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Am J Physiol Heart Circ Physiol 279: H1029-H1035, 2000;
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Vol. 279, Issue 3, H1029-H1035, September 2000

Heterogeneity of local myocardial flow and oxidative metabolism

Uwe Schwanke1, Andreas Deussen2, Gerd Heusch1, and Jochen D. Schipke3

1 Department of Pathophysiology, University of Essen Medical School, D-45122 Essen; 2 Institute for Physiology, Department of Medical Science Carl Gustav Carus, Dresden Technical University, D-01307 Dresden; and 3 Department of Surgery, Research Group of Experimental Surgery, Heinrich-Heine-University, D-40225 Düsseldorf, Germany


    ABSTRACT
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

In mammalian hearts, local myocardial flow (LMF) varies between 20 and 200% of the mean. It is not clear whether oxidative metabolism has a similar degree of heterogeneity. Therefore, we investigated the relation between LMF and local oxidative metabolism in isolated rabbit hearts. Buffer oxygenation with 18O2 resulted in labeled myocardial oxidation water (H218O). In four hearts, myocardial oxygen consumption (MVO2) was calculated from the H218O production and compared with that calculated according to Fick. In eight additional hearts, LMF was measured using microspheres. Coronary venous H218O kinetics and local H218O residues were determined and analyzed by mathematical modeling. MVO2 recovery from H218O was >93% compared with that according to Fick. LMF ranged from 1.91 to 11.24 ml · min-1 · g-1, and local H218O residue ranged from 0.41 to 1.04 µmol/g. Both variables correlated (r = 0.62, n = 64, P < 0.001). Measurements in nine hearts were fitted by modeling using capillary permeability-surface area products (PSc) from 2 to 10 ml · min-1 · g-1. With flow-proportional PSc, a 3.33-fold difference in LMF was associated with a 6.45-fold difference in local MVO2. Both LMF and local oxidative metabolism are spatially heterogeneous, and they correlate to one another.

myocardial oxidation water; oxygen-18 labeling; modeling


    INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

THE SPATIAL DISTRIBUTION of local blood flow within mammalian myocardium is heterogeneous, with variations between 20% (low-flow areas) and 200% (high-flow areas) of the mean value (13, 22). Such perfusion patterns can remain stable for at least 15 min in isolated hearts (26).

Myocardial areas in which blood flow is <50% of mean flow cannot compensate for the low flow by increasing their oxygen extraction (13). On the other hand, mathematical modeling suggests that the oxygen concentration in capillaries in which flow is only 20% of the mean flow drops to zero after one-half the capillary length, assuming that cellular oxygen clearance is uniformly distributed (12). One would expect that such conditions would subject low-flow areas to hypoxia and, if prolonged, to necrosis. However, low-flow areas exhibit none of the typical signs of ischemia, such as increased lactate or cytosolic adenosine (33) or increased heat shock protein (HSP)-70 (27) concentrations. Hence, myocardial oxygenation in low-flow areas does not appear to be critical.

Previous studies have already studied the relation between local metabolism and local myocardial flow (LMF). In fact, extraction of iodophenylpentadecanoic acid correlated well with LMF (8, 18), and the local uptake of [3H]deoxyglucose in canine myocardium can only be explained quantitatively if it is assumed that the rate of phosphorylation of deoxyglucose is flow proportional (11).

On average, fatty acids and glucose each contribute ~30% to total myocardial oxidative metabolism under resting conditions (21). Their exact contributions, however, depend on the supply of substrate and the inotropic state, which makes it difficult to estimate local aerobic metabolism on the basis of local fatty acid or glucose uptake.

The most direct measure of local aerobic metabolism is the local rate of production of oxidation water. Because the rate of production is difficult to measure directly, the local residue of oxidation water may serve as a reasonable index of the local production rate. To test whether LMF and local tissue residue of oxidation water are correlated, we used a method of 18O labeling recently developed in our laboratory (31, 32). Because the local 18O-labeled water tissue residue depends on both intracellular production and simultaneous washout from the tissue, a mathematical model analysis (12) was used to account for these effects and to calculate the local rate of oxygen metabolism.


    METHODS
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Experiments on 12 isolated hearts from adult New Zealand White rabbits (age 4-8 mo, body mass 2,800-3,800 g) were performed in accordance with the animal welfare regulations of the German federal authorities, which adhere to the guiding principles of the American Physiological Society.

