Sink Hypothesis Research Articles

Blood‐brain barrier permeable antibodies for Alzheimer’s potential therapeutic and diagnostic applications

AbstractBackgroundAlzheimer’s disease (AD), the most common cause of dementia, is a slowly progressing neurodegenerative disorder that has become an escalating global epidemic with an unbearable emotional and physical strain on patients, loved ones, and their caretakers. So far, there is no cure or treatment to alter its progression or improve cognition. The reasons for such a failure to develop disease modifying therapy for AD can probably be best described, in our opinion, as to the lack of understanding of what the real disease trigger is and the inability of most of the potentially disease modifying agents to cross the blood‐brain barrier ((BBB) in therapeutically effective doses. Mouse monoclonal antibodies (mAbs) against amyloid‐beta (Aβ) and Tau proteins have been employed to normalize the levels of these proteins in the brain despite their low (0.1%) reported brain uptake. Though Aβ levels have been reduced, either through direct target engagement in the brain or most likely due to the so called “Sink Hypothesis” in multiple clinical trials, cognition improvement has not been observed so far. It appears that memory deficit may be related to early events such as mitochondria and lysosomal dysfunction, axonal demyelination, and neuroinflammation, etc. that take place long before symptoms and protein aggregation are observed.MethodTo demonstrate BBB permeability proof‐of‐principle we developed heavy‐chain only antibody for amyloid‐beta (1‐42) by immunizing animals with a synthetic antigen. Blood from the immunized animals was collected and processed using standard published methods to obtain heavy‐chain only antibody, referred to as the SMART* Molecule (SM). [SMART* is an acronym that stands for Specific Molecular Architecture for Recognition and Therapy].ResultLow dose (1.5 mg/Kg) tail‐vein injection of Aβ‐SM in transgenic AD mice exquisitely decorated amyloid‐plaque as shown by tissue histology of brain slices demonstrating potential diagnostic application of Aβ‐SM. One weekly tail‐vein injection with the same low dose for 12 weeks in AD transgenic mice cleared amyloid plaque, oligomeric and fibrillar, by about 60% compared to the untreated mice, thus demonstrating therapeutic potential of Aβ‐SM.ConclusionSMART Molecules, heavy‐chain only antibodies, appear to have therapeutic and diagnostic potential for neurodegenerative diseases of the central nervous system.

Interactions of intrinsically disordered proteins with the unconventional chaperone human serum albumin: From mechanisms of amyloid inhibition to therapeutic opportunities.

Human Serum Albumin (HSA), the most abundant protein in plasma, serves a diverse repertoire of biological functions including regulation of oncotic pressure and redox potential, transport of serum solutes, but also chaperoning of misfolded proteins. Here we review how HSA interacts with a wide spectrum of client proteins including intrinsically disordered proteins (IDPs) such as Aβ, the islet amyloid peptide (IAPP), alpha synuclein and stressed globular proteins such as insulin. The comparative analysis of the HSA chaperone - client interactions reveals that the amyloid-inhibitory function of HSA arises from at least four emerging mechanisms. Two mechanisms (the monomer stabilizer model and the monomer competitor model) involve the direct binding of HSA to either IDP monomers or oligomers, while other mechanisms (metal chelation and membrane protection) rely on the indirect modulation by HSA of other factors that drive IDP aggregation. While HSA is not the only extracellular chaperone, given its abundance, HSA is likely to account for a significant fraction of the chaperoning effects in plasma, thus opening new therapeutic opportunities in the context of the peripheral sink hypothesis.

The Role of Circulating Aβ Seeds in the Progression of Cerebral Amyloidosis.

While understudied, it is suspected that peripheral Aβ peptides affect Alzheimer’s disease (AD)-associated pathological changes in the brain. The peripheral sink hypothesis postulates that the central and peripheral pools of Aβ co-exist in equilibrium. As such, cerebral amyloid levels may be modulated by intervening circulating Aβ. In this commentary, we discuss relevant literature supporting the potential role of peripheral Aβ in exacerbating brain amyloidosis in both humans and mouse models of AD. Moreover, we highlight the need to further understand the mechanisms by which circulating Aβ peptides may reach the brain and contribute to neuropathology. Finally, we discuss the implications of targeting peripheral Aβ as a therapeutic approach in treating AD.

Discerning Bulk and Interfacial Polarons in a Dual Electron Donor/Acceptor Polymer.

