Lee Sang Hoon, Moon Jee Youn, Jung Sung Jun, Kang Jin Gu, Choi Seung Pyo, Jang Jun Ho
Department of Biomedical Science, Graduate School of Biomedical Science; Engineering, Hanyang University, Seoul, Republic of Korea.
Department of Anesthesiology and Pain Medicine, Seoul National University Hospital College of Medicine, Seoul, Republic of Korea.
PLoS One. 2015 Jan 30;10(1):e0117316. doi: 10.1371/journal.pone.0117316. eCollection 2015.
Eugenol has sedative, antioxidant, anti-inflammatory, and analgesic effects, but also serves as an irritant through the regulation of a different set of ion channels. Activation of gamma aminobutyric acid (GABA) receptors on sensory neurons leads to the stabilization of neuronal excitability but contributes to formalin-induced inflammatory pain. In this study, we examined the effect of eugenol on the GABA-induced current in rat trigeminal ganglia (TG) neurons and in human embryonic kidney (HEK) 293 cells expressing the GABAA receptor α1β2γ2 subtype using the whole-cell patch clamp technique. RT-PCR and Western blot analysis were used to confirm the expression of GABAA receptor γ2 subunit mRNA and protein in the TG and hippocampus. Eugenol decreased the amplitude ratio of the GABA-induced current to 27.5 ± 3.2% (p < 0.05) in TG neurons, which recovered after a 3-min washout. In HEK 293 cells expressing the α1β2γ2 subtype, eugenol inhibited GABA-induced currents in a dose-dependent manner. Application of eugenol also decreased the GABA response in the presence of a G-protein blocker. Eugenol pretreatment with different concentrations of GABA resulted in similar inhibition of the GABA-induced current in a non-competitive manner. In conclusion, eugenol inhibits the GABA-induced current in TG neurons and HEK 293 cells expressing the GABAA receptor in a reversible, dose-dependent, and non-competitive manner, but not via the G-protein pathway. We suggest that the GABAA receptor could be a molecular target for eugenol in the modulation of nociceptive information.
丁香酚具有镇静、抗氧化、抗炎和镇痛作用，但通过调节另一组不同的离子通道也可作为一种刺激物。感觉神经元上γ-氨基丁酸（GABA）受体的激活可导致神经元兴奋性的稳定，但会加重福尔马林诱导的炎性疼痛。在本研究中，我们使用全细胞膜片钳技术，研究了丁香酚对大鼠三叉神经节（TG）神经元以及表达GABAA受体α1β2γ2亚型的人胚肾（HEK）293细胞中GABA诱导电流的影响。采用RT-PCR和蛋白质印迹分析来确认TG和海马中GABAA受体γ2亚基mRNA和蛋白质的表达。丁香酚使TG神经元中GABA诱导电流的幅度比降至27.5±3.2%（p<0.05），冲洗3分钟后恢复。在表达α1β2γ2亚型的HEK 293细胞中，丁香酚以剂量依赖性方式抑制GABA诱导的电流。在存在G蛋白阻滞剂的情况下，丁香酚的应用也降低了GABA反应。用不同浓度的GABA对丁香酚进行预处理，以非竞争性方式对GABA诱导电流产生类似的抑制作用。总之，丁香酚以可逆、剂量依赖性和非竞争性方式抑制TG神经元和表达GABAA受体的HEK 293细胞中的GABA诱导电流，但不通过G蛋白途径。我们认为GABAA受体可能是丁香酚调节伤害性信息的分子靶点。
