KR102087492B1 - Composition for treatment or improvement of sciatica that comprising GABA or GABA agonist - Google Patents

Abstract

The present invention relates to a pharmaceutical composition for treating and alleviating sciatica, including GABA or GABA agonist which can be directly injected into ZI of a sciatica patient, and a method of treatment using the composition. As a result of this study, it was confirmed that the signal of GABA, a neurotransmitter, is decreased in the brain's indeterminate zone (ZI, zona incerta) of patients with sciatica. It can be cured.

Description

Composition and treatment for the treatment and alleviation of sciatica, including GABA and GABA agonists. {Composition for treatment or improvement of sciatica that comprising GABA or GABA agonist}

The present invention relates to a pharmaceutical composition for treating and alleviating sciatica, including GABA or GABA agonist which can be directly injected into ZI of sciatica patients, and a method of treatment using the composition. Since the signal of the neurotransmitter GABA is reduced in the zona incerta (ZI, zona incerta) of the brain of patients with sciatica, the present invention includes a pharmaceutical composition that can be injected into ZI to treat the disease and a method of treatment .

Neuropathic pain is usually caused by peripheral nerve damage (e.g., due to postoperative pain or radiation), central nervous system damage (e.g. multiple sclerosis, spinal cord injury), viral infections (e.g. shingles after shingles), tumors, etc. Can be. Neuropathic pain is more severe than other types of chronic pain in severity and duration, and 5% of patients complain of pain despite administration of analgesics. Thus, neuropathic pain is one of the most difficult clinical outcomes and cannot be controlled with standard pain medications and is resistant to numerous drugs. Chronic neuropathic pain due to peripheral nerve injury can be identified as a problem according to the current treatment. Patients with neuropathic pain complain of symptoms such as shooting and burning, pain such as electric shock, tingling, numbness, numbness, numbness of hands and feet, allodynia and hyperalgesia. Chronic constriction injury (CCI) of the sciatic nerve is commonly used to generate a rat model of neuropathic pain. The cause of CCI-induced pain is not known precisely, but chronic neuropathic pain is thought to be due to inhibition of GABA (γ-Aminobutyric acid (GABA)) mediated in the abnormal indeterminate zone (zona incerta, ZI) in the thalamus. Because of the varying degrees of action of GABA in each area of the brain, it is very complex to identify mechanisms for controlling pain states caused by the action of GABA.

Many previous studies have reported an association between brain uncertainty (ZI) and chronic pain. ZI is a GABAergic 'nucleus located in the hypothalamus, which is the granular part of the brain stem, and most ZI neurons are GABA. Interneurons have inhibitory pathways such as GABA-like pathways that affect the balance between excitatory and inhibitory neurons in synaptic transmission of ZI. Recently, Masri et al. Described a novel system for the regulation of stimulus response processes that inhibit the flow of stimulatory or sensory information in the posterior thalamus, such as ZI, which is the cytoplasmic and neurochemically dividing cell population in the thalamus. It was. That is, ZI regulates both spontaneous and induced activity in the posterior thalamus, ZI inactivation reduces response latency in a subset of the postthalamic nucleus, and ZI is a pathway that coordinates the somatosensory and internal sensory receptor systems. Include both (primary and secondary).

Therefore, the study of GABA signaling system, the diagnosis of neuropathic pain, especially sciatica and the development of a method (composition) for treating it is required.

In order to test GABA and GABA sex signals, the present inventors investigated the change of GABA concentration and GABA sex ZI neuron activity in vivo using a rat model of sciatic nerve CCI. Rat sciatic nerve CCI exhibits relatively consistent neuropathic pain behavior that mimics neuropathic pain due to peripheral nerve damage in humans. Pain-related behavior and neuronal activity were assessed after injecting agonists and antagonists against GABA receptors in ZI

The role of GABA in the mediation and perception of pain, Enna SJ et al GABA mechanisms and antinociception in mice with ligated sciatic nerve, Zarrindast MR et al.

