The influence of transmembrane flux limitations on cellular metabolism of purine nucleosides was assessed in whole organ studies. Transcapillary transport of the purine nucleosides adenosine (Ado) and inosine (Ino) via paracellular diffusion through interendothelial clefts in parallel with carrier-mediated transendothelial fluxes was studied in isolated, Krebs-Henseleit-perfused rabbit and guinea pig hearts. After injection into coronary inflow, multiple-indicator dilution curves were obtained from coronary outflow for 90 s for ¹³¹I-labeled albumin (intravascular reference tracer), [³H]arabinofuranosyl hypoxanthine (AraH; extracellular reference tracer and nonreactive adenosine analog), and either [¹⁴C]Ado or [¹⁴C]Ino. Ado or Ino was separated from their degradative products, hypoxanthine, xanthine, and uric acid, in each outflow sample by HPLC and radioisotope counting. Ado and Ino, but not AraH, permeate the luminal membrane of endothelial cells via a saturable transporter with permeability-surface area product PS_ecl and also diffuse passively through interendothelial clefts with the same conductance (PS_g) as AraH. These parallel conductances were estimated via fitting with an axially distributed, multi-pathway, four-region blood-tissue exchange model. PS_g for AraH were ~4 and 2.5 ml · g¹ · min¹ in rabbits and guinea pigs, respectively. In contrast, transplasmalemmal conductances (endothelial PS_ecl) were ~0.2 ml · g¹ · min¹ for both Ado and Ino in rabbit hearts but ~2 ml · g¹ · min¹ in guinea pig hearts, an order of magnitude different. Purine nucleoside metabolism also differs between guinea pig and rabbit cardiac endothelium. In guinea pig heart, 50% of the tracer Ado bolus was retained, 35% was washed out as Ado, and 15% was lost as effluent metabolites; 25% of Ino was retained, 50% washed out, and 25% was lost as metabolites. In rabbit heart, 45% of Ado was retained and 5% lost as metabolites, and 7% of Ino was retained and 3% lost as metabolites. We conclude that endothelial transport of Ado and Ino is a prime determinant of their metabolic fates: where transport rates are high, metabolic transformation is high.