Subcellular metabolic transients and mitochondrial redox waves in heart cells

Abstract

Precise matching of energy supply with demand requires delicately balanced control of the enzymes involved in substrate metabolism. In response to a change in substrate supply, the nonlinear properties of metabolic control may induce complex dynamic behavior. Using confocal imaging of f lavoprotein redox potential and mitochondrial membrane potential, we show that substrate deprivation leads to subcellular heterogeneity of mitochondrial energization in intact cells. The complex spatiotemporal patterns of redox and matrix potential included local metabolic transients, cell-wide coordinated redox transitions, and propagated metabolic waves both within and between coupled cells. Loss of metabolic synchrony during mild metabolic stress reveals that intra-and intercellular control of mitochondrial function involves diffusible cytoplasmic messengers.Nonlinear dynamic control in metabolic pathways plays a crucial role in conferring sensitivity and rapid responses to changes in cellular workload or environmental conditions. Through a combination of allosteric and stoichiometric effects on multiple control points, the enzymatic pathways involved in energy metabolism can undergo large changes in activity in response to small perturbations of key effector molecules (1). Experimental and theoretical studies have demonstrated that, under some conditions, such finely tuned systems may become unstable and display self-organizing oscillations, bistability, or chaotic behavior (reviewed in ref.2). Early observations of metabolic oscillations in yeast (3-5) and our own investigation of oscillations of NADH and sarcolemmal K,ATP current in substrate-deprived heart cells (6) provide evidence that such phenomena can be observed in intact cells. In our previous work, modulation of the amplitude and͞or frequency of the oscillations by external glucose led to the hypothesis that they were driven by a glycolytic oscillator, but the predominantly mitochondrial origin of the NADH f luorescence in cardiomyocytes (7) suggested that metabolic transients in oxidative phosphorylation were also present.To investigate the mitochondrial component of the oscillations, the present study uses the endogenous fluorescence of flavoproteins to image redox oscillations in the mitochondrial matrix. Subcellular imaging reveals a remarkable degree of spatial and temporal heterogeneity in mitochondrial redox and electrical potential, including local transients and propagated metabolic waves. The last observation suggests that communication between mitochondria in the same or neighboring cells is likely to involve diffusible cytosolic messengers acting to synchronize the energy state of the entire population of mitochondria.

Biological chemistry of reactive oxygen and nitrogen and radiation-induced signal transduction mechanisms

“…Additional evidence for this proposal came from measuring radiation-induced mitochondrial membrane depolarization and enhanced permeability of the mitochondrial membrane to the fluorescent small molecule, calcein. This model fits with previous findings from other investigators demonstrating such propagating signals within a cell's mitochondrial pool and as a mechanism involved in Ca 2 þ homeostasis and cellular redox modulation (Bernardi and Petronelli, 1996;Vercesi et al, 1997;Ichas and Mazat, 1998;Romashko et al, 1998;Zorov et al, 2000). Specifically, Zorov et al (2000) proposed that ROSinduced ROS release accompanies the mitochondrial permeability transition and provides a 'self-amplifying' mechanism by which redox signals can be transmitted throughout the cell.…”

“…This too entails a two-step process with an initial nonspecific oxidative event triggering the second but specific redox sensitive step involving propagation of a reversible mitochondrial permeability transition from one mitochondrion to adjacent mitochondria (e.g. Romashko et al, 1998;Zorov et al, 2000;Leach et al, 2001Leach et al, , 2002. This propagation ultimately results in the Ca 2 þ -dependent activation of constitutive NOS and NO signaling (Leach et al, 2002).…”

Mitochondrial Ion Channels in Cardiac Function and Dysfunction

“…Modulation of the cellular action potential by these metabolic oscillations led us to hypothesize that these oscillations could result in arrhythmias if present in the heart after ischaemia-reperfusion. Subsequently, we identified the mitochondria as the source of the oscillations and observed they involved the synchronized depolarization of the mitochondrial network of almost the entire heart cell (Romashko et al 1998). Based on the observation that ΔΨ m depolarization could either occur in individual mitochondria, clusters of mitochondria, or in the whole network, we suggested that the mitochondria may represent a network of coupled oscillators (Romashko et al 1998).…”

Percolation and criticality in a mitochondrial network

“…Although the transition in the mitochondrial spanning cluster appears abrupt, the phase transition should be slow enough to be resolved as waves of depolarization with faster image acquisition and͞or interventions that slow propagation, as described for spontaneous metabolic oscillations (9). Indeed, at higher temporal resolution, depolarization waves could be detected with a speed of 22 m⅐s Ϫ1 (Fig.…”

“…At the subcellular level, adapting energy production to meet a variable metabolic demand depends on the coordinated action of thousands of mitochondria in the cardiac myocyte, which are distributed between myofilaments in a lattice-like network of nonlinear elements. This arrangement is conducive to spatiotemporal patterns of self-organization, for instance, oscillations and͞or propagating waves (9,10). However, it is not known how individual mitochondria interact with their neighbors or how metabolic signals are communicated over cellular distances.…”