointestinal TRPV4 activity enhancer is one or more selected from the group consisting of 4α-PDD, 5,6-EET, RN-1747, and GSK1016790A, The oral skin microcirculation regulator or skin sympathetic nerve activity regulator according to item 19>.
<21> The oral skin microcirculation regulator according to <19>, wherein the TRPV4 activity inhibitor or gastrointestinal TRPV4 activity inhibitor is one or more selected from RN1734, GSK205, and HC067047 Skin sympathetic nerve activity control agent.
<22> In the total amount of the oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent, the content of the active ingredient is 0.001% by mass or more, preferably 0. <16> to <21, which is 01% by mass or more, more preferably 0.1% by mass or more, 10% by mass or less, preferably 1% by mass or less, and more preferably 0.5% by mass or less. > The oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent according to any one of the above.
<23> In the total amount of the oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent, the content of the active ingredient is 0.0001% by mass or more, preferably 0. 001% by mass or more, more preferably 0.01% by mass or more, 1% by mass or less, preferably 0.1% by mass or less, more preferably 0.05% by mass or less, <16> to <22> The oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent according to any one of <22>.

<24> Use of a gastrointestinal TRPV4 activity regulator as an oral skin microcirculation regulator or an oral skin sympathetic nerve activity regulator having an action via afferent vagus nerve activity.
<25> Use of a digestive tract TRPV4 activity regulator for the production of an oral skin microcirculation regulator or an oral skin sympathetic nerve activity regulator having an action via afferent vagus nerve activity.
<26> A method of using a gastrointestinal TRPV4 activity control agent having an action via afferent vagus nerve activity as an oral skin microcirculation control agent or an oral skin sympathetic nerve activity control agent.
<27> A method for controlling skin microcirculation or skin sympathetic nerve activity, which applies a gastrointestinal TRPV4 activity regulator having an action via afferent vagus nerve activity.

<28> The use or method according to any one of <24> to <27>, wherein the gastrointestinal TRPV4 activity regulator is a gastrointestinal TRPV4 activity enhancer.
<29> The <28> item, wherein the gastrointestinal TRPV4 activity enhancer is one or more selected from the group consisting of 4α-PDD, 5,6-EET, RN-1747, and GSK1016790A. Use or method.
<30> The use or method according to any one of <24> to <27>, wherein the gastrointestinal TRPV4 activity regulator is a gastrointestinal TRPV4 activity inhibitor.
<31> The use or method according to <30>, wherein the gastrointestinal TRPV4 activity inhibitor is one or more selected from RN1734, GSK205, and HC067047.
<32> Human or animal desiring to enhance or inhibit skin sympathetic nerve activity or skin microcirculation, human or animal in need of enhancement or inhibition of skin sympathetic nerve activity or skin microcirculation, Or any one of the above <24> to <31>, which is applied to a human who desires the prevention or improvement effect of telangiectasia, the prevention or improvement effect of poor circulation, or the prevention or improvement effect of cold syndrome, or the preliminary group of the above symptoms Use or method according to paragraph.
<33> In the total amount of the oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent, the content of the active ingredient is 0.001% by mass or more, preferably 0. <24> to <32, which is 01% by mass or more, more preferably 0.1% by mass or more, 10% by mass or less, preferably 1% by mass or less, and more preferably 0.5% by mass or less. > Use or method of any one of>.
<34> In the total amount of the above-mentioned oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent, the content of the active ingredient is 0.0001% by mass or more, preferably 0. 001% by mass or more, more preferably 0.01% by mass or more, 1% by mass or less, preferably 0.1% by mass or less, more preferably 0.05% by mass or less, <24> to The use or method according to any one of <32>.

<35> A gastrointestinal TRPV4 activity control agent having an action via afferent vagus nerve activity, which is used for a method for controlling skin microcirculation or skin sympathetic nerve activity.
<36> Use of a gastrointestinal TRPV4 activity regulator having an action via afferent vagus nerve activity for the manufacture of a drug for controlling skin microcirculation or skin sympathetic nerve activity.
<37> Use of a gastrointestinal TRPV4 activity regulator having an action through afferent vagus nerve activity, which is used for a non-therapeutic treatment method for controlling skin microcirculation or skin sympathetic nerve activity.