An object of the present invention is to provide a pharmaceutical composition for treating and alleviating sciatica, including GABA or GABA agonist as an active ingredient.

It is another object of the present invention to provide a sciatica treatment method comprising the step of injecting a composition comprising the above components into the brain uncertain region (ZI).

In order to solve the above problems, the present invention provides a sciatic nerve treatment and palliative pharmaceutical composition comprising a GABA or GABA agonist as an active ingredient.

In another aspect, the present invention provides a method of treating sciatica, comprising injecting a GABA or a pharmaceutical composition comprising a GABA agent as an active ingredient into a brain indeterminate region.

The sciatica treatment and sciatica treatment comprising the GABA or GABA agonist of the present invention or sciatica treatment method comprising the step of injecting the pharmaceutical composition into the indeterminate area of the brain has a GABA or GABA action in relation to the disease I designate specific areas of the brain where I work, and suggest precise treatments for them. Therefore, compared with the known invention, the therapeutic effect on sciatica has excellent features.

1 is a schematic of the overall flow of the present study.
Figure 2 shows the results of measuring the change in PWT (withdrawal threshold) (A) and the change in PWL (foot hide) (B) in the animal model induced CCI. In the case of PWT, the CCI-induced threshold value was lowered over time, and it was confirmed that pain hypersensitivity was induced when the dorsal thigh nerve injury occurred. In the case of PWL, it was confirmed that it decreased over time.
3 shows the results of analyzing neurotransmitters of ZI using in vivo microdialysis.
Figure 4 is a result showing the change in pain hypersensitivity caused by agonist, antagonist or saline in CCI induced animal model and placebo treatment group.
5 shows spontaneous neuronal activity in ZI of CCI-induced and placebo treated animal models.

This study deals with neuropathic pain associated with GABA neuronal activity of neurotransmitters in the inactive region of the brain. Studies have shown that neuronal activity in ZI is significantly lower in rats that cause cyclical striction injury (CCI) in sciatic nerves compared to placebo-treated groups. Therefore, treatment with GABAA agonists reduces PWT and PWL and increases the ignition rate of ZI in a rat model in which the thigh nerves are injured. All these data suggest that inhibition of GABA neuronal activity is a major cause of neuropathic pain by CCI.

Previous studies have suggested neural circuits from ZI to hypothalamic relays including hypothalamic relays. ZI protrudes into a large number of sagittal nuclei and affects tonic-suppressive feed-forward in the thalamus. Decreased neuronal activity in ZI in a rat model of neuropathic pain caused by spinal cord injury has been demonstrated in previous studies. Reduction of ZI activity is associated with a pathological increase in neuronal activity of the somatosensory thalamus nuclei, which is important for processing posterior thalamus and invasive receptive information. The regulation of ZI firing rate and synaptic GABA concentrations may be an effective means of controlling the incerto-thalamic circuit in computerized studies. This indicates that GABAergic regulation in ZI alleviates neuropathic pain in vivo.

This pathway is inhibitory (GABA) and has the potential to act as a regulator of hypothalamic delivery. ZI receives topographically structured projections from the deep layer sensory trigeminal nucleus complexes and dorsal column nuclei of the brain's superiorcolliculus, mainly the brain's superior and pretectal nuclei, contralateral ZI and The same high-level thalamus nucleus is mainly included in GABA neurons. ZI neurons stimulate various brain regions, such as the sagittal nucleus and cerebral cortex, and GABA-determined neurons can induce transient deinhibition in the brain's indeterminate projective regions by microwave stimulation or injection of musimol. The complex active region at the ZI terminus has been found to be very strongly influenced by the ignition properties of thalamic neurons, which are delivered through transmissive cells to various brain cortical regions and are widely projected into the neocortex. The GABA-like ends of ZI selectively stimulate the proximal dendrites of associated neurons, mainly in the higher nucleus of the thalamus. ZI forms the center of the hepatic brain, a narrow area wedge-shaped below the hypothalamus, and produces an immediate response (instinct, excitement, concentration, or post-motor) to sensory stimuli (body and muscle).

