Division of Cardiovascular Surgery, Cardiovascular Center, Far-Eastern Memorial Hospital, New Taipei City 22060, Taiwan; Department of Electrical Engineering, Yuan Ze University, Taoyuan City 32003, Taiwan.
Department of Anesthesiology, Far-Eastern Memorial Hospital, New Taipei City 22060, Taiwan; Department of Mechanical Engineering, Yuan Ze University, Taoyuan City 32003, Taiwan.
Neurotoxicology. 2022 Jul;91:218-227. doi: 10.1016/j.neuro.2022.05.014. Epub 2022 May 25.
The inhibition of the excessive release of glutamate in the brain has emerged as a promising new option for developing therapeutic strategies for neurodegenerative disorders. This study investigated the effect and mechanism of lappaconitine, a diterpenoid alkaloid found in species of Aconitum, on glutamate release in rat cerebral cortex nerve terminals (synaptosomes). Here, we report that in the rat cortical synaptosomal preparation, lappaconitine reduced the K channel blocker 4-aminopyridine (4-AP)-evoked Ca-dependent release of glutamate. The inhibitory effect of lappaconitine on the evoked glutamate release was blocked by the vesicular transporter inhibitor bafilomycin A1 and calcium-chelating agent ethylene glycol tetraacetic acid (EGTA), but was unaffected by exposure to the glutamate transporter inhibitor dl-threo-beta-benzyloxyaspartate (dl-TBOA). The depolarization-induced elevation of cytosolic calcium concentration ([Ca]c) was inhibited by lappaconitine, while the 4-AP-mediated depolarization of the synaptosomal membrane potential was not affected. The inhibition of glutamate release by lappaconitine was markedly decreased in synaptosomes pretreated with the Ca2.3 (R-type) channel blocker SNX-482 or the protein kinase A inhibitor H89. Nevertheless, the lappaconitine-mediated inhibition of glutamate release was not abolished by the intracellular Ca-release inhibitors dantrolene and CGP37157. Lappaconitine also significantly decreased the 4-AP-induced phosphorylation of PKA and SNAP-25, a presynaptic substrate for PKA. Our data suggest that lappaconitine reduces Ca influx through R-type Ca channels, subsequently reducing the protein kinase A cascade to inhibit the evoked glutamate release from rat cerebral cortex nerve terminals.
脑内谷氨酸释放的抑制作用已成为开发神经退行性疾病治疗策略的一种有前途的新选择。本研究探讨了乌头碱，一种存在于乌头属植物中的二萜生物碱，对大鼠大脑皮质神经末梢（突触体）中谷氨酸释放的影响和作用机制。在这里，我们报告在大鼠皮质突触体制备中，乌头碱可减少 K 通道阻断剂 4-氨基吡啶（4-AP）诱导的 Ca 依赖性谷氨酸释放。乌头碱对诱发谷氨酸释放的抑制作用被囊泡转运抑制剂巴弗洛霉素 A1 和钙螯合剂乙二醇四乙酸（EGTA）阻断，但不受谷氨酸转运抑制剂 dl-threo-β-苄氧基天冬氨酸（dl-TBOA）的影响。乌头碱抑制去极化诱导的细胞溶质钙离子浓度升高([Ca]c)，而 4-AP 介导的突触体膜电位去极化不受影响。用 Ca2.3（R 型）通道阻断剂 SNX-482 或蛋白激酶 A 抑制剂 H89 预处理突触体后，乌头碱对谷氨酸释放的抑制作用明显降低。然而，细胞内 Ca 释放抑制剂丹曲林和 CGP37157 并不能消除乌头碱介导的谷氨酸释放抑制作用。乌头碱还显著降低了 4-AP 诱导的 PKA 和 SNAP-25 的磷酸化，PKA 的一种突触前底物。我们的数据表明，乌头碱通过减少 R 型 Ca 通道的钙内流，随后减少蛋白激酶 A 级联反应，抑制大鼠大脑皮质神经末梢诱发的谷氨酸释放。
