Phosphate metabolite concentrations and ATP hydrolysis potential in normal and ischaemic hearts

Abstract

To understand how cardiac ATP and CrP remain stable with changes in work rate - a phenomenon that has eluded mechanistic explanation for decades - data from (31)phosphate-magnetic resonance spectroscopy ((31)P-MRS) are analysed to estimate cytoplasmic and mitochondrial phosphate metabolite concentrations in the normal state, during high cardiac workstates, during acute ischaemia and reactive hyperaemic recovery. Analysis is based on simulating distributed heterogeneous oxygen transport in the myocardium integrated with a detailed model of cardiac energy metabolism. The model predicts that baseline myocardial free inorganic phosphate (P(i)) concentration in the canine myocyte cytoplasm - a variable not accessible to direct non-invasive measurement - is approximately 0.29 mm and increases to 2.3 mm near maximal cardiac oxygen consumption. During acute ischaemia (from ligation of the left anterior descending artery) P(i) increases to approximately 3.1 mm and ATP consumption in the ischaemic tissue is reduced quickly to less than half its baseline value before the creatine phosphate (CrP) pool is 18% depleted. It is determined from these experiments that the maximal rate of oxygen consumption of the heart is an emergent property and is limited not simply by the maximal rate of ATP synthesis, but by the maximal rate at which ATP can be synthesized at a potential at which it can be utilized. The critical free energy of ATP hydrolysis for cardiac contraction that is consistent with these findings is approximately -63.5 kJ mol(-1). Based on theoretical findings, we hypothesize that inorganic phosphate is both the primary feedback signal for stimulating oxidative phosphorylation in vivo and also the most significant product of ATP hydrolysis in limiting the capacity of the heart to hydrolyse ATP in vivo. Due to the lack of precise quantification of P(i) in vivo, these hypotheses and associated model predictions remain to be carefully tested experimentally.

Figure 1. Diagram of model used to simulate cardiac tissue oxygen transport and energy metabolism

Oxygen is transported via advection in capillaries, diffusion into cardiomyocytes from capillaries through interstitium, and reduced into water via the complex IV reaction in mitochondria. Cellular energy metabolism is simulated by a computer model of mitochondrial tricarboxylic acid cycle, oxidative phosphorylation, metabolite transport and electrophysiology (Wu et al. 2007b).

Figure 2. Steady-state energetics phosphate metabolites as functions of myocardial oxygen consumption

A, model-predicted steady-state CrP/ATP level plotted as a function of oxygen consumption rate, , and compared to experimental data from canine heart in vivo. B, model-predicted steady-state ΔP_i/CrP plotted as a function of . C, model-predicted steady-state free energy of ATP hydrolysis, −ΔG_ATPase, is plotted as a function of . D, model-predicted steady-state cytoplasmic inorganic phosphate concentration, [P_i]_c and ADP, [ADP]_c, plotted as a functions of . E, model-predicted steady-state cytoplasmic ATP, [ATP]_c, plotted as a function of . F, model-predicted steady-state cytoplasmic CrP [CrP]_c, plotted as a function of . For all simulations, is varied by varying the rate of ATP hydrolysis, J_ATPase, in the cytoplasm. Experimental data are obtained from the following sources: ○, Zhang et al. (2005) (dobutamine + dopamine); ▵, Zhang et al. (2003), ◊, Gong et al. (2003) (dobutamine + dopamine); ▵, Ochiai et al. (2001) (dobutamine + dopamine); ▿, Gong et al. (1999), □, Bache et al. (1999) (dobutamine, dobutamine +dopamine). Here protocols used to elevate work load from baseline are indicated in parentheses. The values of J_ATPase corresponding to baseline and maximal , 0.36 and 1.2 mmol s⁻¹ (l cell)⁻¹, respectively, are indicated in A. Error bars indicate standard error.

Figure 3. Sample ³¹P-MRS spectra during transient occlusion and recovery

Spectra from baseline, occlusion and recovery periods are shown. A ³¹P-MRS pulse train of 28 scans was generated with repetition time of 6 s. The number of transients is one (NT = 1) for each spectrum. The resonance peaks for CrP, P_i and the 3 resonances from ATP are identified. One of the four spectra acquired during baseline conditions for a single experiment is shown, indicated as time point 0. Following the baseline period, complete LAD occlusion was initiated and maintained for the following 36 s, during which the 6th to 10th spectra were obtained. The 6th, 8th and 10th spectra (at the 12, 24 and 36 s time points during LAD occlusion, respectively) are shown demonstrating the progressive increase of the P_i and decrease of CrP during LAD occlusion. After the 10th spectrum was obtained, the occluder was released and the 11th to 28th spectra were acquired in the following 108 s. The first 3 spectra during reperfusion are shown demonstrating the fast recovery of CrP and P_i levels during the reperfusion.

Figure 4. Energetic phosphate metabolites and myoglobin saturation during ischaemia and recovery

A, the mean time course of experimentally determined coronary blood flow (with the total occlusion starting at t= 24 s and ending at t= 60 s). Data points correspond to mean of nine experiments; error bars indicate standard error. B, model simulations compared to experimental observations of CrP (normalized to baseline value) and ΔP_i/CrP. Model-predicted time courses of myoglobin saturation S_Mb (C), ATP hydrolysis flux, J_ATPase, and free energy, ΔG_ATPase (D), cytoplasmic creatine phosphate and inorganic phosphate concentrations, [CrP]_c, and [P_i]_c (E), and relative levels of cytoplasmic P_i, ADP and ATP (F) are illustrated for this experiment. Continuous lines in B, D and E represent the model prediction using the ATP hydrolysis flux expression of eqn (1), and dashed lines represent the model prediction assuming constant ATP hydrolysis flux.

Figure 5. Cardiac energetic phosphate metabolite levels and myoglobin saturation during coronary hypoperfusion

A, model-predicted steady-state ATP levels (normalized to baseline value) as a function of coronary blood flow. B, model-predicted steady-state CrP levels (normalized to ATP) (CrP/ATP). C, model-predicted steady-state ΔP_i/CrP, as a function of coronary blood flow. D, model-predicted steady-state myoglobin saturation, S_Mb, as a function of coronary blood flow. Experimental data obtained from Zhang et al. (2001). In C, data are divided into epicardium (epi), midwall (mid) and endocardium (endo), as indicated in the figure. Error bars indicate standard error.

Figure 6. Regulation of ATP synthesis and hydrolysis in vivo

A, the phenomenological relationship of eqn (1) between ATP hydrolysis rate and mass-action ratio ATP]_c/([ADP]_c[Pi]_c) is plotted as continuous lines for different steady-state work rates. The different continuous lines correspond to different values of X_AtC, corresponding to three different work rates, as indicated on the figure. Also plotted (dashed line) is the relationship between the steady-state ATP synthesis rate (the flux through adenine nucleotide translocase J_ANT) and ATP]_c/([ADP]_c[Pi]_c) predicted by the integrated model. B, control of ATP synthesis by inorganic phosphate. With [ATP]_c fixed at 10 mm, the model-predicted relationship between J_ANT and cytoplasmic ADP and P_i is plotted. Both [ADP]_c and [P_i]_c are varied over the predicted in vivo ranges for these variables. The continuous line traces the predicted in vivo values over the range of cardiac work rates studied. The three points correspond to the baseline, moderate and high work rates illustrated in A. C, variation in mitochondrial membrane potential with cardiomyocyte work rate. The model-predicted ΔΨ is plotted for the conditions described for panel B. The values of J_ANT and ΔΨ plotted in panels B and C correspond to average tissue values.