Skip Navigation

European Journal of Heart Failure 2007 9(8):754-761; doi:10.1016/j.ejheart.2007.03.011
This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrowRequest Permissions
Right arrow Disclaimer
Google Scholar
Right arrow Articles by Keweloh, B.
Right arrow Articles by Hermann, H.-P.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Keweloh, B.
Right arrow Articles by Hermann, H.-P.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

© 2007 European Society of Cardiology

Influence of pyruvate on economy of contraction in isolated rabbit myocardium

Boris Keweloha,b, Paul M.L. Janssenb,1, Ulf Siegelb, Nicolin Datzb, Oliver Zeitzb,2 and Hans-Peter Hermannb,*

a Franz-Volhard-Klinik, Universitätsklinikum Charité, Berlin, Germany
b Abteilung Kardiologie und Pneumologie, Universität Göttingen, Göttingen, Germany

* Corresponding author. Georg-August-Universität Göttingen, Abteilung Kardiologie und Pneumologie, Robert-Koch-Str. 40, D-37075 Göttingen, Germany. Tel.: +49 551 392920; fax: +49 551 398918. E-mail address: phermann{at}med.uni-goettingen.de


   Abstract
Top
 Notes
 Abstract
1. Introduction
 2. Materials and methods
 3. Results
 4. Discussion
 References
 
Background: Treatment of acute heart failure frequently requires positive-inotropic stimulation. However, there is still no inotropic agent available, which combines a favourable haemodynamic profile with low expenditure for energy metabolism. Pyruvate exhibits positive inotropic effects in vitro and in patients with heart failure. The effect on myocardial energy metabolism however remains unclear, but is meaningful in light of a clinical application.

Aims and methods: We investigated the influence of pyruvate on contractility and oxygen consumption in isolated isometric contracting rabbit myocardium compared to β-adrenergic stimulation with isoproterenol.

Results: Pyruvate (30 mM) increased developed force from 18.7±4.1 to 50.8±12.1 mN/mm2 (n=10, p<0.01). Force-time integral (FTI) increased by 329%, oxygen consumption assessed by diffusion-microelectrode technique increased from 2.86±0.30 mlO2/min*100 g to 6.28±1.28 mlO2/min*100 g (n=7, p<0.05). Economy of myocardial contraction calculated as the ratio of total FTI to oxygen consumption remained unchanged. In contrast, while isoproterenol (10 µM) produced a comparable increase in developed force from 21.4±8.3 to 67.3±15mN/mm2 (n=7, p<0.01), FTI increased only by 260% and MVO2 increased from 2.96±0.43 to 6.12±1.01 mlO2/min*100 g (n=7, p<0.01); thus, economy decreased by 23% (n=7, p<0.05).

Conclusion: Pyruvate does not impair economy of myocardial contraction while isoproterenol decreases economy. Regarding energy expenditure, pyruvate appears superior to isoproterenol for the purpose of positive inotropic stimulation.

Key Words: Pyruvate • Heart failure • Energetics • Inotropic therapy • Oxygen consumption

Received August 30, 2006; Revised January 31, 2007; Accepted March 8, 2007


   1. Introduction
Top
 Notes
 Abstract
 1. Introduction
2. Materials and methods
 3. Results
 4. Discussion
 References
 
The concept of treating heart failure with positive inotropic agents was considered controversial during the last decade. With the exception of digitalis, which improves morbidity but not mortality, and which is still in use for the treatment of symptomatic heart failure, most positive inotropic drugs fail to improve or even worsen long-term outcome in heart failure [1,2]. However, positive inotropic agents are frequently inevitable in the treatment of acute heart failure and cardiogenic shock in order to maintain or re-establish the failing circulation and to ensure minimal perfusion pressure. A number of major side effects of currently available β-adrenergic agonists and phosphodiesterase (PDE) inhibitors strongly limit them for short term use: First, β-adrenergic agonists and phosphodiesterase inhibitors markedly decrease economy of contraction; they elevate oxygen demand and substrate metabolism disproportionate relative to the increase in contractility [3,4]. This results in a state of energy depletion with a risk of superimposed ischaemia and subsequent cell death. Second, β-adrenergic agonists increase intracellular calcium concentration disproportionate relative to force generation, which may be associated with induction of harmful arrhythmias. Third, most β-adrenergic agonists exhibit vasoconstrictive properties in peripheral vasculature, which additionally impair the perfusion of vital organs and can rapidly result in multiorgan failure [5,6].

