Hypoxia in brain may lead to cell death by apoptosis and
necrosis. Concomitant is the formation of
purine nucleosides, e.g.
adenosine, a powerful endogenous
neuroprotectant. Despite vigorous studies, many aspects of the mechanisms involved in
purine-based protection are still unclear. In this study, we wanted to investigate the effect of
purine nucleosides on cellular responses to chemical
hypoxia. O(2)-sensitive neuronal
pheochromocytoma (PC12)-cells, which are widely used as a model system for sympathetic ganglion-like neurons, were subjected to chemical
hypoxia induced with
rotenone, an inhibitor of mitochondrial complex I.
Adenosine and its relatives
guanosine and
inosine were tested for their neuroprotective capability to improve neurite outgrowth and viability. In addition, cell lysates were analyzed for
mitogen-activated-protein-kinases (MAPK) activation by anti-active and anti-total MAPKinase immunoblotting.
Adenosine,
guanosine and
inosine significantly inhibited the loss of viability after hypoxic insult. In combination with
NGF,
purine nucleosides also partially rescued neurite outgrowth. The
MEK-1/-2 inhibitor
PD098059 inhibited
purine nucleoside-mediated protection up to 85.23% and also markedly decreased neurite formation induced by
NGF and
purine nucleosides in hypoxic cells. Immunoblot analysis revealed a strong activation of MAPKinase upon incubation of cells with
adenosine,
guanosine or
inosine. In combination with
NGF an additive effect was observed. Results suggested that activation of the MAPKinase pathway plays a vital role in
purine nucleoside-mediated protection of neuronal cells following hypoxic insult.