kjpp The Korean Journal of Physiology & Pharmacology Korean J Physiol Pharmacol 1226-4512 2093-3827 Korean J Physiol Pharmacol 10.4196/kjpp.2011.15.1.17 kjpp-15-17 Original Article Quercetin Inhibits α3β4 Nicotinic Acetylcholine Receptor-Mediated Ion Currents Expressed in Xenopus Oocytes LeeByung-Hwan ∗ HwangSung-Hee ∗ ChoiSun-Hye ∗ ShinTae-Joon KangJiyeon LeeSang-Mok NahSeung-Yeol Department of Physiology, College of Veterinary Medicine and Bio/Molecular Informatics Center, Konkuk University, Seoul 143-701, Korea Corresponding to: Seung-Yeol Nah, Department of Physiology, College of Veterinary Medicine and Bio/Molecular Informatics Center, Konkuk University, 1, Hwayang-dong, Gwangjin-gu, Seoul 143-701, Korea. (Tel) 82-2-450-4154, (Fax) 82-2-450-3037, (E-mail) synah@konkuk.ac.kr

These authors contributed equally to this work.

 022011 18022011 15 1 17 22 27122010 17012011 18012011 Copyright © 2011 Korean J Physiol Pharmacol 2011 This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. Abstract

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-HT3A and α7 nicotinic acetylcholine receptors. In the present study, we examined quercetin effects on heteromeric neuronal α3β4 nicotinic acetylcholine receptor channel activity expressed in Xenopus oocytes after injection of cRNA encoding bovine neuronal α 3 and β4 subunits. Treatment with acetylcholine elicited an inward peak current (IACh) in oocytes expressing α3β4 nicotinic acetylcholine receptor. Co-treatment with quercetin and acetylcholine inhibited IACh in oocytes expressing α3β4 nicotinic acetylcholine receptors. The inhibition of IACh by quercetin was reversible and concentration-dependent. The half-inhibitory concentration (IC50) of quercetin was 14.9±0.8 μ M in oocytes expressing α3β4 nicotinic acetylcholine receptor. The inhibition of IACh by quercetin was voltage-independent and non-competitive. These results indicate that quercetin might regulate α3β4 nicotinic acetylcholine receptor and this regulation might be one of the pharmacological actions of quercetin in nervous systems.

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Chemical structure of quercetin (A) and effect of quercetin (Que) in oocytes expressing α3β4 nicotinic acetylcholine receptors. Quercetin itself had no effect on IACh in oocytes expressing α3β4 nicotinic acetylcholine receptors (B).

Effect of quercetin (Que) on IACh in oocytes expressing α3β4 nicotinic acetylcholine receptors. (A) Acetylcholine (ACh, 100 μM) was first applied and then acetylcholine was co- or pre-applied with quercetin (Que, 30 μM). Thus, co- and pre-application of quercetin with acetylcholine inhibited IACh. The resting membrane potential of oocytes was about –35 mV and oocytes were voltage-clamped at a holding potential of –80 mV prior to drug application. Traces are representative of six separate oocytes from three different frogs. (B) Co- or pre-application of quercetin did not affect differently on IACh. (C) IACh in oocytes expressing α3β4 nicotinic acetylcholine receptors was elicited at –80 mV holding potential with indicated time in the presence of 100 μM acetylcholine and then the indicated concentration of quercetin was co-applied with acetylcholine. (D) % Inhibition by quercetin of IACh was calculated from the average of the peak inward current elicited by acetylcholine alone before quercetin and the peak inward current elicited by acetylcholine alone after co-application of quercetin with acetylcholine. The continuous line shows the curve fitted according to the equation. y/ymax=[Quercetin]/[Quercetin] + K1/2), where ymax, the maximum inhibition (97.8±1.7%, mean±S.E.M.) and K1/2 is the concentration for half-maximum inhibition (14.9±0.8 μM, mean±S.E.M.), and [Quercetin] is the concentration of quercetin. Each point represents the mean±S.E.M. (n=9∼12 from three different frogs).

Current-voltage relationship and voltage-independent inhibition by quercetin. (A) Current-voltage relationships of IACh inhibition by quercetin (Que) in α3β4 nicotinic acetylcholine receptors. Representative current-voltage relationships were obtained using voltage ramps of –100 to +60 mV for 300 ms at a holding potential of –80 mV. Voltage steps were applied before and after application of 100 μM acetylcholine in the absence or presence of 20 μM quercetin. (B) Voltage-independent inhibition of IACh in the α3β4 nicotinic acetylcholine receptors by quercetin. Inset; the values were obtained from the receptors in the absence or presence of 20 μM quercetin at the indicated membrane holding potentials.

Concentration-dependent effects of acetylcholine on quercetin-mediated inhibition of IACh. (A) The representative traces were obtained from α3β4 nicotinic acetylcholine receptors expressed in oocytes. IACh of the upper and lower panels were elicited from concentration of 30 μM ACh and 1 mM ACh at a holding potential of –80 mV, respectively. (B) Concentration-response relationships for ACh in the α3β4 nicotinic acetylcholine receptors treated with ACh (3∼1,000 μM) alone or with ACh plus co-application of 20 μM quercetin. The IACh of oocytes expressing the α3β4 nicotinic acetylcholine receptors was measured using the indicated concentration of ACh in the absence (☐) or presence (❍) of 20 μM quercetin (Que). Oocytes were exposed to ACh alone or to ACh with quercetin. Oocytes were voltage-clamped at a holding potential of –80 mV. Each point represents the mean± S.E.M. (n=9∼12/group).