In the search for alternative inotropic agents for the treatment of acute heart failure, it has recently been shown that the glycolytic intermediate pyruvate exhibits favourable haemodynamic properties in patients with congestive heart failure [7,8]. Exogenous pyruvate produces positive inotropic effects in normal, hypoxic and post-ischaemic myocardium in isolated rabbit-and swine hearts [9,10], in isolated myocardial trabeculae from rabbits [11] and humans [12] and in the intact anaesthetised dog [13-15]. Pyruvate has numerous molecular effects, which may contribute to its inotropic action [16]. These include a reduction of inorganic phosphate [10,17,18], a decrease in hydrogen ion concentration [9,19] and a modulation of the cytosolic redox state [9,20,21]. The additional direct supply of the mitochondria with an energetic substrate increases ATP production by oxidative phosphorylation [18] leading to an increase in the cytosolic phosphorylation potential and positive inotropism [9]. Furthermore, data obtained in isolated myocardium indicate additional mechanisms of action including changes in myofilament calcium sensitivity and/or changes in cross-bridge cycling kinetics [19,22]. The energetic consequences of the positive inotropic effect are currently unknown. However, this may be of paramount importance in light of a potential clinical application since supply of oxygen and energetic substrates is limited in most forms of heart failure [23]. Thus, the goal of the present study was to investigate the influence of pyruvate on myocardial contractility and oxygen consumption in isolated multicellular myocardial preparations.


   2. Materials and methods
Top
 Notes
 Abstract
 1. Introduction
 2. Materials and methods
3. Results
 4. Discussion
 References
 
2.1. Muscle preparations and mechanical measurements
Female White New-Zealand rabbits weighing between 1.5 and 2.5 kg were anaesthetised with thiopental (50 mg/kg) via an ear vein after heparinization (1000 I.U.). Hearts were rapidly dissected and retrogradely perfused through the aorta with a Krebs-Henseleit (K-H) solution. Right ventricular trabeculae were dissected as previously described [24,25]. Muscle preparations were carefully dissected with the aid of a stereo microscope and dimensions were measured at 40x magnification for normalisation of force values by cross-sectional area. Average muscle cross-sectional area was 0.16±0.02 mm2 (n=38). Dissected trabeculae were kept in oxygenated cardioplegic solution containing (mM): 120 NaCl, 5 KCl, 2 MgSO4, 1.2 NaH2PO4, 20 NaHCO3, 10 Glucose, 0.25 CaCl2 and 20 2,3-butanedione monoxime (BDM). BDM has been shown to protect the myocardium during muscle preparation and to be completely reversible after washout [26]. The muscle preparations were transferred to the muscle chamber containing oxygenated (95% O2, 5% CO2) Krebs-Henseleit solution at 37 °C. Muscles were mounted using two blocks of ventricular or valvular tissue in the experimental set-up between a basket-shaped extension of a force transducer and a hook connected to a micro-displacement device [24,25]. Superfusion with K-H solution (at 37 °C) was started and calcium concentration was raised from 0.25 to 1.25 mM in steps of 0.25 mM every 2-5 min. After the 1.0 mM concentration was reached, stimulation was started through 5 ms asymmetric pulses at 20% above threshold voltage (typically 2-4 V) at 1.0 Hz. At 1.25 mmol/L [Ca2+] the muscle was carefully stretched in several steps until developed force reached a maximum and further stretching did not result in an additional increase in developed force or diastolic force exceeded 20% of developed force [25,27]. The muscles were left contracting under these conditions for at least one additional hour to equilibrate. Force was registered with a force transducer (KG4, Scientific Instruments, Heidelberg, FRG), and force parameters were obtained online and from the recorded charts. Intact rabbit muscles were discarded from the experiment when either developed force at 2 Hz stimulation frequency was <10 mN/mm2 or loss of force during the experiment exceeded 15% per h. Diastolic force (Fdia) was defined as the lowest force during the stimulus interval, developed force (Fdev) as the difference between peak systolic force and diastolic force. Total force-time integral (FTI) was defined as the sum of developed force-time integral (FTIsys) and diastolic force-time integral (FTIdias), where FTIsys was defined as the area between the systolic twitch force curve and the diastolic force curve, and FTIdias the area between the diastolic force level and the force measured after addition of 20 mM BDM to inhibit crossbridge cycling [28]. For calculation of economy total force-time integral was used as described previously [28]. Total FTI includes bottom square integrals reflecting active diastolic tension. Since there was no significant active diastolic tension during the course of any experiment in this study, there was no significant difference in the calculated values of economy of contraction if systolic FTI was used instead of total FTI (data not shown). FTI is expressed in (mN s/min mm2).

2.2. Oxygen consumption measurements
Oxygen consumption measurements were obtained using a recently developed and validated diffusion-microelectrode technique [28]. The experimental set up consisted of a closed muscle chamber with a central heating unit, in- and outlets for the perfusion solution, for the polarographic oxygen electrode and for a temperature probe. To obtain oxygen consumption measurements of isometrically contracting muscle, the perfusion with oxygenated solution was stopped for an interval of 10 to 15 s. At a pre-defined distance from the muscle surface, the time-dependent decrease of oxygen partial pressure was measured and recorded. Muscle oxygen consumption was calculated by comparison of the obtained curves to theoretically pre-calculated curves with given oxygen consumption and defined muscle cross-sectional areas [28]. Muscle preparations for which the appropriate pre-calculated curves predicted hypoxia at the muscle core were excluded from the study.

