Depletion Of High-energy Phosphates Research Articles

Time Course of High-Energy Phosphate Depletion During Cold Storage of Human Heart Grafts Using the Celsior Solution.

The aim of this study was to provide insight into high-energy phosphate compound concentration dynamics under realistic clinical cold-storage conditions using the Celsior solution in seven heart grafts discarded from transplantation. The hearts of seven local donors (three males, four females, age 37 ± 17years, height 175 ± 5cm, weight 75 ± 9kg) initially considered for transplantation and eventually discarded were submitted to a Magnetic Resonance Spectroscopy observation in a clinical Magnetic Resonance Imaging scanner over at least 9h. The grafts remained in their sterile container at 4°C during the entire examination. Hence, Phosphocreatine (PCr), adenosine triphosphate (ATP), inorganic phosphate (Pi) and intracellular pH were recorded non-destructively at a 30-minute interval. With the ischemic time Ti, the concentration ratios decreased at PCr/ATP = 1.68-0.0028·Tis, Pi/ATP = 1.38 + 0.0029·Tis, and intracellular pH at 7.43-0.0012·Tis. ATP concentration remained stable for at least 9h and did not decrease as long as phosphocreatine was detectable. Acidosis remained moderate. In addition to the standard parameters assessed at the time of retrieval, Magnetic Resonance Spectroscopy can provide an assesment of the metabolic status of heart grafts before transplantation. These results show how HEPC metabolites deplete during cold storage. Although many parameters determine graft quality during cold storage, the dynamics of HEPC and intracellular pH may be helpful in the development of strategies aiming at extending the ischemic time.

Status of Mitochondrial Oxidative Phosphorylation during the Development of Heart Failure.

Mitochondria are specialized organelles, which serve as the "Power House" to generate energy for maintaining heart function. These organelles contain various enzymes for the oxidation of different substrates as well as the electron transport chain in the form of Complexes I to V for producing ATP through the process of oxidative phosphorylation (OXPHOS). Several studies have shown depressed OXPHOS activity due to defects in one or more components of the substrate oxidation and electron transport systems which leads to the depletion of myocardial high-energy phosphates (both creatine phosphate and ATP). Such changes in the mitochondria appear to be due to the development of oxidative stress, inflammation, and Ca2+-handling abnormalities in the failing heart. Although some investigations have failed to detect any changes in the OXPHOS activity in the failing heart, such results appear to be due to a loss of Ca2+ during the mitochondrial isolation procedure. There is ample evidence to suggest that mitochondrial Ca2+-overload occurs, which is associated with impaired mitochondrial OXPHOS activity in the failing heart. The depression in mitochondrial OXPHOS activity may also be due to the increased level of reactive oxygen species, which are formed as a consequence of defects in the electron transport complexes in the failing heart. Various metabolic interventions which promote the generation of ATP have been reported to be beneficial for the therapy of heart failure. Accordingly, it is suggested that depression in mitochondrial OXPHOS activity plays an important role in the development of heart failure.

Skeletal muscle abnormalities in heart failure with preserved ejection fraction.

Almost half of all heart failure (HF) disease burden is due to HF with preserved ejection fraction (HFpEF). The primary symptom in patients with HFpEF, even when well compensated, is severe exercise intolerance and is associated with their reduced quality of life. Recently, studies showed that HFpEF patients have multiple skeletal muscle (SM) abnormalities, and these are associated with decreased exercise intolerance. The SM abnormalities are likely intrinsic to the HFpEF syndrome, not a secondary consequence of an epiphenomenon. These abnormalities are decreased muscle mass, reduced type I (oxidative) muscle fibers, and reduced type I-to-type II fiber ratio as well as a reduced capillary-to-fiber ratio, abnormal fat infiltration into the thigh SM, increased levels of atrophy genes and proteins, reduction in mitochondrial content, and rapid depletion of high-energy phosphate during exercise with markedly delayed repletion of high-energy phosphate during recovery in mitochondria. In addition, patients with HFpEF have impaired nitric oxide bioavailability, particularly in the microvasculature. These SM abnormalities may be responsible for impaired diffusive oxygen transport and/or impaired SM oxygen extraction. To date, exercise training (ET) and caloric restriction are some of the interventions shown to improve outcomes in HFpEF patients. Improvements in exercise tolerance following aerobic ET are largely mediated through peripheral SM adaptations with minimal change in central hemodynamics and highlight the importance of targeting SM to improve exercise intolerance in HFpEF. Focusing on the abnormalities mentioned above may improve the clinical condition of patients with HFpEF.

