| Biochemical characterization of NADPH-diaphorase in the chick embryo retina: stimulation by calcium ions and inhibition by arginine analogs. | |
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MedLine Citation:
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PMID: 7581050 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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Nitric oxide is an important intercellular messenger in the central nervous system. NADPH-diaphorase, reported to be identical to nitric oxide synthase, is present in specific groups of cells in several neural tissues, including the retina. We determined NADPH-diaphorase activity in homogenates of the chick embryo retina. The enzyme activity was measured spectrophotometrically at 585 nm after incubating retinal total homogenates (100-150 micrograms protein) with 1 mM NADPH and 0.5 mM nitroblue tetrazolium in 50 mM Tris buffer, pH 8.1, at 37 degrees C. NADPH-diaphorase was detected in 14-day old retinas and 53-65% of the enzyme activity was inhibited by 3 mM NG-nitro-L-arginine (NARG), the arginine analog. One mM L-N5-(1-iminoethyl)ornithine (NIO) was the most potent inhibitor (63% inhibition) while 3 mM NG-nitro-L-arginine methyl ester (NAME) (33% inhibition) and 1 mM NG-monomethyl-L-arginine acetate (NMMA) (14% inhibition) were less effective. Enzyme activity was increased by 48% by 2 mM calcium chloride, an effect reversed by 1 mM EGTA or EDTA. Basal enzyme levels were also partially inhibited by the chelators, indicating the presence of calcium-dependent and -independent isoforms of nitric oxide synthase in the retina. The results show that the NADPH-diaphorase assay is simple and sensitive and that the different isoforms of nitric oxide synthase expressed in chick retinal cells during development can be demonstrated. |
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Authors:
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M H de-Faria; J L do-Nascimento; R Paes-de-Carvalho |
Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't |
Journal Detail:
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Title: Brazilian journal of medical and biological research = Revista brasileira de pesquisas médicas e biológicas / Sociedade Brasileira de Biofísica ... [et al.] Volume: 28 ISSN: 0100-879X ISO Abbreviation: Braz. J. Med. Biol. Res. Publication Date: 1995 Feb |
Date Detail:
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Created Date: 1995-11-24 Completed Date: 1995-11-24 Revised Date: 2006-11-15 |
Medline Journal Info:
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Nlm Unique ID: 8112917 Medline TA: Braz J Med Biol Res Country: BRAZIL |
Other Details:
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Languages: eng Pagination: 252-5 Citation Subset: IM |
Affiliation:
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Departamento de Neurobiologia, Universidade Federal Fluminense, Niterói, RJ, Brasil. |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Animals Arginine / analogs & derivatives Calcium / pharmacology Chick Embryo Enzyme Activation NADPH Dehydrogenase / metabolism* Nitric Oxide Synthase / metabolism* Retina / enzymology* Time Factors |
| Chemical | |
Reg. No./Substance:
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74-79-3/Arginine; 7440-70-2/Calcium; EC 1.14.13.39/Nitric Oxide Synthase; EC 1.6.99.1/NADPH Dehydrogenase |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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