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{{MITOEAGLE}}
{{MITOEAGLE}}
[[File:OXPHOS-coupled energy cycles.jpg|right|300px|thumb|OXPHOS-coupled energy cycles. From [[Gnaiger_2014_MitoPathways#Chapter_1._Real-time_OXPHOS_analysis |Gnaiger 2014 MitoPathways]].]]
::: <big>'''Building blocks of mitochondrial physiology'''</big> '''MitoEAGLE recommendations Part 1.'''
__TOC__
 
<br />
 
{{MitoEAGLE preprint 1 Phases}}
 


::: '''Scope of MITOEAGLE publications: respiratory control'''
== Open invitation ==
::::* Target a broad audience – also the new generation
::::* List of terms including historical terms; abbreviations (mtDNA, mt to abbreviate mitochondr*); OXPHOS capacity versus State 3 (discuss saturating ADP/Pi .. concentrations)
::::* Scientific terminology should be general and platform independent - meet the demands of all working groups.


:::: ''This manuscript on ‘The protonmotive force and respiratory control’ is a position statement in the frame of COST Action CA15203 MitoEAGLE. The list of coauthors evolved from MitoEAGLE Working Group Meetings and a bottom-up spirit of COST: This is an open invitation to scientists and students to join as coauthors, to provide a balanced view on mitochondrial respiratory control, a fundamental introductory presentation of the concept of the protonmotive force, and a critical discussion on reporting data of mitochondrial respiration in terms of metabolic flows and fluxes. We plan a series of follow-up reports by the expanding MitoEAGLE Network, to increase the scope of consensus-oriented recommendations and facilitate global communication and collaboration.''


== Mitochondrial respiratory control: a conceptual perspective on coupling states ==
::::* ''We continue to invite comments and suggestions on the MitoEAGLE preprint (phase 2; until '''October 12'''), particularly if you are an early career investigator adding an open future-oriented perspective, or an established scientist providing a balanced historical basis. Your critical input into the quality of the manuscript will be most welcome, improving our aims to be educational, general, consensus-oriented, and practically helpful for students working in mitochondrial respiratory physiology.''
::: '''MITOEAGLE recommendations Part 1.'''


::: '''MITOEAGLE Terminology Committee''' - first draft prepared by [[Gnaiger E |Erich Gnaiger]] 2017-04-18
::::* '''''Please feel free to focus on a particular section in terms of direct input and references, while evaluating the entire scope of the manuscript from the perspective of your expertise.'''''


::: '''Journal''': Int J Biochem Cell Biol (W Koopman will be the new editor); Open Access is a requirement
::::* Your comments will be largely posted on the discussion page of the MitoEAGLE preprint website. If you prefer to submit comments in the format of a referee's evaluation rather than a contribution as a coauthor, I will be glad to distribute your views to the updated list of coauthors for a balanced response. We would ask for your consent on this open bottom-up procedure.


=== Abstract ===
:::: ''We organize a MitoEAGLE session linked to our series of communications at the MiP''conference'' Nov 2017 in Hradec Kralove in close association with the MiP''society'' (where you hopefully will attend) and at EBEC 2018 in Budapest.''
:::: Clarity of concepts and consistency of nomenclature is a signature of the quality of a research area across its specializations, aimed at facilitating translational communication and teaching. The expanding field of mitochondrial respiratory physiology can benefit from the harmonization of nomenclature on mitochondrial respiratory states and control parameters. Peter Mitchell’s protonmotive, chemiosmotic force across the inner mitochondrial membrane, Δ''p''<sub>mt</sub>, establishes the link between electron transfer and phosphorylation of ADP to ATP, and between the chemical (pH difference, ΔpH) and electric (mt-membrane potential difference, Δ''Ψ''<sub>mt</sub>) components of energy transformation. This unifying concept provides the framework upon which a consistent terminology on mitochondrial physiology and bioenergetics can be based. IUPAC guidelines are followed for general terms of physical chemistry, extended by concepts of nonequilibrium thermodynamics and open systems. The differential nomenclature of classical bioenergetics (numerical differentiation of experimental protocol-linked respiratory States 1, 2, 3, 4 and 5) is incorporated into a concept-driven constructive terminology to address the basic meaning of a respiratory state and direct attention from the experimental ‘how’ to the concentual ‘why’. [[LEAK respiration |LEAK]] states are evaluated to study resting respiration, ''L'', compensating mainly for the proton leak. [[Oxidative phosphorylation |OXPHOS]] capacity, ''P'', is measured at saturating levels of ADP and inorganic phosphate to obtain kinetic references values for diagnostic applications. The ETS state differentiates the oxidative capacity of the [[electron transfer system]], ''E'', from OXPHOS capacity, revealing the limitation of P by the phosphorylation system. Development of databases on mitochondrial respiratory control requires application of strictly defined terms for comparison of respiratory states.
:::::::::::: » '''[[MiP2017_Hradec_Kralove_CZ]]'''  


:::: ''I thank you in advance for your feedback.''
:::: ''With best wishes,''