<38> The digestive tract TRPV4 activity control agent or the use according to any one of <35> to <37>, wherein the digestive tract TRPV4 activity control agent is a digestive tract TRPV4 activity enhancer.
<39> The <38> item, wherein the gastrointestinal TRPV4 activity enhancer is one or more selected from the group consisting of 4α-PDD, 5,6-EET, RN-1747, and GSK1016790A. Of gastrointestinal TRPV4 activity regulator or use.
<40> The gastrointestinal TRPV4 activity regulator or the use according to any one of <35> to <37>, wherein the gastrointestinal TRPV4 activity regulator is a gastrointestinal TRPV4 activity inhibitor.
<41> The gastrointestinal TRPV4 activity regulator or use according to <40>, wherein the gastrointestinal TRPV4 activity inhibitor is one or more selected from RN1734, GSK205, and HC067047.
<42> Human or animal desiring to enhance or inhibit skin sympathetic nerve activity or skin microcirculation, human or animal in need of enhancement or suppression of skin sympathetic nerve activity or skin microcirculation, Or any one of the above <35> to <41>, applied to a human who desires prevention or improvement effect of telangiectasia, prevention or improvement of poor circulation, or prevention or improvement effect of cold syndrome, or a preliminary group of the above symptoms The digestive tract TRPV4 activity control agent or use according to Item.
<43> The digestive tract TRPV4 activity control agent or the use according to any one of <35> to <42>, wherein the digestive tract TRPV4 activity control agent is applied in the form of a pharmaceutical composition.
<44> The content of the gastrointestinal TRPV4 activity control agent is 0.001% by mass or more, preferably 0.01% by mass or more, more preferably 0, as the solid content concentration (in terms of solid content) in the total amount of the agent. 1% by mass or more, 10% by mass or less, preferably 1% by mass or less, more preferably 0.5% by mass or less, or the digestive tract TRPV4 activity control agent or use according to item <43> .
<45> The gastrointestinal TRPV4 activity control agent or the use according to any one of <35> to <42>, wherein the gastrointestinal TRPV4 activity control agent is applied in the form of food, beverage, or feed.
<46> The content of the gastrointestinal tract TRPV4 activity control agent is 0.0001% by mass or more, preferably 0.001% by mass or more, more preferably 0 as solid content concentration (solid content conversion) in the total amount of the agent. 0.01% by mass or more, 1% by mass or less, preferably 0.1% by mass or less, more preferably 0.05% by mass or less, The digestive tract TRPV4 activity control agent according to <45>, Or use.

  Hereinafter, although the present invention is explained in detail based on an example, the present invention is not limited to this.

[Test Example 1] Effect of oral administration of TRPV4 activity enhancer on rat afferent vagus nerve Rats (SHR, male, 11 weeks old) were fasted for 18 hours and then urethane (manufactured by Wako Pure Chemical Industries, Ltd.) , 1 g / kg body weight, i.p.) under anesthesia and exposing the right subdiaphragmatic vagus nerve to the method of Niijima et al. (Niijima A., et al., Physiol. Behav., 1991, vol.49 (5), p.1025-1028), afferent nerve activity was recorded. From the start of surgery to the end of measurement of afferent nerve activity, a decrease in body temperature was prevented with a heat retention device.

Specifically, after inserting a polyethylene catheter (Becton Dickinson) filled with heparin (Nipro, 100 units / mL) into the right femoral vein, a midline incision was made while securing the airway in the supine position, and the left and right subdiaphragmatic vagus nerve And the right subdiaphragmatic vagus nerve was detached from the esophagus. After exfoliating the vagus nerve, the right subdiaphragm vagus nerve is cut directly under the diaphragm (central side), and a Teflon (registered trademark) coated bipolar stainless wire electrode (As634, manufactured by Cooner Wire) is used as the central cutting terminal. They were connected and insulated and fixed using silicon (Sil-Gel 604, manufactured by Wacker) to obtain afferent vagus nerve activity.
Next, a Nipro nutrition catheter (Nipro Corporation) was orally inserted into the stomach for material administration. Then, in order to prevent an increase in gastric pressure more than necessary due to the administration material, a silicon tube having an inner diameter of 3 mm and an outer diameter of 5 mm was inserted from the stomach pylorus to promote excretion of the stomach contents. After acclimation for 1 hour or more after the operation, the stomach was washed with 10 mL of physiological saline before material administration. Then, after resting for 15 minutes or more, a known TRPV4 activity inhibitor (HC067074, manufactured by Sigma-aldrich, 4 mmol / kg body weight) dissolved in 0.05% NaCl aqueous solution containing 1% DMSO, and known TRPV4 activity enhancement The agent (GSK1016790A, manufactured by Sigma-aldrich, 0.4 μmol / kg body weight) was orally administered (4 mL / kg body weight) at 1 mL / min. As a control, 0.05M NaCl aqueous solution containing 1% DMSO was used. After completion of the test, a saturated potassium chloride aqueous solution was intravenously administered through a femoral vein catheter and euthanized.

