• Applied Microscopy
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• 제42권1호
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• pp.27-33
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• 2012

Transient receptor potential ankyrin 1 (TRPA1)은 $17^{\circ}C$보다 낮은 유해한 온도 및 자극적인 화합물에 의해 활성화되며 통각을 조절한다. 그러나 TRPA1에 의한 통각정보가 어떻게 처리되는지에 대한 정보는 많이 알려져 있지 않다. 본 연구에서는 흰쥐의 삼차신경절에서 TRPA1을 발현하는 신경세포의 특성을 규명하기 위해서, 면역형광기법을 사용하여 TRPA1을 발현하는 신경세포에서 다른 통각수용기들에서 발현되며, 특징적인 기능을 수행하는 수용기인 transient receptor potential vanilloid 1 (TRPV1)와 $P2X_3$와의 발현양상을 조사하였다. TRPA1을 발현하는 신경세포에서 열감각수용기이며, 통각표지자인 TRPV1과의 공존을 조사해 본 결과, TRPA1 면역양성 신경세포 중에서 58.8% (328/558)가 TRPV1을 동시에 발현하였으며, 41.2% (230/558)가 TRPA1만 발현하고 TRPV1을 발현하지 않았다. TRPA1을 발현하는 신경세포 중 TRPV1을 동시에 발현하는 신경세포는 대부분 작거나 중간크기였다. 또한 TRPA1과 조직의 손상, 그리고 염증 시 분비되는 ATP와 결합하는 $P2X_3$와의 공존을 조사해 본 결과, TRPA1 면역양성 신경세포 중에서 26.1% (310/1186)의 신경세포에서 $P2X_3$을 동시에 발현하였으며, 73.9% (876/1186)의 신경세포에서 TRPA1만 발현하였다. TRPA1을 발현하는 신경세포 중 $P2X_3$을 동시에 발현하는 신경세포는 대부분 작거나 중간크기였다. 이러한 결과는 TRPA1을 발현하는 신경세포가 TRPV1 또는 $P2X_3$를 동시에 발현함으로써 동일한 신경세포가 구강안면영역에서의 냉통각 및 열통각을 조절할 뿐 아니라, 냉통각 및 염증성동통을 동시에 전달하는 등 하나의 신경세포가 여러 가지 통각의 전달에 관여하는 것을 시사한다.

• 대한화장품학회지
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• 제44권1호
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• pp.15-21
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• 2018

UV 뿐 아니라 가시광선, 적외선에 의해 발생된 열로 인해 피부 온도 상승, MMP-1의 증가에 따른 피부 노화가 진행된다. 따라서 열에 의한 노화에서 피부 온도 조절은 노화 억제에 중요한 핵심 요소이다. 일시적인 수용체 전위 통로인 TRPM8은 멘솔 수용체(CMR1)로써 $25^{\circ}C$ 이하의 온도에서 활성화되고 시원한 감각을 발생시키는 냉 수용체로 보고되어 있다. TRPM8 조절을 통해 시원한 감각과 피부 온도를 조절하는 연구가 활발히 진행되고 있다. 본 연구에서는 천연물인 영지버섯을 이용하여 냉 수용체인 TRPM8 발현에 어떠한 영향을 주는지 확인하였다. 영지버섯추출물 및 용매 분획물의 TRPM8 발현에 대한 영향을 측정한 결과, 영지버섯추출물, n-hexane 분획물 및 water 분획물에서 농도의존적으로 TRPM8 발현이 증가함을 확인하였다. Hex 분획물에서 유효성분을 찾고자 크로마토그래피를 실시하여 1개의 화합물을 분리하였으며 $^1H$ 및 $^{13}C$ NMR spectrum 분석을 통하여 화학구조를 동정하였다. 분리된 화합물은 에르고스테롤로 TRPM8 발현 증가에 효과가 있음을 확인하였다. 결과를 토대로, 영지버섯추출물 및 에르고스테롤은 화장품 분야에서 새로운 쿨링 소재로서 개발가능성이 있다고 사료된다.

