Adenosine, through the A1 receptor (A1R), is an endogenous anticonvulsant. Development of adenosine receptor agonists as antiseizure drugs has been hampered by cardiac effects. A moderately A1R-selective agonist, MRS5474, has been reported to suppress seizures without appreciable cardiac action (Tosh et al., 2012, J Med Chem, 55:8075). We aimed at understanding its mechanism of action. MRS5474 (50-500nM) was devoid of effect upon rodent excitatory synaptic signals in hippocampal slices, except when hyperexcitability was previously induced in vivo or ex vivo. This contrasted with the effect of other A1R agonists. MRS5474 (50nM) inhibited GAT-1 mediated GABA uptake (47±6.5%, n=15, p<0.05)), an action not blocked by an A1R antagonist, but blocked by an A3R antagonist, MRS1523 (10μM), and mimicked by an A3R agonist, MRS5698 (100nM, %inhibition: 48±11%, n=8, p>0.05 vs MRS5474). A3R was overexpressed (P<0.05) in human hippocampal samples from epileptic patients undergoing surgery. MRS5474 induced a concentration-dependent potentiation (P<0.05, n=8-15) of GABA currents in oocytes micro-transplanted with human hippocampal membranes prepared from epileptic patients but not in those prepared from non-epileptic individuals (P>0.05, n=10). The action of MRS5474 on GABAergic currents was blocked by the A3R antagonist. We thus identified a drug that activates A3R and has selective actions on epileptic hippocampus. This underscores A3R as a promising target for development of antiseizure medications. Acknowledgments: PTDC/MED-FAR/30933/2017, H2020 GA 952455-EpiEpiNet, iMM-BI-4-2022. Tatiana Morais, Sandra Vaz, Sara Xapelli, Claudia Valente, Maria José Diógenes (FMUL/iMM/UL) for discussions, and Eleonora Aronica (Univ. Amsterdam), Beatriz Bordadágua, Alexandre Rainha-Campos, Carla Bentes (CHLN) for human samples.