The afferent vagus nerve activity uses a preamplifier (JC-101B, manufactured by Nihon Kohden) and a high-sensitivity amplifier (MEG-5100, manufactured by Nihon Kohden) with a built-in bandpass filter (low cut: 150 Hz, high cut: 1 kHz). The sample was amplified and connected to a polygraph system (LEG-1000, manufactured by Nihon Kohden) and a thermal array recorder (RTA-1100, manufactured by Nihon Kohden), and data was recorded at a sampling rate of 4 k / s.
Afferent vagus nerve activity was obtained only when excitement of nerve activity was observed during euthanasia by intravenous administration of saturated potassium chloride aqueous solution. The nerve activity value after 20 minutes after euthanasia by administration of a saturated potassium chloride aqueous solution to the femoral vein was defined as noise. In addition, for the analysis of afferent vagus nerve activity, the value of nerve activity minus noise was used. For analysis of afferent vagus nerve activity, the average value of the integrated nerve activity for 14.4 seconds at each time point was used, and the rate of change with respect to rest (average value for 5 minutes before administration every minute). (%) Was calculated.
The obtained results are shown in FIG. In FIG. 1, the afferent vagus nerve activity is represented by (mean value ± standard error). The test of the difference of the average value between groups performed ANOVA, and then performed Student-Newman-Keuls test.

As shown in FIG. 1, the oral administration of the TRPV4 activity enhancer significantly increased the afferent vagus nerve activity at 3 to 5 minutes and 15 minutes after the administration of the TRPV4 activity enhancer compared to the control administration group. It was increased.
Moreover, the enhancement of afferent vagus nerve activity due to the action of the TRPV4 activity enhancer was significantly suppressed by the combined administration of the TRPV4 activity inhibitor.

[Test Example 2] Effect of oral administration of TRPV4 activity enhancer on skin sympathetic nerve activity via afferent vagus nerve Rats (SHR, male, 10 weeks old) were treated with pentobarbital (Kyoritsu Pharmaceutical Co., Ltd., 50 mg / kg). Under the anesthesia of body weight, i.p.), a midline incision operation is performed, and the connecting branch of the subdiaphragm vagus nerve parallel to the esophagus is excised, and the subdiaphragm vagus nerve on the central side from the liver branch is removed from the esophagus About 5 mm was excised bilaterally. After excision, the incision was sutured with a suture. Control rats underwent sham surgery with the same procedure except excision of the vagus nerve.
The skin sympathetic nerve activity shown below was measured after a recovery period of 1 week or more after the operation. In addition, from the start of surgery to the end of the measurement of skin sympathetic nerve activity, a decrease in body temperature was prevented with a heat retaining device.

Rats subjected to surgery were fasted for 18 hours, and then heparin (Nipro, 100 unit / mL) under anesthesia with urethane (Wako Pure Chemical Industries, 1 g / kg body weight, ip). A filled polyethylene catheter (Becton Dickinson) was inserted into the right femoral vein. Thereafter, an incision was made in the outer thigh region to expose the tibial nerve containing the skin sympathetic nerve governing the skin blood flow and sweat glands on the heel, and cut on the distal side (toe side). Next, the distal end of the cut tibial nerve is connected to a Teflon (registered trademark) -coated bipolar stainless steel wire electrode (As634, manufactured by Cooner Wire), and silicon (Sil-Gel 604, manufactured by Wacker) The tibial nerve activity was acquired with insulation fixation.
Next, a Nipro nutrition catheter (Nipro Corporation) was orally inserted into the stomach for material administration. Then, after acclimatization for 1 hour or more, a known TRPV4 enhancer (GSK1016790A, manufactured by Sigma-aldrich, 0.4 μmol / kg body weight) dissolved in 0.05% NaCl aqueous solution containing 1% DMSO is 0.5 mL / min. Orally (4 mL / kg body weight). The control received 0.05M NaCl aqueous solution containing 1% DMSO.

