Adenosine vasoconstricts preglomerular arterioles via adenosine A₁receptors. Because adenosine also activates adenosine A₂receptors, its overall renal vascular actions are complex and not fully understood. The present study was performed to determine the relative contributions of adenosine A₁ and A_2a receptors to the responsiveness of the renal microvasculature to adenosine. Afferent and efferent arteriolar diameters were monitored in vitro using the blood-perfused rat juxtamedullary nephron preparation. Basal afferent and efferent arteriolar diameters averaged 17.1 ± 0.5 (n = 35) and 17.8 ± 0.5 (n = 20) μm, respectively. Superfusion with 0.1 and 1 μmol/l adenosine did not significantly alter afferent and efferent arteriolar diameters; however, 10 μmol/l adenosine significantly reduced afferent and efferent arteriolar diameters (−8.2 ± 0.8 and −5.7 ± 0.6%, respectively). The afferent and efferent arteriolar vasoconstrictor responses to adenosine waned at a dose of 100 μmol/l, such that diameters returned to values not significantly different from control within 2 min. During adenosine A₁ receptor blockade with 8-noradamantan-3-yl-1,3-dipropylxanthine (KW-3902: 10 μmol/l), 10 and 100 μmol/l adenosine significantly increased afferent diameter by, respectively, 8.1 ± 1.2 and 13.7 ± 1.3% (n = 14) and efferent arteriolar diameter by 6.4 ± 1.3 and 9.3 ± 1.2% (n = 8). The afferent and efferent arteriolar vasodilatory responses to adenosine in the presence of KW-3902 were significantly attenuated by addition of the adenosine A_2a receptor antagonist 1,3-dipropyl-7-methyl-8-(3,4-dimethoxystyryl)xanthine (KF-17837: 15 μmol/l, n = 7 and 6, respectively). The addition of KF-17837 alone significantly enhanced afferent (n = 15) and efferent (n = 6) arteriolar vasoconstrictor responses to 1, 10, and 100 μmol/l adenosine. These results indicate the presence of adenosine A₁ and A_2a receptors on afferent and efferent arterioles of juxtamedullary nephrons, such that adenosine A_2a receptor-mediated vasodilation partially buffers adenosine-induced vasoconstriction in both pre- and postglomerular segments of the renal microvasculature.