2.3. Calculation of economy of contraction
Oxygen consumption is a reliable index for myocardial energy turnover since in the absence of ischaemia there is almost no anaerobic glycolysis in the myocardium. Even recovery metabolism in the myocardium occurs almost exclusively under aerobic conditions [29]. The force-time integral of an isometric contraction results from the summation of all individual cross-bridge interactions consuming most of the energy during a contraction-relaxation cycle. The isometric force time integral was chosen because it has been shown to correlate closely with myocardial energy consumption [28,30,31]. Thus, economy of isometric contraction can be approximated as the ratio of force-time integral (FTI) to oxygen consumption (MVO2) [32]. For practical purpose, in the present study the energetic economy of active isometric contraction was determined as the inverse slope of the linear correlation between myocardial oxygen consumption and total force time integral of the muscle. Energy expenditure for resting metabolism was measured directly as oxygen consumption without electrical stimulation. Energy expenditure for basal metabolism was obtained in absence of electrical stimulation and after addition of 20 mM BDM for inhibition of residual cross-bridge cycling [26,28].

2.4. Experimental protocol
Twitch contractions in all experiments were measured starting at a calcium concentration of 2.5 mM and 1 Hz stimulation (control). Force, twitch timing parameters and oxygen consumption were measured at each concentration after reaching steady state conditions.

2.4.1. Sodium pyruvate
The dose-dependent effect of increasing pyruvate concentrations (0, 0.3, 1, 3, 10, 30 mM sodium pyruvate) on force generation and twitch timing parameters was investigated in 10 muscle preparations. In seven additional experiments, the effect of rising concentrations of sodium-pyruvate on myocardial oxygen consumption was studied: Starting from 2.5 mM calcium and 1 Hz stimulation frequency force- and oxygen values were measured at stimulation frequencies of 0, 0.5, 1, 2 and 3 Hz in order to obtain a wide range of data for calculation of economy of contraction (control). After addition of pyruvate (0.3, 1, 3, 10, 30 mM), force and oxygen measurements were repeated at 0, 0.5, 1, 2 and 3 Hz. After addition of the highest dose of pyruvate, the osmotic pressure and pH of the solution were measured to ensure that the observed effects were not due to changes in osmotic pressure or pH. At the end of each experiment, oxygen consumption was measured after addition of 20 mM BDM in order to inhibit residual cross-bridge cycling.

2.4.2. Sodium chloride
To exclude that increasing sodium concentration would cause the observed inotropic and energetic effects of sodium-pyruvate, force and oxygen consumption measurements were obtained before and after addition of 30 mM sodium chloride in six muscle preparations.

2.4.3. Cyclopiazonic acid
To determine the influence of pyruvate-induced stimulation of sarcoplasmic reticulum calcium ATPase on resting metabolism and economy of contraction, sarcoplasmic reticulum Ca2+ ATPase was inhibited with cyclopiazonic acid [33] (30 µM CPA, n=6; 1 mM CPA, n=2). Force and oxygen consumption were measured at 2.5 mM [Ca2+] and 0, 1 and 3 Hz following addition of CPA with repeated measurements. Thereafter, CPA was washed out by changing perfusion solutions in order to investigate the reversibility of the effect. Subsequently, 30 mM pyruvate was administered and force and oxygen measurements were obtained at 0, 1 and 3 Hz. Again, CPA was added and oxygen and force measurements were repeated at 0, 1 and 3 Hz.

2.4.4. Isoproterenol
To investigate the influence of isoproterenol on oxygen consumption, seven experiments were performed: starting from 2.5 mM [Ca2+] and 1 Hz stimulation frequency, force and oxygen were measured at stimulation frequencies of 0, 0.5, 1, 2 and 3 Hz in order to obtain a wide range of data for calculation of economy of contraction (control). After addition of isoproterenol (0.01, 0.1, 1 and 10 µM) force and oxygen measurements were repeated. At the highest concentration of 10 µM isoproterenol measurements were repeated with 0, 0.5, 1, 2 and 3 Hz stimulation frequency. At the end of each experiment, oxygen consumption was measured after addition of 20 mM BDM in order to inhibit residual cross-bridge cycling.

2.5. Data analysis and statistics
Data were collected by custom designed programs, written in LabView (National Instruments, Texas) and were analysed both on-line and off-line. Data are expressed as mean±S.E.M. unless stated otherwise. Significance between groups or interventions was performed by one-way ANOVA and by Student's t-test for paired data where applicable. A two-tailed value for p<0.05 was considered significant.


   3. Results
Top
 Notes
 Abstract
 1. Introduction
 2. Materials and methods
 3. Results
4. Discussion
 References
 
3.1. Sodium pyruvate
Following addition of sodium pyruvate a transient decrease of developed force (Fdev) lasting for 3 to 4 min which preceded the pronounced positive inotropic effect could be observed in all experiments. Developed force of contraction dose-dependently increased under influence of pyruvate, as shown in Fig. 1; steady state conditions at each concentration were achieved after approximately 10 min. 30 mM pyruvate increased Fdev by 172% compared to control at 2.5 mM calcium (from 18.7±4.2 to 50.8±12.1 mN/mm2; n=10, p<0.01). Diastolic force decreased only slightly from 8.3±2.7 to 6.0±1.6 mN/mm2 at 30 mM pyruvate (p=n.s.). Twitch timing parameters were dose-dependently prolonged upon application of pyruvate (Table 1).


