Adenosine Triphosphate Demand Research Articles

Effects of Time-Dependent Adenosine Triphosphate Consumption Caused by Neuron Firing on Adenosine Triphosphate Concentrations in Synaptic Boutons Containing and Lacking a Stationary Mitochondrion.

The precise mechanism behind the supply of adenosine triphosphate (ATP) to approximately half of the presynaptic release sites in axons that lack a stationary mitochondrion is not fully understood. This paper presents a mathematical model designed to simulate the transient ATP concentration in presynaptic en passant boutons. The model is utilized to investigate how the ATP concentration responds to increased ATP demand during neuronal firing in boutons with a stationary mitochondrion and those without one. The analysis suggests that neuron firing may cause oscillations in the ATP concentrations, with peak-to-peak amplitudes ranging from 0.06% to 5% of their average values. However, this does not deplete boutons lacking a mitochondrion of ATP; for physiologically relevant values of model parameters, their concentration remains approximately 3.75 times higher than the minimum concentration required for synaptic activity. The variance in average ATP concentrations between boutons containing a stationary mitochondrion and those lacking one ranges from 0.3% to 0.8%, contingent on the distance between the boutons. The model indicates that diffusion-driven ATP transport is rapid enough to adequately supply ATP molecules to boutons lacking a stationary mitochondrion.

Disrupting Na+ ion homeostasis and Na+/K+ ATPase activity in breast cancer cells directly modulates glycolysis in vitro and in vivo

BackgroundGlycolytic flux is regulated by the energy demands of the cell. Upregulated glycolysis in cancer cells may therefore result from increased demand for adenosine triphosphate (ATP), however it is unknown what this extra ATP turnover is used for. We hypothesise that an important contribution to the increased glycolytic flux in cancer cells results from the ATP demand of Na+/K+-ATPase (NKA) due to altered sodium ion homeostasis in cancer cells.MethodsLive whole-cell measurements of intracellular sodium [Na+]i were performed in three human breast cancer cells (MDA-MB-231, HCC1954, MCF-7), in murine breast cancer cells (4T1), and control human epithelial cells MCF-10A using triple quantum filtered 23Na nuclear magnetic resonance (NMR) spectroscopy. Glycolytic flux was measured by 2H NMR to monitor conversion of [6,6-2H2]d-glucose to [2H]-labelled l-lactate at baseline and in response to NKA inhibition with ouabain. Intracellular [Na+]i was titrated using isotonic buffers with varying [Na+] and [K+] and introducing an artificial Na+ plasma membrane leak using the ionophore gramicidin-A. Experiments were carried out in parallel with cell viability assays, 1H NMR metabolomics of intracellular and extracellular metabolites, extracellular flux analyses and in vivo measurements in a MDA-MB-231 human-xenograft mouse model using 2-deoxy-2-[18F]fluoroglucose (18F-FDG) positron emission tomography (PET).ResultsIntracellular [Na+]i was elevated in human and murine breast cancer cells compared to control MCF-10A cells. Acute inhibition of NKA by ouabain resulted in elevated [Na+]i and inhibition of glycolytic flux in all three human cancer cells which are ouabain sensitive, but not in the murine cells which are ouabain resistant. Permeabilization of cell membranes with gramicidin-A led to a titratable increase of [Na+]i in MDA-MB-231 and 4T1 cells and a Na+-dependent increase in glycolytic flux. This was attenuated with ouabain in the human cells but not in the murine cells. 18FDG PET imaging in an MDA-MB-231 human-xenograft mouse model recorded lower 18FDG tumour uptake when treated with ouabain while murine tissue uptake was unaffected.ConclusionsGlycolytic flux correlates with Na+-driven NKA activity in breast cancer cells, providing evidence for the ‘centrality of the [Na+]i-NKA nexus’ in the mechanistic basis of the Warburg effect.

Integrated Functions of Cardiac Energetics, Mechanics, and Purine Nucleotide Metabolism.

