Document Detail


Citrulline immunohistochemistry for demonstration of NOS activity in vivo and in vitro.
MedLine Citation:
PMID:  10944418     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Nitric oxide (NO), a biomolecule with major cytotoxic potency, is generated by NO synthases (NOS) utilizing l-arginine as substrate and citrulline is formed as a "side product." In brain tissue, citrulline is considered to be produced exclusively by NOS, due to the incomplete urea cycle in the brain. We aimed to characterize NOS activity by citrulline immunostaining in different cell types of the brain under in situ conditions and in slice and culture experiments. NOS-positive neurons and activated microglial cells were the most prominent citrulline-positive structures. Lack of citrulline immunoreaction in neurons of nNOS knockout mice emphasizes the dependency of citrulline positivity on NOS activity, and likewise there was no citrulline staining after application of the NOS inhibitors 7-nitroindazole and NIL. Interestingly, only a portion of NOS-containing neurons costained for citrulline. The inhibition of argininosuccinate synthetase by alpha-methyl-dl-aspartate increased the number of citrulline-positive cells, apparently due to reduction of the turnover rate of citrulline. Cells positive for NOS but negative for citrulline may indicate that the enzyme is either not activated or inhibited by cellular control mechanisms. The fact that not all citrulline-positive cells were NOS positive may be explained by an insufficient detection sensitivity or by disparate sites of citrulline production and recycling. The present results show that citrulline immunocytochemistry offers a viable and convenient means for studying NOS activity at the single-cell level to elicit its posttranslational control under physiological and pathophysiological conditions.
Authors:
G Keilhoff; M Reiser; A Stanarius; E Aoki; G Wolf
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Publication Detail:
Type:  In Vitro; Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Nitric oxide : biology and chemistry / official journal of the Nitric Oxide Society     Volume:  4     ISSN:  1089-8603     ISO Abbreviation:  Nitric Oxide     Publication Date:  2000 Aug 
Date Detail:
Created Date:  2000-09-07     Completed Date:  2000-09-07     Revised Date:  2008-11-21    
Medline Journal Info:
Nlm Unique ID:  9709307     Medline TA:  Nitric Oxide     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  343-53     Citation Subset:  IM    
Copyright Information:
Copyright 2000 Academic Press.
Affiliation:
Institute of Medical Neurobiology, Otto-von-Guericke University of Magdeburg, Leipziger Strasse 44, Magdeburg, D-39120, Germany. Gerburg.Keilhoff@medizin.uni-magdeburg.de
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MeSH Terms
Descriptor/Qualifier:
Animals
Antigens, Surface / immunology
Argininosuccinate Synthase / antagonists & inhibitors
Brain / cytology,  enzymology*
Cells, Cultured
Citrulline / analysis*,  immunology
Corpus Striatum / chemistry,  cytology,  enzymology
Interferon-gamma / pharmacology
Lipopolysaccharides / pharmacology
Male
Mice
Mice, Knockout
Microscopy, Fluorescence
N-Methylaspartate / analogs & derivatives*,  pharmacology
Neuroglia / chemistry,  drug effects,  enzymology
Neurons / chemistry,  drug effects,  enzymology
Nitric Oxide Synthase / antagonists & inhibitors,  immunology,  metabolism*
Nitric Oxide Synthase Type I
Nitric Oxide Synthase Type II
Rats
Rats, Wistar
Chemical
Reg. No./Substance:
0/Antigens, Surface; 0/Lipopolysaccharides; 372-75-8/Citrulline; 6384-92-5/N-Methylaspartate; 82115-62-6/Interferon-gamma; 866-73-9/2-methylaspartic acid; EC 1.14.13.39/Nitric Oxide Synthase; EC 1.14.13.39/Nitric Oxide Synthase Type I; EC 1.14.13.39/Nitric Oxide Synthase Type II; EC 1.14.13.39/Nos1 protein, mouse; EC 1.14.13.39/Nos1 protein, rat; EC 1.14.13.39/Nos2 protein, mouse; EC 1.14.13.39/Nos2 protein, rat; EC 6.3.4.5/Argininosuccinate Synthase

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