Mechanism of guanosine-induced neuroprotection in rat hippocampal slices submitted to oxygen–glucose deprivation

Guanine derivates have been implicated in many relevant extracellular roles, such as modulation of glutamate transmission, protecting neurons against excitotoxic damage. Guanine derivatives are spontaneously released to the extracellular space from cultured astrocytes during oxygen–glucose deprivation (OGD) and may act as trophic factors, glutamate receptors blockers or glutamate transport modulators, thus promoting neuroprotection. The aim of this study was to evaluate the mechanisms involved in the neuroprotective role of the nucleoside guanosine in rat hippocampal slices submitted to OGD, identifying a putative extracellular binding site and the intracellular signaling pathways related to guanosine-induced neuroprotection. Cell damage to hippocampal slices submitted to 15 min of OGD followed by 2 h of reperfusion was decreased by the addition of guanosine (100 μM) or guanosine-5′-monophosphate (GMP, 100 μM). The neuroprotective effect of guanosine was not altered by the addition of adenosine receptor antagonists, nucleosides transport inhibitor, glutamate receptor antagonists, glutamate transport inhibitors, and a non-selective Na+ and Ca2+ channel blocker. However, in a Ca2+-free medium (by adding EGTA), guanosine was ineffective. Nifedipine (a Ca2+ channel blocker) increased the neuroprotective effect of guanosine and 4-aminopyridine, a K+ channel blocker, reversed the neuroprotective effect of guanosine. Evaluation of the intracellular signaling pathways associated with guanosine-induced neuroprotection showed the involvement of PKA, PKC, MEK and PI-3K pathways, but not CaMKII. Therefore, this study shows guanosine is acting via K+ channels activation, depending on extracellular Ca2+ levels and via modulation of the PKA, PKC, MEK and/or PI-3K pathways.