Activation of adenosine A(1) receptors (A(1)R) can inhibit arginine vasopressin (AVP)-induced cAMP formation in isolated cortical and medullary collecting ducts. To assess the in vivo consequences of the absence of A(1)R, we performed experiments in mice lacking A(1)R (A(1)R(-/-)). We assessed the effects of the vasopressin V(2) receptor (V(2)R) agonist 1-desamino-8-d-arginine vasopressin (dDAVP) on cAMP formation in isolated inner medullary collecting ducts (IMCD) and on water excretion in conscious water-loaded mice. dDAVP-induced cAMP formation in isolated IMCD was significantly greater ( approximately 2-fold) in A(1)R(-/-) compared with wild-type mice (WT) and, in contrast to WT, was not inhibited by the A(1)R agonist N6-cyclohexyladenosine. A(1)R(-/-) and WT had similar basal urinary excretion of vasopressin, expression of aquaporin-2 protein in renal cortex and medulla, and acute increases in urinary flow rate and electrolyte-free water clearance in response to the V(2)R antagonist SR121463 or acute water loading; the latter increased inner medullary A(1)R expression in WT. Dose dependence of dDAVP-induced antidiuresis after acute water loading was not different between the genotypes. However, A(1)R(-/-) had greater inner medullary expression of cyclooxygenase-1 under basal conditions and of the P2Y(2) and EP(3) receptor in response to water loading compared with WT mice. Thus vasopressin-induced cAMP formation is enhanced in isolated IMCD of mice lacking A(1)R, but the adenosine-A(1)R/V(2)R interaction demonstrated in vitro is likely compensated in vivo by multiple mechanisms, a number of which can be "uncovered" by water loading.