Mitochondria play a critical role in mediating the cellular response to oxidants formed during acute and chronic cardiac dysfunction. It is widely assumed that, as cells are subjected to stress, mitochondria are capable of drawing upon a ‘reserve capacity’ which is available to serve the increased energy demands for maintenance of organ function, cellular repair or detoxification of reactive species. This hypothesis further implies that impairment or depletion of this putative reserve capacity ultimately leads to excessive protein damage and cell death. However, it has been difficult to fully evaluate this hypothesis since much of our information about the response of the mitochondrion to oxidative stress derives from studies on mitochondria isolated from their cellular context. Therefore the goal of the present study was to determine whether ‘bioenergetic reserve capacity’ does indeed exist in the intact myocyte and whether it is utilized in response to stress induced by the pathologically relevant reactive lipid species HNE (4-hydroxynonenal). We found that intact rat neonatal ventricular myocytes exhibit a substantial bioenergetic reserve capacity under basal conditions; however, on exposure to pathologically relevant concentrations of HNE, oxygen consumption was increased until this reserve capacity was depleted. Exhaustion of the reserve capacity by HNE treatment resulted in inhibition of respiration concomitant with protein modification and cell death. These data suggest that oxidized lipids could contribute to myocyte injury by decreasing the bioenergetic reserve capacity. Furthermore, these studies demonstrate the utility of measuring the bioenergetic reserve capacity for assessing or predicting the response of cells to stress.