Purine nucleotides play central roles in energy metabolism in the heart. Most fundamentally, the free energy of hydrolysis of the adenine nucleotide adenosine triphosphate (ATP) provides the thermodynamic driving force for numerous cellular processes including the actin-myosin crossbridge cycle. Perturbations to ATP supply and/or demand in the myocardium lead to changes in the homeostatic balance between purine nucleotide synthesis, degradation, and salvage, potentially affecting myocardial energetics and, consequently, myocardial mechanics. Indeed, both acute myocardial ischemia and decompensatory remodeling of the myocardium in heart failure are associated with depletion of myocardial adenine nucleotides and with impaired myocardial mechanical function. Yet there remain gaps in the understanding of mechanistic links between adenine nucleotide degradation and contractile dysfunction in heart disease. The scope of this article is to: (i) review current knowledge of the pathways of purine nucleotide depletion and salvage in acute ischemia and in chronic heart disease; (ii) review hypothesized mechanisms linking myocardial mechanics and energetics with myocardial adenine nucleotide regulation; and (iii) highlight potential targets for treating myocardial metabolic and mechanical dysfunction associated with these pathways. It is hypothesized that an imbalance in the degradation, salvage, and synthesis of adenine nucleotides leads to a net loss of adenine nucleotides in both acute ischemia and under chronic high-demand conditions associated with the development of heart failure. This reduction in adenine nucleotide levels results in reduced myocardial ATP and increased myocardial inorganic phosphate. Both of these changes have the potential to directly impact tension development and mechanical work at the cellular level. © 2024 American Physiological Society. Compr Physiol 14:5345-5369, 2024.

The role of photorespiration in preventing feedback regulation via ATP synthase in Nicotiana tabacum.

Photorespiration consumes substantial amounts of energy in the forms of adenosine triphosphate (ATP) and reductant making the pathway an important component in leaf energetics. Because of this high reductant demand, photorespiration is proposed to act as a photoprotective electron sink. However, photorespiration consumes more ATP relative to reductant than the C3 cycle meaning increased flux disproportionally increases ATP demand relative to reductant. Here we explore how energetic consumption from photorespiration impacts the flexibility of the light reactions in nicotiana tabacum. Specifically, we demonstrate that decreased photosynthetic efficiency (ϕII ) at low photorespiratory flux was related to feedback regulation at the chloroplast ATP synthase. Additionally, decreased ϕII at high photorespiratory flux resulted in the accumulation of photoinhibition at photosystem II centers. These results are contrary to the proposed role of photorespiration as a photoprotective electron sink. Instead, our results suggest a novel role of ATP consumption from photorespiration in maintaining ATP synthase activity, with implications for maintaining energy balance and preventing photodamage that will be critical for plant engineering strategies.

Plasticity of mitochondrial function safeguards phosphorylating respiration during in vitro simulation of rest-phase hypothermia.

Many animals downregulate body temperature to save energy when resting (rest-phase hypothermia). Small birds that winter at high latitudes have comparatively limited capacity for hypothermia and so pay large energy costs for thermoregulation during cold nights. Available evidence suggests this process is fueled by adenosine triphosphate (ATP)-dependent mechanisms. Most ATP is produced by oxidative phosphorylation in the mitochondria, but mitochondrial respiration may be lower during hypothermia because of the temperature dependence of biological processes. This can create conflict between increased organismal ATP demand and a lower mitochondrial capacity to provide it. We studied this in blood cell mitochondria of wild great tits (Parus major) by simulating rest-phase hypothermia via a 6°C reduction in assay temperature in vitro. The birds had spent the night preceding the experiment in thermoneutrality or in temperatures representing mild or very cold winter nights, but night temperatures never affected mitochondrial respiration. However, across temperature groups, endogenous respiration was 14% lower in hypothermia. This did not reflect general thermal suppression of mitochondrial function because phosphorylating respiration was unaffected by thermal state. Instead, hypothermia was associated with a threefold reduction of leak respiration, from 17% in normothermia to 4% in hypothermia. Thus, the coupling of total respiration to ATP production was 96% in hypothermia, compared to 83% in normothermia. Our study shows that the thermal insensitivity of phosphorylation combined with short-term plasticity of leak respiration may safeguard ATP production when endogenous respiration is suppressed. This casts new light on the process by which small birds endure harsh winter cold and warrants future tests across tissues in vivo.

Exposure to low temperature prepares the turtle brain to withstand anoxic environments during overwintering.

