Monophasic action potentials, recorded with contact non-suction electrodes, have been used both clinically and experimentally. However, to date no systematic microelectrode validation studies have been done to underlying myocardial cell populations from different myocardial regions with different transmembrane potential profiles. In the present study transmembrane action potential properties, recorded with standard microelectrodes, were compared with monophasic action potentials recorded with contact electrodes in three different (endocardium, epicardium, and free running Purkinje fibre) isolated canine preparations during pacing and during spontaneous automatic activity. The mean transmembrane durations at 50% and 90% repolarisations (APD50 and APD90) of 19-30 cells at a monophasic action potential recording site was not statistically significant from monophasic action potential duration in all three tissue preparations studied. However, in endocardial preparations, composed of superficial (1-2 cell layers) Purkinje fibres with deeper ventricular muscle cells, the APD50 (139(17) ms) and APD90 (181(26) ms) of monophasic action potentials more closely reflected (but not significantly different) the underlying deeper ventricular muscle cells (APD50 134(14) ms and APD90 167(15) ms) rather than the mean transmembrane action potential durations of the underlying most superficial Purkinje fibres (166(22) ms for APD50 and 210(30) ms for APD90) (p less than 0.025). Tetrodotoxin (TTX) at 1 x 10(-6) mol.litre-1 shortened Purkinje fibre action potential duration and slightly lengthened that of ventricular muscle. Simultaneously recorded monophasic action potential showed an intermediate change in action potential duration. Incremental pacing and applied single premature stimuli resulted in similar degrees of shortening of action potential duration for both monophasic action potential and transmembrane potential in all three preparations. In endocardial preparations, barium chloride (4 mmol.litre-1) superfusion induced early afterdepolarisations, and spontaneous phase 4 depolarisations (n = 6) in both Purkinje and ventricular muscle cells giving rise to spontaneous automatic activity. These abnormal automatic activities were accurately detected by simultaneous monophasic action potential recordings. Suppression of automaticity by verapamil (0.2-0.5 micrograms.ml-1) as confirmed by transmembrane action potential recordings were similarly detected by monophasic action potential recordings (ABSTRACT TRUNCATED AT 250 WORDS)