=== Introduction ===
:::: Every study of mitochondrial function and disease in human tissues and cells is faced with '''e'''volution, '''a'''ge, '''g'''ender, '''l'''ifestyle and '''e'''nvironment (EAGLE) as essential background conditions characterizing the individual patient, subject, study group, species, tissue or – to some extent - cell line. This range of factors is too wide to be accommodated in any single project on mitochondrial respiration. Only a large and well coordinated network can manage to generate the necessary type, quality and number of consistent data to address the complexity of EAGLE. The global MITOEAGLE network is a strategic innovation to develop harmonization protocols towards generating a rigorously monitored database on mitochondrial respiratory function. A quality control and data management system (QMS) is necessary to interrelate results obtained in a large number of studies, to interpret pathological phenotypes, and to set results into the multidimensional context of EAGLE.
:::: Reliability and comparability of quantitative results depends on accuracy of measurement under well defined conditions. To relate results of studies carried out by different research groups further requires reporting in a conceptually meaningful framework, assigning values expressed in common units (the measurements) to specific attributes (the rows in a table). If the attribute is well defined and understood (‘well defined experimental conditions’), it does not matter which names or symbols are given to the rows in the table of a database. But if terminology is vague or ambiguous when characterizing the type of result, then comparison of even the most accurate data can lead to confusion and may turn valuable signals into wasteful noise. Clarification of some fundamental terms used in studies of mitochondrial respiratory control, therefore, represents a basic component of a QMS. Even if standardization of nomenclature remains a goal of the optimist and is out of reach in the real world, harmonization of the technical jargon will improve the awareness of the intricate meaning of divergent scientific vocabulary. The MITOEAGLE Terminology Committee aims at accomplishing the ambitious goal to harmonize, unify and thus simplify the terminology in the field of mitochondrial physiology.


=== Respiratory coupling states: from bioenergetics to mitochondrial physiology ===
:::: ''Erich Gnaiger''
::::::: ''‘Every professional group develops its own technical jargon for talking about matters of critical concern. .. People who know a word can share that idea with other members of their group, and a shared vocabulary is part of the glue that holds people together and allows them to create a shared culture’'' (Miller 1991).
:::: ''Chair Mitochondrial Physiology Society''  - http://www.mitophysiology.org
:::: ''Chair COST Action MitoEAGLE'' - http://www.mitoeagle.org
:::: ''Medical University of Innsbruck, Austria''


==== Constructive versus differential terminologies ====
__TOC__
:::: Five classical states of mitochondrial respiration and cytochrome redox states have been introduced by Chance and Williams (1955; 1956). Table 1 explains a protocol with isolated mitochondria (imt) in a closed respirometric chamber:
::::* '''State 1''' is obtained after addition of imt to isotonic respiration medium containing inorganic phosphate but no adenylates, specifically no ADP, and no fuel substrates (CHNO).
::::* '''State 2''' is induced by addition of a high level of ADP, which stimulates transiently respiration on the basis of endogenous fuel substrates, followed by a low respiratory activity limited by fuel substrate availability.
::::* '''State 3''': Respiratory fuel substrates are added to initiate State 3. Respiration is stimulated while the ADP level is still high and supports coupled energy transformation in oxidative phosphorylation.
::::* '''State 4''': If the imt preparation is of high quality, devoid of ATPase activity and well coupled, gradual depletion of ADP by phosphorylation to accumulating ATP leads to a sudden decline of oxygen uptake in the transition from State 3 to State 4, when a maximum mt-membrane potential and the ATP/ADP ratio are maintained. State 4 respiration is limited by physiological uncoupling (mainly the intrinsic proton leak). A repeated titration of high ADP can restore active respiration in State 3.
::::* '''State 5''' is the anoxic state after the aerobic-anoxic transition, with zero respiration in a closed respirometric system. Ideal closed respirometers without any backdiffusion of oxygen into the chamber are difficult to obtain in practice, whence this steady state (State 5) is characterized by the oxygen-limited respiration compensating for the oxygen backdiffusion at near-zero oxygen levels.


:::: It has been suggested to extend the differential nomenclature (States 1 to 5) by a concept-driven constructive terminology on coupling states (Gnaiger 2009). In a constructive theory, the meanings are expressed by terms which carry information on the concept they represent (Miller 1991). The concept-driven terminology must be (i) general and not restricted to a particular experimental protocol with imt in a closed oxygraph, (ii) applicable to any mt-preparation, (iii) describe and guide studies with any experimental platform using a wide variety of experimental protocols, and (iv) extend the focus of classical bioenergetics on coupling mechanisms and coupling sites of oxidative phosphorylation to perspectives on mitochondrial physiology and pathology with diagnostic applications. This does not lead simply to a system of synonyms, but to a conceptual differentiation between the classical respiratory states which have been entirely sufficient to describe a specific bioenergetic protocol on one hand, and the extended constructive framework of the theory of mitochondrial physiology on the other hand.
::: '''Scope'''
::::* Target a broad audience – introduce the new generation of investigators.
::::* List of terms including historical terms; abbreviations (mtDNA, mt to abbreviate mitochondr*); OXPHOS capacity versus State 3 (discuss saturating ADP/Pi .. concentrations).
::::* Protonmotive force: not a force defined in physics, but an ''isomorphic'' force of statistical and nonequilibrium thermodynamics.
::::* Flux and flow: clarification of normalization.
::::* Scientific terminology should be general and platform-independent, meeting the demands of all working groups.


