We tested whether the brain and kidney respond differently to cardiopulmonary bypass (CPB) and to changes in perfusion conditions during CPB. Therefore, in ovine CPB, we assessed regional cerebral oxygen saturation (rSO₂) by near-infrared spectroscopy and renal cortical and medullary tissue oxygen tension (PO₂), and, in some protocols, brain tissue PO₂, by phosphorescence lifetime oximetry. During CPB, rSO₂ correlated with mixed venous SO₂ (r = 0.78) and brain tissue PO₂ (r = 0.49) when arterial PO₂ was varied. During the first 30 min of CPB, brain tissue PO₂, rSO₂ and renal cortical tissue PO₂ did not fall, but renal medullary tissue PO₂ did. Nevertheless, compared with stable anaesthesia, during stable CPB, rSO₂ (66.8 decreasing to 61.3%) and both renal cortical (90.8 decreasing to 43.5 mm Hg) and medullary (44.3 decreasing to 19.2 mm Hg) tissue PO₂ were lower. Both rSO₂ and renal PO₂ increased when pump flow was increased from 60 to 100 mL kg⁻¹ min⁻¹ at a target arterial pressure of 70 mm Hg. They also both increased when pump flow and arterial pressure were increased simultaneously. Neither was significantly altered by partially pulsatile flow. The vasopressor, metaraminol, dose-dependently decreased rSO₂, but increased renal cortical and medullary PO₂. Increasing blood haemoglobin concentration increased rSO₂, but not renal PO₂. We conclude that both the brain and kidney are susceptible to hypoxia during CPB, which can be alleviated by increasing pump flow, even without increasing arterial pressure. However, increasing blood haemoglobin concentration increases brain, but not kidney oxygenation, whereas vasopressor support with metaraminol increases kidney, but not brain oxygenation.