Figure 01
View larger version (16K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 1 (A) Influence of pyruvate on twitch contraction of a rabbit ventricular trabecula. With increasing pyruvate concentrations (0, 0.3, 1, 3, 10 and 30 mM) developed force dramatically increases, while diastolic force remains almost unchanged. Twitch duration is concentration-dependently prolonged; both time-to-peak tension and relaxation time are prolonged at increasing pyruvate concentrations. Basic stimulation frequency was 1 Hz, extracellular Ca2+ concentration was 2.5 mM. Dimensions of this preparation were 240 µm wide, 200 µm thick and 3.1 mm long. (B) Concentration-dependent effect of pyruvate on developed force and diastolic force. Developed force increases with higher pyruvate concentrations while diastolic force remains unchanged. The small decrease in diastolic force is still present after washout of pyruvate and thus due to passive lengthening of the preparation over time. After washout of pyruvate (30 min) developed force rapidly decreases to control level. * denotes a significant (p<0.05) difference vs. control (0 mM pyruvate).

 


View this table:
[in this window]
[in a new window]

  		Table 1 Force and twitch timing parameters after addition of 0, 3 and 30 mM pyruvate from 10 isolated rabbit myocardial trabeculae

 
Under influence of pyruvate oxygen consumption at 1 Hz stimulation frequency increased by 120% from 2.86±0.30 mlO2/min*100 g at control to 6.28±1.28 mlO2/min*100 g at 30 mM pyruvate (n=7, p<0.05). The systolic force time integral (FTIsys) increased in parallel by 329% from 25.5±6 (control) to 109±31 mN s/min mm2 (30 mM pyruvate) at 1 Hz stimulation frequency (p<0.05, n=7) and increased by 35% from 228±21 (control) to 307±40 mN s/min mm2 (30 mM pyruvate) at 2 Hz stimulation frequency. There was no change in economy of active myocardial contraction, calculated as the inverse slope of the MVO2/FTI relation (54±6, 57±8, 62±8, 58±6 mlO2/mN min at 0, 0.3, 3, 30 mM pyruvate respectively, n.s.). Fig. 2 shows the relationship between oxygen consumption and FTI under influence of pyruvate for one typical experiment (A, top panel) and an average of all seven experiments (B, bottom panel).


Figure 02
View larger version (19K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 2 (A) Relationship between myocardial oxygen consumption (MVO2) and total force time integral (FTI) with increasing pyruvate concentration (0, 0.3, 3 and 30 mM) from one typical experiment. The inverse slope of the regression lines reflects economy of isometric contraction; thus, an increasing slope indicates decreasing economy. Of note, there is no significant alteration of the slope of the MVO2/FTI relationship under influence of pyruvate. (B) Economy of contraction calculated from the inverse slope of regression lines of all experiments showing no significant change under increasing pyruvate concentrations (n=7, p=n.s.).

 
Resting metabolism dose-dependently increased after addition of pyruvate up to 200% of control which is shown in Fig. 4, top panel (2.24±0.35, 2.26±0.33, 2.62±0.44, 3.26±0.48, 4.74±0.86 and 6.71±1.45 mlO2/min*100 g, at 0, 0.3, 1, 3, 10 and 30 mM pyruvate respectively; n=7, p<0.05 for 10 and 30 mM pyruvate). Basal metabolism at 30 mM pyruvate was measured 35% lower than resting metabolism (n=7, p<0.05). All effects of sodium-pyruvate were completely reversible, after washout of the substance force and oxygen values returned to 90 to 130% of baseline in approximately 30 min.

Addition of 30 mM sodium-pyruvate neither changed the pH of the K-H-solution (7.43±0.04 vs. 7.41±0.03, n=7, n.s.) nor the osmotic pressure (302±10 vs. 314±12 mosM/l, n=7, n.s.).

3.2. Isoproterenol
Isoproterenol dose-dependently increased developed force: at the highest concentration of isoproterenol (10 µM) an average increase of 215% of control could be observed (from 21.4±8.3 to 67.3±15.0 mN/mm2; n=7, p<0.01). Diastolic force did not significantly change.

