Capillary Electrophoresis Analysis of Nitric Oxide Synthase Related Metabolites in Single Identified Neurons
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- 作者：Philip D. Floyd ; Leonid L. Moroz ; Rhanor Gillette ; and Jonathan V. Sweedler
- 刊名：Analytical Chemistry
- 出版年：1998
- 出版时间：June 1, 1998
- 年：1998
- 卷：70
- 期：11
- 页码：2243 - 2247
- 全文大小：82K
- 年卷期：v.70,no.11(June 1, 1998)
- ISSN：1520-6882
文摘
Intracellular concentrations of L-citrulline (Cit) anditsmetabolites are related to nitric oxide synthase (NOS)activity, an enzyme producing the intercellular messengerNO in animal tissues including the nervous system. Acapillary electrophoresis system using laser-induced fluorescence detection is described, and methods are developed to monitor the levels of L-arginine (Arg), Cit,andrelated molecules in identified neurons of the marineslugs, Pleurobranchaea californica andAplysia californica. The limits of detection for Arg, Cit,L-argininosuccinate, L-ornithine, and L-arginine phosphaterangefrom 50 amol to 17 fmol (5 nM to 17 
M in the neuronsunder study); these detection limits are significantlylowerthan actual intracellular levels of the metabolites,allowingthe direct assay of single cells. The levels of NOSmetabolites in individual neurons varied from 6 (Arg) and4 mM (Cit) in putative NOS-containing neurons down to<1 M (undetectable) levels in many putative NOS-negative cells. The Arg/Cit ratio is independent ofcellvolume, correlates with NADPH-diaphorase staining, andappears to be a characteristic parameter for the presenceof NOS activity in identified neurons.
M in the neuronsunder study); these detection limits are significantlylowerthan actual intracellular levels of the metabolites,allowingthe direct assay of single cells. The levels of NOSmetabolites in individual neurons varied from 6 (Arg) and4 mM (Cit) in putative NOS-containing neurons down to<1 M (undetectable) levels in many putative NOS-negative cells. The Arg/Cit ratio is independent ofcellvolume, correlates with NADPH-diaphorase staining, andappears to be a characteristic parameter for the presenceof NOS activity in identified neurons.