• BMB Reports
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• 제41권11호
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• pp.803-807
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• 2008

We investigated the expression of muscarinic acetylcholine receptors (mAChRs) and their possible involvement in the regulation of cell proliferation in four colon cancer cell lines (SNU-61, SNU-81, SNU-407, and SNU-1033) derived from Korean colon carcinoma patients. A ligand binding assay showed that all four cell lines expressed mAChRs. Treatment of the four cell lines with the cholinergic agonist carbachol led to the activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2). In SNU-407 cells, carbachol significantly stimulated cell proliferation, which could be abolished by the muscarinic antagonist atropine and the ERK1/2 kinase inhibitor PD98059. These results indicate that mAChRs specifically mediate the proliferation of SNU-407 colon cancer cells via the ERK1/2 pathway.

• The Korean Journal of Physiology and Pharmacology
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• 제8권6호
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• pp.313-317
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• 2004

The present study were designed to characterize the action mechanisms of acetylcholine (ACh)-induced endothelium-dependent relaxation in arteries precontracted with high $K^+$(70 mM). For this, we simultaneously measured both muscle tension and cytosolic free $Ca^{2+}$ concentration $([Ca^{2+}]_i)$, using fura-2, in endothelium-intact, rabbit carotid arterial strips. In the artery with endothelium, high $K^+$ increased both $[Ca^{2+}]_i$ and muscle tension whereas ACh $(10{\mu}M)$ significantly relaxed the muscle and increased $[Ca^{2+}]_i$. In the presence of $N^G$-nitro-L-arginine (L-NAME, 0.1 mM), ACh increased $[Ca^{2+}]_i$ without relaxing the muscle. In the artery without endothelium, high $K^+$ increased both $[Ca^{2+}]_i$ and muscle tension although ACh was ineffective. 4-DAMP (10 nM) or atropine $(0.1{\mu}M)$ abolished ACh-induced increase in $[Ca^{2+}]_i$ and relaxation. The increase of $[Ca^{2+}]_i$ and vasorelaxation by ACh was siginificantly reduced by either $3{\mu}M$ gadolinium, $10{\mu}M$ lanthanum, or by $10{\mu}M$ SKF 96365. These results suggest that in rabbit carotid artery, ACh-evoked relaxation of 70 mM $K^+$-induced contractions appears to be mediated by the release of NO. ACh-evoked vasorelaxation is mediated via the $M_3$ subtype, and activation of the $M_3$ subtype is suggested to stimulate nonselective cation channels, leading to increase of $[Ca^{2+}]_i$ in endothelial cells.

• 농업생명과학연구
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• 제45권6호
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• pp.213-226
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• 2011

아세틸콜린 분해효소(acetylcholine esterase, AChE) 억제제는 아세틸콜린 함량을 높여 콜린성 neuron을 활성화함으로써 기억 능력의 개선 및 치매 개선을 가져와 현재 다양한 AChE 억제제들이 개발되어 사용되고 있다. 본 연구에서는 AChE에 대한 억제 활성을 갖는 천연물을 다양한 식물추출물 및 에센스오일로부터 탐색하였으며, 탐색한 추출물의 scopolamine으로 기억손상을 유발한 쥐의 기억력 개선 활성을 치매 치료제로 사용하고 있는 donepizil과 비교 분석하였다. 그 결과 자몽(Citrus paradisi) 유래의 에센스 오일이 AChE 억제 활성이 가장 높아 20 ug/ml의 농도로 처리하였을 때 90% 이상의 효소 억제 활성을 나타내었다. 수동회피 실험 결과, 자몽 유래의 에센스오일(100 mg/kg, p.o.)을 투여한 쥐는 치매 치료제로 사용하고 있는 donepizile (0.5 mg/kg)을 투여한 쥐와 유사한 latency time을 나타내어 인지기능이 개선되었다. 또한, 수중미로 시험 결과, 자몽 유래 에센스오일(100 mg/Kg, p.o.)을 투여한 쥐는 donepizile(0.5 mg/kg)을 투여한 쥐와 유사한 latency time을 나타내어 인지기능이 개선되었다. 이상의 결과로부터 자몽 유래 에센스오일은 매우 효과적으로 기억력을 개선하여 인지기능을 개선해 줄 수 있는 안전하고 효과적인 후보물질이라고 사료된다.