In most vertebrates, anoxia drastically reduces the production of the essential adenosine triphosphate (ATP) to power its many necessary functions, and, consequently, cell death occurs within minutes. However, some vertebrates, such as the painted turtle (Chrysemys picta bellii), have evolved the ability to survive months without oxygen by simultaneously decreasing ATP supply and demand, surviving the anoxic period without any apparent cellular damage. The impact of anoxia on the metabolic function of painted turtles has received a lot of attention. However, the impact of low temperature has received less attention and the interactive effect of anoxia and temperature even less. In the present study, we investigated the interactive impacts of reduced temperature and severe hypoxia on the electrophysiological properties of pyramidal neurons in painted turtle cerebral cortex. Our results show that an acute reduction in temperature from 20 to 5°C decreases membrane potential, action potential width and amplitude, and whole-cell conductance. Importantly, acute exposure to 5°C considerably slows membrane repolarization by voltage-gated K+ channels. Exposing pyramidal cells to severe hypoxia in addition to an acute temperature change slightly depolarized membrane potential but did not alter action potential amplitude or width and whole-cell conductance. These results suggest that acclimation to low temperatures, preceding severe environmental hypoxia, induces cellular responses in pyramidal neurons that facilitate survival under low oxygen concentrations. In particular, our results show that temperature acclimation invokes a change in voltage-gated K+ channel kinetics that overcomes the acute inhibition of the channel.

Obesity modifies the energetic phenotype of dilated cardiomyopathy.

AimsWe sought to determine if myocardial energetics could distinguish obesity cardiomyopathy as a distinct entity from dilated cardiomyopathy.Methods and resultsSixteen normal weight participants with dilated cardiomyopathy (DCMNW), and 27 with DCM and obesity (DCMOB), were compared to 26 normal weight controls (CTLNW). All underwent cardiac magnetic resonance imaging and 31P spectroscopy to assess function and energetics. Nineteen DCMOB underwent repeat assessment after a dietary weight loss intervention. Adenosine triphosphate (ATP) delivery through creatine kinase (CK flux) was 55% lower in DCMNW than in CTLNW (P = 0.004), correlating with left ventricular ejection fraction (LVEF, r = 0.4, P = 0.015). In contrast, despite similar LVEF (DCMOB 41 ± 7%, DCMNW 38 ± 6%, P = 0.14), CK flux was two-fold higher in DCMOB (P < 0.001), due to higher rate through CK [median kf 0.21 (0.14) vs. 0.11 (0.12) s−1, P = 0.002]. During increased workload, the CTLNW heart increased CK flux by 97% (P < 0.001). In contrast, CK flux was unchanged in DCMNW and fell in DCMOB (by >50%, P < 0.001). Intentional weight loss was associated with positive left ventricular remodelling, with reduced left ventricular end-diastolic volume (by 8%, P < 0.001) and a change in LVEF (40 ± 9% vs. 45 ± 10%, P = 0.002). This occurred alongside a fall in ATP delivery rate with weight loss (by 7%, P = 0.049).ConclusionsIn normal weight, DCM is associated with reduced resting ATP delivery. In obese DCM, ATP demand through CK is greater, suggesting reduced efficiency of energy utilization. Dietary weight loss is associated with significant improvement in myocardial contractility, and a fall in ATP delivery, suggesting improved metabolic efficiency. This highlights distinct energetic pathways in obesity cardiomyopathy, which are both different from dilated cardiomyopathy, and may be reversible with weight loss.

Effect of hypoxia on purine metabolism of human skeletal muscle cells

Mammals experience some degree of hypoxia during their lifetime. In response to hypoxic challenge, mammalian cells orchestrate specific responses at transcriptional and posttranslational level which lead to changes in the purine metabolites in order to cope with threatening conditions. The aim of this study was to evaluate the response of the enzymes involved in the purine metabolism of human muscle cells to hypoxic conditions. Muscle cells in culture were exposed to hypoxia and the enzymatic activity of inosine monophosphate dehydrogenase (IMPDH), xanthine oxidase (XO), purine nucleoside phosphorylase (PNP) and hypoxanthine guanine phosphoribosyl transferase (HGPRT) as well as their transcript expression were quantified under normoxic and hypoxic conditions. Purine metabolite (hypoxanthine (HX), xanthine (X), uric acid (UA), inosine monophosphate (IMP), inosine, nicotinamide adenine dinucleotide (NAD+), adenosine, adenosine monophosphate (AMP), adenosine diphosphate (ADP), adenosine triphosphate (ATP), guanosine diphosphate (GDP) and guanosine triphosphate (GTP)) concentrations were also quantified. Significant reduction of IMPDH activity and HX and IMP concentrations (p &lt; 0.05) were observed after hypoxia, suggesting a decrease of de novo synthesis of purines. After hypoxia a global reduction of transcripts was observed, suggesting a reduction of the metabolic machinery of purine metabolism to new steady states that balance ATP demand and ATP supply pathways.

