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应用全细胞膜片钳记录大鼠新鲜分离背根神经节(DRG)神经元GABA-激活电流,观察咖啡因对GABA-激活电流(IGABA)的调制作用。结果显示:大部分受检细胞(97.4%,113/116)对外加GABA敏感。1-1000μmol/LGABA引起一剂量依赖性、有明显去敏感作用的内向电流。预加咖啡因(0.01-100μmol/L)30s后再加GABA能明显抑制GABA(100μmol/L)激活电流的幅值。预加咖啡因后GABA量效曲线明显下移;GABA-激活电流的最大值较之对照下降约57%;而Kd值(30μmol/L)几乎不变。该结果提示此种抑制为非竞争性的。预加氨茶碱(theophylline)亦可明显抑制GABA激活电流,同一浓度(10μmol/L)下氨茶碱的抑制作用较咖啡因的抑制作用强。预加安定(diazepam,1μmol/L)对GABA(10μmol/L)激活电流有增强作用,而预加咖啡因(10μmol/L)有拮抗安定增强IGABA的作用。胞内透析H-8后,几乎可以完全消除咖啡因对IGABA的抑制作用。本结果表明咖啡因在初级传入末稍可能产生对抗突触前抑制的效应。
Whole-cell patch-clamp was used to record the GABA-activated current of freshly isolated DRG neurons in rats. The modulation of GABA-activated current (IGABA) by caffeine was observed. The results showed that most of the tested cells (97.4%, 113/116) were sensitive to GABA. 1-1000μmol / LGABA caused a dose-dependent, significant desensitization of the inward current. Pretreatment with caffeine (0.01-100μmol / L) for 30s followed by GABA significantly inhibited the amplitude of GABA (100μmol / L) activation current. After pre-addition of caffeine, the dose-response curve of GABA decreased significantly. The maximum value of GABA-activated current decreased about 57% compared with the control. However, the Kd value (30μmol / L) remained almost unchanged. This result suggests that this inhibition is noncompetitive. Pretreatment with theophylline also markedly inhibited GABA activation, and the inhibitory effect of aminophylline was stronger than that of caffeine at the same concentration (10 μmol / L). Pretreatment with diazepam (1μmol / L) enhanced the activation current of GABA (10μmol / L), whereas pretreatment with caffeine (10μmol / L) antagonized diazepam and enhanced the activity of IGABA. Intracellular dialysis of H-8 almost completely abolished the inhibitory effect of caffeine on IGABA. This result shows that caffeine may have an effect against presynaptic inhibition at the primary afferent end.