The active layer of organic solar cells typically possesses a complex morphology, with amorphous donor/acceptor mixed domains present in addition to purer, more crystalline domains. These crystalline domains may represent an energy sink for free charges that aids charge separation and suppresses bimolecular recombination. The first step in exploiting this behavior is the identification and characterization of charges located in these different domains. Herein, the generation and recombination of both bulk and interfacial polarons are demonstrated in the dual electron donor/acceptor polymer XIND using transient absorption spectroscopy. The absorption spectra of XIND bulk polarons, present in pristine polymer domains, are clearly distinguishable from those of polarons present at the donor/acceptor interface. Furthermore, it is shown that photogenerated polarons are transferred from the interface to the bulk. These findings support the energy sink hypothesis and offer a way to maximize morphology relationships to enhance charge generation and suppress recombination.

The sink or SWIM hypothesis; a roadmap from pathology to work instability

Purpose: Selecting the most appropriate health-related work outcome to evaluate an intervention can be fraught with difficulty. To aid clinicians in navigating this problem we have developed a model, which illustrates how pathology can affect specific measureable quantities, such as work instability.Methods: Using a modified-Delphi procedure, a panel of experts met initially to analyze the content of 95 health-related work outcome measures and organize the identified areas of measurement into a coherent model, complemented by a narrative review of the literature. This initial model underwent two rounds of stakeholder-based feedback, the results of which were incorporated in the final expert panel meeting to produce the States-traits Work Instability Model (SWIM).Results: The States-traits Work Instability Model (SWIM) illustrates how changes to an individual’s physical and psychological states and traits might affect their work-related performance, well-being and self-efficacy. Moreover, each concept utilized in the model was specifically selected as it represents a measurable quantity, for which there are tools available.Conclusion: The SWIM is arguably the first holistic model of work that is based on both the clinical realities of vocational rehabilitation, sociological research, and is born from analyzing the basis of practical measurements.Implications for RehabilitationWork Instability • Work instability has multiple causes many of which are amenable to intervention • The model clarifies the measureable domains of vocational rehabilitation interventions, which is of particular benefit for services working with people with disability at work who are struggling to remain in work • The model conceptualizes how the potential areas for intervention may be related based on evidence available in the literature.

Deep Soil Conditions Make Mediterranean Cork Oak Stem Growth Vulnerable to Autumnal Rainfall Decline in Tunisia

Tree rings provide fruitful information on climate features driving annual forest growth through statistical correlations between annual tree growth and climate features. Indices built upon tree growth limitation by carbon sequestration (source hypothesis) or drought-driven cambial phenology (sink hypothesis) can be used to better identify underlying processes. We used both analytical frameworks on Quercus suber, a sparsely studied species due to tree ring methodological issues, and growing on a favorable sub-humid Mediterranean climate and deep soil conditions in Tunisia (North Africa). Statistical analysis revealed the major role of autumnal rainfall before the growing season on annual tree growth over the 1918–2008 time series. Using a water budget model, we were able to explain the critical role of the deep soil water refill during the wet season in affecting both the drought onset controlling growth phenology and the summer drought intensity affecting carbon assimilation. Analysis of recent climate changes in the region additionally illustrated an increase in temperatures enhancing the evaporative demand and advancing growth start, and a decline in rainfalls in autumn, two key variables driving stem growth. We concluded on the benefits of using process-based indices in dendrochronological analysis and identified the main vulnerability of this Mediterranean forest to autumnal rainfall decline, a peculiar aspect of climate change under summer-dry climates.

IS THE PERIPHERAL SINK HYPOTHESIS PHYSIOLOGICALLY FEASIBLE? EVIDENCE FROM MODEL-BASED ASSESSMENT OF THE AMYLOID PATHWAY

IS THE PERIPHERAL SINK HYPOTHESIS PHYSIOLOGICALLY FEASIBLE? EVIDENCE FROM MODEL-BASED ASSESSMENT OF THE AMYLOID PATHWAY

Detecting tau in serum of transgenic animal models after tau immunotherapy treatment

In the attempt to elucidate if the “peripheral sink hypothesis” could be a potential mechanism of action for tau removal in passive immunotherapy experiments, we have examined tau levels in serum of chronically injected JNPL3 and Tg4510 transgenic animals. Measurement of tau in serum of mice treated with tau antibodies is challenging because of the antibody interference in sandwich enzyme-linked immunosorbent assays. To address this issue, we have developed a heat-treatment protocol at acidic pH to remove interfering molecules from serum, with excellent recovery of tau. The present data show that pan-tau and conformational antibodies do increase tau in mouse sera. However, these concentrations in serum do not consistently correlate with reductions of tau pathology in brain, suggesting that large elevations of tau species measured in serum are not predictive of efficacy. Here, we describe a reliable method to detect tau in serum of transgenic animals that have undergone tau immunotherapy. Levels of tau in human serum are less than the sensitivity of current assays, although artifactual signals are common. The method may be useful in similarly treated humans, a situation in which false positive signals are likely.