Under influence of isoproterenol (10 µM) oxygen consumption at 1 Hz stimulation frequency increased by 107% from 2.96±0.43 mlO2/min*100 g at control to 6.12±1.0 mlO2/min*100 g (p<0.01, n=7). The systolic force time integral (FTIsys) increased in parallel by 263% from 131±39 mN s/min mm2 to 476±142 mN s/min mm2 (p<0.05, n=7) at 1 Hz stimulation frequency but did not further increase at 2 Hz (763±237 mN s/min mm2 at control, 772±161 mN s/min mm2 at 10 µm isoproterenol, n.s.). Accordingly, in contrast to pyruvate we observed under influence of isoproterenol a significant decrease in economy of active myocardial contraction of 23% of control (157±50 vs. 121±43 mlO2/mN min at 0 and 10 µM isoproterenol, n=7, p<0.05). Fig. 3 shows the relationship between oxygen consumption and FTI under influence of isoproterenol for one typical experiment (A, top panel) and the decrease of economy of contraction in average of all 7 experiments (B, bottom panel).


Figure 03
View larger version (16K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 3 (A) Relationship between myocardial oxygen consumption (MVO2) and total force time integral (FTI) under influence of isoproterenol (10 µM) from one typical experiment. The inverse slope of the regression lines reflects economy of isometric contraction. Of note, the slope of the regression line markedly increases under influence of isoproterenol indicating decreased economy of contraction. (B) Economy of contraction of all experiments showing a decrease of 23% of control with 10 µM isoproterenol (n=7, *p<0.05).

 
Resting metabolism did not change significantly after addition of isoproterenol as shown in Fig. 4B, bottom panel.


Figure 04
View larger version (20K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 4 (A) Influence of pyruvate on resting- and basal oxygen consumption (n=7). * denotes a significant (p<0.05) difference vs. control, § denotes a significant (p<0.05) difference vs. 30 mM pyruvate. Resting oxygen consumption is measured in absence of stimulation. The right column (BDM-2,3-butanedione monoxime) shows basal oxygen consumption after addition of 20 mM BDM to inhibit cross bridge cycling and EC-coupling at the end of each experiment. (B) Influence of isoproterenol on resting oxygen consumption (n=7, p=n.s.).

 
3.3. Sodium chloride
At 1 Hz stimulation frequency 30 mM sodium-chloride was added to the solution. No effect on developed force and oxygen consumption could be observed (Fdev 15.1±2.6 mN/mm2 at control, 15.8±2.6 mN/mm2 at 30 mM NaCl; MVO2 1.50±0.20 mlO2/min*100 g at control, 1.49±0.21 mlO2/min*100 g at 30 mM sodium chloride, n=6, p=n.s.). Thus, the inotropic and energetic effects of pyruvate were not due to an overall rise in sodium concentration.

3.4. Cyclopiazonic acid
To investigate whether the observed two-fold increase in resting metabolism under high concentrations of pyruvate was due to an increased cytosolic phosphorylation potential primarily resulting in an activation of the sarcoplasmic reticulum Ca2+ ATPase, we measured oxygen consumption and force development under influence of pyruvate and inhibition of SR-Ca2+ ATPase by cyclopiazonic acid (CPA). Developed force at 1 Hz stimulation frequency decreased by 56% from 6.2±0.8 to 3.5±0.5 mN/mm2 when 30 µM CPA alone was added to the K-H solution (Fig. 5). Resting metabolism did not change significantly (from 1.00±0.08 at control to 1.08±0.17 mlO2/min*100 g after 30 µM CPA; n=6, p=n.s.). Following 30 mM pyruvate, addition of 30 µM CPA reduced developed force by 46% (from 10.7 to 5.8±0.6 mN/mm2) while again did not influence resting metabolism (Fig. 5). Calculated economy of myocardial contraction remained unchanged (data not shown). Thus, the increase in resting metabolism could not be attributed to an activation of SR-Ca2+ ATPase.


Figure 05
View larger version (20K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Fig. 5 Influence of pyruvate on (A) developed force and (B) oxygen consumption before and after inhibition of sarcoplasmic reticulum Ca2+ ATPase by cyclopiazonic acid (CPA). Of note, CPA decreases developed force in control contractions and under influence of pyruvate (A), while oxygen consumption for resting metabolism remains unchanged, even if elevated under influence of pyruvate (B), suggesting that the energetic effect of pyruvate is not due to activation of SR Ca2+ ATPase. * denotes a significant (p<0.05) difference vs. control. § denotes a significant (p<0.05) difference vs. 30 mM pyruvate.

 

   4. Discussion
Top
 Notes
 Abstract
 1. Introduction
 2. Materials and methods
 3. Results
 4. Discussion
References
 
The present study was performed to investigate the effect of pyruvate on myocardial energy metabolism. We demonstrated that pyruvate exhibits positive inotropic effects, enhances resting metabolism at high concentrations, but most importantly does not adversely affect economy of myocardial contraction. This is in clear contrast to the β-adrenergic agonist isoproterenol, which significantly decreased economy of myocardial contraction.