CCI of the sciatic nerve is widely used in peripheral chronic neuropathic models. This causes stable hypersensitivity pain at least one month after injury. After inducing CCI of the sciatic nerve, animal models spontaneously show abnormal postures (ie, toe collection, plantar flexion, extraversion, etc.) in the damaged hind paw to check for the presence of pain or repeated tremors of the plantar foot. They showed or protected and licked the soles of their feet. CCI affects the formation of maladaptive synaptic circuits in the spinal dorsal horn, individually or synergistically. These neurological hypersensitivity reactions or central sensitization enhance the spontaneous or induced neurological response to external stimuli provided to the peripheral receptive zone, lower the threshold by activity, increase the size of the peripheral receptive zone, and post response It is characterized by increasing activity.

As discussed, ZI neurons are stimulated by the activity of the thalamus nucleus. Previous studies have shown that neuropathic pain involves the reduction of ZI neurons. Delivery of reduced GABAergic inhibition is responsible for the development of neuropathic pain. Thus, the underlying mechanism for sciatic nerve CCI is under study through the inhibition of gradual apoptosis of GABA neurons in ZI as seen by CCI lesions. Microdialysis has demonstrated that neuronal activity of ZI is controlled by GABA treatment because inactivation of ZI neurons is associated with inhibition of GABA neuronal neurons. GABA and serotonin levels were found to be very low in sciatic nerve CCI-induced animal models. In animals with neuropathic pain, serotonin levels in the ventral thalamus decreased. Presumably, the mechanisms that mediate these changes include changes in the nervous system of GABA, and it can be assumed that treatment with these neurotransmitters, GABA and serotonin, can cure neuropathic pain.

GABA activity outside synapses hyperpolarizes neurons while blocking these receptors to promote ignition. However, other studies have suggested that stimulation of successive cells in the thalamus is associated with nociceptive pain and that stimulation is associated with inhibition of nociceptor information. The transmission of GABA sex in ZI is complex. Cha et al. Said that inactivation of ZI neurons can be suppressed by injecting musimol into ZI, but the results by Moon et al. Are different. It is possible when targeting the ZI complex itself or a posterior thalamus. Petronilho et al. Reported that injection of glutamate with ZI can produce anti-nociceptive solubility. They used an acute pain model to investigate transient antinociceptive efficacy after glutamate administration, but used a chronic pain model in this study. They focused on pain inhibition pathways that descend into the spinal cord along the dorsolateral funiculus using serotonin-, a-adrenergic, and endogenous oipoid-based mechanisms, but we have GABA Emphasis was placed on the suppression of the thalamus by the influence of.

The cause of neuropathic pain has not yet been identified, but chronic neuropathic pain was assumed to be due to abnormal suppression in the thalamus. GABA transmission is known to control the state of excitement and pain, but the underlying mechanisms are extensive, and because GABA neurotransmission is very complex, the study of the underlying mechanism is necessary. Spinothalamic tracts, which deliver important signals for local pain, and spinoreticular tracts, which are involved in the emotional aspects of pain, are the major pain pathways. ZI receives dense nociceptive input through the spinal column and sends exclusively tonic suppression feed forward to the thalamus nucleus, particularly the posterior nucleus of the hypothalamus. ZI generally strongly inhibits the thalamus nucleus, but neuropathic pain after CCI induction is believed to be due to decreased neuronal activity of ZI. Thus, inhibition of ZI activity is associated with a pathological increase in neuronal activity in the posterior thalamus, a somatosensory thalamus important for processing pain information.

As a result of the above research, the present inventors have newly discovered that the action by the neurotransmitter GABA in the ZI region is important in order to suppress neuropathic pain, and the agonist of GABA which can improve the concentration of GABA. The present invention has been completed in that the symptom of neuropathic pain can be alleviated by injecting ZI into ZI.

Accordingly, the present invention provides a pharmaceutical composition for treating and alleviating sciatica, comprising GABA or GABA agonist as an active ingredient, which is injected into ZI.

