Adenosine is released in response to cerebral ischemia when it is believed to exert a neuroprotective influence by inhibiting glutamate release via activation of adenosine A₁ receptors. To correlate adenosine production during in vitro ischemia with excitatory synaptic transmission we have measured directly extracellular adenosine whilst recording glutamatergic field excitatory postsynaptic potentials (fEPSPs) from area CA1 of the rat hippocampal slice. Slices were prepared from 16−23 day old rats killed humanely by cervical dislocation and maintained at 32 °C in aCSF gassed with 95 % O₂,5 % CO₂ (pH 7.4). In vitro ischemia was induced via oxygen/glucose deprivation (OGD; replaced by 95 % N₂,5 % CO₂ and 10 mM sucrose, respectively). In control conditions the fEPSPs were fully depressed by 99 ± 2 % (mean ± SEM, n = 4) after 6.25 min of OGD, whereas in the presence of 8-CPT (1 µM, n = 4), a competitive A₁ receptor antagonist, the fEPSPs were only depressed by 32 ± 3 % (unpaired t-test, p = 0.0012) demonstrating that the depression of glutamatergic transmission was largely dependent on A₁ receptor activation. To measure directly adenosine release in response to OGD an enzyme-based adenosine sensor was placed on the surface of area CA1 (Dale et al., 2000). In control experiments after exposure to OGD of 6.25 minutes (n = 3) the adenosine released 5 minutes after OGD was 13.88 ± 2.2 µM, with an estimated IC₅₀ = 0.64 ± 0.4 µM, after which transmission slowly recovered to 77 ± 0.3 % after 1 hr of re-oxygenation. To examine the basis of the protracted recovery of the fEPSP, 1 µM 8-CPT (n = 3) was applied to the slice 22 minutes after reoxygenation after which the fEPSP rapidly increased to close to baseline values (ie 100 %, of which 20 % of the increase can be attributed to basal (ie non OGD-induced) adenosine), compared to 23 ± 1 % of control immediately prior to 8-CPT. 8-CPT had no effect on extracellular adenosine. When two periods of OGD (8.5 − 9.5 min; n= 4) were given, recovery of the fEPSP after the first measured 63 ± 4 % at 40 min, whereas after only 26 min of recovery the second OGD resulted in 85 ± 1 % recovery of the fEPSP. This was associated with reduced adenosine release during the second OGD (12.1 ± 1.8 µM) compared to the first (20 ± 4 µM). These data suggest that elevated extracellular adenosine can account for the protracted recovery of transmission after OGD and that the accelerated recovery of synaptic transmission after repeated OGD can be due to adenosine depletion.