• Applied Biological Chemistry
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• 제50권2호
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• pp.127-131
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• 2007

일련의 새로운 3-benzylidenemyosmine 유도체들의 구조 변화와 미국 바퀴벌래(Periplaneta. americana L.)의 nicotin acetylcholine 수용체 (nAChRs) 사이의 결합 친화력 상수에 관한 정량적인 구조와 활성과의 관계를 분자 홀로그램(H) QSAR 방법으로 검토하였다. 친화력 상수에 관하여 가장 양호한 HQSAR 모델은 분자조각 크기 5${\sim}$8 bin 조건에서 유도된 모델(IV-2)이었다. HQSAR 모델(VI-2)은 높은 예측성(q$^2$=0.507)과 상관성(r$^2_{nev.}$=0.944)에 근거하여 양호한 통계값들을 나타내었다. 그리고 HQSAR 기여도로부터 결합 친화력 상수는 분자내 anabaseine 고리에 의존적이었으며 결합 친화력성이 높은 화합물들이 최적화된 모델(VI-2)에 의하여 설계되었다.

• The Korean Journal of Physiology and Pharmacology
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• 제15권1호
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• pp.17-22
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• 2011

Quercetin mainly exists in the skin of colored fruits and vegetables as one of flavonoids. Recent studies show that quercetin, like other flavonoids, has diverse pharmacological actions. However, relatively little is known about quercetin effects in the regulations of ligand-gated ion channels. In the previous reports, we have shown that quercetin regulates subsets of homomeric ligand-gated ion channels such as glycine, 5-$HT_{3A}$ and ${\alpha}7$ nicotinic acetylcholine receptors. In the present study, we examined quercetin effects on heteromeric neuronal ${\alpha}3{\beta}4$ nicotinic acetylcholine receptor channel activity expressed in Xenopus oocytes after injection of cRNA encoding bovine neuronal ${\alpha}3$ and ${\beta}4$ subunits. Treatment with acetylcholine elicited an inward peak current ($I_{ACh}$) in oocytes expressing ${\alpha}3{\beta}4$ nicotinic acetylcholine receptor. Co-treatment with quercetin and acetylcholine inhibited $I_{ACh}$ in oocytes expressing ${\alpha}3{\beta}4$ nicotinic acetylcholine receptors. The inhibition of $I_{ACh}$ by quercetin was reversible and concentration-dependent. The half-inhibitory concentration ($IC_{50}$) of quercetin was $14.9{\pm}0.8\;{\mu}M$ in oocytes expressing ${\alpha}3{\beta}4$ nicotinic acetylcholine receptor. The inhibition of $I_{ACh}$ by quercetin was voltage-independent and non-competitive. These results indicate that quercetin might regulate ${\alpha}3{\beta}4$ nicotinic acetylcholine receptor and this regulation might be one of the pharmacological actions of quercetin in nervous systems.

• Environmental Analysis Health and Toxicology
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• 제18권1호
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• pp.21-25
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• 2003