Acute Capsaicin and Exercise Performance in Humans: Potential Neuromuscular Mechanisms

The pungent bioactive ingredient in peppers (Capsaicin; CAP) has been widely investigated for its activation of the transient receptor potential vanilloid 1 (TRPV1) channel on sensory neurons. It has been shown to improve mitochondrial biogenesis, adenosine triphosphate (ATP) production and vascular function. In animal models has been demonstrated that increases exercise performance and decreases fatigue development. However, whether acute oral CAP improves endurance performance and the potential underlying mechanisms remains elusive. This study aimed to evaluate the potential ergogenic and anti-fatigue effects of acute oral CAP supplementation in healthy humans. In a double-blind, placebo-controlled, cross-over design, 10 young healthy males performed constant-load cycling exercise (294±55W; 85% of peak power output) to exhaustion, after acute ingestion of placebo (PL; 1000mg fiber pill) and CAP (780mg pill) capsules. Fatigue was assessed on the dominant quadriceps with the interpolated twitch technique, via changes in supramaximal electrical stimulation pre- vs. post-exercise, during isometric maximal voluntary contractions (MVC) and at rest. Cardiopulmonary parameters (heart rate, cardiac output, oxygen consumption, ventilation) were assessed throughout the exercise. No statistically significant differences were detected in the muscle contractile functions, despite a trend (P=0.07) for a decreased time to exhaustion during CAP (PL: 369±85 vs. CAP: 296±64 sec). All the values (except muscle voluntary activation (%VMA)) significantly decreased from pre- to post-exercise in both conditions. The exercise-induced attenuation of the intrinsic muscle contractile properties (Maximal rate of force development (MRFD) and Maximal relaxation rate (MRR) of the resting twitch force) were mitigated, in part, by CAP. Notably, in the PL condition MRR was seemingly reduced by 47±33%, while only of 29±68% in CAP; contrarily, MRFD decreased similarly of 47±23% and of 40±27% in PL and CAP, respectively. In 9 participants, the resting twitch (estimate of peripheral muscle fatigue) pre- vs. post-exercise decreased less with CAP (PL: 44±17 vs. CAP: 37±13%). CAP did not influence the MVC nor %VMA. The cardiopulmonary responses were similar between the conditions (P>0.05). In contrast to prior work, acute oral capsaicin did not improve endurance performance in the present study. CAP did elicit a tendency for faster relaxation rate, which might suggest an enhanced activation of the sarcoendoplasmic reticulum calcium ATPase (SERCA) pumps, thereby increasing the rate of Ca2+ reuptake and relaxation. However, this altered Ca2+ handling might also increase ATP demand. The tendency for CAP to attenuate the exercise-induced reduction in muscle function, or fatigue, might provide a mechanistic basis for the previously demonstrated beneficial effects of CAP. Further work is needed in larger samples, to further determine the optimal dosing and loading strategies, to better understand the potential ergogenic effects of capsaicin.

Contrasting strategies of hypoxic cardiac performance and metabolism in cichlids and armoured catfish.

