Glutamate and aspartate measurements in individual planaria by rapid capillary electrophoresis
Glutamate and aspartate measurements in individual planaria by rapid capillary electrophoresis
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DOI：
10.1016/j.vascn.2010.08.002
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发表时间：
2011-01-01
影响因子：
1.9
通讯作者：
Shackman, Jonathan G.
中科院分区：
文献类型：
--
作者：
Vyas, Chandni A.;Rawls, Scott M.;Shackman, Jonathan G.
介绍：Planaria提出了一个独特的模式生物研究原始中枢神经系统。哺乳动物的主要兴奋性神经递质，谷氨酸和天冬氨酸，先前已通过高压液相色谱法（HPLC）在涡虫测量。一个更快的提取和分析程序，使用毛细管电泳（CE）的开发，确认这些氨基酸的存在下，在单一的涡虫匀浆。方法：在盐酸/乙腈中均质化和离心单独的涡虫后，用萘-2，3-二甲醛（NDA）衍生谷氨酸和天冬氨酸。使用毛细管电泳-激光诱导荧光（CE-LIF）测定标记的氨基酸。结果：CE-LIF电泳图在不到1分钟内生成。谷氨酸和天冬氨酸的量分别为1200 +/-500和1900 +/-700 pmol/mg-真涡虫（n=22）。谷氨酸和天冬氨酸的加标平均回收率分别为96%和91%。讨论内容：高通量方法提供了在发育或刺激条件下定量兴奋性神经递质变化的能力。监测多种神经递质水平的能力提供了将行为反应与涡虫中的生化变化相关联的机会。(C)2010年爱思唯尔公司All rights reserved.
Introduction: Planaria present a unique model organism for studying primitive central nervous systems. The major mammalian excitatory neurotransmitters, glutamate and aspartate, have previously been measured in planaria via high pressure liquid chromatography (HPLC). A faster extraction and analysis procedure using capillary electrophoresis (CE) was developed which confirms the presence of these amino acids in single planaria homogenates. Method: Following homogenization and centrifugation of individual planaria in hydrochloric acid/acetonitrile, glutamate and aspartate were derivatized with naphthalene-2, 3-dicarboxaldehyde (NDA). The labeled amino acids were measured using capillary electrophoresis with laser-induced fluorescence (CE-LIF). Results: CE-LIF electropherograms were generated in less than 1 min. The mean +/- S.D. amounts of glutamate and aspartate were 1200 +/- 500 and 1900 +/- 700 pmol/mg-planarian (n=22), respectively. Spiked average recoveries of glutamate and aspartate were 96% and 91%, respectively. Discussion: The high-throughput method provides the ability to quantitate changes in excitatory neurotransmitters under developmental or stimulatory conditions. The capability to monitor multiple neurotransmitter levels offers the opportunity to correlate behavioral responses with biochemical changes in planaria. (C) 2010 Elsevier Inc. All rights reserved.