[[File:EPL-free and excess.jpg|right|300px|thumb|Capacities of the electron transfer system, oxidative phosphorylation and resting LEAK respiration (ETS, OXPHOS, LEAK) and four-compartmental OXPHOS model. (i) Capacity of the ETS module, ''E'', in the noncoupled state, generating the protonmotive force, Δ''p''<sub>mt</sub>. OXPHOS capacity, ''P'', is partitioned into (ii) the dissipative LEAK component, ''L'' (disspation of Δ''p''<sub>mt</sub>), and (iii) the free OXPHOS capacity, ''≈P=P-L'' (energy conversion driven by Δ''p''<sub>mt</sub>). If ''≈P'' is limited by the capacity of the phosphorylation system, then (iv) the apparent ETS excess capacity, ''ExP=E-P'', is available to drive coupled processes other than phosphorylation without competing with ATP production. Free divided by total ETS capacity, ''≈E/E'', is the ETS coupling efficiency. Free divided by total OXPHOS capacity, ''≈P/P'', is the OXPHOS coupling efficiency. From [[Gnaiger 2014 MitoPathways]].]]
:::::::::::: ''‘Every professional group develops its own technical jargon for talking about matters of critical concern. .. People who know a word can share that idea with other members of their group, and a shared vocabulary is part of the glue that holds people together and allows them to create a shared culture’'' (Miller 1991).
==== Three fundamental coupling states of mitochondrial preparations ====
::::* '''ETS, state ''E''''': Noncoupled state at optimal uncoupler concentration for maximum oxygen flux as a measure of electron transfer system (ETS) capacity, in the presence of fuel substrates and oxygen.
::::* '''OXPHOS, state ''P''''': ADP-activated state at maximum oxygen flux as a measure of the capacity for oxidative phosphorylation (OXPHOS), supported by saturating ADP and inorganic phosphate concentrations and in the presence of fuel substrates and oxygen. The definition of State 3 lacks a fundamental attribute of OXPHOS capacity. 'High ADP' in State 3 is a concentration of ADP specifically selected to allow measurememt of a State 3 to State 4 transition of isolated mitochondria in a closed respirometric system. Starting at oxygen levels near air saturation, the 'high ADP' concentration added must be low enough to allow phosphorylation to ATP at a coupled oxygen consumption that does not lead to oxygen depletion during the transition to State 4. In contrast, OXPHOS capacity requires evaluation of kinetically saturating ADP concentrations, which are usually an order of magnitude higher than 'high ADP'.
::::* '''LEAK, state ''L''''': Resting state of non-phosphorylating respiration when oxygen flux is maintained mainly to compensate for the proton leak in the absence of ATP synthesis, either due to absence or depletion of ADP at a maximum ATP/ADP ratio, or due to inhibition of phosphorylation by inhibitor such as oligomycin or atractyloside, in the presence of fuel substrates and oxygen.
::::* '''ROX''': Residual oxygen consumption (ROX) is not characteristic of a coupling state, but represents a baseline for correction of mitochondrial respiration in the presence of oxygen and the absence of fuel substrates or the addition of inhibitors of the electron transfer system.


:::: In kinetic studies an infinite number of intermediary steady states can be established (i) between LEAK an OXPHOS, using ADP-regenerating enzyme systems or steady-state ADP injections, (ii) between LEAK and ETS or OXPHOS and ETS, by uncloupler titrations, and (iii) between any of the three fundamental coupling states and ROX, by inhibitor titrations.
=== Three fundamental coupling states of mitochondrial preparations and residual oxygen consumption ===
::::* '''[[OXPHOS]]'''
::::* '''[[Electron transfer pathway]]'''
::::* '''[[LEAK respiration]]'''
::::* '''[[ROX]]'''


=== Rates and states ===
=== Classical terminology for isolated mitochondria ===
::::* Units (important for a database); analogous to electic terms: Flow [C.s-1]; Flux [C.s-1.m-2]; Rate (?)
:::::::::::: ''‘When a code is familiar enough, it ceases appearing like a code; one forgets that there is a decoding mechanism. The message is identical with its meaning’'' (Hofstadter 1979).
:::: Metabolic fluxes and steady-state variables are measured in defined mitochondrial respiratory states. Strictly, steady states can be obtained only in open systems, in which internal transformations (e.g. oxygen consumption or enthalpy change of chemical reactions) are instantaneously compensated by external flows (oxygen supply or heat flow across the system boundaries), such that changes do not occur in the system but only in the environment. Mitochondrial respiratory states measured in closed systems may satisfy the criteria of pseudo-steady states for limited periods of time, when the changes occurring in the system (e.g. oxygen concentration; ADP concentration; mt-membrane potential) do not exert significant effects on the metabolic fluxes (respiration, phosphorylation of ADP to ATP). Such pseudo-steady states require saturation levels of substrates to be maintained and thus depend on the kinetics of the processes under investigation.
::::* '''[[State 2]]'''
::::* '''[[State 3]]'''
::::* '''[[State 4]]'''