The heart of tropical fishes is a particularly useful model system in which to investigate mechanisms of hypoxic tolerance. Here we focus on insights gained from two groups of fishes, cichlids and armoured catfishes. Cichlids respond to hypoxia by entering a sustained hypometabolism with decreased heart performance to match whole animal circulatory needs. Heart rate is decreased along with protein turnover to reduce adenosine triphosphate demand. This occurs despite the inherent capacity for high levels of cardiac power development. Although highly hypoxic tolerant at the whole animal level, the heart of cichlids does not have high constitutive activities of glycolytic enzymes compared to other species. Information is conflicting with respect to changes in glycolytic gene expression and enzyme activity following hypoxic exposure with some studies showing increases and others decreases. In contrast to cichlids, species of armoured catfish, that are routinely exposed to water of low oxygen content, do not display hypoxic bradycardia. Under hypoxia there are early changes in glucose trafficking suggestive of activation of glycolysis before lactate accumulation. Thereafter, heart glycogen is mobilized and lactate accumulates in both heart and blood, in some species to very high levels. Heart performance under hypoxia is enhanced by defense of intracellular pH. A functional sarcoplasmic reticulum and binding of hexokinase to the outer mitochondrial membrane may also play a role in cardioprotection. Maintenance of heart performance under hypoxia may relate to a tradeoff between air breathing via a modified stomach and circulatory demands for digestion.

Improvement of Oxidative Stress and Mitochondrial Dysfunction by β-Caryophyllene: A Focus on the Nervous System.

Mitochondrial dysfunction results in a series of defective cellular events, including decreased adenosine triphosphate (ATP) production, enhanced reactive oxygen species (ROS) output, and altered proteastasis and cellular quality control. An enhanced output of ROS may damage mitochondrial components, such as mitochondrial DNA and elements of the electron transport chain, resulting in the loss of proper electrochemical gradient across the mitochondrial inner membrane and an ensuing shutdown of mitochondrial energy production. Neurons have an increased demand for ATP and oxygen, and thus are more prone to damage induced by mitochondrial dysfunction. Mitochondrial dysfunction, damaged electron transport chains, altered membrane permeability and Ca2+ homeostasis, and impaired mitochondrial defense systems induced by oxidative stress, are pathological changes involved in neurodegenerative disorders. A growing body of evidence suggests that the use of antioxidants could stabilize mitochondria and thus may be suitable for preventing neuronal loss. Numerous natural products exhibit the potential to counter oxidative stress and mitochondrial dysfunction; however, science is still looking for a breakthrough in the treatment of neurodegenerative disorders. β-caryophyllene is a bicyclic sesquiterpene, and an active principle of essential oils derived from a large number of spices and food plants. As a selective cannabinoid receptor 2 (CB2) agonist, several studies have reported it as possessing numerous pharmacological activities such as antibacterial (e.g., Helicobacter pylori), antioxidant, anti-inflammatory, analgesic (e.g., neuropathic pain), anti-neurodegenerative and anticancer properties. The present review mainly focuses on the potential of β-caryophyllene in reducing oxidative stress and mitochondrial dysfunction, and its possible links with neuroprotection.

Frequency-Dependent Energy Demand of Dendritic Responses to Deep Brain Stimulation in Thalamic Neurons: A Model-Based Study.

Thalamic deep brain stimulation (DBS) generates excitatory postsynaptic currents and action potentials (APs) by triggering large numbers of synaptic inputs to local cells, which also activates axonal spikes to antidromically invade the soma and dendrites. To maintain signaling, the evoked dendritic responses require metabolic energy to restore ion gradients in each dendrite. The objective of this study is to estimate the energy demand associated with dendritic responses to thalamic DBS. We use a morphologically realistic computational model to simulate dendritic activity in thalamocortical (TC) relay neurons with axonal intracellular stimulation or DBS-like extracellular stimulation. We determine the metabolic cost by calculating the number of adenosine triphosphate (ATP) expended to pump Na+ and Ca2+ ions out of each dendrite. The ATP demand of dendritic activity exhibits frequency dependence, which is determined by the number of spikes in the dendrites. Each backpropagating AP from the soma activates a spike in the dendrites, and the dendritic firing is dominated by antidromic activation of the soma. High stimulus frequencies decrease dendritic ATP cost by reducing the fidelity of antidromic activation. Synaptic inputs and stimulus-induced polarization govern the ATP cost of dendritic responses by facilitating/suppressing antidromic activation, which also influences the ATP cost by depolarizing/hyperpolarizing each dendrite. These findings are important for understanding the synaptic signaling energy in TC relay neurons and metabolism-dependent functional imaging data of thalamic DBS.

A Novel Method to Synthesize Phosphocreatine and Phosphocreatine Prodrugs.

