TRANSIENT MITOCHONDRIAL DEPOLARIZATIONS REFLECT FOCAL SARCOPLASMIC RETICULAR CALCIUM-RELEASE IN SINGLE-RAT CARDIOMYOCYTES

Digital imaging of mitochondrial potential in single rat cardiomyocyte s revealed transient depolarizations of mitochondria discretely locali zed within the cell, a phenomenon that we shall call ''flicker.'' Thes e events were usually highly localized and could be restricted to sing le mitochondria, but they could also be more widely distributed within the cell, Contractile waves, either spontaneous or in response to dep olarization with 50 mM K+, were associated with propagating waves of m itochondrial depolarization, suggesting that propagating calcium waves are associated with mitochondrial calcium uptake and consequent depol arization, Here we demonstrate that the mitochondrial flicker was dire ctly related to the focal release of calcium from sarcoplasmic reticul ar (SR) calcium stores and consequent uptake of calcium by local mitoc hondria. Thus, the events were dramatically reduced by (a) depletion o f SR calcium stores after long-term incubation in EGTA or thapsigargin (500 nM); (b) buffering intracellular calcium using BAPTA-AM loading; (c) blockade of SR calcium release with ryanodine (30 mu M); and (d) blockade of mitochondrial calcium uptake by microinjection of diaminop entane pentammine cobalt (DAPPAC), a novel inhibitor of the mitochondr ial calcium uniporter. These observations demonstrate that focal SR ca lcium release results in calcium microdomains sufficient to promote lo cal mitochondrial calcium uptake, suggesting a tight coupling of calci um signaling between SR release sites and nearby mitochondria.