=== Intact cells versus mitochonddrial preparations ===


=== Normalization of flux and flow ===
=== States and rates ===
::::* '''[[Force |Force, ''F''<sub>tr</sub>]]'''
::::* '''[[External flow |External flow, ''I''<sub>ext</sub>]]'''
::::* '''[[Internal flow |Internal flow, ''I''<sub>int</sub>]]'''
::::* '''[[Open system]]'''
::::* '''[[Respiratory state]]'''


=== References ===
=== Normalization: fluxes and flows ===
::::* Altmann R (1894) Die Elementarorganismen und ihre Beziehungen zu den Zellen. Zweite vermehrte Auflage. Verlag Von Veit & Comp, Leipzig:160 pp, 34 Tafeln. - [[Altmann 1894 Verlag Von Veit & Comp |»Bioblast link«]]
::::* '''[[Extensive quantity]]'''
::::* Chance B, Williams GR (1955) Respiratory enzymes in oxidative phosphorylation. I. Kinetics of oxygen utilization. J Biol Chem 217:383-93. - [[Chance 1955 J Biol Chem-I |»Bioblast link«]]
::::* '''[[Oxygen flow |Oxygen flow, ''I''<sub>O2</sub>]]'''
::::* Chance B, Williams GR (1955) Respiratory enzymes in oxidative phosphorylation. III. The steady state. J Biol Chem 217:409-27. - [[Chance 1955 J Biol Chem-III |»Bioblast link«]]
::::* '''[[Oxygen flux |Oxygen flux, ''J''<sub>O2</sub>]]'''
::::* Chance B, Williams GR (1956) The respiratory chain and oxidative phosphorylation. Adv Enzymol17:65-134. - [[Chance 1956 Adv Enzymol Relat Subj Biochem |»Bioblast link«]]
::::* '''[[Mitochondrial marker]]'''
::::* Cohen ER, Cvitas T, Frey JG, Holmström B, Kuchitsu K, Marquardt R, Mills I, Pavese F, Quack M, Stohner J, Strauss HL, Takami M, Thor HL (2008) Quantities, Units and Symbols in Physical Chemistry, IUPAC Green Book, 3rd Edition, 2nd Printing, IUPAC & RSC Publishing, Cambridge. - [[Cohen 2008 IUPAC Green Book |»Bioblast link«]]
::::* '''[[Specific quantity]]'''
::::* Estabrook R (1967) Mitochondrial respiratory control and the polarographic measurement of ADP:O ratios. Meth Enzymol 10:41-7. - [[Estabrook 1967 Methods Enzymol |»Bioblast link«]]
::::* '''[[Intensive quantity]]'''
::::* Galluzzi L, Vitale I, Abrams JM, Alnemri ES, Baehrecke EH, Blagosklonny MV, Dawson TM, Dawson VL, El-Deiry WS, Fulda S, Gottlieb E, Green DR, Hengartner MO, Kepp O, Knight RA, Kumar S, Lipton SA, Lu X, Madeo F, Malorni W, Mehlen P, Nuñez G, Peter ME, Piacentini M, Rubinsztein DC, Shi Y, Simon HU, Vandenabeele P, White E, Yuan J, Zhivotovsky B, Melino G, Kroemer G (2012) Molecular definitions of cell death subroutines: recommendations of the Nomenclature Committee on Cell Death 2012. Cell Death Differ 19:107-20. - [[Galluzzi 2012 Cell Death Differ |»Bioblast link«]]
::::* Gnaiger E (1993) Nonequilibrium thermodynamics of energy transformations. Pure Appl Chem 65:1983-2002. - [[Gnaiger 1993 Pure Appl Chem |»Bioblast link«]]
::::* Gnaiger E (1993) Efficiency and power strategies under hypoxia. Is low efficiency at high glycolytic ATP production a paradox? In: Surviving Hypoxia: Mechanisms of Control and Adaptation. Hochachka PW, Lutz PL, Sick T, Rosenthal M, Van den Thillart G (eds) CRC Press, Boca Raton, Ann Arbor, London, Tokyo:77-109. - [[Gnaiger 1993 Hypoxia |»Bioblast link«]]
::::* Gnaiger E (2009) Capacity of oxidative phosphorylation in human skeletal muscle. New perspectives of mitochondrial physiology. Int J Biochem Cell Biol 41:1837-45. - [[Gnaiger 2009 Int J Biochem Cell Biol |»Bioblast link«]]
::::* Gnaiger E (2014) Mitochondrial pathways and respiratory control. An introduction to OXPHOS analysis. 4th ed. Mitochondr Physiol Network 19.12. OROBOROS MiPNet Publications, Innsbruck:80 pp. - [[Gnaiger_2014_MitoPathways#Chapter_2._Respiratory_states:_coupling_control |»Bioblast link«]]
::::* Gnaiger E, Méndez G, Hand SC (2000) High phosphorylation efficiency and depression of uncoupled respiration in mitochondria under hypoxia. Proc Natl Acad Sci U S A 97:11080-5. - [[Gnaiger 2000 Proc Natl Acad Sci U S A |»Bioblast link«]]
::::* International Union of Biochemistry and Molecular Biology: Recommendations for terminology and databases for biochemical thermodynamics. - [http://www.chem.qmul.ac.uk/iubmb/thermod2/ »Open Access«]
::::* Hatefi Y, Haavik AG, Fowler LR, Griffiths DE (1962) Studies on the electron transfer system. XLII. Reconstitution of the electron transfer system. J Biol Chem 237:2661-9. - [[Hatefi 1962 J Biol Chem |»Bioblast link«]]
::::* International Union of Biochemistry (1981) Symbolism and terminology in enzyme kinetics. - [http://www.chem.qmul.ac.uk/iubmb/kinetics/ »Open Access«]
::::* Lemieux H, Blier PU, Gnaiger E (2017) Remodeling pathway control of mitochondrial respiratory capacity by temperature in mouse heart: electron flow through the Q-junction in permeabilized fibers. Sci Rep (in press). - [[Lemieux 2017 bioRxiv |»Preprint Bioblast link«]]
::::* Miller GA (1991) The science of words. Scientific American Library New York:276 pp. - [[Miller 1991 Scientific American Library |»Bioblast link«]]
::::* Mitchell P (1961) Coupling of phosphorylation to electron and hydrogen transfer by a chemi-osmotic type of mechanism. Nature 191:144-8. - [[Mitchell 1961 Nature |»Bioblast link«]]
::::* Mitchell P (1966) Chemiosmotic coupling in oxidative and photosynthetic phosphorylation. Glynn Research Ltd, Bodmin:192 pp. - ''The Grey Book'' 1.
::::* Mitchell P (1968) Chemiosmotic coupling and energy transduction. Glynn Research Ltd, Bodmin:111 pp. - ''The Grey Book'' 2.
::::* Mitchell P, Moyle J (1967) Respiration-driven proton translocation in rat liver mitochondria. Biochem J 105:1147-62. - [[Mitchell 1967 Biochem J |»Bioblast link«]]


