Mp. Skatchkov et al., QUANTIFICATION OF SUPEROXIDE RADICAL FORMATION IN INTACT VASCULAR TISSUE USING A CYPRIDINA LUCIFERIN ANALOG AS AN ALTERNATIVE TO LUCIGENIN, Biochemical and biophysical research communications (Print), 248(2), 1998, pp. 382-386
Lucigenin has been frequently used for the chemiluminescent detection
of superoxide (O-.(2)-) in intact tissue. More recent studies, however
, revealed that lucigenin per se causes formation of O-.(2)- raising d
oubt about this probe to detect reliably O-.(2)-. We therefore tested
a more recently described chemiluminescence probe (2-methyl-6-phenyl-3
,7-dihydroimidazol[1,2-alpha] pyrazine-3-one (CLA)) to estimate the ab
ility of vascular tissue to generate O-.(2)- as an alternative to luci
genin. In a cell free system as well as in vascular tissue, CLA-enhanc
ed chemiluminescence was dose dependently inhibited by superoxide dism
utase (SOD), vitamin C and sodium nitroprusside (SNP). Electron spin r
esonance studies revealed that lucigenin (250 mu M) but not CLA (1 mu
M) caused extra O-.(2)- production in vascular tissue. Stimulation of
vessels with NADH (200 mu M) increased CLA enhanced chemiluminescence,
which was inhibited by low concentrations of superoxide dismutase (20
U/ml). Endothelial removal as well as the nitric oxidase-synthase inh
ibitor increased CLA chemiluminescence in vessels. We conclude that CL
A is a sensitive and specific chemiluminescence probe to detect O-.(2)
- production ire intact vascular tissue. (C) 1998 Academic Press.