• The Korean Journal of Physiology and Pharmacology
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• 제28권5호
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• pp.413-422
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• 2024

Group I metabotropic glutamate receptors (mGluRs) modulate postsynaptic neuronal excitability and epileptogenesis. We investigated roles of group I mGluRs on low extracellular Mg²⁺ concentration ([Mg²⁺]_o)-induced epileptiform activity and neuronal cell death in the CA1 regions of isolated rat hippocampal slices without the entorhinal cortex using extracellular recording and propidium iodide staining. Exposure to Mg²⁺-free artificial cerebrospinal fluid can induce interictal epileptiform activity in the CA1 regions of rat hippocampal slices. MPEP, a mGluR 5 antagonist, significantly inhibited the spike firing of the low [Mg²⁺]_o-induced epileptiform activity, whereas LY367385, a mGluR1 antagonist, did not. DHPG, a group 1 mGluR agonist, significantly increased the spike firing of the epileptiform activity. U73122, a PLC inhibitor, inhibited the spike firing. Thapsigargin, an ER Ca²⁺-ATPase antagonist, significantly inhibited the spike firing and amplitude of the epileptiform activity. Both the IP₃ receptor antagonist 2-APB and the ryanodine receptor antagonist dantrolene significantly inhibited the spike firing. The PKC inhibitors such as chelerythrine and GF109203X, significantly increased the spike firing. Flufenamic acid, a relatively specific TRPC 1, 4, 5 channel antagonist, significantly inhibited the spike firing, whereas SKF96365, a relatively non-specific TRPC channel antagonist, did not. MPEP significantly decreased low [Mg²⁺]_o DMEM-induced neuronal cell death in the CA1 regions, but LY367385 did not. We suggest that mGluR 5 is involved in low [Mg²⁺]_o-induced interictal epileptiform activity in the CA1 regions of rat hippocampal slices through PLC, release of Ca²⁺ from intracellular stores and PKC and TRPC channels, which could be involved in neuronal cell death.

• The Korean Journal of Physiology and Pharmacology
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• 제16권2호
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• pp.139-144
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• 2012

It has been reported that activation of metabotropic glutamate receptor 1 (mGluR1) can mediate endocannabinoid-induced short-term depression of synaptic transmission in cerebellar parallel fiber (PF)-Purkinje cell (PC) synapse. mGluR1 has signaling pathways involved in intracellular calcium increase which may contribute to endocannabinoid release. Two major mGluR1-evoked calcium signaling pathways are known: (1) slow-kinetic inward current carried by transient receptor potential canonical (TRPC) channel which is permeable to $Ca^{2+}$; (2) $IP_3$-induced calcium release from intracellular calcium store. However, it is unclear how much each calcium source contributes to endocannabinoid signaling. Here, we investigated whether calcium influx through mGluR1-evoked TRPC channel contributes to endocannabinoid signaling in cerebellar Purkinje cells. At first, we applied SKF96365 to inhibit TRPC, which blocked endocannabinoid-induced short-term depression completely. However, an alternative TRP channel inhibitor, BTP2 did not affect endocannabinoid-induced short-term depression although it blocked mGluR1-evoked TRPC currents. Endocannabinoid signaling occurred normally even though the TRPC current was mostly blocked by BTP2. Our data imply that TRPC current does not play an important role in endocannabinoid signaling. We also suggest precaution in applying SKF96365 to inhibit TRP channels and propose BTP2 as an alternative TRPC inhibitor.

• 대한본초학회지
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• 제33권4호
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• pp.77-85
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• 2018

Objective : This study was intended to examine the effects of water extract of Prunellae Spica (PS), which is a herb with 'cold' nature based on hot and cold theory of traditional Korean medicine. Methods : Hyperthyroidism was induced in SD rats by LT4 (0.5 mg/kg, i.p.) daily for four weeks. After 2 weeks of LT4 injection, rats were divided randomly into four groups; normal, LT4-induced hyperthyroid control, PS extract (500 mg/kg, p.o.)-treated group, and propylthiouracil (PTU, 10 mg/kg, s.c.)-treated positive group. After 2 weeks of drug treatment, all rats were sacrificed and harvested blood samples and thyroid tissues. The changes of body weight, food and water intake, and body temperature were measured weekly. Serological markers were analyzed in sera using an enzyme-based assay, and thyroid tissues were stained with Hematoxylin & Eosin (H&E). Brain and dorsal root ganglion (DRG) tissues were isolated and analyzed the expression of transient receptor potential (TRP) channels by Western blot. Results : PS extract administration attenuated the loss of body weight and the increase of body temperature in LT4-induced hyperthyroidism rats. PS extract increased the level of thyroid stimulating hormone (TSH) and decreased tiiodothyronine (T3) and tetraiodothyronine (T4). In action mechanism, PS extract regulated the expression of transient receptor potential channel subfamily V member 1 (TRPV1) and transient Receptor Potential channel subfamily M member 8 (TRPM8), the thermoregulators. Conclusion : To conclude, PS extract can improve the symptoms of hyperthyroidism through regulation of the thyroid hormones imbalance and thermoregulation via TRP channels.

