Effect of the mitochondrial membrane potential on the absorbance of the reduced form of cytochrome c oxidase.

The effect of mitochondrial membrane potential (ΔΨ) on the absorbance of the reduced cytochrome c oxidase (COX) was evaluated in isolated rabbit heart mitochondria using integrating sphere optical spectroscopy. Maximal reduction of the mitochondrial cytochromes was achieved by either blowing nitrogen to remove oxygen, or by adding cyanide. Gradual depolarization of ΔΨ by adding increasing concentrations of uncoupler resulted in an increase of up to 50 % in the absorbance of cytochrome aa under nitrogen saturation, and of 25 % with cyanide.

Lactate oxidation in Paracoccus denitrificans.

Paracoccus denitrificans has a classical cytochrome-dependent electron transport chain and two alternative oxidases. The classical transport chain is very similar to that in eukaryotic mitochondria. Thus, P.

Cardiac nitric oxide scavenging: role of myoglobin and mitochondria.

Vascular production of nitric oxide (NO) regulates vascular tone. However, highly permeable NO entering the cardiomyocyte would profoundly impact metabolism and signalling without scavenging mechanisms. The purpose of this study was to establish mechanisms of cardiac NO scavenging.

Increased mitochondrial free Ca during ischemia is suppressed, but not eliminated by, germline deletion of the mitochondrial Ca uniporter.

Mitochondrial Ca overload is proposed to regulate cell death via opening of the mitochondrial permeability transition pore. It is hypothesized that inhibition of the mitochondrial Ca uniporter (MCU) will prevent Ca accumulation during ischemia/reperfusion and thereby reduce cell death. To address this, we evaluate mitochondrial Ca in ex-vivo-perfused hearts from germline MCU-knockout (KO) and wild-type (WT) mice using transmural spectroscopy.

Spectroscopic identification of the catalytic intermediates of cytochrome c oxidase in respiring heart mitochondria.

The catalytic cycle of cytochrome c oxidase (COX) couples the reduction of oxygen to the translocation of protons across the inner mitochondrial membrane and involves several intermediate states of the heme a-Cu binuclear center with distinct absorbance properties. The absorbance maximum close to 605 nm observed during respiration is commonly assigned to the fully reduced species of hemes a or a (R). However, by analyzing the absorbance of isolated enzyme and mitochondria in the Soret (420-450 nm), alpha (560-630 nm) and red (630-700 nm) spectral regions, we demonstrate that the Peroxy (P) and Ferryl (F) intermediates of the binuclear center are observed during respiration, while the R form is only detectable under nearly anoxic conditions in which electrons also accumulate in the higher extinction coefficient low spin a heme.

Multiomics analyses reveal early metabolic imbalance and mitochondrial stress in neonatal photoreceptors leading to cell death in Pde6brd1/rd1 mouse model of retinal degeneration.

Retinal diseases exhibit extensive genetic heterogeneity and complex etiology with varying onset and severity. Mutations in over 200 genes can lead to photoreceptor dysfunction and/or cell death in retinal neurodegeneration. To deduce molecular pathways that initiate and/or drive cell death, we adopted a temporal multiomics approach and examined molecular and cellular events in newborn and developing photoreceptors before the onset of degeneration in a widely-used Pde6brd1/rd1 (rd1) mouse, a model of autosomal recessive retinitis pigmentosa caused by PDE6B mutations.

Metabolic design in a mammalian model of extreme metabolism, the North American least shrew (Cryptotis parva).

Mitochondrial adaptations are fundamental to differentiated function and energetic homeostasis in mammalian cells. But the mechanisms that underlie these relationships remain poorly understood. Here, we investigated organ-specific mitochondrial morphology, connectivity and protein composition in a model of extreme mammalian metabolism, the least shrew (Cryptotis parva).

Energy homeostasis is a conserved process: Evidence from Paracoccus denitrificans' response to acute changes in energy demand.

Paracoccus denitrificans is a model organism for the study of oxidative phosphorylation. We demonstrate a very high respiratory capacity compared to mitochondria when normalizing to cytochrome aa3 content even in the absence of alternative terminal oxidases. To gain insight into conserved mechanisms of energy homeostasis, we characterized the metabolic response to K+ reintroduction.

Monitoring mitochondrial calcium and metabolism in the beating MCU-KO heart.

Optical methods for measuring intracellular ions including Ca revolutionized our understanding of signal transduction. However, these methods are not extensively applied to intact organs due to issues including inner filter effects, motion, and available probes. Mitochondrial Ca is postulated to regulate cell energetics and death pathways that are best studied in an intact organ.

Assessment of Lung Structure and Regional Function Using 0.55 T MRI in Patients With Lymphangioleiomyomatosis.

Objectives: Contemporary lower-field magnetic resonance imaging (MRI) may offer advantages for lung imaging by virtue of the improved field homogeneity. The aim of this study was to evaluate the utility of lower-field MRI for combined morphologic imaging and regional lung function assessment. We evaluate low-field MRI in patients with lymphangioleiomyomatosis (LAM), a rare lung disease associated with parenchymal cysts and respiratory failure.

Oxygen-enhanced functional lung imaging using a contemporary 0.55 T MRI system.

