MIPS521 is a positive allosteric modulator of adenosine A1 receptor (A1AR). MIPS521 also has a lower A1R allosteric affinity (pKB=4.95). MIPS521 exhibits pain-relieving effects in vivo. MIPS521 (compound 13o) (3-10 μM) improves the ability of R-PIA to promote A1AR-mediated ERK1/2 phosphorylation.MIPS521 (0.3-30 μM; pretreament for 10 min, co-treatment for 30 min) produces a concentration-dependent potentiation of signalling by ADO in an inhibition of cAMP assay (expressed as a percentage of the inhibition of 3 μM forskolin-mediated cAMP) in CHO cells. MIPS521 (1-30 μg in 10 μL; intrathecal administration) reverses mechanical hyperalgesia in rats, promoting robust antinociception.MIPS521 (10 μg in 10 μL; intrathecal administration) significantly reduces spontaneous pain in a conditioned place preference model.MIPS521 (1-30 μg in 10 μL; intrathecal administration) reduces eEPSCs in spinal cord from nerve-injured rats, with a pEC50 of 6.9. The maximum MIPS521-induced decrease in synaptic current amplitude is significantly greater in nerve-injured rats than in sham surgery controls.