One and one is not always two: hypo- and hyper-additive effects of the chemoreflex during exercise.

One and one is not always two: hypo- and hyper-additive effects of the chemoreflex during exercise.

The Respiratory Compensation Point and the Deoxygenation Break Point Are Not Valid Surrogates for Critical Power and Maximum Lactate Steady State.

The Respiratory Compensation Point and the Deoxygenation Break Point Are Not Valid Surrogates for Critical Power and Maximum Lactate Steady State.

The Respiratory Compensation Point and the Deoxygenation Break Point Are Valid Surrogates for Critical Power and Maximum Lactate Steady State.

The Respiratory Compensation Point and the Deoxygenation Break Point Are Valid Surrogates for Critical Power and Maximum Lactate Steady State.

Magnesium induces preconditioning of the neonatal brain via profound mitochondrial protection

Magnesium sulphate (MgSO4) given to women in preterm labor reduces cerebral palsy in their offspring but the mechanism behind this protection is unclear, limiting its effective, safe clinical implementation. Previous studies suggest that MgSO4 is not neuroprotective if administered during or after the insult, so we hypothesised that MgSO4 induces preconditioning in the immature brain. Therefore, we administered MgSO4 at various time-points before/after unilateral hypoxia-ischemia (HI) in seven-day-old rats. We found that MgSO4 treatment administered as a bolus between 6 days and 12 h prior to HI markedly reduced the brain injury, with maximal protection achieved by 1.1 mg/g MgSO4 administered 24 h before HI. As serum magnesium levels returned to baseline before the induction of HI, we ascribed this reduction in brain injury to preconditioning. Cerebral blood flow was unaffected, but mRNAs/miRNAs involved in mitochondrial function and metabolism were modulated by MgSO4. Metabolomic analysis (H+-NMR) disclosed that MgSO4 attenuated HI-induced increases in succinate and prevented depletion of high-energy phosphates. MgSO4 pretreatment preserved mitochondrial respiration, reducing ROS production and inflammation after HI. Therefore, we propose that MgSO4 evokes preconditioning via induction of mitochondrial resistance and attenuation of inflammation.

Fatigability, Exercise Intolerance, and Abnormal Skeletal Muscle Energetics in Heart Failure.

Among central and peripheral factors contributing to exercise intolerance (EI) in heart failure (HF), the extent to which skeletal muscle (SM) energy metabolic abnormalities occur and contribute to EI and increased fatigability in HF patients with reduced or preserved ejection fraction (HFrEF and HFpEF, respectively) are not known. An energetic plantar flexion exercise fatigability test and magnetic resonance spectroscopy were used to probe the mechanistic in vivo relationships among SM high-energy phosphate concentrations, mitochondrial function, and EI in HFrEF and HFpEF patients and in healthy controls. Resting SM high-energy phosphate concentrations and ATP flux rates were normal in HFrEF and HFpEF patients. Fatigue occurred at similar SM energetic levels in all subjects, consistent with a common SM energetic limit. Importantly, HFrEF New York Heart Association class II-III patients with EI and high fatigability exhibited significantly faster rates of exercise-induced high-energy phosphate decline than did HFrEF patients with low fatigability (New York Heart Association class I), despite similar left ventricular ejection fractions. HFpEF patients exhibited severe EI, the most rapid rates of high-energy phosphate depletion during exercise, and impaired maximal oxidative capacity. Symptomatic fatigue during plantar flexion exercise occurs at a common energetic limit in all subjects. HFrEF and HFpEF patients with EI and increased fatigability manifest early, rapid exercise-induced declines in SM high-energy phosphates and reduced oxidative capacity compared with healthy and low-fatigability HF patients, suggesting that SM metabolism is a potentially important target for future HF treatment strategies.