Local anaesthetic systemic toxicity

Access the CPD MCQs for this issue online here ### Key points Well-placed local anaesthetics (LAs) can yield great clinical benefits. But systemic toxicity related to their use can be devastating. The highly publicized case in 2004 of Mayra Cabrera, a theatre nurse who died shortly after delivery of her baby boy when her epidural infusion of bupivacaine was mistakenly connected to her i.v. line, reminds us to be vigilant and to learn from such rare events. While prevention is clearly the most important element in avoiding morbidity and mortality associated with local anaesthetic systemic toxicity (LAST), such cases still occur despite best practice. Knowing how to manage these uncommon events is vital. In this review, we cover LA toxicity, its incidence, clinical features, risk factors, prevention, and management and examine i.v. lipid emulsion (ILE) therapy in more detail. LAST has been recognized for more than a hundred years, but the precise incidence is currently unknown. In 1928, the American Medical Association reported 40 deaths attributable to LAs.1 Cocaine was responsible for half of these deaths, but procaine was also implicated. These findings prompted the search for less toxic agents. Lidocaine, first synthesized in 1944, was the first amide LA to be used clinically. However, in 1979, the potentially fatal toxicity of amide LAs was highlighted by Albright.2 More recently, studies from 1993 …

Survival following massive amitriptyline overdose: The use of intravenous lipid emulsion therapy and the occurrence of acute respiratory distress

While survival has been reported after ingestion of 4.25 g amitriptyline, for which intravenous lipid emulsion (ILE) therapy was administered, we recently managed a patient who made a full recovery after a 4.8 g amitriptyline overdose. Interesting features of the case were a marked response to ILE and the subsequent development of acute respiratory distress syndrome (ARDS). Our patient had taken 96 amitriptyline 50mg tablets (total 4.8 g or 80mg/kg), an uncertain amount of zopiclone, alcohol and venlafaxine. In the emergency department the patient was hypotensive and unresponsive (GCS 3), with a prolonged QTc identified on ECG. Rapid sequence induction was performed followed by transfer to the intensive care unit for full supportive therapy. Further management and key parameters are illustrated in Figure 1. Hypotension was unresponsive to fluid therapy, and noradrenaline therapy was required. Intravenous bicarbonate therapy and mild hyperventilation (with tidal volumes 6ml/kg) were utilised, aiming to achieve a pH of 7.55. Three grand mal seizures occurred which were treated with a midazolam bolus followed by an infusion. Approximately 12 h after the drug ingestion, with escalating noradrenaline requirements, ILE was administered. The subsequent improvement in blood pressure and reduction in noradrenaline can be seen in Figure 1. Case reports detailing the successful use of ILE for the treatment of toxicity of lipophilic drugs including tricyclic antidepressants are well documented. A recent prospective, randomised, controlled trial found no benefit of ILE compared to bicarbonate in a pig model of amitriptyline toxicity. A rabbit model found conflicting results. Various mechanisms have been suggested for the mechanism of action of ILE, but the lipid sink hypothesis describes how ILE may provide a large binding site for the drug, removing it from effective circulation. Given the pKa of amitriptyline of 9.4, prior alkalinisation, as was undertaken in this case, might be expected to augment the effect of ILE by decreasing the ionised portion of the drug. On day 3 of their care, our patient developed criteria of ARDS with a PaO2/FiO2 ratio of 70mmHg by day 5. Proning was required, tracheostomy was performed and a prolonged respiratory wean ensued with 40 days of invasive ventilation. There was no evidence of pulmonary aspiration at the scene of the overdose, during intubation or on the initial chest radiograph, though micro-aspiration cannot be excluded. Ultimately, Streptococcus pneumonia and mixed coliforms were grown from bronchoalveolar lavage of the patient’s lungs; however, concern was raised that ILE could have been linked to the ARDS. The patients subsequently made a full physical recovery. Acute respiratory distress syndrome has been reported in three patients in a case series of nine patients treated with ILE but, as the authors acknowledge, critically ill patients are at risk of ARDS. Furthermore, ARDS has been reported as a complication of tricyclic overdose. Nonetheless, case series into the use of lipid infusion in ARDS and septicaemia have linked it to worsening oxygenation. We believe this to be one of the highest reported overdoses of amitriptyline that has been survived and that the use of ILE contributed to this outcome. Our experience demonstrates the potential use of ILE outside the setting of cardiac arrest or cardiovascular collapse but highlights concerns regarding its potential association with ARDS.

