TY - JOUR
T1 - Imaging of nitric oxide in a living vertebrate using a diaminofluorescein probe
AU - Lepiller, Sandrine
AU - Laurens, Véronique
AU - Bouchot, André
AU - Herbomel, Philippe
AU - Solary, Eric
AU - Chluba, Johanna
N1 - Funding Information:
We thank Alain Pugin and David Wendehenne for helpful discussions, Frank Ménétrier (Microscopy Facility of No. 100 Federative Institute of Research), Claude Humbert (Common Facilities in Biology, University of Burgundy) for technical assistance, and Klaus-Dieter Kröncke for advice during this project and critical reading of the manuscript. This work was supported by grants from the Ligue Contre le Cancer Côte d’Or, Nièvre, Saône et Loire, the Association pour la Recherche contre le Cancer (ARC), and the Conseil Régional de Bourgogne.
PY - 2007/8/15
Y1 - 2007/8/15
N2 - Numerous approaches have been described to identify nitric oxide (NO), a free radical involved in various physiological and pathophysiological processes. One of these approaches is based on the use of chemical probes whose transformation by NO generates highly fluorescent derivatives, permitting detection of NO down to nanomolar concentrations. Here, we show that the cell-permeant diaminofluorophore 4-amino-5-methylamino-2′-7′-difluorofluorescein diacetate (DAF-FM-DA) can be used to detect NO production sites in a living vertebrate, the zebrafish Danio rerio. The staining pattern obtained in larvae includes the bulbus arteriosus, forming bones, the notochord, and the caudal fin. The specificity of the signal was confirmed by its decrease in animals exposed to a NO scavenger or a NO synthase inhibitor and its increase in the presence of a NO donor. Using this method, NO production was observed to change along development in the notochord and the caudal fin whereas it remained stable in the bulbus arteriosus. Local changes in NO production in response to stressful conditions were also detected by this method. Altogether, labeling with DAF-FM DA is an efficient method to monitor changes in NO production in live zebrafish under physiological as well as pathophysiological conditions, suggesting applications to drug screening and molecular pharmacology.
AB - Numerous approaches have been described to identify nitric oxide (NO), a free radical involved in various physiological and pathophysiological processes. One of these approaches is based on the use of chemical probes whose transformation by NO generates highly fluorescent derivatives, permitting detection of NO down to nanomolar concentrations. Here, we show that the cell-permeant diaminofluorophore 4-amino-5-methylamino-2′-7′-difluorofluorescein diacetate (DAF-FM-DA) can be used to detect NO production sites in a living vertebrate, the zebrafish Danio rerio. The staining pattern obtained in larvae includes the bulbus arteriosus, forming bones, the notochord, and the caudal fin. The specificity of the signal was confirmed by its decrease in animals exposed to a NO scavenger or a NO synthase inhibitor and its increase in the presence of a NO donor. Using this method, NO production was observed to change along development in the notochord and the caudal fin whereas it remained stable in the bulbus arteriosus. Local changes in NO production in response to stressful conditions were also detected by this method. Altogether, labeling with DAF-FM DA is an efficient method to monitor changes in NO production in live zebrafish under physiological as well as pathophysiological conditions, suggesting applications to drug screening and molecular pharmacology.
KW - DAF-FM DA
KW - Development
KW - Imaging
KW - Nitric oxide
KW - Vertebrate
KW - Zebrafish
UR - http://www.scopus.com/inward/record.url?scp=34447345949&partnerID=8YFLogxK
U2 - 10.1016/j.freeradbiomed.2007.05.025
DO - 10.1016/j.freeradbiomed.2007.05.025
M3 - Article
C2 - 17640572
AN - SCOPUS:34447345949
SN - 0891-5849
VL - 43
SP - 619
EP - 627
JO - Free Radical Biology and Medicine
JF - Free Radical Biology and Medicine
IS - 4
ER -