The hearts were connected to a temperature-controlled (38°C), modified Langendorff apparatus and were perfused with Krebs-Henseleit (KH) buffer. The KH composition was (in mM) 90 NaCl, 30 NaHCO3, 4 KCl, 1 Na2HPO4, 0.5 MgSO4, 2.5 CaCl2, 2.2 pyruvate, and 11.1 glucose. A servo-controlled roller pump (WM-505; Watson Marlow, Falmouth, UK) was used to adjust coronary perfusion pressure to 80 mmHg.

Equilibration of KH buffer was performed with 95% O2-5% CO2. In a second perfusion system, 500 ml of KH buffer were equilibrated with 95% N2-5% CO2 for the removal of residual oxygen. This buffer was then equilibrated with recirculating 18O2. The experimental setup permitted switching between the perfusion media during the experiment.

A latex balloon (HSE no. 12-14; Hugo Sachs Elektronik, March-Hugstetten, Germany) was inserted into the left ventricle via the mitral valve. The balloon was connected to an artificial systemic circuit that was separated from the perfusion circuit and contained two valves to mimic the aortic and the mitral valves, a windkessel, and an ultrasonic flow probe for assessing cardiac output. Left ventricular pressure (TC-500; Millar Instruments, Houston, TX), aortic pressure (Statham P23; Gould Nicolet, Erlensee, Germany), and cardiac output (T-206; Transonic Systems, Ithaca, NY) were measured in the systemic circuit. Coronary arterial pressure (Statham P23; Gould Nicolet), coronary flow (T-206; Transonic Systems), and coronary arterial and venous oxygen partial pressures (MT1-AC15; L. Eschweiler, Kiel, Germany) were measured in the perfusion circuit.

To validate our 18O-labeling technique, global myocardial oxygen consumption [MVO2 (ml · min-1 · g-1)] was calculated according to Fick's principle as
M<A><AC>V</AC><AC>˙</AC></A><SC>o</SC><SUB>2</SUB><IT>=&agr;</IT>O<SUB><IT>2</IT></SUB><IT>·</IT>[(Pa<SUB><SC>O</SC><SUB><IT>2</IT></SUB></SUB><IT>−</IT>Pv<SUB><SC>O</SC><SUB><IT>2</IT></SUB></SUB>)<IT>/760</IT>]<IT>·</IT>CF
with alpha O2 = 0.024 ml O2 · ml-1 · 760 mmHg-1, where PaO2 and PvO2 are coronary arterial and venous oxygen partial pressures (mmHg), respectively, and CF is normalized coronary flow (ml · min-1 · g-1). This value was compared with the sum of the total H218O tissue residues plus the coronary venous H218O discharge in four hearts, collected over a period of 240 s.

In eight additional hearts, red microspheres were used to measure LMF. After sonication (Sonorex RK156; Bandelin Electronic, Berlin, Germany) and vigorous shaking (Vibrofix VF1; Janke & Kunkel, Staufen, Germany), the spheres were injected into the perfusion line immediately above the aortic root.

Experimental protocol. After stabilization, baseline hemodynamics were assessed over 15 min. Four hearts were then perfused with 18O-equilibrated solution for 240 s. During 18O perfusion, coronary venous effluent was collected at 0, 10, 20, 30, 40, 50, 60, 120, 180, and 240 s.

In eight hearts, 120,000 red microspheres were injected after measurement of baseline hemodynamics to assess LMF concomitantly with local oxidative metabolism. The KH buffer equilibrated with 95% O2-5% CO2 was replaced by the 18O2-equilibrated buffer for 240 s, and in five of these eight experiments, samples were also taken from the coronary venous effluent at 0, 10, 20, 30, 40, 50, 60, 120, 180, and 240 s. These five hearts and the four hearts mentioned above were used for mathematical model analysis of global oxidative metabolism (n = 9 hearts). At the end of the protocol, all hearts were immersed in liquid nitrogen.