Revisiting the peripheral sink hypothesis: inhibiting BACE1 activity in the periphery does not alter β-amyloid levels in the CNS.

Aggregation of amyloid beta (Aβ) peptides and the subsequent neural plaque formation is a central aspect of Alzheimer's disease. Various strategies to reduce Aβ load in the brain are therefore intensely pursued. It has been hypothesized that reducing Aβ peptides in the periphery, that is in organs outside the brain, would be a way to diminish Aβ levels and plaque load in the brain. In this report, we put this peripheral sink hypothesis to test by investigating how selective inhibition of Aβ production in the periphery using a β-secretase (BACE)1 inhibitor or reduced BACE1 gene dosage affects Aβ load in the brain. Selective inhibition of peripheral BACE1 activity in wild-type mice or mice over-expressing amyloid precursor protein (APPswe transgenic mice; Tg2576) reduced Aβ levels in the periphery but not in the brain, not even after chronic treatment over several months. In contrast, a BACE1 inhibitor with improved brain disposition reduced Aβ levels in both brain and periphery already after acute dosing. Mice heterozygous for BACE1, displayed a 62% reduction in plasma Aβ40, whereas brain Aβ40 was only lowered by 11%. These data suggest that reduction of Aβ in the periphery is not sufficient to reduce brain Aβ levels and that BACE1 is not the rate-limiting enzyme for Aβ processing in the brain. This provides evidence against the peripheral sink hypothesis and suggests that a decrease in Aβ via BACE1 inhibition would need to be carried out in the brain. Aggregation of amyloid beta (Aβ) peptides in the brain is a central aspect of Alzheimer's disease. In this study, we demonstrate that inhibition of Aβ formation by BACE1 inhibitors needs to be carried out in the brain and that reduction of Aβ in the periphery is not sufficient to reduce brain Aβ levels. This information is useful for developing future Aβ-targeting therapies for Alzheimer's disease.

Membrane protein stability as the energy sink for sorting in the periplasm (224.3)

We will present novel measurements of folding free energies for the transmembrane proteins PagP and OmpW from E. coli. Our data were collected in the same lipid bilayer and buffer system we previously employed to determine those parameters for E. coli OmpLA. From a biophysical perspective, our results suggest that the stabilities of these proteins may be strongly correlated to the water‐to‐bilayer transfer free energy of the lipid‐facing residues in their transmembrane regions. From a biological perspective, our findings suggest that the folding free energies for these membrane proteins may be the thermodynamic sink that establishes an energy gradient across the periplasm. This potential thus drives the sorting of outer membrane proteins by chaperones to the outer membranes in living bacteria. Binding free energies of these outer membrane proteins with periplasmic chaperones support this energy sink hypothesis.

Primate study deals a fresh blow to the peripheral sink hypothesis

Primate study deals a fresh blow to the peripheral sink hypothesis

Sustained peripheral depletion of amyloid-β with a novel form of neprilysin does not affect central levels of amyloid-β

Alzheimer's disease is characterized by the accumulation of amyloid deposits in the brain and the progressive loss of cognitive functions. Although the precise role of amyloid-β in disease progression remains somewhat controversial, many efforts to halt or reverse disease progression have focussed on reducing its synthesis or enhancing its removal. It is believed that brain and peripheral soluble amyloid-β are in equilibrium and it has previously been hypothesized that a reduction in peripheral amyloid-β can lower brain amyloid-β, thereby reducing formation of plaques predominantly composed of insoluble amyloid-β; the so-called peripheral sink hypothesis. Here we describe the use of an amyloid-β degrading enzyme, the endogenous metallopeptidase neprilysin, which is fused to albumin to extend plasma half-life and has been engineered to confer increased amyloid-β degradation activity. We used this molecule to investigate the effect of degradation of peripheral amyloid-β on amyloid-β levels in the brain and cerebrospinal fluid after repeated intravenous dosing for up to 4 months in Tg2576 transgenic mice, and 1 month in rats and monkeys. This molecule proved highly effective at degradation of amyloid-β in the periphery but did not alter brain or cerebrospinal fluid amyloid-β levels, suggesting that the peripheral sink hypothesis is not valid and is the first time that this has been demonstrated in non-human primates.