The GABA agonists include, but are not limited to, benzodiazepines, diazepam, chlorodiazepoxide, barbitruic acid, picrotoxin, steroids, musimol, and the like, and preferably mucimol.

The pharmaceutical composition of the present invention may include an appropriate amount of salts and buffer solutions, in order to maintain the activity of the active ingredient to the maximum, and includes a pharmaceutically acceptable carrier and excipients, lubricants, wetting agents, emulsifiers, preservatives and the like. It can be included without limitation. In addition, the formulations may be in the form of solutions, suspensions or emulsions in oils or aqueous media.

The present invention also provides a method of treating neuropathic pain, comprising injecting GABA or GABA agonists into ZI of the brain.

The injection includes intravenous infusion, oral or subcutaneous administration, direct injection into the brain, but is not limited to the method, preferably means direct injection into ZI of the brain. In addition, a specialized drug injection device may be used for direct injection in the brain, and specifically, includes an Ommaya reservoir and a drug injection catheter.

In addition, in the treatment method of the present invention, a suitable dosage of the drug may be applied to the method of administration of the pharmaceutical composition, according to the formulation method, mode of administration, age, weight, sex, morbidity, food, Depending on factors such as time of administration, route of administration, rate of excretion, and response sensitivity, a skilled practitioner will usually be able to readily determine and prescribe a dosage effective for the desired treatment or prevention. Although not limited thereto, the GABA or GABA agonist of the invention is at a concentration of 0.1 to 1.0 mmol / μl, with a single dose of 1 μl to 5 μl. In an embodiment of the present invention, the GABA or GABA agonist was injected at a concentration of 0.5 mmol / μl (low concentration) or 5 mmol / μl (high concentration), with a single dose of 2 μl. When injected less than the concentration and the amount of the above range, the effect on pain treatment is insignificant, and when injected over the above range, the GABA response may be excessively induced and side effects may occur.

Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are only for illustrating the present invention in more detail, it will be apparent to those skilled in the art that the scope of the present invention is not limited to these examples.

Experimental Materials and Experimental Methods

1. Selection and design of experimental animals

All animal experiments were conducted in accordance with approved national guidelines set by the Chungbuk National University Animal Experiment Ethics Review Committee. Aseptic animal surgical equipment, gloves and sterile curtains and swabs were used in accordance with the guidelines of the International Association of Pain of Study. The skin was disinfected with 70% ethanol and the fur was shaved at the surgical site and covered with sterile drape.

Sprague Dawley male rats (250-300 g, Korea Biolink, Korea, Negative Group) were bred under standard housing conditions (12:12 hour contrast cycle) and in accordance with the guidelines of the National Institutes of Health on Animal Welfare and free food and water in the laboratory. It was dietary. Each rat was housed in a separate cage with flat paper bedding to prevent airway obstruction, following random and double-blind controlled instructions. The timeline of the experiment is as shown in FIG. Sample size was set by PASS 14 software. (NCSS Inc, USA, UT, Kaysville.alpha = 0.05, power = 0.8)

2. CCI Surgery

In this study, we used CCI (n = 20) and falsely treated (n = 10) mice as controls. All animals underwent surgery on the same day. Cages were randomly selected from all pools and grouped. The cages were temporarily numbered and mice were numbered by the registrar. All animals received the same anesthesia, preparation, sedation, and preoperative procedures and were blinded after surgery. For this study, animals were removed from cages and permanently assigned to archives.

3. Behavior Analysis

Mechanical hyperalgesia was measured with a Dynamic Plantar Aesthesiometer (Model 37450, Ugo Basile, Varese, Italy). Mice (n = 30) were placed in a Plexiglas cage (20 × 20 × 14 cm) with a grid bottom and allowed to rest for at least 40 minutes. CCI lesions were subjected to mechanical stimulation by continuously increasing pressure (5 g / s) on the ipsilateral and contralateral forefoot heel region. Physiological saline, mucimol, GABA agonist (0.5 mmol / μL [0.28525 μg] or 5 mmol / μL [2.8525 μg]) and GABA antagonist bicuculline (0.5 μmol / μL [0.0009185 μg] or 5 μmol / μL [0.009185 μg] ]) Were injected 2μL each, and then Paw Withdrawal Threshold (PWT) and Paw Withdrawal Latency (PWL) were measured. Minimum and maximum concentrations were determined based on known content.

