We have reported previously that peroxynitrite stimulates L-arginine release from astrocytes, but the mechanism responsible for such an effect remains elusive. To
explore this issue, we studied the regulation of
L-[3
H]arginine transport by either exogenous or endogenous peroxynitrite in glial cells. A 2-fold peroxynitritemediated stimulation of L-arginine release in C6 cells
was found to be Na -independent, was prevented by 5
mM L-arginine and, although only in the presence of Na ,
was blocked by 5 mM L-alanine or L-leucine. Peroxynitrite-mediated stimulation of L-arginine uptake was
trans-stimulated by 10 mM L-arginine and was inhibited
in a dose-dependent fashion (ki of 40 M) by the system
y inhibitor N-ethylmaleimide in C6 cells. Endogenous
production of peroxynitrite in lipopolysaccharidetreated astrocytes triggered an increased L-arginine
transport activity without affecting Cat1 L-arginine
transporter mRNA levels. However, Western blot analyses of peroxynitrite-treated astrocytes and C6 glial cells
revealed a 3-nitrotyrosinated anti-Cat1-immunopositive
band, strongly suggesting peroxynitrite-mediated Cat1
nitration. Furthermore, peroxynitrite stimulation of Larginine release was abolished in fibroblast cells homozygous for a targeted inactivation of the Cat1 gene.
Finally, peroxynitrite-triggered L-arginine released
from astrocytes was efficiently taken up by neurons in
an insert-based co-culture system. These results
strongly suggest that peroxynitrite-mediated activation
of the Cat1 transporter in glial cells may serve as a
mechanism focused to replenish L-arginine in the neighboring neurons.