Numerous molecular effects of pyruvate have been postulated to contribute to its inotropic action. These include an increase in cytosolic phosphorylation potential [9,17,18], a reduction of inorganic phosphate [9,17,18,21], a decrease in hydrogen ion concentration [9,19] and a modulation of the cytosolic redox state [9,20,21]. Pyruvate enters the cell via a proton-monocarboxylate symporter and is directly transported into the mitochondria via a monocarboxylate transporter located in the mitochondrial membrane [34]. In studies in isolated rat ventricular myocytes the inhibition of the mitochondrial outer membrane transport system by {alpha}-cyano-4-hydroxycinnamate abolished the positive inotropic effect of pyruvate [18,35]. Elimination of glucose from the perfusion medium however could not significantly affect the positive inotropism of pyruvate. Thus, externally added pyruvate does not mainly affect energetics of cytosolic glycolysis and the positive inotropic effect of pyruvate does not depend on the presence of glucose in the perfusion medium.

The present study contributes two primary findings to our current knowledge. First, pyruvate was shown not to deteriorate economy of myocardial contraction. This is beneficial in light of a potential clinical application and in clear contrast to β-adrenergic agonists, which increase oxygen consumption disproportionate to contractility [32]. The "oxygen wasting" effect of β-adrenergic agonists may result from increased calcium cycling, compensating for decreased calcium sensitivity of the contractile proteins [27,36-38], from phosphorylation of proteins involved in calcium cycling [39] and from enhanced metabolic futile cycles [37]. Second, we observed a significant increase in resting metabolism at high concentrations of pyruvate and a smaller non-significant enhancement at lower concentrations. This could be of major concern in the clinical setting especially in patients with heart failure of ischaemic aetiology. However, intracoronary pyruvate concentrations between 3-6 mM [7] have been shown to produce a 25%-increase in cardiac output, whereas a significantly enhanced resting metabolism in the current study was only observed at higher concentrations of between 10 and 30 mM. The increase in resting metabolism at high concentrations was not due to increased ATP consumption of the sarcoplasmic reticulum calcium pump since inhibition of the SR-Ca2+ ATPase with CPA did not abolish the observed effect. A rise in overall sodium concentration, which could increase the amount of ATP needed by the sarcolemmal sodium-potassium pump was not responsible for the elevated resting metabolism either, since no elevation of resting oxygen consumption was observed after addition of 30 mM sodium chloride.

We previously hypothesised that the positive inotropic effect of pyruvate may be due in part to mechanisms downstream of sarcoplasmic reticulum calcium handling [12,19,22]. The present study reveals more evidence to support this theory. We observed that despite inhibition of sarcoplasmic reticulum calcium reuptake, addition of pyruvate still resulted in a clear positive inotropic effect. This indicates that the positive inotropic effect of pyruvate cannot be solely due to increased sarcoplasmic reticulum calcium handling. Other processes, located downstream (i.e. cross-bridge cycling kinetics and/or myofilament calcium sensitivity) may also play a significant, or possibly even a primary, role in pyruvate-induced inotropism.

In conclusion, pyruvate has positive inotropic effects, increases oxygen consumption and resting metabolism, but unlike the β-adrenergic agonist isoproterenol, it does not change economy of active myocardial contraction. These observed energetically favourable properties of pyruvate might further qualify it as a positive inotropic agent for the treatment of acute heart failure and cardiogenic shock. However, further investigations are necessary to elucidate the underlying mechanisms of action and to determine a more feasible method of administration, since treatment of patients with pyruvate is currently limited by the prerequisite for intracoronary infusion [7,8]. Because the haemodynamic effect in vivo requires pyruvate concentrations above 1 mM and since pyruvate is rapidly metabolised, high doses would be necessary if pyruvate was to be given intravenously. Such high intravenous doses however, would result in sodium overload and hyperosmolarity if the sodium salt of pyruvic acid was used as in the present study. However, other formulations of pyruvate may be developed. Even if intracoronary application was the only route of effective administration of pyruvate, this treatment could still be useful in patients with acute heart failure refractory to conventional therapy if a catheter laboratory was available.


   Notes
Top
 Notes
Abstract
 1. Introduction
 2. Materials and methods
 3. Results
 4. Discussion
 References
 
1 Current address: Department of Physiology and Cell Biology, The Ohio State University, 304 Hamilton Hall, 1645 Neil Avenue, Columbus, OH 43210-1218, USA. Back

2 Current address: Universitätsklinikum Hamburg-Eppendorf, Klinik und Poliklinik für Augenheilkunde, Martinistr. 52, D-20246 Hamburg, Germany. Back


   References
Top
 Notes
 Abstract
 1. Introduction
 2. Materials and methods
 3. Results
 4. Discussion
 References
 

