Active and passive electrical properties of sheep cardiac Purkinje fibres were determined using conventional microelectrode techniques. In addition, oxygen tension was monitored immediately adjacent to the fibres with oxygen sensing microelectrodes. Oxygen was withdrawn for 130 min in a glucose-free solution (PO2 less than 15 mmHg), glycogen breakdown inhibited by 2-4-deoxyglucose and the following effects observed: (i) a decrease in action potential duration by as much as 80%; (ii) a decrease in maximal rate of rise of the action potential (Vmax) by 52%; (iii) an increase in internal longitudinal resistance (ri) by 176%; and (iv) a decrease in conduction velocity (theta) by 47%. The magnitude of the effects depended on PO2 (range: 4 to 46 mmHg). In particular, ri was steeply dependent upon the PO2 of the medium (glycogenolysis also inhibited by 2-4-deoxyglucose): ri increased to 470% above control when PO2 was reduced to 4 mmHg versus only a 47% increase in ri at PO2 = 11 mmHg. These effects were largely reversible after re-oxygenation and washout of 2-4-deoxyglucose although the time course of recovery of ri lagged behind that of action potential duration. Similar to previous studies, practically no effect of hypoxia was observed on the electrical properties of Purkinje fibres when glycogenolysis was not inhibited. The measured decrease of theta correlated relatively well with that predicted from linear cable theory using the changes of (i) Vmax and (ii) ri. No significant changes in membrane capacitance and only minor changes in action potential amplitude developed during combined hypoxia and inhibition of glycolysis.