Adenosine triphosphate (ATP) is the energy currency of the body; it takes part in various and indispensable metabolic processes for the maintenance of cell homeostasis, degrading to its hydrolysis product, adenosine diphosphate (ADP). Efficient ways to restore ATP are therefore necessary in the cells. When the cell lacks energy due to ischemic conditions or high ATP demand, phosphocreatine gives its phosphate group to ADP that converts to ATP, in a reaction catalyzed by the enzyme creatine kinase. For this reason, phosphocreatine is utilized as a pharmacological treatment in human diseases that involve a failure of the cellular energy, most notably in coronary artery disease. Commercially available phosphocreatine is currently synthesized using different methods, each of one characterized by a rather low yield of the final product, probably due to the low reactivity of the guanylating reagent. The aim of this work is to overcome the drawbacks of the synthetic methods currently employed, devising a novel synthetic route to obtain phosphocreatine and phosphocreatine prodrugs in higher yields and purity. To obtain a higher yield of the final product and a lower number of sub-products, this method utilizes a new guanylating agent characterized by high reactivity, endowed with a protecting group t-Boc on one of the two nitrogen atoms of the guanidinic function and a protected phosphate on the other one; that compound is then conjugated with an opportune secondary amine. The obtained product is cleaved first with acidic conditions to obtain the phosphocreatine prodrug (phosphocreatine ethyl ester) and then with an enzymatic method to obtain the phosphocreatine. Have been obtained in good yield and purity as demonstrated by HPLC and mass spectrometry analysis. This novel synthetic route permits to obtain the phosphocreatine molecule in higher yield and purity compared to the methods currently employed with a combination of chemical and enzymatic methods.

Osteoblast-like MC3T3-E1 Cells Prefer Glycolysis for ATP Production but Adipocyte-like 3T3-L1 Cells Prefer Oxidative Phosphorylation.

Mesenchymal stromal cells (MSCs) are early progenitors that can differentiate into osteoblasts, chondrocytes, and adipocytes. We hypothesized that osteoblasts and adipocytes utilize distinct bioenergetic pathways during MSC differentiation. To test this hypothesis, we compared the bioenergetic profiles of preosteoblast MC3T3-E1 cells and calvarial osteoblasts with preadipocyte 3T3L1 cells, before and after differentiation. Differentiated MC3T3-E1 osteoblasts met adenosine triphosphate (ATP) demand mainly by glycolysis with minimal reserve glycolytic capacity, whereas nondifferentiated cells generated ATP through oxidative phosphorylation. A marked Crabtree effect (acute suppression of respiration by addition of glucose, observed in both MC3T3-E1 and calvarial osteoblasts) and smaller mitochondrial membrane potential in the differentiated osteoblasts, particularly those incubated at high glucose concentrations, indicated a suppression of oxidative phosphorylation compared with nondifferentiated osteoblasts. In contrast, both nondifferentiated and differentiated 3T3-L1 adipocytes met ATP demand primarily by oxidative phosphorylation despite a large unused reserve glycolytic capacity. In sum, we show that nondifferentiated precursor cells prefer to use oxidative phosphorylation to generate ATP; when they differentiate to osteoblasts, they gain a strong preference for glycolytic ATP generation, but when they differentiate to adipocytes, they retain the strong preference for oxidative phosphorylation. Unique metabolic programming in mesenchymal progenitor cells may influence cell fate and ultimately determine the degree of bone formation and/or the development of marrow adiposity. © 2018 American Society for Bone and Mineral Research.

Event-Associated Oxygen Consumption Rate Increases ca. Five-Fold When Interictal Activity Transforms into Seizure-Like Events In Vitro.

Neuronal injury due to seizures may result from a mismatch of energy demand and adenosine triphosphate (ATP) synthesis. However, ATP demand and oxygen consumption rates have not been accurately determined, yet, for different patterns of epileptic activity, such as interictal and ictal events. We studied interictal-like and seizure-like epileptiform activity induced by the GABAA antagonist bicuculline alone, and with co-application of the M-current blocker XE-991, in rat hippocampal slices. Metabolic changes were investigated based on recording partial oxygen pressure, extracellular potassium concentration, and intracellular flavine adenine dinucleotide (FAD) redox potential. Recorded data were used to calculate oxygen consumption and relative ATP consumption rates, cellular ATP depletion, and changes in FAD/FADH2 ratio by applying a reactive-diffusion and a two compartment metabolic model. Oxygen-consumption rates were ca. five times higher during seizure activity than interictal activity. Additionally, ATP consumption was higher during seizure activity (~94% above control) than interictal activity (~15% above control). Modeling of FAD transients based on partial pressure of oxygen recordings confirmed increased energy demand during both seizure and interictal activity and predicted actual FAD autofluorescence recordings, thereby validating the model. Quantifying metabolic alterations during epileptiform activity has translational relevance as it may help to understand the contribution of energy supply and demand mismatches to seizure-induced injury.

