Imaging the nanomolar range of nitric oxide with an amplifier-coupled fluorescent indicator in living cells.

Nitric oxide (NO) is a small uncharged free radical that is involved in diverse physiological and pathophysiological mechanisms. NO is generated by three isoforms of NO synthase, endothelial, neuronal, and inducible ones. When generated in vascular endothelial cells, NO plays a key role in vascular tone regulation, in particular. Here, we describe an amplifier-coupled fluorescent indicator for NO to visualize physiological nanomolar dynamics of NO in living cells (detection limit of 0.1 nM). This genetically encoded high-sensitive indicator revealed that approximately 1 nM of NO, which is enough to relax blood vessels, is generated in vascular endothelial cells even in the absence of shear stress. The nanomolar range of basal endothelial NO thus revealed appears to be fundamental to vascular homeostasis.

[1]  F. Murad,et al.  Stimulation of guanylate cyclase by sodium nitroprusside, nitroglycerin and nitric oxide in various tissue preparations and comparison to the effects of sodium azide and hydroxylamine. , 1977, Journal of cyclic nucleotide research.

[2]  S. Moncada,et al.  Nitric oxide release accounts for the biological activity of endothelium-derived relaxing factor , 1987, Nature.

[3]  L. Ignarro,et al.  Endothelium-derived relaxing factor produced and released from artery and vein is nitric oxide. , 1987, Proceedings of the National Academy of Sciences of the United States of America.

[4]  J. Beavo,et al.  Inhibition and stimulation of photoreceptor phosphodiesterases by dipyridamole and M&B 22,948. , 1989, Molecular pharmacology.

[5]  S. Moncada,et al.  Characterization of three inhibitors of endothelial nitric oxide synthase in vitro and in vivo , 1990, British journal of pharmacology.

[6]  D. Wink,et al.  New nitric oxide-releasing zwitterions derived from polyamines , 1993 .

[7]  S. Moncada,et al.  The L-arginine-nitric oxide pathway. , 1993, The New England journal of medicine.

[8]  S. Moncada,et al.  Visualisation of nitric oxide generated by activated murine macrophages. , 1996, Biochemical and biophysical research communications.

[9]  T. Malinski,et al.  Nitric oxide measurement using electrochemical methods. , 1996, Methods in Enzymology.

[10]  S. Fujii,et al.  In vivo EPR detection and imaging of endogenous nitric oxide in lipopolysaccharide-treated mice , 1996, Nature Biotechnology.

[11]  S. Kawahara,et al.  Detection and imaging of nitric oxide with novel fluorescent indicators: diaminofluoresceins. , 1998, Analytical chemistry.

[12]  R. Busse,et al.  Characterization of NS 2028 as a specific inhibitor of soluble guanylyl cyclase , 1998, British journal of pharmacology.

[13]  Y. Umezawa,et al.  Fluorescent indicators for cyclic GMP based on cyclic GMP-dependent protein kinase Ialpha and green fluorescent proteins. , 2000, Analytical chemistry.

[14]  L. Ignarro.  Nitric oxide : biology and pathobiology , 2000 .

[15]  H Kojima,et al.  Bioimaging of nitric oxide with fluorescent indicators based on the rhodamine chromophore. , 2001, Analytical chemistry.

[16]  J. Corbin,et al.  Cyclic nucleotide phosphodiesterases: relating structure and function. , 2001, Progress in nucleic acid research and molecular biology.

[17]  C. Napoli,et al.  Nitric oxide and atherosclerosis. , 2001, Nitric oxide : biology and chemistry.

[18]  T. Nagano,et al.  Bioimaging of nitric oxide. , 2002, Chemical reviews.

[19]  Y. Umezawa,et al.  Production of PtdInsP3 at endomembranes is triggered by receptor endocytosis , 2003, Nature Cell Biology.

[20]  Yasuteru Urano,et al.  Highly sensitive fluorescence probes for nitric oxide based on boron dipyrromethene chromophore-rational design of potentially useful bioimaging fluorescence probe. , 2004, Journal of the American Chemical Society.

[21]  A. Friebe,et al.  Nitric oxide-sensitive guanylyl cyclase: structure and regulation , 2004, Neurochemistry International.

[22]  F. Murad,et al.  Regulation of nitric oxide and soluble guanylyl cyclase , 2004, Brain Research Bulletin.

[23]  Yasuteru Urano,et al.  Highly sensitive near-infrared fluorescent probes for nitric oxide and their application to isolated organs. , 2005, Journal of the American Chemical Society.