The influence of environmental factors on the abundance and recruitment of the sand crab Emerita analoga (Stimpson 1857): Source–sink dynamics?

The sandcrab Emerita analoga is the dominant species inhabiting sandy beaches along the Pacific coast of the American continent. In our study, 10 sandy beaches were sampled seasonally from 2006 to 2011, including coastal planktonic sampling from 2006 to 2008. Two major population cores were detected, the first one in the northern part of the study area and the second in the area immediately to the south of the Itata River mouth. Zoeal stages were found along the entire coastal zone. Highest densities and recruitment were found during spring and summer of each year. PLS regression indicated that source–sink habitat proxies correlated positively with morphodynamic parameters; while beach slope and total organic matter were negatively correlated. These results agree with the source–sink hypothesis, finding higher densities of adults, recruits and cohort recurrence on open coast beaches with milder physical dynamics. Furthermore, a hypoxic event and a mega-earthquake/tsunami negatively affected recruitment at the inter-annual scale.

Membrane protein thermodynamic stability may serve as the energy sink for sorting in the periplasm

Thermodynamic stabilities are pivotal for understanding structure-function relationships of proteins, and yet such determinations are rare for membrane proteins. Moreover, the few measurements that are available have been conducted under very different experimental conditions, which compromises a straightforward extraction of physical principles underlying stability differences. Here, we have overcome this obstacle and provided structure-stability comparisons for multiple membrane proteins. This was enabled by measurements of the free energies of folding and the m values for the transmembrane proteins PhoP/PhoQ-activated gene product (PagP) and outer membrane protein W (OmpW) from Escherichia coli. Our data were collected in the same lipid bilayer and buffer system we previously used to determine those parameters for E. coli outer membrane phospholipase A (OmpLA). Biophysically, our results suggest that the stabilities of these proteins are strongly correlated to the water-to-bilayer transfer free energy of the lipid-facing residues in their transmembrane regions. We further discovered that the sensitivities of these membrane proteins to chemical denaturation, as judged by their m values, was consistent with that previously observed for water-soluble proteins having comparable differences in solvent exposure between their folded and unfolded states. From a biological perspective, our findings suggest that the folding free energies for these membrane proteins may be the thermodynamic sink that establishes an energy gradient across the periplasm, thus driving their sorting by chaperones to the outer membranes in living bacteria. Binding free energies of these outer membrane proteins with periplasmic chaperones support this energy sink hypothesis.

Stem galls drain nutrients and decrease shoot performance in Diplusodon orbicularis (Lythraceae)

Insect galls are important nutrient sinks in the plant, ultimately affecting its reproductive success. We assessed the influence of a stem galler on the survival of plant shoots and whether these are able to concentrate nutrients in the gall’s tissues; thus testing the nutrient sink hypothesis. We measured gall sizes and nutrient concentrations in a Brazilian Cerrado plant, Diplusodon orbicularis, and its Cecidomyiidae stem galler. Galls grew larger on thicker shoots. Also, these shoots suffered smaller mortality from gall attack, while thinner shoots were commonly associated with the death of the shoot distal to the gall. Apical shoots suffered higher mortality, while basal shoots suffered lower mortality and were proportionally less attacked. Galled tissues were more nutrient rich than non-galled tissues. The nutrients P, Ca, and Mg were more concentrated inside the galls when compared to tissues in non-galled stems. Gallers also seem to be able to reduce toxic Al concentration in the plant tissues, as Al occurred in smaller concentrations inside the galls than out of them. Although stem gallers feed on tissues of low nutritional value, these are in a favourable position to intercept flowing nutrients and water. The death of small galled shoots possibly is due to the lack of essential nutrients and energy drained, or water restriction in them.

CXCL5 regulates chemokine scavenging and pulmonary host defense to bacterial infection.

The chemokine sink hypothesis pertaining to erythrocyte Duffy Antigen Receptor for Chemokines (DARC) during inflammation has received considerable attention, but lacks direct in vivo evidence. Here we demonstrate, using mice with a targeted deletion in CXCL5, that CXCL5 bound erythrocyte DARC and impaired its chemokine scavenging in blood. CXCL5 increased the plasma concentrations of CXCL1 and CXCL2 in part through inhibiting chemokine scavenging, impairing chemokine gradients and desensitizing CXCR2, which led to decreased neutrophil influx to the lung, increased lung bacterial burden and mortality in an Escherichia coli pneumonia model. In contrast, CXCL5 exerted a predominant role in mediating neutrophil influx to the lung during inflammation after LPS inhalation. Platelets and lung resident cells were the sources of homeostatic CXCL5 in blood and inflammatory CXCL5 in the lung respectively. This study presents a paradigm whereby platelets and red cells alter chemokine scavenging and neutrophil-chemokine interaction during inflammation.