MRS Evidence of Adequate O₂ Supply in Human Skeletal Muscle at the Onset of Exercise.

At exercise onset, intramuscular oxidative energy production responds relatively slowly in comparison with the change in adenosine triphosphate demand. To determine whether the slow kinetics of oxidative adenosine triphosphate production is due to inadequate O2 supply or metabolic inertia, we studied the kinetics of intramyocellular deoxygenation (deoxy-myoglobin (Mb)) and metabolism (phosphocreatine (PCr)) using proton (1H) and phosphorus (31P) magnetic resonance spectroscopy in six healthy subjects (33 ± 5 yr). Specifically, using dynamic plantarflexion exercise, rest to exercise and recovery were assessed at both 60% of maximum work rate (moderate intensity) and 80% of maximum work rate (heavy intensity). At exercise onset, [PCr] fell without delay and with a similar time constant (τ) at both exercise intensities (approximately 33 s). In contrast, the increase in deoxy-Mb was delayed at exercise onset by 5-7 s, after which it increased with kinetics (moderate τ = 37 ± 9 s; heavy τ = 29 ± 6 s) that was not different from τPCr (P > 0.05). At cessation, deoxy-Mb recovered without time delay and more rapidly (τ = ∼20 s) than PCr (τ = ∼33 s) (P < 0.05). Using a unique combination of in vivo magnetic resonance spectroscopy techniques with high time resolution, this study revealed a delay in intramuscular deoxygenation at the onset of exercise and rapid reoxygenation kinetics upon cessation. Together, these data imply that intramuscular substrate-enzyme interactions, and not O2 availability, determine the exercise onset kinetics of oxidative metabolism in healthy human skeletal muscles.

Control of glycolytic flux in directed biosynthesis of uridine-phosphoryl compounds through the manipulation of ATP availability

Adenosine triphosphate (ATP), the most important energy source for metabolic reactions and pathways, plays a vital role in control of metabolic flux. Considering the importance of ATP in regulation of the glycolytic pathway, the use of ATP-oriented manipulation is a rational and efficient route to regulate metabolic flux. In this paper, a series of efficient ATP-oriented regulation methods, such as changing ambient temperature and altering reduced nicotinamide adenine dinucleotide (NADH), was developed. To satisfy the different demand for ATP at different phases in directed biosynthesis of uridine-phosphoryl compounds, a multiphase ATP supply regulation strategy was also used to enhance to yield of target metabolites.

Hypoxic conformance of metabolism in primary rat hepatocytes: A model of hepatic hibernation

Following prolonged hypoxia, mammalian cells invoke adaptive mechanisms to enhance oxygen delivery and promote energy conservation. We previously reported that hepatocytes subjected to prolonged moderate hypoxia (PO2 = 20-50 mmHg for > 3 hours) demonstrated a reversible inhibition of cellular respiration with maintenance of cell viability, associated with a decrease in mitochondrial adenosine triphosphate (ATP) synthesis; acute hypoxia (similar PO2 for < 30 minutes) did not induce a similar suppression of respiration and ATP synthesis. In the current study, using an in vitro model of primary rat hepatocytes, we measured the changes in metabolic demand for ATP during hypoxic conformance, and tested whether viability is maintained by preferentially suppressing nonessential processes while sustaining processes essential for maintaining cell homeostasis. In addition, the rate of recovery of oxygen consumption and ATP concentrations following reoxygenation after prolonged and acute hypoxia/anoxia was compared. Oxygen consumption and ATP concentrations decreased during prolonged hypoxia compared with acute hypoxia. However, ouabain-inhibitable respiration did not decrease during prolonged hypoxia, indicating that membrane Na+/K+ ATPase activity, an essential process for cell viability, was maintained. In contrast, ATP-dependent glucuronidation and sulfation of acetaminophen, deemed "non-essential" processes, were decreased significantly compared with normoxic cells. After reoxygenation, cells exposed to prolonged moderate hypoxia demonstrated a more rapid recovery of respiration compared to acute hypoxia/anoxia. This "hepatic hibernation" during prolonged moderate hypoxia may represent an anticipatory adaptation that seeks to maintain cell viability while delaying or preventing the onset of lethal hypoxia, and facilitates rapid recovery after the resumption of normoxia.