Acetylation: A potential “regulating valve” of cardiac energy metabolism during atrial fibrillation

Atrial fibrillation (AF), the most common sustained cardiac arrhythmia, has a close pathophysiological association with energy metabolism [1,2]. Impaired atrial myocardial energy status has been demonstrated in atrial tissue of animal AF models and AF patients [2–4]. The performances of impaired energy status related to reduction of adenine nucleotides, a depletion of high energy phosphates, reduced activities of energy metabolism enzymes, decreasing in the cellular energy potential and alteration of regulator in mitochondrial oxidative metabolism [2–4].

BGP-15, a PARP-inhibitor, prevents imatinib-induced cardiotoxicity by activating Akt and suppressing JNK and p38 MAP kinases

In this study, we investigate the cardiotoxic effects of the well-known cytostatic agent imatinib mesylate (Gleevec), and presented evidence for the cardioprotective effect of BGP-15 which is a novel insulin sensitizer. The cardiotoxic effect of imatinib mesylate was assessed in Langendorff rat heart perfusion system. The cardiac high-energy phosphate levels (creatine phosphate (PCr) and ATP) were monitored in situ by (31)P NMR spectroscopy. The protein oxidation, lipid peroxidation, and the activation of signaling pathways were determined from the freeze-clamped hearts. Prolonged treatment of the heart with imatinib mesylate (20 mg/kg) resulted in cardiotoxicity, which were characterized by the depletion of high-energy phosphates (PCr and ATP), and significantly increased protein oxidation and lipid peroxidation. Imatinib mesylate treatment-induced activation of MAP kinases (including ERK1/2, p38, and JNK) and the phosphorylation of Akt and GSK-3beta. BGP-15 (200 μM) prevented the imatinib mesylate-induced oxidative damages, attenuated the depletion of high-energy phosphates, altered the signaling effect of imatinib mesylate by preventing p38 MAP kinase and JNK activation, and induced the phosphorylation of Akt and GSK-3beta. The suppressive effect of BGP-15 on p38 and JNK activation could be significant because these kinases contribute to the cell death and inflammation in the isolated perfused heart.

Protection of DFP-induced oxidative damage and neurodegeneration by antioxidants and NMDA receptor antagonist

Prophylactic agents acutely administered in response to anticholinesterases intoxication can prevent toxic symptoms, including fasciculations, seizures, convulsions and death. However, anticholinesterases also have long-term unknown pathophysiological effects, making rational prophylaxis/treatment problematic. Increasing evidence suggests that in addition to excessive cholinergic stimulation, organophosphate compounds such as diisopropylphosphorofluoridate (DFP) induce activation of glutamatergic neurons, generation of reactive oxygen (ROS) and nitrogen species (RNS), leading to neurodegeneration. The present study investigated multiple affectors of DFP exposure critical to cerebral oxidative damage and whether antioxidants and NMDA receptor antagonist memantine provide neuroprotection by preventing DFP-induced biochemical and morphometric changes in rat brain. Rats treated acutely with DFP (1.25 mg/kg, s.c.) developed onset of toxicity signs within 7–15 min that progressed to maximal severity of seizures and fasciculations within 60 min. At this time point, DFP caused significant ( p < 0.01) increases in biomarkers of ROS (F 2-isoprostanes, F 2-IsoPs; and F 4-neuroprostanes, F 4-NeuroPs), RNS (citrulline), and declines in high-energy phosphates (HEP) in rat cerebrum. At the same time, quantitative morphometric analysis of pyramidal neurons of the hippocampal CA1 region revealed significant ( p < 0.01) reductions in dendritic lengths and spine density. When rats were pretreated with the antioxidants N- tert-butyl-α-phenylnitrone (PBN, 200 mg/kg, i.p.), or vitamin E (100 mg/kg, i.p./day for 3 days), or memantine (18 mg/kg, i.p.), significant attenuations in DFP-induced increases in F 2-IsoPs, F 4-NeuroPs, citrulline, and depletion of HEP were noted. Furthermore, attenuation in oxidative damage following antioxidants or memantine pretreatment was accompanied by rescue from dendritic degeneration of pyramidal neurons in the CA1 hippocampal area. These findings closely associated DFP-induced lipid peroxidation with dendritic degeneration of pyramidal neurons in the CA1 hippocampal area and point to possible interventions to limit oxidative injury and dendritic degeneration induced by anticholinesterase neurotoxicity.