=== List of selected terms and symbols ===
::::* '''[[Coupled respiration]]'''
::::* '''[[Dyscoupled respiration]]'''
::::* '''[[Isolated mitochondria |Isolated mitochondria, imt]]'''
::::* '''Mitochondria, mt''' (Greek mitos: thread; chondros: granule) are small structures within cells, which function in cell respiration as powerhouses or batteries. Mitochondria belong to the bioblasts of Richard Altmann (1894). Abbreviation: mt, as generally used in mtDNA. Singular: mitochondrion (bioblast); plural: mitochondria (bioblasts).
::::* '''[[Mitochondrial inner membrane]]'''
::::* '''[[Mitochondrial outer membrane]]'''
::::* '''[[Mitochondrial matrix]]'''
::::* '''[[Mitochondrial membrane potential]]
::::* '''Mitochondrial preparations, mtprep''' are isolated mitochondria (imt), tissue homogenate (thom), mechanically or chemically permeabilized tissue (permeabilized fibres, pfi) or permeabilized cells (pce). In these preparations the cell membranes are either removed (imt and smtp) or mechanically (thom) and chemically permeabilized (pfi), while the mitochondrial functional integrity and to a large extent the mt-structure is maintained.
::::* '''[[Mitochondrial respiration]]'''
::::* '''[[Noncoupled respiration]]'''
::::* '''[[Oligomycin |Oligomycin, Omy]]'''
::::* '''[[Permeabilized tissue or cells |Permeabilized tissue or cells, pti, pce]]'''
::::* '''[[Phosphorylation system]]'''
::::* '''[[Proton leak]]'''
::::* '''[[Proton pump]]'''
::::* '''[[Proton slip]]'''
::::* '''Protonmotive force'''
::::* '''[[Tissue homogenate |Tissue homogenate, thom]]'''
::::* '''[[Uncoupler |Uncoupler, U]]'''


== Mitochondrial respiratory control: pathway states in mt-preparations and intact cells ==
 
::: '''MITOEAGLE recommendations Part 2.'''
== Mitochondrial pathways and respiratory control in mt-preparations ==
::: '''Building blocks of mitochondrial physiology Part 2.'''
 
:::::::::::: ''‘It is essential to define both the substrate and ADP levels in order to identify the steady-state condition of the mitochondria during the experiment’'' (Chance and Williams, 1956).


::::* The mitochondrial respiratory system
::::* The mitochondrial respiratory system
::::* Substrates and inhibitors
::::* Substrates and inhibitors
::::* Switch to pathway-related nomenclature instead of enzyme-linked terminology (N/NS/S versus CI/CI+II/CII)
::::* Switch to pathway-related nomenclature instead of enzyme-linked terminology (N/NS/S versus CI/CI&II/CII)
 
 
== Mitochondrial respiratory control: cell respiration ==
::: '''Building blocks of mitochondrial physiology Part 3.'''
 