Control of maximum metabolic rate in humans: dependence on performance phenotypes.

Borrowing from metabolic control analysis the concept of control coefficients or ci values, defined as fractional change in MMR/fractional change in the capacity of any given step in ATP turnover, we used four performance phenotypes to compare mechanisms of control of aerobic maximum metabolic rate (MMR): (i) untrained sedentary (US) subjects, as a reference group against which to compare (ii) power trained (PT), (iii) endurance trained (ET), and (iv) high altitude adapted native (HA) subject groups. Sprinters represented the PT group; long distance runners illustrated the ET group; and Andean natives represented the HA group. Numerous recent studies have identified contributors to control on both the adenosine triphosphate (ATP) supply side and the ATP demand side of ATP turnover. From the best available evidence it appears that at MMR all five of the major steps in energy delivery (namely, ventilation, pulmonary diffusion, cardiac output, tissue capillary--mitochondrial O2 transfer, and aerobic cell metabolism per se) approach an upper functional ceiling, with control strength being distributed amongst the various O2 flux steps. On the energy demand side, the situation is somewhat simplified since at MMR approximately 90% of O2-based ATP synthesis is used for actomyosin (AM) and Ca2+ ATPases; at MMR these two ATP demand rates also appear to be near an upper functional ceiling. In consequence, at MMR the control contributions or ci values are distributed amongst all seven major steps in ATP supply and ATP demand pathways right to the point of fatigue. Relative to US (the reference group), in PT subjects at MMR control strength shifts towards O2 delivery steps (ventilation, pulmonary diffusion, and cardiac output); here physiological regulation clearly dominates MMR control. In contrast in ET and HA subjects at MMR control shifts towards the energy demand steps (AM and Ca2+ ATPases), and more control strength is focussed on tissue level ATP supply and ATP demand. One obvious advantage of the ET and HA biochemical-level control is improved metabolite homeostasis. Additionally, with some reserve capacity in the O2 delivery steps, the focussing of control on ATP turnover at the tissue level has allowed nature to improve on an 'endurance machine' design.

Can any metabolites partially alleviate fatigue manifestations at the cross-bridge?

During exercise, intracellular homeostasis depends on the matching of adenosine triphosphate (ATP) supply and ATP demand. Metabolites play a useful role in communicating the extent of ATP demand to the metabolic supply pathways. During fatigue from high-intensity exercise, a major change in the intracellular milieu of skeletal muscle is not ATP depletion but metabolite accumulation that affects the actomyosin cross-bridge interaction. The resulting reduction in myosin ATPase activity, cross-bridge turnover rate, and velocity of contraction can be considered a useful downregulation of ATP demand. Although maximal force is reduced, it is reduced less than myosin ATPase activity. In combination, efficiency of force production at the cross-bridge is thus enhanced. This is a second useful role for metabolites during fatigue because the total ATP cost per unit of force is partially reduced. Theoretical models predict that ADP may alleviate some effects of fatigue by further enhancing cross-bridge efficiency, thus providing a third useful role for metabolite accumulation. Recent experimental evidence reviewed here suggests that this occurs when ATP concentration is dramatically reduced. Single-fiber chemical analyses of fatigued muscle show lower ATP concentrations than other methods, but whether the appropriate microenvironments for effective competition by ADP for the nucleotide binding site occur around some or all of the cross-bridges remains technically difficult to prove at this time. During fatigue, muscle activation is also decreased, a response that potentially has the greatest effect on ATP demand-supply matching. I propose that the mismatch between the expected force production relative to muscle activation and the reduced force production from inorganic phosphate accumulation is the peripheral signal for reduced activation and is therefore the fourth useful role of metabolites in alleviating fatigue.