Muscle metabolic responses to exercise above and below the “critical power” assessed using31P-MRS

We tested the hypothesis that the asymptote of the hyperbolic relationship between work rate and time to exhaustion during muscular exercise, the "critical power" (CP), represents the highest constant work rate that can be sustained without a progressive loss of homeostasis [as assessed using (31)P magnetic resonance spectroscopy (MRS) measurements of muscle metabolites]. Six healthy male subjects initially completed single-leg knee-extension exercise at three to four different constant work rates to the limit of tolerance (range 3-18 min) for estimation of the CP (mean +/- SD, 20 +/- 2 W). Subsequently, the subjects exercised at work rates 10% below CP (<CP) for 20 min and 10% above CP (>CP) for as long as possible, while the metabolic responses in the contracting quadriceps muscle, i.e., phosphorylcreatine concentration ([PCr]), P(i) concentration ([P(i)]), and pH, were estimated using (31)P-MRS. All subjects completed 20 min of <CP exercise without duress, whereas the limit of tolerance during >CP exercise was 14.7 +/- 7.1 min. During <CP exercise, stable values for [PCr], [P(i)], and pH were attained within 3 min after the onset of exercise, and there were no further significant changes in these variables (end-exercise values = 68 +/- 11% of baseline [PCr], 314 +/- 216% of baseline [P(i)], and pH 7.01 +/- 0.03). During >CP exercise, however, [PCr] continued to fall to the point of exhaustion and [P(i)] and pH changed precipitously to values that are typically observed at the termination of high-intensity exhaustive exercise (end-exercise values = 26 +/- 16% of baseline [PCr], 564 +/- 167% of baseline [P(i)], and pH 6.87 +/- 0.10, all P < 0.05 vs. <CP exercise). These data support the hypothesis that the CP represents the highest constant work rate that can be sustained without a progressive depletion of muscle high-energy phosphates and a rapid accumulation of metabolites (i.e., H(+) concentration and [P(i)]), which have been associated with the fatigue process.

Depletion of high energy phosphates implicates post-exercise mortality in carp and trout; an in vivo31P-NMR study

As in vivo 31P-Nuclear Magnetic Resonance spectroscopy is currently the state of the art method to measure continuously intracellular pH (pH i) and energy status of muscle tissue, we used this method to study the recovery from exhaustive exercise. The biochemical changes during recovery are not well understood and it was suggested that post-exercise mortality could be caused by low pH i; other studies however indicate that energy depletion might be more important. To analyse the mechanism of post-exercise recovery pH i, ATP, P i, and PCr must be measured at the same time, which is possible using in vivo 31P-NMR. Common carp and rainbow trout of about 100 g were exercised to exhaustion in a swim tunnel. After swimming 10 h at 1.5 body lengths (BL)/s (aerobic control), 50% of the fish were forced to swim at 6 BL/s until exhaustion. Recovery of energy rich phosphates was found to be faster in carp (1.2–1.9 h) than in trout (1.5–2.3 h). The same applied for the recovery from acidosis, which took 1.75 h in carp and 5.75 h in trout. In parallel experiments the energy phosphates and lactate levels were measured in liver, red muscle, and white muscle. Exhaustion caused a significant drop in the energy status of red and white muscle tissue of trout and carp (corroborates NMR data), while no change at all was observed in liver tissue. The lactate levels were increased in the muscle but not in liver and blood. While all experimental animals looked healthy after exhaustion, 40–50% of the carp as well as trout died during the recovery phase. The energy status of those individuals measured by 31P-NMR was much lower than that of the survivors, while in contrast there was no difference in pH i. Thus, it appears that not acidosis but depletion of high energy phosphates disabled muscle function and therefore may have been the cause of death of the non-survivors.

Abstract 1571: Hibernating Myocardium has Reduced High Energy Phosphate Levels and a Preconditioning-Like Effect with Slower ATP Depletion in Response to Acute Ischemia