Behavioral tests were performed 2 hours after drug administration, with each value representing the mean of three independent measurements.

4. Insertion of the guide cannula

Guide cannula was implanted into mice 4 days before the experiment to perform microdialysis and to administer GABA-based drugs. Vertical incisions were made along the midline of the skull to expose bregma and lambda. Micro guide tubes (Eicom, Kyoto, Japan) for recording and drug injection were fixed using two bone screws and acrylic resin (Ortho-Jet; Lang Dental, Wheeling, IL, USA). After anesthesia with 15 mg / kg tiletamine / zolazepam and 9 mg / kg xylazine, a stainless steel guide cannula (outer diameter, 0.5 mm, AG-8, Eicom) was inserted stereotically at the following coordinates of ZI: Paxinos and Watson 2.8 mm at midline and 6.8 mm below brain surface according to atlas.

After inserting the guide cannula as above, the dummy cannula (outer diameter 0.5 mm) was carefully inserted.

5. Biomicrodialysis

The day before microdialysis, the inner stylet was replaced with a dialysis probe with a 1.0 mm long semipermeable membrane (outer diameter, 0.31 mm; A-I-8-02; Eicom).

Connect a 2-channel fluid rotator (SSU-20, Eicom) to the probe inlet and outlet and artificial cerebrospinal fluid (147 mM NaCl, 4 mM KCl, 1.2 mM CaCl ₂ , 0.9 mM MgCl ₂ ) using a pump to 1 μL / min The probe was washed by injection at the rate of. After equilibration overnight, 36 μL of dialysate was collected every 30 minutes from a glass bottle containing ethylenediamine tetraacetic acid (200 μM) and acetic acid solution (40 μM) in the same volume (36 μL) for dialysis. Samples were stored at -70 ° C until samples were collected. GABA, glutamic acid and serotonin were measured in the placebo treated group (n = 10) and the CCI induced group (n = 10), respectively.

6. Microinjection Procedure

Drug or saline was injected into ZI using a microdialysis probe protected by a telescoping steel tube system. The tube assembly was inserted into the guide cannula prior to microinjection and the needle was allowed to protrude 3 mm from the guide cannula tip.

Since the area of the ZI (~ 3 mm ²⁾ greater than the area of the anterior nucleus (~ ² mm 2) it was injected at a rate of 0.25 μL / min. The solution was delivered at the same rate for 2 minutes and after the procedure was completed, the injection needle was removed for 20 seconds.

7. In Vitro Extracellular Recordings

After 10 days after sciatic nerve injury, placebo-treated CCI mice (6 per group) were anesthetized with 15 mg / kg tiletamine / zolazepam and 9 mg / kg xylazine. Extracellular recordings were obtained via quartz insulated carbon electrodes (Kation Scientific, Minneapolis, MN, USA) at ZI. This study uses a digital Lynx SX data acquisition system (Neuralynx Inc., Bozeman, MT, USA) to digitize waveforms (40 kHz), and the unit with dual threshold and principal component analysis (PCA), which uses Neuralynx SpikeSort 3D software. To sort it offline. The NeuroExplorer software (Neuralynx) was used to generate fire rate (spikes / second) and twitch time histogram (PSTH) to confirm that it was recorded in a single unit.