Tibial nerve activity is amplified using a preamplifier (JC-101B, manufactured by Nihon Kohden) and a high-sensitivity amplifier (MEG-5100, manufactured by Nihon Kohden) with a built-in band-pass filter (low cut: 50 Hz, high cut: 1 kHz). The data was output to a multipurpose data acquisition system (PowerLab 4/30, manufactured by AD Instruments) and recorded at a sampling rate of 10 k / s. The acquired tibial nerve activity is further removed using a band-pass filter (low cut: 250 Hz, high cut: 400 Hz) of LabChart analysis software (ver 7.0, ADInstruments), and the obtained tibial nerve activity is sympathetic to the skin. Nervous activity.
Cutaneous sympathetic nerve activity is reduced by intravenous administration of hexamethonium bromide (13.5 mg / kg body weight, manufactured by Wako Pure Chemical Industries, Ltd.), an autonomic ganglion inhibitor, before euthanasia. In addition, it was obtained only when excitement of neural activity was observed during euthanasia by intravenous administration of a saturated aqueous potassium chloride solution. The nerve activity value after 20 minutes after euthanasia by intravenous administration of a saturated potassium chloride aqueous solution was defined as noise. Moreover, the nerve activity value which subtracted the noise was used for the analysis of the skin sympathetic nerve activity. For the analysis of the skin sympathetic nerve activity, the average value of the integrated nerve activity for 1 minute at each time point is used, and the change rate (%) with respect to the rest time (average value for 10 minutes before administration every minute). Was calculated.
The obtained results are shown in FIG. In FIG. 2, the skin sympathetic nerve activity is represented by (mean value ± standard error). The test of the difference of the average value between groups performed the F test, followed by Student's t test in the case of equal variance, and Welch's t test in the case of unequal variance.

As shown in FIG. 2 (A), when a TRPV4 activity enhancer was administered in the sham-operated group of rats, the skin sympathetic nerve activity was enhanced compared to the control administration. In particular, the skin sympathetic nerve activity was significantly enhanced at 1 minute, 5 minutes, 20 minutes, and 30 minutes after administration of the TRPV4 activity enhancer.
Moreover, the enhancement of the skin sympathetic nerve activity by the action of the TRPV4 activity enhancer was not observed in the rats subjected to vagus nerve excision as shown in FIG. 2 (B). Therefore, it became clear that the increase in skin sympathetic nerve activity by administration of the TRPV4 activity enhancer is via the vagus nerve.

[Test Example 3] Effect of oral administration of TRPV4 activity enhancer on skin microcirculation via afferent vagus nerve Rats (SHR, male, 10 weeks old) were treated with pentobarbital (Kyoritsu Pharmaceutical Co., Ltd., 50 mg / kg body weight). , I.p.) Under anesthesia under anesthesia, excision of the connecting branch of the subdiaphragmatic vagus nerve parallel to the esophagus and removal of the central subpharyngeal vagus nerve from the liver branch from the esophagus About 5mm was resected. After excision, the incision was sutured with a suture. Control rats underwent sham surgery with the same procedure except excision of the vagus nerve.
Skin microcirculation can be evaluated from the total blood flow of arterioles, capillaries, and venules in the skin. Therefore, in order to evaluate the skin microcirculation, the skin blood flow shown below was measured after a recovery period of 1 week or more after the operation.

Rats that underwent surgery were fasted for 18 hours and then contacted laser Doppler blood flow meter (ALF21, Advanced) under anesthesia with urethane (manufactured by Wako Pure Chemical Industries, 1 g / kg body weight, ip) The total skin blood flow of the arterioles, capillaries and venules of the epidermis on the back of the starboard was measured.
Next, a Nipro nutrition catheter (Nipro Corporation) was orally inserted into the stomach for material administration. Then, after acclimatization for 1 hour or more, a known TRPV4 enhancer (GSK1016790A, manufactured by Sigma-aldrich, 0.4 μmol / kg body weight) dissolved in 0.05% NaCl aqueous solution containing 1% DMSO is 0.5 mL / min. Orally (4 mL / kg body weight). The control received 0.05M NaCl aqueous solution containing 1% DMSO.

Skin blood flow was recorded at a sampling rate of 1 k / s using a multipurpose data acquisition system (PowerLab 4/30, manufactured by AD Instruments). For the analysis, the average value of skin blood flow for 1 minute at each time point was used to calculate the rate of change (%) with respect to resting time (average value for 10 minutes before administration every minute).
The obtained results are shown in FIG. In FIG. 3, the skin blood flow was expressed as (mean value ± standard error). The test of the difference of the average value between groups performed the F test, followed by Student's t test in the case of equal variance, and Welch's t test in the case of unequal variance.

As shown in FIG. 3 (A), in the sham-operated rat, when the TRPV4 activity enhancer was administered, the skin blood flow was reduced compared to the control administration. In particular, at 3 minutes, 5 minutes, and 10 minutes after administration of the TRPV4 activity enhancer, skin blood flow was significantly reduced.
In addition, a decrease in blood flow due to the action of the TRPV4 activity enhancer was not observed in rats subjected to vagus nerve excision as shown in FIG. Therefore, it was clarified that by oral administration, the gastrointestinal tract TRPV4 was enhanced and the skin blood flow decreased via the vagus nerve.