Background. Pigs with a chronic LAD stenosis develop hibernating myocardium with contractile dysfunction and reduced resting flow. We hypothesized that this reduced energy utilization preserves ATP and protects the heart from irreversible injury during acute ischemia. Methods. Pigs with hibernating myocardium were studied 3-months after instrumentation with a 1.5 mm proximal LAD stenosis (n=7). Hibernating myocardium was confirmed by reduced LAD wall thickening (2.4±0.2 vs.6.1±0.6 mm in remote, p&lt;0.05) and reduced resting flow (0.95±0.15 vs. 1.66±0.29 ml/min/g in remote, p&lt;0.05) without infarction. Subendocardial samples were rapidly excised from propofol anesthetized pigs and serial depletion of high energy phosphates quantified by HPLC (in μmol/g dry weight) during simulated total ischemia in vitro (37°C). Results. At Baseline, ATP and ADP were significantly reduced in the hibernating LAD region in comparison to controls (n=8, Table ), with preserved CP/ATP and ATP/ADP ratios. During simulated ischemia, hibernating myocardium displayed a markedly reduced rate of ATP depletion (ΔATP), with ATP levels at 20 min significantly higher than control (Table ). Higher ATP levels were maintained throughout 80 min of ischemia. Interestingly, identical preservation of high energy phosphates occurred in the remote normally perfused region of hearts with hibernating myocardium (Table ). Conclusion. These data indicate that there is a balanced reduction in high energy phosphate levels in hibernating myocardium with chronic protection from ischemia manifested by a slower rate of ATP depletion during acute ischemia. This preconditioning-like effect is global suggesting that it arises from stimuli that are not directly related to ischemia. This raises the possibility that stretch from cyclical elevations in LV filling pressure or a circulating factor released from the heart in response to ischemia can protect the heart in chronic coronary artery disease.

Abstract 2626: Intramyocardial High-Energy Phosphate Depletion during Ventricular Fibrillation Correlates with Electrocardiographic Change

Background: Ventricular fibrillation (VF) is highly energy-wasteful, producing no perfusion. High-energy phosphates (HEPs) deplete rapidly during VF, adding to metabolic/ischemic stress and adversely affecting VF. We have shown that the scaling exponent (ScE) of the VF waveform predicts rescue shock outcome and can be used to guide decision-making during CPR. Hypotheses: We hypothesized that intramyocardial HEPs would rapidly diminish during untreated VF, and that this decrease would correlate with changes in the ScE. Methods: We anesthetized and instrumented 18 domestic swine (26.6 ± 1.9 kgs). We systematically assigned them (6 per group) to 5, 10, or 15 minutes of untreated VF, which was induced with a transthoracic shock. At the predetermined timepoint open-chest biopsies were taken from the left ventricle. Biopsied tissue samples were immediately flash frozen in a slurry of dry ice and isopentane (-90 o C), and HEPs were extracted with 1% trichloroacetic acid. Levels of ATP and ADP were measured luminometrically with an ADP/ATP ratio kit. ATP and ADP levels were assayed in duplicate, and the average recorded (reported in nmol/mg myocardial tissue). Fifteen seconds of ECG signal was digitally recorded (for ScE analysis) immediately prior to biopsy. ScE values were paired with ATP and ADP for calculation of linear regression. HEPs vs. time were compared with ANOVA. Results: Mean ATP/ADP were 1.20/0.21 at 5 min, 0.94/0.12 at 10 min, and 0.45/0.11 at 15 min. ATP correlated with ScE (r=.63, p=0.009) but ADP did not (r=.35, p=0.17). Both ATP (p=0.004) and ADP (p=0.024) differed over time. Conclusions: HEPs rapidly diminish during untreated VF. ATP depletion correlates with ScE change.

Localized therapeutic hypothermia in the brain for the treatment of ischemic stroke

stroke is, according to the World Health Organization ([10][1]), the third most common cause of death in the developed countries, exceeded only by coronary heart disease and cancer. Annually, 15 million people worldwide suffer a stroke. Of these 15 million people, 5.5 million die and 5 million are