• Molecules and Cells
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• 제37권3호
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• pp.257-263
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• 2014

A mammalian cell renovates itself by autophagy, a process through which cellular components are recycled to produce energy and maintain homeostasis. Recently, the abundance of gap junction proteins was shown to be regulated by autophagy during starvation conditions, suggesting that transmembrane proteins are also regulated by autophagy. Transient receptor potential vanilloid type 1 (TRPV1), an ion channel localized to the plasma membrane and endoplasmic reticulum (ER), is a sensory transducer that is activated by a wide variety of exogenous and endogenous physical and chemical stimuli. Intriguingly, the abundance of cellular TRPV1 can change dynamically under pathological conditions. However, the mechanisms by which the protein levels of TRPV1 are regulated have not yet been explored. Therefore, we investigated the mechanisms of TRPV1 recycling using HeLa cells constitutively expressing TRPV1. Endogenous TRPV1 was degraded in starvation conditions; this degradation was blocked by chloroquine (CLQ), 3MA, or downregulation of Atg7. Interestingly, a glucocorticoid (cortisol) was capable of inducing autophagy in HeLa cells. Cortisol increased cellular conversion of LC3-I to LC-3II, leading autophagy and resulting in TRPV1 degradation, which was similarly inhibited by treatment with CLQ, 3MA, or downregulation of Atg7. Furthermore, cortisol treatment induced the colocalization of GFP-LC3 with endogenous TRPV1. Cumulatively, these observations provide evidence that degradation of TRPV1 is mediated by autophagy, and that this pathway can be enhanced by cortisol.

• 한방비만학회지
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• 제18권1호
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• pp.10-18
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• 2018

Objectives: Metabolic syndrome is correlated with increased cardiovascular risk and characterized by several factors, including visceral obesity, hypertension, insulin resistance, and dyslipidemia. Several members of a large family of nonselective cation entry channels, e.g., transient receptor potential (TRP) melastatin 7 (TRPM7) channels have been associated with the development of cardiovascular diseases. The purpose of this study was to investigate the effects of Dangkwisoo-san, ginger and curcumin on TRPM7 channel. Methods: Human embryonic kidney (HEK) 293 cells stably transfected with the TRPM7 expression vectors were maintained in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum (FBS), 1% penicillin/streptomycin, $5{\mu}g/mL$ blasticidin, and 0.4 mg/mL zeocin in a humidified 20% $O_2$/10% $CO_2$ atmosphere at $37^{\circ}C$. Whole-cell patch clamp recordings were obtained using an Axopatch 700B amplifier and pClamp v.10.4 software, and signals were digitalized at 5 kHz using Digidata 1422A. Results: Dangkwisoo-san extract (100, 200, 300, 400, and $500{\mu}g/mL$) inhibited the outward and inward TRPM7 whole-cell currents at dose dependent manner and the half maximal inhibitory concentration $(IC)_{50}$ of Dangkwisoo-san was $218.3{\mu}g/mL$. Also, ginger extract (100, 200, 300, 400, and $500{\mu}g/mL$) inhibited the outward and inward of TRPM7 whole-cell currents in a dose dependent manner and the $IC_{50}$ of ginger was $877.2{\mu}g/mL$. However, curcumin had no effects on TRPM7 whole-cell currents. Conclusions: These results suggest that both Dangkwisoo-san and ginger have good roles to inhibit the TRPM7 channel, suggesting that Dangkwisoo-san and ginger are considered one of the candidate agents for the treatment of metabolic syndrome such as cardiovascular disease.

• 한국산학기술학회논문지
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• 제12권1호
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• pp.312-317
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• 2011

캡사이신 채널로 알려진 바닐로이드 수용체 TRPV1 (캡사이신채널, Transient Receptor Potential Vanilloid 1)은 통증발현에서 중요한 역할을 하는 것으로 알려져 있다. 하지만 TRPV1의 활성조절에 관여하는 단백질에 대하여는 알려진 바가 많지 않다. 최근 rat TRPV1과 직접적으로 결합하는 단백질을 탐색하여 mouse Rab11-FIP3 (rab11-family interaction protein 3)가 rat TRPV1과 직접적으로 결합한다는 것이 보고되었다. Rab11은 여러 가지의 세포내 이동에 관여하는 것으로 보고되었다. 그러므로 Rab11-FIP3과의 결합을 통해 TRPV1의 세포막으로의 이동에 관여할 것으로 추측할 수 있다. 본 연구에서는 전에 보고된 연구가 mouse와 rat 이라는 다른 종의 단백질끼리의 결합이기 때문에 같은 종에서의 상호작용을 확인하고 Rab11-FIP3의 TRPV1의 세포막으로의 이동에서의 역할을 알아보고자 현재까지 동정되지 않은 rat의 Rab11-FIP3의 유전자를 GenBank 서열을 바탕으로 rat 뇌의 RNA 로부터 cDNA 를 클로닝하여 유전자를 분리하고 TRPV1 과의 관계를 세포생물학적으로 알아보았다. 연구결과 rat의 Rab11-FIP3는 489개의 아미노산 서열을 가지고 있으며 human과는 80%, mouse와는 90% 이상 아미노산 서열의 상동성을 보였다. 조직별 분포는 심장, 뇌, 간, 콩팥, 정소에서 발현되고 있는 것을 northern blot assay와 western blot assay 로 확인하였다. rat 의 뇌조직에서 TRPV1 과 Rab11-FIP3 단백질이 결합하여 colocalize 하는 것을 면역화학방법으로 확인하였다. 이 결합은 같은 family 의 TRPV2 와는 결합하지 않는 특이적 결합이므로 Rab11-FIP3 가 TRPV1 과 상호작용하여 세포막으로의 이동에 관여할 것이라는 것을 시사한다.

