The aim of this study was to examine the effects of hypoxia (PO2 = 34 mm Hg), acidosis (pH 6.8), lactate (10 mM), glucose removal, hyperkalaemia (8 mM, and lysophosphatidylcholine (5-50-mu-M), alone and in combination, on paced sheep Purkinje fibre action potentials in an attempt to find a combination that simulated the electrophysiological changes associated with the delayed phase of ischaemia-induced arrhythmias. Modification of the physiological salt solution to produce a combination of acidosis, lactate, and lysophosphatidylcholine (5-mu-M) reduced the resting membrane potential and the maximum rate of depolarisation and prolonged action potential duration, these effects being stable from 60 to 120 min of superfusion. Hypoxia, glucose removal, or elevation of the extracellular concentration of potassium alone or in combination with the above factors caused action potential shortening. Thus, the electrophysiological changes associated with the delayed phase of ischaemia-induced arrhythmias in vivo can be simulated in vitro by a combination of acidosis, lactate, and lysophosphatidylcholine.