Intravenous immunoglobulin (IVIg) Gammagard liquid contains anti-RAGE IgG and sLRP

Baxter's IVIg preparation Gammagard Liquid (GGL) has shown encouraging results in a phase II trial treating patients with mild Alzheimer's disease (AD) [Relkin et al, Neurobiol. Aging, 2008]. GGL, which is manufactured from large pools of human plasma donations, has been shown to contain amyloid-β conformer-specific IgG autoantibodies that could be involved in the beneficial effects observed in the phase II study [O'Nuallain et al, Biochemistry, 2008]. Here we asked the question whether GGL contains other antibodies and soluble proteins that can modulate the amyloid-β pathway apart from these naturally occurring anti-amyloid-β IgGs. In particular, we checked for the presence of antibodies against the receptor for advanced glycation end products (RAGE), a receptor involved in the amyloid-β influx from plasma into the brain, and for the presence of soluble low density lipoprotein receptor-related protein (sLRP), a high-affinity Aβ binding protein that can modulate amyloid-β efflux from the brain into plasma. We investigated 20 lots of GGL. We checked for the presence of anti-RAGE IgG by a direct ELISA using a synthetic peptide. For the sLRP measurement, we used plate-immobilized recombinant receptor-associated protein (RAP) to capture sLRP from the sample and detected the bound sLRP with a monoclonal anti-LRP and an anti-mouse IgG peroxidase [Quinn et al, J. Biol. Chem., 1997]. Furthermore, we ran inhibition studies for checking the specificity of our assays. We detected naturally occurring anti-RAGE IgG in all GGL lots investigated and measured sLRP levels in the μg range (up to 40 μg/mL) with only moderate lot-to-lot variations among the different GGL lots. Inhibition with the RAGE peptide and RAP proved both assays to be specific. Anti-RAGE antibodies present in GGL could block RAGE and inhibit the uptake of amyloid-β peptide by the brain, whereas sLRP also present in GGL could lower the amyloid-β burden of the brain consistent with the peripheral sink hypothesis. These findings that GGL contains anti-RAGE IgG and sLRP broaden the possible mode of action of GGL in AD beyond the effects of anti-amyloid-β antibodies.

Effects Of Mitochondrial Depolarization On Cardiac Electrical Activity In An Integrated Multiscale Model Of The Myocardium

Metabolic or oxidative stress can trigger the abrupt collapse or oscillation of mitochondrial membrane potential, which activates KATP current and alters the cardiac action potential. This mechanism can introduce both temporal and spatial dispersion of electrical excitability in the form of a “metabolic sink”, leading to heterogeneous conduction in the tissue, reentrant tachyarrhythmias, or fibrillation. To quantitatively study this mechanism, a 2D monodomain model of the myocardium (5×5cm2; 200 micron resolution) was developed, comprised of 63,000 nodes, each representing an integrated cellular model of cardiac excitation-contraction coupling, mitochondrial energetics, and ROS-induced ROS release (ECME-RIRR). Oxidative stress was initiated in a central circular zone of the tissue by increasing the fractional mitochondrial ROS production (shunt) during oxidative phosphorylation from 2% to 10%. Model simulations show that mitochondrial dynamics bifurcate during stimulation at 1 Hz and complete depolarization of αΨm ensues in the sink zone. Within the metabolic sink, sarcolemmal KATP currents increase, action potentials dramatically shorten, and the refractory period is abbreviated. These effects are enhanced by increasing the KATP density. In addition, fast and irregular electric activity (ventricular fibrillation) in the electrically paced tissue is observed when an S2 stimulus is introduced within or near the border of the metabolic sink, but only when the sink exceeds a critical size (r>0.4cm). Phase singularity analysis indicates that the fibrillatory activity is initiated at sites close the border zone. The results emphasize the power of integrating cellular electrophysiological, Ca2+ handling, and metabolic subsystems into a multiscale model to simulate emergent macroscopic phenomena in the heart. Moreover, the results provide a proof-of-concept of the metabolic sink hypothesis and a new tool to study its role in arrhythmogenesis and sudden cardiac death.