::: '''Living cells versus mitochondrial preparations'''
::::* '''[[Living cells |Living cells, ce]]'''
::::* '''[[Basal respiration]]'''
::::* '''[[Cell respiration]]'''
::::* '''[[Resting metabolic rate]]'''
::::* '''[[ROUTINE respiration |ROUTINE state, state ''R'']]''': ROUTINE respiration of intact, viable cells is regulated according to physiological activity, at intracellular non-saturating ADP levels. ''R'' increases under various conditions of activation. When incubated in culture medium, cells maintain a ROUTINE level of activity, ''R'' (ROUTINE mitochondrial respiration; corrected for residual oxygen consumption due to oxidative side reactions). ROUTINE activity may include aerobic energy requirements for cell growth and is thus fundamentally different from the definition of basal metabolic rate (BMR). When incubated for short experimental periods in a medium devoid of fuel substrates, the cells respire solely on endogenous substrates at the corresponding state of ROUTINE activity, e<sub>''R''</sub> (e, endogenous substrate supply).
 
:::: It is difficult to stimulate living cells to maximum OXPHOS activity, since ADP and inorganic phosphate do not equilibrate across intact plasma membranes, and thus saturating concentrations of these metabolites can hardly be achieved in living cells. LEAK and ET-pathway states, however, can be induced in viable cells with application of inhibitors of the phosphorylation system and uncouplers, respectively, due to the fact that cell membranes are highy permeable for these substances. External fuel substrates are taken up by living cells to various extents, and intracellular metabolism of exogenous and endogenous substrates supports mitochondrial respiration with a physiological substrate supply. In contrast, mt-preparations depend on the external supply of fuel substrates which support the electron transfer-pathway with reducing equivalents. ET-pathway competence of external substrates is required for all coupling states of mt-preparations (''L, P, E'') and depends on (i) transport of substrates across the inner mt-membrane or oxidation by dehydrogenases located on the outer face of the inner mt-membrane (e.g. glycerophosphate dehydrogenase complex, CGpDH), (ii) oxidation in the mt-matrix (TCA cycle dehydrogenases and other matrix dehydrogenases, e.g. mtGDH) or on the inner face of the inner mt-membrane (succinate dehydrogenase, CII), (iii) oxidation of substrates without accumulation of inhibitory endproducts (e.g. oxaloacetate inhibiting succinate dehydrogenase; NADH and oxaloacetate inhibiting malate dehydrogenase), and (iv) electron transfer through the membrane-bound ET-pathway (mETS). Endproducts must be either easily exported from the matrix across the inner mt-membrane (e.g. malate formed from succinate via fumarate), or metabolized in the TCA cycle (e.g. malate-derived oxaloacetate forming citrate in the presence of external pyruvate&malate).
 
 
== Mitochondrial respiratory control: coupling control ratios and control factors ==
::: '''Building blocks of mitochondrial physiology Part 4.'''
 
::::* '''[[Coupling control factor |Coupling control factor, ''CCF'']]'''
::::* '''[[Coupling-control ratio |Coupling-control ratio, ''CCR'']]'''
 
::::* '''[[E-L coupling efficiency|E-L coupling efficiency, ''j<sub>≈E</sub>'']]''': ''j<sub>≈E</sub>'' = ''≈E/E'' = (''E-L'')/''E'' = 1-''L/E''
::::* '''[[LEAK-control ratio |LEAK-control ratio, ''L/E'']]'''
 
::::* '''[[Excess E-P capacity factor |Excess ''E-P'' capacity factor, ''j<sub>ExP</sub>'']]''': ''j<sub>Exp</sub>'' = (''E-P'')/''E'' = 1-''P/E''
::::* '''[[OXPHOS-control ratio |OXPHOS-control ratio, ''P/E'']]'''
 
::::* '''[[OXPHOS-coupling efficiency |OXPHOS-coupling efficiency, ''j<sub>≈P</sub>'']]''': ''j<sub>≈P</sub>'' = ''≈P/P'' = (''P-L'')/''P'' = 1-''L/P''
::::* '''[[L/P coupling control ratio |''L/P'' coupling control ratio, ''L/P'']]'''
::::* '''[[Respiratory acceptor control ratio |Respiratory acceptor control ratio, RCR]]''': RCR = State 4/State 3
 
::::* '''[[NetOXPHOS control ratio |netOXPHOS control ratio, ''≈P/E'']]''': ''≈P/E'' = (''P-L'')/''E''
 
::::* '''[[Uncoupling-control ratio |Uncoupling-control ratio, UCR]]''': UCR = ''E/R''




== Action ==
== Action ==
::::» '''WG1 Action''' - [[WG1 MITOEAGLE protocols, terminology, documentation]]: Standard operating procedures and user requirement document: Protocols, terminology, documentation
::::» '''WG1 Action''' - [[WG1 MitoEAGLE protocols, terminology, documentation]]: Standard operating procedures and user requirement document: Protocols, terminology, documentation
::::» [[MITOEAGLE protocols, terminology, documentation |WG1 Project application]]
::::» [[MitoEAGLE protocols, terminology, documentation |WG1 Project application]]
::::» Gnaiger E, Aasander Frostner E, Abdul Karim N, Abumrad NA, Acuna-Castroviejo D, Adiele RC et al (2019) Mitochondrial respiratory states and rates. MitoFit Preprint Arch doi:10.26124/mitofit:190001. - [[Gnaiger 2019 MitoFit Preprints |»Bioblast link«]]


 
::::» [[MitoEAGLE: Respiratory states |Pre-publication: Mitochondrial respiratory control states]]
::::» [[MITOEAGLE: Respiratory states |Pre-publication: Mitochondrial respiratory control states]]
::::» [[MitoPedia: Respiratory control ratios]]
::::» [[MitoPedia: Respiratory control ratios]]
::::» [[MitoPedia: SUIT]]
::::» [[MitoPedia: SUIT]]
::::» 2017-07 [[MiPschool Obergurgl 2017]]
::::» 2017-03 [[MITOEAGLE Barcelona 2017]]
::::» 2016-11 [[MITOEAGLE 2016 Verona IT]]

Latest revision as of 23:40, 25 January 2021


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COST Action CA15203 (2016-2021): MitoEAGLE
Evolution-Age-Gender-Lifestyle-Environment: mitochondrial fitness mapping


The protonmotive force and respiratory control


Building blocks of mitochondrial physiology MitoEAGLE recommendations Part 1.