  1. Lubsen J., Just H., Hjalmarsson A.C., et al. Effect of pimobendan on exercise capacity in patients with heart failure: main results from the Pimobendan in Congestive Heart Failure (PICO) trial. Heart (1996) 76:223–231.[Abstract/Free Full Text]
  2. Uretsky B.F., Jessup M., Konstam M.A., et al. Multicenter trial of oral enoximone in patients with moderate to moderately severe congestive heart failure. Lack of benefit compared with placebo. Enoximone Multicenter Trial Group. Circulation (1990) 82:774–780.[Abstract/Free Full Text]
  3. Hasenfuss G., Holubarsch C., Heiss H.W., et al. Myocardial energetics in patients with dilated cardiomyopathy. Influence of nitroprusside and enoximone. Circulation (1989) 80:51–64.[Abstract/Free Full Text]
  4. Hasenfuss G., Mulieri L.A., Allen P.D., Just H., Alpert N.R. Influence of isoproterenol and ouabain on excitation-contraction coupling, cross-bridge function, and energetics in failing human myocardium. Circulation (1996) 94:3155–3160.[Abstract/Free Full Text]
  5. Colucci W.S., Wright R.F., Braunwald E. New positive inotropic agents in the treatment of congestive heart failure. Mechanisms of action and recent developments (first of two parts). New Engl J Med (1986) 314:290–299.[Web of Science][Medline]
  6. Colucci W.S., Wright R.F., Braunwald E. New positive inotropic agents in the treatment of congestive heart failure. Mechanisms of action and recent developments (second of two parts). New Engl J Med (1986) 314:349–358.[Web of Science][Medline]
  7. Hermann H.P., Pieske B., Schwarzmuller E., Keul J., Just H., Hasenfuss G. Haemodynamic effects of intracoronary pyruvate in patients with congestive heart failure: an open study. Lancet (1999) 353:1321–1323.[CrossRef][Web of Science][Medline]
  8. Hermann H.P., Arp J., Pieske B., et al. Improved systolic and diastolic myocardial function with intracoronary pyruvate in patients with congestive heart failure. Eur J Heart Fail (2004) 6:213–218.[Abstract/Free Full Text]
  9. Bünger R., Mallet R.T., Hartman A. Pyruvate enhanced phosphorylation potential and inotropism in normoxic and postischemic isolated working heart. Near-complete prevention of reperfusion contractile failure. Eur J Biochem (1988) 180:221–233.[Web of Science]
  10. Liedtke A.J., Nellis S.H. Effects of buffered pyruvate on regional cardiac function in moderate, short-term ischemia in swine heart. Circ Res (1978) 43:189–199.[Free Full Text]
  11. Hermann H.P., Zeitz O., Keweloh B., Hasenfuss G., Janssen P.M.L. Pyruvate potentiates inotropic effects of isoproterenol and Ca(2+) in rabbit cardiac muscle preparations. Am J Physiol (Heart Circ Physiol) (2000) 279(2):H702–H708.[Abstract/Free Full Text]
  12. Hermann H.P., Zeitz O., Lehnart S.E., et al. Potentition of β-adrenergic inotropic response by pyruvate in failing human myocardium. Cardiovasc Res (2002) 53:116–123.[Abstract/Free Full Text]
  13. Mentzer R.M. Jr., van Whylen D.G., Sodhi J., et al. Effect of pyruvate on regional ventricular function in normal and stunned myocardium. Ann Surg (1989) 209:629–633.[Web of Science][Medline]
  14. Yanos J., Patti M.J., Stanko R.T. Hemodynamic effects of intravenous pyruvate in the intact, anesthetized dog. Crit Care Med (1994) 22:844–850.[Web of Science][Medline]
  15. Zweier J.L., Jacobus W.E. Substrate induced alterations of high energy phosphate metabolism and contractile function in the perfused heart. J Biol Chem (1987) 262:8015–8021.[Abstract/Free Full Text]
  16. Mallet R.T., Sun J., Knott E.M., Sharma A.B., Olivencia-Yurvati A.H. Metabolic cardioprotection by pyruvate: recent progress. Exp Biol Med (2005) 230:435–443.[Abstract/Free Full Text]
  17. Mallet R.T., Bünger R. Energetic modulation of cardiac inotropism and sarcoplasmic reticular Ca2+ uptake. Biochim Biophys Acta (1994) 1224:22–32.[Medline]
  18. Martin B.J., Valdivia H.H., Bünger R., Lasley R.D., Mentzer R.M. Jr. Pyruvate augments calcium transients and cell shortening in rat ventricular myocytes. Am J Physiol (Heart Circ Physiol) (1998) 274(1 Pt 2):H8–H17.[Abstract/Free Full Text]
  19. Hasenfuss G., Maier L.S., Hermann H.P., et al. Influence of pyruvate on contractile performance and Ca2+-cycling in isolated failing human myocardium. Circulation (2002) 105:194–199.[Abstract/Free Full Text]
  20. Laughlin M.R., Heineman F.W. The relationship between phosphorylation potential and redox state in the isolated working rabbit heart. J Mol Cell Cardiol (1994) 26:1525–1536.[CrossRef][Web of Science][Medline]
