Since forty years, it is known that long-chain polyunsaturated fatty acids of the series n-3 (PUFAs) have cardioprotective effects by preventing cardiac arrhythmias1. The main n-3 PUFAs are eicosapentaenoic acid (C20:5 n-3, EPA) and docosahexaenoic acid (C22:6 n-3, DHA) that are highly peroxidable due to the presence of the skipped dienes2. The effects of n-3 PUFA on cardiac function are still debated, notably because of the lack of information on the mechanisms involved3. For example, it is not really known which the active lipid is: the PUFA or one of its oxygenated metabolites. A diet enriched in n-3 PUFAs (mainly fish-based), leads to enrichment in these fatty acids of cardiac cell membranes. Our hypothesis is that, during an infarct, the oxidative stress and the generation of reactive oxygen species might be responsible for an oxidation of membrane-bound PUFAs and the oxygenated metabolites generated might modulate the activity of ionic channels to exert anti-arrhythmic effects4. We thus decided to investigate the influence of the peroxidation of DHA on its potentially anti-arrhythmic properties. In this study, we applied DHA free acid or DHA methyl ester (OMe) on freshly isolated mouse ventricular myocytes without or with α-tocopherol (Vitamin E, to prevent oxidation) or hydrogen peroxide (to enhance oxidation). We investigated, using a photometric system, calcium transients (using the ratiometric calcium fluorescent dye Indo-1) and cell shortening of electrically stimulated myocytes. By stimulating β-adrenergic pathways with 10 nM isoproterenol, it is possible to observe the occurence of arrhythmic events. We observed that DHA free acid reduced the percentage of arrhythmic cells but do not DHA-OMe. The effects of DHA are correlated with the peroxidation of the fatty acid since α-tocopherol prevented the anti-arrhythmic effects while hydrogen peroxide enhanced them. In resting cells, single RyR channels spontaneously open with a very low frequency. An increased frequency of these events can cause propagating calcium waves that can lead to arrhythmias. The spontaneous calcium released events named sparks, can be observed by confocal microscopy. Here again, we observed that the frequency of calcium sparks is negatively correlated with the oxidative status of the DHA solution. These results suggest thus that rather than DHA itself, it is one or more non-enzymatic oxygenated metabolites derived from DHA that are potentially anti-arrhythmic such as neuroprostanes and/or neurofuranes4.