카바메이트계 농약인 카보푸란은 어류에 대한 독성이 매우 높으며, 낮은 농도에서 어류의 척추기형이나 행동이상을 유발한다. 이러한 카보푸란의 독성기전을 밝히기 위한 일환으로 신경물질대사와 관련이 깊은 acetylcholine esterase(AChE)와 monoamine oxidase(MAO)에 미치는 농약의 효과를 송사리(Oryzias latipes; Medaka fish)를 이용하여 평가하였다. Medaka fish에 대한 카보푸란의 반수치사농도(LC$_{50}$)는 2.5 ppm이었으며, 1 ppb 카보푸란에 24시간 노출된 경우, AChE 효소활성이 머리와 몸통부위에서 각각 30, 20%씩 감소되었다. 한편, MAO 효소활성은 카보푸란의 농도가 증가함에 따라 머리부위에서는 감소한 반면, 몸통부위에서는 증가하는 경향을 보였다. 특히 카보푸란의 농도가 1 ppb에서도 송사리의 MAO 효소활성이 영향을 받는 것으로 나타나, 카보푸란에 의한 송사리의 행동이상은 AChE 활성 뿐 아니라 MAO활성의 변화에 의한 복합적인 효과일 가능성이 높다.

• The Korean Journal of Physiology and Pharmacology
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• 제8권4호
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• pp.219-225
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• 2004

The pelvic ganglia provide autonomic innervations to the various urogenital organs, such as the urinary bladder, prostate, and penis. It is well established that both sympathetic and parasympathetic synaptic transmissions in autonomic ganglia are mediated mainly by acetylcholine (ACh). Until now, however, the properties of ACh-induced currents and its receptors in pelvic ganglia have not clearly been elucidated. In the present study, biophysical characteristics and molecular nature of nicotinic acetylcholine receptors (nAChRs) were studied in sympathetic and parasympathetic major pelvic ganglion (MPG) neurons. MPG neurons isolated from male rat were enzymatically dissociated, and ionic currents were recorded by using the whole cell variant patch clamp technique. Total RNA from MPG neuron was prepared, and RT-PCR analysis was performed with specific primers for subunits of nAChRs. ACh dose-dependently elicited fast inward currents in both sympathetic and parasympathetic MPG neurons $(EC_{50};\;41.4\;{\mu}M\;and\;64.0\;{\mu}M,\;respectively)$. ACh-induced currents showed a strong inward rectification with a reversal potential near 0 mV in current-voltage relationship. Pharmacologically, mecamylamine as a selective antagonist for ${\alpha}3{\beta}4$ nAChR potently inhibited the ACh-induced currents in sympathetic and parasympathetic neurons $(IC_{50};\;0.53\;{\mu}M\;and\;0.22\;{\mu}M,\;respectively)$. Conversely, ${\alpha}-bungarotoxin$, ${\alpha}-methyllycaconitine$, and $dihydro-{\beta}-erythroidine$, which are known as potent and sensitive blockers for ${\alpha}7$ or ${\alpha}4{\beta}2$ nAChRs, below micromolar concentrations showed negligible effect. RT-PCR analysis revealed that ${\alpha}3$ and ${\beta}4$ subunits were predominantly expressed in MPG neurons. We suggest that MPG neurons have nAChRs containing ${\alpha}3$ and ${\beta}4$ subunits, and that their activation induces fast inward currents, possibly mediating the excitatory synaptic transmission in pelvic autonomic ganglia.

• BMB Reports
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• 제28권2호
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• pp.155-161
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• 1995

A nicotinic acetylcholine receptor (nAchR) isolated from the electric tissues of Torpedo californica has been reconstituted into a vesicle comprising a bifunctional azo-ligand (Bae 1) compound, and a liposome containing phospholipids and cholesterol (1 : 1, w/w). The liposome-mediated reconstituted receptor showed a concentration-dependent response to cholinergic drugs in a lithium ion flux assay. This liposome-mediated reconstituted nAchR was immobilized onto an electrode using various synthetic polymers which were tested for their response to the cholinergic ligands. The immobilized nAchR not only exhibited a linear response to a wide range of cholinergic ligand concentrations but also retained an operational stability which lasted for longer than 6 days. Thus, this result provides a basis for application of the immobilized nAchR-based biosensor in detecting cholinergic ligands in vitro.

