Inactivation of membrane surface ecto-5′-nucleotidase by sodium nitroprusside in C6 glioma cells

Ecto-5′-nucleotidase (nt5e),a predominant enzyme that produces extracellular adenosine from amp,plays an important role in a variety of physiological and pathophysiological processes. this study was performed to identify agents that affect nt5e activity using c6 glioma cells. when cells were incubated with sodium nitroprusside (snp),phorbol 12-myristate 13- acetate,forskolin,lipopolysaccharide,or interferon-γ,only snp inhibited nt5e activity in a time- and concentration-dependent manner (ic 50 = 1.2 μm). the inhibitory effect of snp was long-lasting even after snp washout; and its action was not mimicked by nitric oxide generating agents,8-bromo cyclic gmp,ferricyanide,ferrocyanide,or sodium cyanide. snp did not change nt5e mrna level or membrane surface protein expression. similar to snp,fe 2+ inhibited nt5e activity,but to a lesser extent. although fe 2+ is known to increase oxidative stress,fe 2+-mediated oxidative stress was not involved in snp inhibition of nt5e because the inhibition of nt5e by snp was not affected by superoxide dismutase and catalase. in contrast,addition of zn 2+,an essential metal co-factor of nt5e activity,prevented snp from inhibiting nt5e. these results suggest that snp disrupts a critical zn 2+-dependent enzyme activity and might be useful as a pharmacological tool for inhibiting nt5e. ©the japanese pharmacological society.