【作者】 王薇；

【导师】 刘烈炬；

【作者基本信息】 华中科技大学 ， 生理学， 2010， 博士

【摘要】 尽管大麻素对TRPV1受体的抑制作用可以解释大麻素在外周的抗痛觉过敏和抗伤害感受效应,但大麻素抑制初级感觉神经元上TRPV1受体的机制目前还不清楚。因此,我们使用内源性大麻素anandamide (AEA)和人工合成的大麻素WIN55,212-2 (WIN)研究大麻素如何对三叉神经节神经元上TRPV1受体进行调制。首先,我们研究AEA和WIN对Icap调制的浓度依赖性：此量效曲线呈“钟”形,并且AEA和WIN对Icap调制的浓度阈值非常低(低于10-15 M)。AEA和WIN对Icap的最大抑制浓度分别为10-10M和10-9M,此时,AEA和WIN对Icap的最大抑制率分别为89±7.5%和95±1.6%。当AEA和WIN的浓度高于10-SM,AEA和WIN对Icap的抑制作用被完全翻转。本实验中,我们检测了一些细胞内信号转导通路来探究AEA和WIN对Icap的抑制作用的胞内信号转导机制。我们发现,WIN对Icap的抑制作用可以被PKA系统特异性拮抗剂H-89和KT5720完全翻转。同样,WIN对Icap的抑制作用也可被PKC系统特异性拮抗剂BIM和staurosporine完全翻转。我们也发现PKG系统拮抗剂PKGi可部分翻转WIN对Icap的抑制作用,而G蛋白系统拮抗剂GDP-βs和PLC拮抗剂U73122则不影响WIN对Icap的抑制作用。实验表明细胞内一些信号转导通路,如PKA和PKC,参与了大麻素对Icap的抑制作用。本实验应用全细胞膜片钳技术,在大鼠三叉神经节神经元上探讨渗透压对5-HT3受体的调制作用。结果显示：(1)实验中大部分受检细胞(75%)对胞外给予5—HT(3-300μM)敏感,可记录到具有浓度依赖性的内向电流,该电流可被5—HT3受体特异性拮抗剂ICS 205-930(1μM)阻断。(2)实验中反复给予100μM 5-HT,间隔3分钟,发现5-HT3受体激活电流无衰减现象。(3)实验中预加低渗细胞外液(260mOsm),对5-HT3受体激活电流无影响。(4)预加高渗细胞外液(348mOsm),对5-HT3受体激活电流无影响。以上结果表明,渗透压对大鼠三叉神经节神经元上5-HT3受体没有调制作用。更多还原

【Abstract】 Although the inhibitory effect of cannabinoids on TRPV1 channel may explain the efficacy of peripheral cannabinoids in antihyperalgesia and antinociceptive actions, the mechanism for cannabinoid-induced inhibition of TRPV1 in primary sensory neurons is not understood. Therefore, we explored how anandamide (AEA, an endogenous cannabinoid) and WIN55,212-2 (WIN, a synthetic cannabinoid) inhibited TRPV1 in rat trigeminal ganglion neurons. The concentration dependence of the effect of AEA and WIN on Icap was studied:a "bell" shaped concentration dependent curve was obtained with very low threshold (less than 10-15 M for both AEA and WIN). The maximal inhibitions were at concentration 10-10M and 10-9M, and at these concentrations the Icap was reduced by 89±7.5% and 95±1.6% respectively. When the concentration of AEA or WIN was higher than 10-5M, inhibitory effects were completely reversed. In this study several intracellular signaling transduction pathways were tested to study whether they were involved in the inhibitory effects of AEA or WIN on capsaicin-induced current (Icap).It was newly found that the inhibitory effect of WIN on Icap was completely reversed by PKA antagonists H-89 and KT5720 as well as by PKC antagonists BIM and staurosporine. It was also found that the inhibitory effect was partly reversed by PKG antagonist PKGi, while G-protein antagonist GDP-βs and PLC antagonist U73122 had no effect on the inhibitory effect. It suggests that several intracellular signaling transduction pathways underlie the inhibitory effects of cannabinoids on Icap,including PKA and PKC systems. This experiment aimed to investigate the modulation of osmolality on 5-HT-activated currents(I5-HT) in primary sensory neurons. Whole cell-patch clamp recording was performed on cultured SD rat trigeminal ganglion (TG) neurons. (1) The majority of examined neurons (75%) were sensitive to 5-HT (3-300μM).5-HT3 receptor activated inward currents in a concentration-dependent manner, and I5-HT3 were blocked by ICS 205-930 (1μM), a selective antagonist of the 5-HT3 receptor. (2) 100μM 5-HT was repeated exerted for every three minut, and 5-HT3 receptor activated inward currents had no change. (3) When 260mOsm was preapplied extracellularly,5-HT3 receptor activated inward currents had no change. (4) When 348mOsm was preapplied extracellularly,5-HT3 receptor activated inward currents also had no change. It suggests that osmolality has no modulation on 5-HT3 receptor activated current.更多还原