8. Neurotransmitter Analysis

LC-MS was used to detect GABA, glutamate and serotonin. Chromatographic analysis was performed using an Agilent 1100 high speed liquid chromatography (HPLC) system (Agilent Technologies, Santa Clara), which included a G1322A degasser, a G1311A quaternary pump, a G1313A well-plate autosampler, and a G1316A thermostated column. , CA, USA). A G1946D mass spectrometer (Agilent Technologies) using an electrospray source interface was used for MS detection. Data collection and analysis was performed using Agilent ChemStation for LC / MS detection (version B.02.01). Perform chromatographic analysis on an Agilent XDB-C18 column (3.0 mm x 50 mm, 1.8 μm), eluting acetonitrile: 0.1% aqueous formic acid solution (24:76, v / v) as mobile phase at a rate of 0.3 mL / min It was. The column temperature was maintained at 25 ° C., autosampler at 4 ° C., and volume at 5 μl. Analysis time was 4.5 minutes per sample. The HPLC system was connected to the mass spectrometer via an electrospray ionization (ESI) interface. The mass conditions of ESI were optimized in anion detection mode as follows: capillary 3,500V, nebulizer 40 psi, dry gas 8L / min, gas temperature 350 ° C. and fragment 80V. Selected ion monitoring was used and fragment transition was m / z 511.1 for curculigoside and m / z 579.1 for naringin.

9. Histological Validation of Electrode and Probe Placement

After the experiment was completed, the electrode and probe site were post-validated. Electrolytic lesions were made prior to cardiac perfusion by passing 0.05 mA current through the recorded electrode for 10 seconds. Mice were completely anesthetized with tiletamine / zolazepam and perfused with 0.1 M phosphate buffered saline through 4% paraformaldehyde through the heart. Mice were epidural perfused together with physiological saline and 10% formalin and saline. Brains were extracted and further fixed in 10% formalin for 24 hours before sectioning. Microtome cryostat (Microm, Walldorf, Germany) was used to cut the coronal plane (5 μm) through the hypothalamus and stain the target site of the brain with neutral red. Electrodes and probes were placed through an optical microscope based on Paxinos and Watson's brain maps.

10. Statistical Analysis

The analysis was performed with a statistical software program Prism 7.0 (GraphPad Software, Inc., La Jolla, CA, USA) and a bidirectional analysis of behavior (ANOVA) via a non-test t-test of microdialysis concentrations. To investigate the effect of the drug after injection, one-way analysis of variance (ANOVA) tests on behavioral and neurological activity records were performed each time. P <0.05 was considered significant in all experiments. Data represent 'mean ± standard deviation' (SD) for behavioral tests and 'mean ± standard error' of the mean for neuronal signals and biochemical analyses.

Result Analysis

1. Identify the behavior of hypersensitivity to pain

To evaluate the susceptibility to neuropathic pain in the CCI-induced group (n = 20) and gastrointestinal treatment group (n = 10), behavioral analysis was conducted to find PWT and PWL. After more than 10 days after surgery, the PWT of the ipsilateral hind paw in the CCI-induced group was 16.84 ± 4.98 g (repeat bidirectional ANOVA, F = 51.21, P =) at 35.66 ± 2.85 (mean ± standard deviation) compared to the opposite hind paw and gastrointestinal animals. 0.0001) (Figure 2A). Consistent with previous reports, 36-38 animals with sciatic nerve injury showed decreased PWT in response to postoperative mechanical stimulation.

In animals treated more than 10 days postoperatively, the PWL of the ipsilateral hind limb of CCI-induced rats was 6.81 ± 1.57 to 3.31 ± 1.01 (repeated bidirectional ANOVA, F = 65.99, P = 0.0001) compared to the opposite hind and thigh hind limbs. It was confirmed that the decrease to (Fig. 2B). Importantly, the PWT and PWL of CCI induced rats were significantly lower than those of gastrotreated rats for the same time. Gastrointestinal rats showed no difference in PWT and PWL during this period. (Figure 2)

2. In Vivo Microdialysis of ZI

To assess changes in neurotransmitter levels, microdialysis probes were implanted into ZI of CCI treated group (n = 10) and gastrointestinal treatment group (n = 10), respectively. GABA levels were measured in the two groups of rats, with 2.31 ± 0.61 μg / L in the CCI group and 5.38 ± 0.43 μg / L in the gastrointestinal group (P <0.05), which significantly lower GABA levels in the CCI group. It can be seen that (Fig. 3). Glutamic acid levels were similar in the CCI and Gastrointestinal groups, and serotonin levels in the CCI and GCI groups were 0.25 ± 0.07 μg / L for the CCI group and 0.46 ± 0.05 for the Gastrointestinal group. As μg / L (P <0.05), the CCI treated group showed significantly lower serotonin levels.