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» Manuscript phases and versions

Manuscript phases and versions - an open-access apporach

COST Action MitoEAGLE
This manuscript on ‘Mitochondrial respiratory states and rates’ is a position statement in the frame of COST Action CA15203 MitoEAGLE. The list of coauthors evolved beyond phase 1 in the bottom-up spirit of COST.
The global MitoEAGLE network made it possible to collaborate with a large number of coauthors to reach consensus on the present manuscript. Nevertheless, we do not consider scientific progress to be supported by ‘declaration’ statements (other than on ethical or political issues). Our manuscript aims at providing arguments for further debate rather than pushing opinions. We hope to initiate a much broader process of discussion and want to raise the awareness on the importance of a consistent terminology for reporting of scientific data in the field of bioenergetics, mitochondrial physiology and pathology. Quality of research requires quality of communication. Some established researchers in the field may not want to re-consider the use of jargon which has become established despite deficiencies of accuracy and meaning. In the long run, superior standards will become accepted. We hope to contribute to this evolutionary process, with an emphasis on harmonization rather than standardization.
  • Phase 1: The protonmotive force and respiratory control
» The protonmotive force and respiratory control - Discussion
» MitoEAGLE preprint 2017-09-21 - Discussion
  • Phase 2: Mitochondrial respiratory states and rates: Building blocks of mitochondrial physiology Part 1
» MitoEAGLE Task Group States and rates - Discussion
  • Phase 4: Journal submission
  • Target: CELL METABOLISM, aiming at indexing by The Web of Science and PubMed.
Coauthors
  • 2017-09-21 Version 01: 105 coauthors
  • 2017-10-15 Version 10: 131 coauthors
  • 2018-01-18 Version 20: 168 coauthors
  • 2018-02-26 Version 30: 225 coauthors
  • 2018-08-20 Version 40: 350 coauthors - EBEC Poster
  • 2018-10-17 Version 44: 426 coauthors - MiPschool Tromso-Bergen 2018
  • 2018-12-12 Version 50: 517 coauthors - Submission to the preprint server bioRxiv not successful
  • 2019-02-12 Preprint version 1: 530 coauthors
  • 2019-03-15 Preprint version 2: 533 coauthors
  • 2019-04-24 Preprint version 3: 533 coauthors
  • 2019-05-20 Preprint version 4: 542 coauthors
  • 2019-07-24 Preprint version 5: 612 coauthors
  • 2019-08-30 Preprint version 6: 622 coauthors - Preprint publication doi:10.26124/mitofit:190001.v6
  • BEC 2020.1. - Gnaiger Erich et al ― MitoEAGLE Task Group (2020) Mitochondrial physiology. Bioenerg Commun 2020.1. doi:10.26124/bec:2020-0001.v1. - »Bioblast link«



Open invitation

This manuscript on ‘The protonmotive force and respiratory control’ is a position statement in the frame of COST Action CA15203 MitoEAGLE. The list of coauthors evolved from MitoEAGLE Working Group Meetings and a bottom-up spirit of COST: This is an open invitation to scientists and students to join as coauthors, to provide a balanced view on mitochondrial respiratory control, a fundamental introductory presentation of the concept of the protonmotive force, and a critical discussion on reporting data of mitochondrial respiration in terms of metabolic flows and fluxes. We plan a series of follow-up reports by the expanding MitoEAGLE Network, to increase the scope of consensus-oriented recommendations and facilitate global communication and collaboration.
  • We continue to invite comments and suggestions on the MitoEAGLE preprint (phase 2; until October 12), particularly if you are an early career investigator adding an open future-oriented perspective, or an established scientist providing a balanced historical basis. Your critical input into the quality of the manuscript will be most welcome, improving our aims to be educational, general, consensus-oriented, and practically helpful for students working in mitochondrial respiratory physiology.
  • Please feel free to focus on a particular section in terms of direct input and references, while evaluating the entire scope of the manuscript from the perspective of your expertise.
  • Your comments will be largely posted on the discussion page of the MitoEAGLE preprint website. If you prefer to submit comments in the format of a referee's evaluation rather than a contribution as a coauthor, I will be glad to distribute your views to the updated list of coauthors for a balanced response. We would ask for your consent on this open bottom-up procedure.
We organize a MitoEAGLE session linked to our series of communications at the MiPconference Nov 2017 in Hradec Kralove in close association with the MiPsociety (where you hopefully will attend) and at EBEC 2018 in Budapest.
» MiP2017_Hradec_Kralove_CZ
I thank you in advance for your feedback.
With best wishes,


