Radiochemical HPLC detection of arginine metabolism: measurement of nitric oxide synthesis and arginase activity in vascular tissue.

Nitric Oxide : Biology and Chemistry
Joseph P de BonoNicholas J Alp

Abstract

Nitric oxide (NO) plays a key role in vascular homeostasis. Accurate measurement of NO production by endothelial nitric oxide synthase (eNOS) is critical for the investigation of vascular disease mechanisms using genetically modified animal models. Previous assays of NO production measuring the conversion of arginine to citrulline have required homogenisation of tissue and reconstitution with cofactors including NADPH and tetrahydrobiopterin. However, the activity and regulation of NOS in vivo is critically dependant on tissue levels of these cofactors. Therefore, understanding eNOS regulation requires assays of NO production in intact vascular tissue that do not depend on the addition of exogenous cofactors and have sufficient sensitivity and specificity. We describe a novel technique, using radiochemical detection of arginine to citrulline conversion, to measure NO production within intact mouse aortas, without exogenous cofactors. We demonstrate the presence of arginase activity in mouse aortas which has the potential to confound this assay. Furthermore, we describe the use of N-hydroxy-nor-L-arginine (nor-NOHA) to inhibit arginase and permit specific detection of NO production in intact mouse tissue. Using this technique we...Continue Reading

References

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Citations

Mar 25, 2009·The Journal of Pharmacology and Experimental Therapeutics·Galen M PieperJeannette Vásquez-Vivar
May 6, 2008·American Journal of Physiology. Heart and Circulatory Physiology·Dhananjay K KaulMary E Fabry
Nov 27, 2008·The European Journal of Neuroscience·Timothy M FountaineRichard Wade-Martins
Jul 6, 2014·British Journal of Pharmacology·C CsonkaT Csont
Apr 14, 2009·Journal of Molecular and Cellular Cardiology·Chee-Wan LeeDavid J Paterson
Nov 26, 2015·PloS One·David I FraserJames S Ballantyne
Aug 13, 2009·Toxicon : Official Journal of the International Society on Toxinology·Kaline WaismamSandra H P Farsky
Oct 7, 2011·Surgery·Sandra L KavalukasAdrian Barbul
Mar 30, 2018·Cardiovascular Research·Gillian DouglasKeith M Channon
Apr 26, 2008·Circulation Research·Paul M Vanhoutte
Sep 4, 2009·Proceedings. Biological Sciences·F GarofaloS Imbrogno
Jul 7, 2015·Journal of Basic and Clinical Physiology and Pharmacology·Prabhakaran Prabha PreethaThankappan Rajamohan
Jul 17, 2012·Eukaryotic Cell·João B VicenteUpinder Singh
May 4, 2021·The Ocular Surface·Shyam Sunder TummanapalliMark D P Willcox

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