• 한국산학기술학회논문지
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• 제12권7호
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• pp.3096-3102
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• 2011

바닐로이드 수용체 TRPV1(transient receptor potential vanilloid 1)은 캡사이신, pH, 열 등의 통증 유발물질에 의해 활성화되는 비특이적 양이온 채널로서 통증발현에 핵심적인 막 단백질이다. TRPV1의 막 수송에 관한 연구가 미미한 가운데 FIP3(family of Rab11 interacting protein 3)가 TRPV1 채널과 결합하여 막수송에 관여한다고 보고되었다. FIP3는 Rab11과 결합하는 단백질인데 최근 Rab11 단백질이 여러 채널 단백질의 막수송에 직접적으로 또는 간접적으로 중요하다고 보고되었다. 그러므로 본 연구에서는 Rab11이 TRPV1의 막 수송에서의 역할을 알아보기 위하여 세포 생물학적, 생화학적으로 알아보았다. 공촛점 현미경을 통하여 확인한 결과 Rab11은 실제로 세포내에서 TRPV1과 동일한 위치에서 발현되어 있음을 확인하였다. 하지만 생화학적인 방법인 GST-pulldown을 실시하였을 때 TRPV1과 Rab11간에는 서로 직접적인 결합은 하지 않는 것으로 나타났다. 비록 직접적인 결합은 하지 않지만 Rab11이 TRPV1의 막 수송에 관여한다고 가정하고 Rab11의 TRPV1의 막수송에서의 역할을 더 자세히 알아보기 위하여 세포내 Rab11a의 발현을 siRNA를 사용하여 Rab11a의 발현을 50%수준으로 저해한 후 TRPV1의 세포막으로의 이동을 알아본 결과 Rab11 발현 저해 시 세포막에 이동된 TRPV1이 현저히 감소함을 확인할 수 있었다. 이 결과로부터 Rab11이 아마도 FIP3을 포함하는 방법으로 TRPV1의 막 수송에 영향을 주는 것으로 결론지을 수 있다.

• The Korean Journal of Pain
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• 제34권3호
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• pp.262-270
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• 2021

Background: Transient receptor potential vanilloid 1 (TRPV1) is a non-selective cation channel implicated in pain sensation in response to heat, protons, and capsaicin (CAPS). It is well established that TRPV1 is involved in mechanical allodynia. This study investigates the effect of Ononis spinosa (Fabaceae) in CAPS-induced mechanical allodynia and its mechanism of action. Methods: Mechanical allodynia was induced by the intraplantar (ipl) injection of 40 ㎍ CAPS into the left hind paw of male Wistar rats. Animals received an ipl injection of 100 ㎍ O. spinosa methanolic leaf extract or 2.5% diclofenac sodium 20 minutes before CAPS injection. Paw withdrawal threshold (PWT) was measured using von Frey filament 30, 90, and 150 minutes after CAPS injection. A molecular docking tool, AutoDock 4.2, was used to study the binding energies and intermolecular interactions between O. spinosa constituents and TRPV1 receptor. Results: The ipsilateral ipl injection of O. spinosa before CAPS injection increased PWT in rats at all time points. O. spinosa decreased mechanical allodynia by 5.35-fold compared to a 3.59-fold decrease produced by diclofenac sodium. The ipsilateral pretreatment with TRPV1 antagonist (300 ㎍ 4-[3-Chloro-2-pyridinyl]-N-[4-[1,1-dimethylethyl] phenyl]-1-piperazinecarboxamide [BCTC]) as well as the β2-adrenoreceptor antagonist (150 ㎍ butoxamine) attenuated the action of O. spinosa. Depending on molecular docking results, the activity of the extract could be attributed to the bindings of campesterol, stigmasterol, and ononin compounds to TRPV1. Conclusions: O. spinosa alleviated CAPS-induced mechanical allodynia through 2 mechanisms: the direct modulation of TRPV1 and the involvement of β2 adrenoreceptor signaling.