Mutations of Arginine 222 in Pre-transmembrane Domain I of Mouse 5-HT3A Receptor Abolish 20(R)- But Not 20(S)-Ginsenoside Rg3 Inhibition of 5-HT-Mediated Ion Currents

Byung-Hwan Lee; Jun-Ho Lee; In-Soo Yoon; Joon-Hee Lee; Sun-Hye Choi; Tae-Joon Shin; Mi Kyung Pyo; Woo-Sung Choi; Sang-Mok Lee; Yoongho Lim; Hyewhon Rhim; Seung-Yeol Nah

doi:10.1248/bpb.30.1721

Regular Articles

Mutations of Arginine 222 in Pre-transmembrane Domain I of Mouse 5-HT_3A Receptor Abolish 20(R)- But Not 20(S)-Ginsenoside Rg₃ Inhibition of 5-HT-Mediated Ion Currents

Byung-Hwan Lee, Jun-Ho Lee, In-Soo Yoon, Joon-Hee Lee, Sun-Hye Choi, Tae-Joon Shin, Mi Kyung Pyo, Woo-Sung Choi, Sang-Mok Lee, Yoongho Lim, Hyewhon Rhim, Seung-Yeol Nah

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Byung-Hwan Lee
Ginsentology Research Laboratory and Department of Physiology, College of Veterinary Medicine, Konkuk University
Jun-Ho Lee
Ginsentology Research Laboratory and Department of Physiology, College of Veterinary Medicine, Konkuk University
In-Soo Yoon
Ginsentology Research Laboratory and Department of Physiology, College of Veterinary Medicine, Konkuk University
Joon-Hee Lee
Ginsentology Research Laboratory and Department of Physiology, College of Veterinary Medicine, Konkuk University
Sun-Hye Choi
Ginsentology Research Laboratory and Department of Physiology, College of Veterinary Medicine, Konkuk University
Tae-Joon Shin
Ginsentology Research Laboratory and Department of Physiology, College of Veterinary Medicine, Konkuk University
Mi Kyung Pyo
Ginsentology Research Laboratory and Department of Physiology, College of Veterinary Medicine, Konkuk University
Woo-Sung Choi
Ginsentology Research Laboratory and Department of Physiology, College of Veterinary Medicine, Konkuk University
Sang-Mok Lee
Ginsentology Research Laboratory and Department of Physiology, College of Veterinary Medicine, Konkuk University
Yoongho Lim
Bio/Molecular Informatics Center, Konkuk University
Hyewhon Rhim
Life Sciences Division, KIST
Seung-Yeol Nah
Ginsentology Research Laboratory and Department of Physiology, College of Veterinary Medicine, Konkuk University

Keywords: ginseng, ginsenoside stereoisomer, 5-HT_3A receptor, ligand-gated ion channel, Xenopus oocyte

JOURNAL FREE ACCESS

2007 Volume 30 Issue 9 Pages 1721-1726

DOI https://doi.org/10.1248/bpb.30.1721

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Abstract

Ginsenosides, active ingredients of Panax ginseng, exist as stereoisomers depending on the position of the hydroxyl group on carbon-20; i.e. 20(R)-ginsenoside and 20(S)-ginsenoside are epimers. We previously investigated the structure–activity relationship of the ginsenoside Rg₃ stereoisomers, 20-R-protopanaxatriol-3-[O-β-D-glucopyranosyl (1→2)-β-glucopyranoside], (20(R)-Rg₃) and 20-S-protopanaxatriol-3-[O-β-D-glucopyranosyl (1→2)-β-glucopyranoside], (20(S)-Rg₃) in regulating 5-HT_3A receptor-mediated ion currents (I_5-HT) expressed in Xenopus oocytes and found that 20(S)-Rg₃ rather than 20(R)-Rg₃ was more stronger inhibitor of I_5-HT. In the present study, we further investigated the effects of 20(R)-Rg₃ and 20(S)-Rg₃ on mouse 5-HT_3A receptor channel activity after site-directed mutations of 5-HT_3A receptor facilitation site, which is located at pre-transmembrane domain I (pre-TM1). 5-HT_3A receptor was expressed in Xenopus oocytes, and I_5-HT was measured using two-electrode voltage clamp technique. In wild-type, both 20(R)-Rg₃ and 20(S)-Rg₃ inhibited I_5-HT with concentration-dependent and reversible manner. Induction of 5-HT_3A receptor facilitation by point mutations of pre-TM1 amino acid residue R222 to R222A, R222D, R222E or R222T not only decreased EC₅₀ values for I_5-HT compared to wild-type but also abolished 20(R)-Rg₃-induced inhibition of I_5-HT. Those mutations also shifted the IC₅₀ values by 20(S)-Rg₃ into right direction by 2- to 4-folds compared with wild-type. These results indicate that 5-HT_3A receptor facilitation differentially affects 20(R)-Rg₃- and 20(S)-Rg₃-mediated 5-HT_3A receptor channel regulation.

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