Viewing a Stressful Episode of ER

See related article, pages 1186–1193 How cardiac cells sense, respond, and adapt to acute and chronic changes of their metabolic and environmental milieu remain among the most enigmatic and fundamental questions in contemporary cardiovascular biology and medicine. With more than 700 000 deaths and 6 million hospital discharges annually in the U.S., ischemic heart disease remains a major public health problem. If prompt therapy is delayed, myocardial injury from depletion of high-energy phosphates from inhibition of oxidative phosphorylation is inevitable. Ischemia/reperfusion (I/R) also unleashes a cascade of cellular and molecular events whose sequalae may sustain organ function at diminished capacity but, beyond a critical balance, threatens the organism’s survival. Reactive oxygen and nitrogen species (ROS; RNS), released from the mitochondria and other sources, alter the tertiary and quaternary structures of proteins, exposing their hydrophobic residues to allow conformational tendencies toward protein misfolding and aggregation.1 Nevertheless, highly sophisticated schemes equipped with molecular sensors, rapid response pathways, and adaptor mechanisms have evolved to mitigate the accumulation of unfolded proteins in a compartment specific manner, principally the cytoplasm and endoplasmic reticulum (ER). Although the cardiovascular field has paid considerably more attention to the cytoprotective mechanisms of heat shock proteins in response to I/R, termed the “classical” heat shock response, a parallel series of events has been unfolding at a breathtaking pace in the ER, with arguably considerable excitement to warrant a sneak preview of the article published in this issue of Circulation Research . To place into perspective the current buzz being garnered by the ER requires a brief discussion of more familiar themes in the field, which were developed thanks to the pioneering efforts of cardiovascular scientists, and others, working on the complementary stress response network, best exemplified by heat shock “stress” proteins and their transcriptional regulators. First discovered by …

Does blood flow restriction enhance hypertrophic signaling in skeletal muscle?

in the last decade, there was a dramatic increase in our knowledge of the molecular events that accompany changes in skeletal muscle mass in response to resistance exercise training or to chronic unloading. For example, the role of myostatin in limiting skeletal muscle growth during development was

AMP-Activated Protein Kinase and Type 2 Diabetes

AMP-activated protein kinase (AMPK) is an enzyme that works as a fuel gauge, being activated in situations of high-energy phosphate depletion. Upon activation, AMPK functions to restore cellular ATP by modifying diverse metabolic pathways. AMPK is activated robustly by skeletal muscle contraction and myocardial ischemia, and may be involved in the stimulation of glucose transport and fatty acid oxidation produced by these stimuli. In liver, activation of AMPK results in enhanced fatty acid oxidation and in decreased production of glucose, cholesterol, and triglycerides. Recent studies have shown that AMPK is the cellular mediator for many of the metabolic effects of drugs such as metformin and thiazolidinediones, as well as the insulin sensitizing adipocytokines leptin and adiponectin. These data, along with evidence from studies showing that chemical activation of AMPK in vivo with 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR) improves blood glucose concentrations and lipid profiles, make this enzyme an attractive pharmacological target for the treatment of type 2 diabetes and other metabolic disorders.

Metalloproteinase Inhibitor Counters High-Energy Phosphate Depletion and AMP Deaminase Activity Enhancing Ventricular Diastolic Compliance in Subacute Heart Failure

Cardiac matrix metalloproteinases (MMPs) stimulated by the sympathomimetic action of angiotensin II (AII) exacerbate chamber diastolic stiffening in models of subacute heart failure. Here we tested the hypothesis that MMP inhibition prevents such stiffening by favorably modulating high-energy phosphate (HEP) stores more than by effects on matrix remodeling. Dogs were administered AII i.v. for 1 week with tachypacing superimposed in the last two days (AII+P; n = 8). A second group (n = 9) underwent the same AII+P protocol but was preceded by oral treatment with an MMP inhibitor PD166793 [(S)-2-(4-bromo-biphenyl-4-sulfonylamino-3-methyl butyric acid] 1 week before and during the AII+P period. Pressure-volume analysis was performed in conscious animals, and myocardial tissue was subjected to in vitro and in situ zymography, collagen content, and HEP analysis (high-performance liquid chromatography). As reported previously, AII+P activated MMP9 and MMP2 and specifically exacerbated diastolic stiffening (+130% in chamber stiffness). PD166793 cotreatment prevented these changes, although myocardial collagen content, subtype, and cross-linking were unaltered. AII+P also reduced ATP, free energy of ATP hydrolysis (DeltaG(ATP)), and phosphocreatine while increasing free [ADP], AMP catabolites (nucleoside-total purines), and lactate. PD166793 reversed most of these changes, in part due to its inhibition of AMP deaminase. MMP activation may influence cardiac diastolic function by mechanisms beyond modulation of extracellular matrix. Interaction between MMP activation and HEP metabolism may play an important role in mediating diastolic dysfunction. Furthermore, these data highlight a potential major role for increased AMP deaminase activity in diastolic dysfunction.