Previous studies showed increased extracellular nucleotides during renal ischemia-reperfusion. While nucleotides represent the main source for extracellular adenosine and adenosine signaling contributes to renal protection from ischemia, we hypothesized a role for ecto-nucleoside-triphosphate-diphosphohydrolases (E-NTPDases) in renal protection. We used a model of murine ischemia-reperfusion and in situ ischemic preconditioning (IP) via a hanging weight system for atraumatic renal artery occlusion. Initial studies with a nonspecific inhibitor of E-NTPDases (POM-1) revealed inhibition of renal protection by IP. We next pursued transcriptional responses of E-NTPDases (E-NTPDase1-3, and 8) to renal IP, and found a robust and selective induction of E-NTPDase1/CD39 transcript and protein. Moreover, based on clearance studies, plasma electrolytes, and renal tubular histology, IP protection was abolished in gene-targeted mice for cd39 whereas increased renal adenosine content with IP was attenuated. Furthermore, administration of apyrase reconstituted renal protection by IP in cd39^-/- mice. Finally, apyrase treatment of wild-type mice resulted in increased renal adenosine concentrations and a similar degree of renal protection from ischemia as IP treatment. Taken together, these data identify CD39-dependent nucleotide phosphohydrolysis in renal protection. Moreover, the present studies suggest apyrase treatment as a novel pharmacological approach to renal diseases precipitated by limited oxygen availability.--Grenz, A., Zhang, H., Hermes, M., Eckle, T., Klingel, K., Huang, D. Y., Müller, C. E., Robson, S. C., Osswald, H., Eltzschig, H. K. Contribution of E-NTPDase1 (CD39) to renal protection from ischemia-reperfusion injury.