In BALB/c mouse thoracic aorta with intact endothelium, adenosine and its analogs produce a dose-response (DR) relaxation with a rank order of potency favoring A2B (NECA>CAD>adenosine>> CGS 21680; Talukder et al. LB4, FASEB J. 13(3), 1999). This was further supported by the fact that the A2B-receptor antagonist, alloxazine shifted DR to right for NECA. However, this study did not investigate the role of endothelium in adenosine-mediated relaxation of mouse aorta. Thus, the present study was conducted to investigate the role of endothelium in adenosine receptor-mediated relaxation. In organ bath studies, in intact endothelium, the EC50s for NECA, CAD and adenosine were found to be 0.05, 1.3 and 3.1 x 10−4 M, respectively. However, in the absence of endothelium, the EC50s for NECA, CAD and adenosine were; 0.098, 1.5 and 0.12 x 10−4 M, respectively indicating a shift in DR to the right. The intactness of endothelium in aortic tissue was tested with acetylcholine. NECA-induced DR relaxation was significantly blocked by L-NAME (10−4 M) in endothelium-intact tissues; whereas no inhibition was found in endothelium-denuded tissues. Moreover, the involvement of NO was confirmed by measuring total nitrites and nitrates (NOx) in endothelium-intact and -denuded aorta. NOx levels with L-NAME (10−4 M) alone and in combination with L-arginine (10−4 M) were 59% and 96%, respectively with respect to control (NECA+PE, p<0.05). These data demonstrate that adenosine-receptor mediated relaxation in mouse aorta is partially dependent on endothelium through adenosine A2B-receptor. (Supported by HL 027339)