• Biomolecules & Therapeutics
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• 제24권4호
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• pp.410-417
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• 2016

Quercetin is a flavonoid usually found in fruits and vegetables. Aside from its antioxidative effects, quercetin, like other flavonoids, has a various neuropharmacological actions. Quercetin-3-O-rhamnoside (Rham1), quercetin-3-O-rutinoside (Rutin), and quercetin-3-(2(G)-rhamnosylrutinoside (Rham2) are mono-, di-, and tri-glycosylated forms of quercetin, respectively. In a previous study, we showed that quercetin can enhance ${\alpha}7$ nicotinic acetylcholine receptor (${\alpha}7$ nAChR)-mediated ion currents. However, the role of the carbohydrates attached to quercetin in the regulation of ${\alpha}7$ nAChR channel activity has not been determined. In the present study, we investigated the effects of quercetin glycosides on the acetylcholine induced peak inward current ($I_{ACh}$) in Xenopus oocytes expressing the ${\alpha}7$ nAChR. $I_{ACh}$ was measured with a two-electrode voltage clamp technique. In oocytes injected with ${\alpha}7$ nAChR copy RNA, quercetin enhanced $I_{ACh}$, whereas quercetin glycosides inhibited $I_{ACh}$. Quercetin glycosides mediated an inhibition of $I_{ACh}$, which increased when they were pre-applied and the inhibitory effects were concentration dependent. The order of $I_{ACh}$ inhibition by quercetin glycosides was Rutin${\geq}$Rham1>Rham2. Quercetin glycosides-mediated $I_{ACh}$ enhancement was not affected by ACh concentration and appeared voltage-independent. Furthermore, quercetin-mediated $I_{ACh}$ inhibition can be attenuated when quercetin is co-applied with Rham1 and Rutin, indicating that quercetin glycosides could interfere with quercetin-mediated ${\alpha}7$ nAChR regulation and that the number of carbohydrates in the quercetin glycoside plays a key role in the interruption of quercetin action. These results show that quercetin and quercetin glycosides regulate the ${\alpha}7$ nAChR in a differential manner.

• 대한약리학회지
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• 제32권2호
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• pp.127-138
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• 1996

The effects of adenosine analogues on the electrically-evoked acetylcholine(ACh) release and the influence of ischemia on the effects were studied in the rat hippocampus. Slices from the rat hippocampus were equilibrated with $0.1{\mu}M$ $[^3H]-choline$ and the release of the labelled product, $[^3H]-ACh$, was evoked by electrical stimulation(3 Hz, 2 ms, 5 $VCm^{-1}$ and rectangular pulses for 2 min), and the influence of various agents on the evoked tritiumoutflow was investigated. Ischemia(10 min with 95% $N_2$ + 5% $CO_2$) increased both the basal and evoked ACh release. These increases were abolished by glucose addition into the superfused medium, and they significantly inhibited either by 0.1 & $0.3{\mu}M$ TTX pretreatment or by removing $Ca^{++}$ in the medium. MK-801($1{\sim}10{\mu}M$), a specific NMDA receptor antagonist, and glibenclamide $(1{\mu}M)$, a $K^+-channel$ inhibitor, did not alter the evoked ACh release and nor did they affect the ischemia-induced increases In ACh release. However, polymyxin B(0.03 mg), a specific protein kinase C inhibitor, significantly inhibited the effects of ischemia on the evoked ACh release. Adenosine and $N^6-cyclopentyladenosine$ decreased the ACh release in a dose dependent manner in ischemic condition, though the magnitude of inhibition was far less than those in normal(normoxic) condition. However, the treatment with $5{\mu}M$ DPCPX, a potent $A_1-adenosine$ receptor antagonist, potentiated the ischemia-effect. These results indicate that the evoked-ACh release is potentiated by ischemia, and this process being most probably mediated by protein kinase C, and that the decreased effect of ACh release mediated by $A_1-adenosine$ receptor is significantly inhibited in ischemic state.