According to the results of Test Examples 1 to 3, it was revealed that skin sympathetic nerve activity and skin microcirculation can be controlled via the afferent vagus nerve by controlling the gastrointestinal TRPV4 activity.
In addition to the efferent nerve that transmits information from the center to the internal organs, the vagus nerve below the diaphragm also plays the role of the afferent nerve that transmits information on the internal organs mainly composed of the digestive tract to the center. As shown in the above test examples, oral administration of a TRPV4 activity enhancer shows an increase in afferent vagal activity. Furthermore, the skin sympathetic nerve activity enhancing action and the skin microcirculation inhibiting action by oral administration of the TRPV4 activity enhancer disappears when the vagus nerve is excised. Therefore, it is considered that only when the TRPV4 activity enhancer acts on the digestive tract TRPV4, it affects the skin sympathetic nerve and the skin microcirculation.

Claims (13)

  In the presence of a candidate substance for an oral regulator of skin sympathetic nerve activity or skin microcirculation, TRPV4 activity control ability is measured, and the candidate substance capable of controlling TRPV4 activity is used as an oral regulator of skin sympathetic nerve activity or skin microcirculation. A screening method for an oral regulator of skin sympathetic nerve activity or skin microcirculation to be selected.   The screening method according to claim 1, wherein the TRPV4 activity is gastrointestinal TRPV4 activity.   The screening method according to claim 2, wherein the control of the gastrointestinal TRPV4 activity controls skin sympathetic nerve activity or skin microcirculation via afferent vagus nerve activity.   A non-therapeutic method of controlling skin sympathetic nerve activity or skin microcirculation by controlling TRPV4 activity by oral administration or ingestion.   5. The method of claim 4, wherein the TRPV4 activity is gastrointestinal TRPV4 activity.   6. The method of claim 5, wherein the control of gastrointestinal TRPV4 activity controls skin sympathetic nerve activity or skin microcirculation via afferent vagus nerve activity.   An oral skin microcirculation control agent or an oral skin sympathetic nerve activity control agent comprising a TRPV4 activity control agent as an active ingredient.   The oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent according to claim 7, wherein the TRPV4 activity control agent is a gastrointestinal TRPV4 activity control agent.   The oral skin microcirculation control agent or oral skin sympathetic nerve activity control agent according to claim 8, wherein the gastrointestinal TRPV4 activity control agent controls skin sympathetic nerve activity or skin microcirculation via afferent vagus nerve activity.   The oral skin microcirculation regulator or oral skin according to any one of claims 7 to 9, wherein the TRPV4 activity regulator or gastrointestinal TRPV4 activity regulator is a TRPV4 activity enhancer or a gastrointestinal TRPV4 activity enhancer. Sympathetic nerve activity control agent.   The TRPV4 activity enhancer or gastrointestinal TRPV4 activity enhancer is 12,13-didecanoic acid 4α-phorbol, 5,6-epoxyeicosatrienoic acid, 1-benzyl-4- (4-chloro-2-nitrophyll) Sulfonylbenzylpiperazine, and N-[(2S) -1- [4-[(2S) 2-[(2,4-dichlorophenyl) sulfonylamino] -3-hydroxypropanoyl] piperazin-1-yl] -4- The oral skin microcirculation control agent or oral skin according to claim 10, which is one or more selected from the group consisting of methyl-1-oxopentan-2-yl] -1-benzothiophene-2-carboxamide Sympathetic nerve activity control agent.   The oral skin microcirculation regulator or oral skin according to any one of claims 7 to 9, wherein the TRPV4 activity regulator or gastrointestinal TRPV4 activity regulator is a TRPV4 activity inhibitor or a gastrointestinal TRPV4 activity inhibitor. Sympathetic nerve activity control agent. The TRPV4 activity inhibitor or gastrointestinal TRPV4 activity inhibitor is 2,4-dichloro-N-propan-2-yl-N- [2- (propan-2-ylamino) ethyl] benzenesulfonamide, N- [4 -[2- [benzyl (methyl) amino] ethyl] phenyl] -5-pyridin-3-yl-1,3-thiazol-2-amine; hydrobromide, and 2-methyl-1- (3-morpholine-4 The oral skin micro of claim 12, which is one or more selected from the group consisting of -ylpropyl) -5-phenyl-N- [3- (trifluoromethyl) phenyl] pyrrole-3-carboxamide. Circulation control agent or oral skin sympathetic nerve activity control agent.

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