3. Effects on withdrawal thresholds of GABA stimulation injections in ZI

To investigate the effect of GABA on pain control in ZI, 2 μL of saline, muscimol (0.5 or 5 mmol / μL), or bicuculline were applied to ZI of rats of CCI treated (n = 10) and gastrointestinal treated (n = 10) rats. (0.5 or 5 μmol / μL) was microinjected using a microdialysis probe at a rate of 0.25 μL per minute. More muscimol injections resulted in increased PWT (one-way ANOVA, F = 3.86, P <0.05) and PWL (one-way ANOVA, F = 3.60, P <0.05) in the ipsilateral hind paw, but for bicuculline, PWT or PWL There was no impact. (Figures 4A and B).

4. Neuronal Activity in ZI After Sciatic Nerve Injury

To assess the effect of sciatic nerve injury on the output of ZI neurons, the neuronal activity of ZI (n = 6) and gastrointestinal (n = 6) rats in the CCI-induced group was recorded. The target ZI is shown in the schematic diagram of the coronal plane (FIG. 5A). The average rate of ignition of ZI neurons was lower in the CCI-induced group than in the gastrointestinal rats (Figure 5B). Moreover, spontaneous ignition rate was significantly lower in CCI-induced group (5.80 ± 0.12spikes / s) than gastrointestinal treatment group (7.73 ± 0.15 spikes / s, unpaired t-test, P <0.001). To analyze the effect of GABA regulation of ZI on neuronal activity, spontaneous firing rates were measured for 30 minutes in gastrointestinal and CCI-induced groups treated with muscimol or bicuculline. The rate of ignition of the gastrointestinal rats was significantly increased after muscimol or bicuculline injection compared to the saline infusion group, which was a control group based on one-way ANOVA every hour (FIG. 5C, P <0.05; Table S1). The rate of ignition increased above the baseline of CCI-induced mice after muscimol injection. On the other hand, after injection of bicuculline, it did not affect the rate of ignition based on one-way ANOVA (Fig. 5D, P <0.05; As demonstrated by PSTH, the overall change in firing rate in the CCI-induced group after GABA drug treatment is shown in FIGS. 5E and F. Neuronal records were monitored from 12 rats.

So far I looked at the center of the preferred embodiment for the present invention. Those skilled in the art will appreciate that the present invention can be implemented in a modified form without departing from the essential features of the present invention. Therefore, the disclosed embodiments should be considered in descriptive sense only and not for purposes of limitation. The scope of the present invention is shown in the appended claims rather than the foregoing description, and all differences within the scope will be construed as being included in the present invention.

Claims (6)

delete delete Inject the GABA agonist muscimol at a concentration of 5.0 mmol / μl into a 2 µl dose in an indeterminate area of the rat model brain, which caused CCI (chronic constriction injury) to the sciatic nerve. As a method of reducing sciatica pain behavior of rat, characterized in that
The indeterminate region is where the GABA-like end selectively stimulates the proximal dendrites of associated neurons in the nucleus of the thalamus, creating an immediate response to sensory stimuli,
Rats in which the thigh nerves were damaged by inducing CCI in the sciatic nerve reduced PWT and PWL, and the neuronal ignition rate in the indeterminate region was reduced to confirm the presence of spontaneous pain in the rat model. Posture, repeated tremors of the soles of the feet and the soles of the feet to protect and lick,
After injecting the GABA agonist muscimol, the rats were treated with mechanical stimulation by applying continuously increasing pressure (5 g / s) to the ipsilateral and contralateral forefoot heel area of the CCI lesion. A method of reducing sciatica pain behavior of rats, characterized by an increase in PWT and PWL in ipsilateral hind paws and an increase in neuronal firing in one-way ANOVA. delete delete delete

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