The purpose of this study was to evaluate oxygen-enhanced pulmonary imaging at 0.55 T with 3D stack-of-spirals ultrashort-T (UTE) acquisition. Oxygen-enhanced pulmonary MRI offers the measurement of regional lung ventilation and perfusion using inhaled oxygen as a contrast agent.

Energy metabolism design of the striated muscle cell.

The design of the energy metabolism system in striated muscle remains a major area of investigation. Here, we review our current understanding and emerging hypotheses regarding the metabolic support of muscle contraction. Maintenance of ATP free energy, so called energy homeostasis, via mitochondrial oxidative phosphorylation is critical to sustained contractile activity, and this major design criterion is the focus of this review.

How hot are single cells?

Large changes in energy metabolism are associated with minimal alterations in surface temperature of isolated mammalian cells.

Mitochondrial DNA segregation and replication restrict the transmission of detrimental mutation.

Although mitochondrial DNA (mtDNA) is prone to accumulate mutations and lacks conventional DNA repair mechanisms, deleterious mutations are exceedingly rare. How the transmission of detrimental mtDNA mutations is restricted through the maternal lineage is debated. Here, we demonstrate that mitochondrial fission, together with the lack of mtDNA replication, segregate mtDNA into individual organelles in the Drosophila early germarium.

Opportunities in Interventional and Diagnostic Imaging by Using High-Performance Low-Field-Strength MRI.

Background Commercial low-field-strength MRI systems are generally not equipped with state-of-the-art MRI hardware, and are not suitable for demanding imaging techniques. An MRI system was developed that combines low field strength (0.55 T) with high-performance imaging technology.

Perfused murine heart optical transmission spectroscopy using optical catheter and integrating sphere: Effects of ischemia/reperfusion.

Tissue transmission optical absorption spectroscopy provides dynamic information on metabolism and function. Murine genetic malleability makes it a major model for heart research. The diminutive size of the mouse heart makes optical transmission studies challenging.

Paradoxical arteriole constriction compromises cytosolic and mitochondrial oxygen delivery in the isolated saline-perfused heart.

The isolated saline-perfused heart is used extensively to study cardiac physiology. Previous isolated heart studies have demonstrated lower tissue oxygenation compared with in vivo hearts based on myoglobin oxygenation and the mitochondrial redox state. These data, consistent with small anoxic regions, suggest that the homeostatic balance between work and oxygen delivery is impaired.

The interactome of intact mitochondria by cross-linking mass spectrometry provides evidence for coexisting respiratory supercomplexes.

Mitochondria exert an immense amount of cytophysiological functions, but the structural basis of most of these processes is still poorly understood. Here we use cross-linking mass spectrometry to probe the organization of proteins in native mouse heart mitochondria. Our approach provides the largest survey of mitochondrial protein interactions reported so far.

Cardiac performance is limited by oxygen delivery to the mitochondria in the crystalloid-perfused working heart.

The left ventricular working, crystalloid-perfused heart is used extensively to evaluate basic cardiac function, pathophysiology, and pharmacology. Crystalloid-perfused hearts may be limited by oxygen delivery, as adding oxygen carriers increases myoglobin oxygenation and improves myocardial function. However, whether decreased myoglobin oxygen saturation impacts oxidative phosphorylation (OxPhos) is unresolved, since myoglobin has a much lower affinity for oxygen than cytochrome c oxidase (COX).

Intracardiac light catheter for rapid scanning transmural absorbance spectroscopy of perfused myocardium: measurement of myoglobin oxygenation and mitochondria redox state.

Absorbance spectroscopy of intrinsic cardiac chromophores provides nondestructive assessment of cytosolic oxygenation and mitochondria redox state. Isolated perfused heart spectroscopy is usually conducted by collecting reflected light from the heart surface, which represents a combination of surface scattering events and light that traversed portions of the myocardium. Reflectance spectroscopy with complex surface scattering effects in the beating heart leads to difficulty in quantitating chromophore absorbance.

Robust fluorescent labelling of micropipettes for use in fluorescence microscopy: application to the observation of a mosquito borne parasite infection.

The ability to monitor micropipette injections with a high-resolution fluorescent microscope has utility for a variety of applications. Herein, different approaches were tested for creating broad-band fluorescently labelled glass micropipettes including: UV cured glass glues, baked glass enamel containing fluorescent dyes as well as nanodiamonds attached during pipette formation in the microforge. The most robust and simplest approach was to use labelled baked enamel on the exterior of the pipette.

Cytochrome c Oxidase Activity Is a Metabolic Checkpoint that Regulates Cell Fate Decisions During T Cell Activation and Differentiation.

T cells undergo metabolic reprogramming with major changes in cellular energy metabolism during activation. In patients with mitochondrial disease, clinical data were marked by frequent infections and immunodeficiency, prompting us to explore the consequences of oxidative phosphorylation dysfunction in T cells. Since cytochrome c oxidase (COX) is a critical regulator of OXPHOS, we created a mouse model with isolated dysfunction in T cells by targeting a gene, COX10, that produces mitochondrial disease in humans.

Power Grid Protection of the Muscle Mitochondrial Reticulum.

Mitochondrial network connectivity enables rapid communication and distribution of potential energy throughout the cell. However, this connectivity puts the energy conversion system at risk, because damaged elements could jeopardize the entire network. Here, we demonstrate the mechanisms for mitochondrial network protection in heart and skeletal muscle (SKM).