Erich Gnaiger
Chair Mitochondrial Physiology Society - http://www.mitophysiology.org
Chair COST Action MitoEAGLE - http://www.mitoeagle.org
Medical University of Innsbruck, Austria
Scope
  • Target a broad audience – introduce the new generation of investigators.
  • List of terms including historical terms; abbreviations (mtDNA, mt to abbreviate mitochondr*); OXPHOS capacity versus State 3 (discuss saturating ADP/Pi .. concentrations).
  • Protonmotive force: not a force defined in physics, but an isomorphic force of statistical and nonequilibrium thermodynamics.
  • Flux and flow: clarification of normalization.
  • Scientific terminology should be general and platform-independent, meeting the demands of all working groups.
‘Every professional group develops its own technical jargon for talking about matters of critical concern. .. People who know a word can share that idea with other members of their group, and a shared vocabulary is part of the glue that holds people together and allows them to create a shared culture’ (Miller 1991).

Three fundamental coupling states of mitochondrial preparations and residual oxygen consumption

Classical terminology for isolated mitochondria

‘When a code is familiar enough, it ceases appearing like a code; one forgets that there is a decoding mechanism. The message is identical with its meaning’ (Hofstadter 1979).


States and rates

Normalization: fluxes and flows

List of selected terms and symbols


Mitochondrial pathways and respiratory control in mt-preparations

Building blocks of mitochondrial physiology Part 2.
‘It is essential to define both the substrate and ADP levels in order to identify the steady-state condition of the mitochondria during the experiment’ (Chance and Williams, 1956).
  • The mitochondrial respiratory system
  • Substrates and inhibitors
  • Switch to pathway-related nomenclature instead of enzyme-linked terminology (N/NS/S versus CI/CI&II/CII)


Mitochondrial respiratory control: cell respiration

Building blocks of mitochondrial physiology Part 3.
Living cells versus mitochondrial preparations
  • Living cells, ce
  • Basal respiration
  • Cell respiration
  • Resting metabolic rate
  • ROUTINE state, state R: ROUTINE respiration of intact, viable cells is regulated according to physiological activity, at intracellular non-saturating ADP levels. R increases under various conditions of activation. When incubated in culture medium, cells maintain a ROUTINE level of activity, R (ROUTINE mitochondrial respiration; corrected for residual oxygen consumption due to oxidative side reactions). ROUTINE activity may include aerobic energy requirements for cell growth and is thus fundamentally different from the definition of basal metabolic rate (BMR). When incubated for short experimental periods in a medium devoid of fuel substrates, the cells respire solely on endogenous substrates at the corresponding state of ROUTINE activity, eR (e, endogenous substrate supply).
It is difficult to stimulate living cells to maximum OXPHOS activity, since ADP and inorganic phosphate do not equilibrate across intact plasma membranes, and thus saturating concentrations of these metabolites can hardly be achieved in living cells. LEAK and ET-pathway states, however, can be induced in viable cells with application of inhibitors of the phosphorylation system and uncouplers, respectively, due to the fact that cell membranes are highy permeable for these substances. External fuel substrates are taken up by living cells to various extents, and intracellular metabolism of exogenous and endogenous substrates supports mitochondrial respiration with a physiological substrate supply. In contrast, mt-preparations depend on the external supply of fuel substrates which support the electron transfer-pathway with reducing equivalents. ET-pathway competence of external substrates is required for all coupling states of mt-preparations (L, P, E) and depends on (i) transport of substrates across the inner mt-membrane or oxidation by dehydrogenases located on the outer face of the inner mt-membrane (e.g. glycerophosphate dehydrogenase complex, CGpDH), (ii) oxidation in the mt-matrix (TCA cycle dehydrogenases and other matrix dehydrogenases, e.g. mtGDH) or on the inner face of the inner mt-membrane (succinate dehydrogenase, CII), (iii) oxidation of substrates without accumulation of inhibitory endproducts (e.g. oxaloacetate inhibiting succinate dehydrogenase; NADH and oxaloacetate inhibiting malate dehydrogenase), and (iv) electron transfer through the membrane-bound ET-pathway (mETS). Endproducts must be either easily exported from the matrix across the inner mt-membrane (e.g. malate formed from succinate via fumarate), or metabolized in the TCA cycle (e.g. malate-derived oxaloacetate forming citrate in the presence of external pyruvate&malate).


Mitochondrial respiratory control: coupling control ratios and control factors

Building blocks of mitochondrial physiology Part 4.


Action

» WG1 Action - WG1 MitoEAGLE protocols, terminology, documentation: Standard operating procedures and user requirement document: Protocols, terminology, documentation
» WG1 Project application
» Gnaiger E, Aasander Frostner E, Abdul Karim N, Abumrad NA, Acuna-Castroviejo D, Adiele RC et al (2019) Mitochondrial respiratory states and rates. MitoFit Preprint Arch doi:10.26124/mitofit:190001. - »Bioblast link«
» Pre-publication: Mitochondrial respiratory control states
» MitoPedia: Respiratory control ratios
» MitoPedia: SUIT
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