Neonatal rat heart myocytes cultured for 3 days in fatty acid-enriched (5 × 10 5 M stearate, oleate, linoleate, linoelaidate or arachidonate at a 3 : 1, fatty acid (FA/albumin ratio) were analyzed by high-pressure liquid chromatography (HPLC) for the FA content of their neutral lipid, glyco-cardiolipid and phospholipid fractions and were tested for their susceptibility to arrhythmias produced by hypoxic/normoxic medium changes. Increases in the relative amount of FA in the above lipid fractions were generally in the order: elaidate = linoelaidate > linoleate = arachidonate > stearate = oleate. Cell morphology was unaffected by such treatment, but the cultures were more susceptible to induced arrhythmias. Cultures enriched with trans-FA, elaidate or linoelaidate, became markedly arrhythmic; arrhythmias in stearate-enriched cultures were of short duration; whereas the other FA produced intermediate increases. Susceptibility to arrhythmias could not be correlated with polyunsaturated fatty acid (PUFA)/saturated fatty acid (SATFA) ratios of FA in the phospholipid fraction.