Controversy exists regarding the effect of A1 adenosine receptor (AR) activation in the kidney during ischemia and reperfusion (I/R) injury. We sought to further characterize the role of A1 ARs in modulating renal function after I/R renal injury using both pharmacological and gene deletion approaches in mice. A1 AR knockout mice (A1KO) or their wild-type littermate controls (A1WT) were subjected to 30 min of renal ischemia. Some A1WT mice were subjected to 30 min of renal ischemia with or without pretreatment with 1,3-dipropyl-8-cyclopentylxanthine (DPCPX) or 2-chrolo-cyclopentyladenosine (CCPA), selective A1 AR antagonist and agonist, respectively. Plasma creatinine and renal histology were compared 24 h after renal injury. A1KO mice exhibited significantly higher creatinines and worsened renal histology compared with A1WT controls following renal I/R injury. A1WT mice pretreated with the A1 AR antagonist or agonist demonstrated significantly worsened or improved renal function, respectively, after I/R injury. In addition, A1WT mice pretreated with DPCPX or CCPA showed significantly increased or reduced markers of renal inflammation, respectively (renal myeloperoxidase activity, renal tubular neutrophil infiltration, ICAM-1, TNF-alpha, and IL-1beta mRNA expression), while demonstrating no differences in indicators of apoptosis. In conclusion, we demonstrate that endogenous or exogenous preischemic activation of A1 ARs protects against renal I/R injury in vivo via mechanisms leading to decreased necrosis and inflammation.