  21. Scholz T.D., Laughlin M.R., Balaban R.S., Kupriyanov V.V., Heineman F.W. Effect of substrate on mitochondrial NADH, cytosolic redox state, and phosphorylated compounds in isolated hearts. Am J Physiol (Heart Circ Physiol) (1995) 268:H82–H91.[Abstract/Free Full Text]
  22. Maier L.S., Braunhalter J., Horn W., Weichert S., Pieske B. The role of SR Ca(2+)-content in blunted inotropic responsiveness of failing human myocardium. J Mol Cell Cardiol (2002) 34:455–467.[CrossRef][Web of Science][Medline]
  23. De Boer R.A., Pinto Y.M., Van Veldhuisen D.J. The imbalance between oxygen demand and supply as a potential mechanism in the pathophysiology of heart failure: the role of microvascular growth and abnormalities. Microcirculation (2003) 10:113–126.[CrossRef][Web of Science][Medline]
  24. Janssen P.M.L., Lehnart S.E., Prestle J., et al. The trabecula culture system: a novel technique to study contractile parameters over a multiday time period. Am J Physiol (Heart Circ Physiol) (1998) 274:H1481–H1488.[Abstract/Free Full Text]
  25. ter Keurs H.E., Rijnsburger W.H., van Heuningen R., Nagelsmit M.J. Tension development and sarcomere length in rat cardiac trabeculae: evidence of length-dependent activation. Circ Res (1980) 46:703–714.[Free Full Text]
  26. Mulieri L.A., Hasenfuss G., Ittleman F., Blanchard E.M., Alpert N.R. Protection of human left ventricular myocardium from cutting injury with 2,3 butanedione monoxime. Circ Res (1989) 65:1441–1449.[Abstract/Free Full Text]
  27. Janssen P.M.L., de Tombe P.P. Protein kinase A does not alter unloaded velocity of sarcomere shortening in skinned rat cardiac trabeculae. Am J Physiol (Heart Circ Physiol) (1997) 273:H2415–H2422.[Abstract/Free Full Text]
  28. Meyer M., Keweloh B., Güth K., et al. Frequency-dependence of myocardial energetics in failing human myocardium as quantified by a new method for the measurement of oxygen consumption in muscle strip preparations. J Mol Cell Cardiol (1998) 30:1459–1470.[CrossRef][Web of Science][Medline]
  29. Challoner D.R. Respiration in myocardium. Nature (1968) 217:78–79.[CrossRef][Medline]
  30. Mc Donald R.H. Jr. Developed tension: a major determinant of myocardial oxygen consumption. Am J Physiol (1966) 210:351–356.[Free Full Text]
  31. Sarnoff S.J., Braunwald E., Welch G.H., Case R.B., Stainsby W.N., Macruz R. Hemodynamic determinants of oxygen consumption of the heart with special reference to the tension-time index. Am J Physiol (1958) 192:148–156.[Web of Science][Medline]
  32. Alpert N.R., Mulieri L.A., Hasenfuss G. Myocardial chemo-mechanical energy transduction. In: The Heart and Cardiovascular System (1992) second edition. New York: Raven Press Ltd. 111–128.
  33. Baudet S., Shaoulian R., Bers D.M. Effects of thapsigargin and cyclopiazonic acid on twitch force and sarcoplasmic reticulum Ca2+ content of rabbit ventricular muscle. Circ Res (1993) 73:813–819.[Abstract/Free Full Text]
  34. Poole R.C., Cranmer S.L., Halestrap A.P., Levi A.J. Substrate and inhibitor specificity of monocarboxylate transport into heart cells and erythrocytes. Further evidence for the existence of two distinct carriers. Biochem J (1990) 269:827–829.[Web of Science][Medline]
  35. Zima A.V., Kockskämper J., Mejia-Alvarez R., Blatter L.A. Pyruvate modulates cardiac sarcoplasmic reticulum Ca2+ release in rats via mitochondria-dependent and -independent mechanisms. J Physiol (2003) 550:765–783.[Abstract/Free Full Text]
  36. Endoh M., Blinks J.R. Actions of sympathomimetic amines on the Ca2+ transients and contractions of rabbit myocardium: reciprocal changes in myofibrillar responsiveness to Ca2+ mediated through alpha- and beta-adrenoceptors. Circ Res (1988) 62:247–265.[Abstract/Free Full Text]
  37. Kurihara S., Konishi M. Effects of beta-adrenoceptor stimulation on intracellular Ca transients and tension in rat ventricular muscle. Pflügers Arch (1987) 409:427–437.[CrossRef][Web of Science][Medline]
  38. Suga H. Ventricular energetics. Physiol Rev (1990) 70:247–277.[Free Full Text]
  39. Rapundalo S.T., Solaro R.J., Kranias E.G. Inotropic responses to isoproterenol and phosphodiesterase inhibitors in intact guinea pig hearts: comparison of cyclic AMP levels and phosphorylation of sarcoplasmic reticulum and myofibrillar proteins. Circ Res (1989) 64:104–111.[Abstract/Free Full Text]

Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?



This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrowRequest Permissions
Right arrow Disclaimer
Google Scholar
Right arrow Articles by Keweloh, B.
Right arrow Articles by Hermann, H.-P.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Keweloh, B.
Right arrow Articles by Hermann, H.-P.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?