Water extraction and processing. This procedure has been described in detail previously (31, 32). The four hearts used for global MVO2 assessment were lyophilized in toto. The extracted tissue water and coronary venous effluent were processed as mentioned below. For local flow/H218O analysis, the eight additional hearts were sliced into basal, median, and apical portions. The right ventricular free wall, septum, and left ventricular free wall were excised from these tissue slices. The basal, median, and apical portions of the left ventricular free wall were further cut into subendocardial and subepicardial layers. A total of eight tissue samples (100 ± 27 mg) was taken from each heart. The water was extracted from the tissue samples during lyophilization, and 7.5-µl aliquots were converted quantitatively to CO2 using the guanidine hydrochloride technique (15). The oxygen isotope ratio (18O/16O) within the CO2 samples was determined by mass spectrometry (type 251; Finnigan MAT, Bremen, Germany). The oxygen isotope ratios were converted from the standard mean ocean water (SMOW) values [per thousand SMOW] (2, 10) to SI units and expressed as local H218O residue of cardiac tissue water per gram wet mass (µmol/g). LMF was measured by red microspheres as described previously (24) and is expressed in milliliters per minute per gram.

Model analysis. The overall configuration of the model (Fig. 1) is a simplification of that described in detail previously (12). In its present form, the model consists of a nonexchanging tube segment, arterial and venous vessels, and a tissue exchange unit arranged in series. The volume of the tube region representing the volume of the perfusion cannula was measured and kept constant for all simulations (0.30 ml/g). The volumes of the arterial (Vart) and venous (Vven) vessels representing nonexchanging arteries and veins were set to 0.03 and 0.07 ml/g, respectively, by using data from the literature (4). The tissue exchange unit comprised an intracapillary and an extracapillary region arranged in a concentric fashion (4, 25). The volume of the intracapillary region (Vc) was assumed to be 0.07 ml/g, and that of the extracapillary region (Ve), which is a composite of endothelial, interstitial, and parenchymal cell regions, was set to 0.63 ml/g (17). Hence, the total tissue water space (Vtiss) amounted to 0.80 ml/g. Because no major differences between local volumes or ultrastructure of regions with high and low flow had been observed previously (17, 33), their Vtiss values were assumed to be identical. Production of 18O-labeled water was described by an extracapillary production term (Pe, in µmol · min-1 · g-1). In the present version of the model (12), Pe is the product of the extracapillary concentration of atomic oxygen (CeO) and the extracapillary clearance rate (GeO): Pe = CeO · GeO.


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  		Fig. 1.   Schematic representation of the 2-region, axially distributed model used to analyze the tissue residue and coronary venous outflow kinetics of 18O-labeled water. A tube region (Vtube), nonexchanging arterial and venous vessel regions (Vart, Vven), and the capillary region (Vc) are arranged in series. Diffusional exchange is permitted only between the capillary and the extracapillary region (Ve). The permeability-surface area product (PSc) is a composite of the diffusional barriers of the capillary wall and the parenchymal cell membrane, indicated by the double line. An extracapillary production term (Pe) describes production of 18O-labeled water. Fc, flow; Cc, intracapillary H218O concentration; Ce, extracapillary H218O concentration. Cven, coronary venous H218O concentration; Cout, effluent H218O concentration. For further details see Model analysis in METHODS.

The exchange of water between the capillary and the extracapillary regions was described by a permeability-surface area product (PSc, in ml · min-1 · g-1), which is equivalent to an interregional clearance. The value of Pe was specified by setting the PSc for oxygen to a constant value of 20 ml · min-1 · g-1 and adjusting the extracapillary clearance rate to give the desired Pe value. The flow (Fc, in ml · min-1 · g-1) was set to the value determined by the local microsphere measurement. Because the experimentally determined arterial oxygen concentration and LMF are used, the model takes regional differences in oxygen appearance time into account. The concentration of 18O-labeled water in the capillary region can be summarized (4, 12) as
V<SUB>c</SUB><IT>·∂</IT>C<SUB>c</SUB><IT>/∂t=</IT>−F<SUB>c</SUB><IT>·L·∂</IT>C<IT>/∂x−</IT>[<IT>PS</IT><SUB>c</SUB><IT>·</IT>(C<SUB>c</SUB><IT>−</IT>C<SUB>e</SUB>)]
and the concentration of 18O-water in the extracapillary region is given by
V<SUB>e</SUB><IT>·∂</IT>C<SUB>e</SUB><IT>/∂t=</IT>P<SUB>e</SUB><IT>−PS</IT><SUB>c</SUB><IT>·</IT>(C<SUB>e</SUB><IT>−</IT>C<SUB>c</SUB>)
where partial C/partial x is the concentration change of 18O-labeled water along the length of the capillary, partial C/partial t is the concentration change as a function of time, and the subscripts c and e denote the capillary and extracapillary regions, respectively. L is an arbitrary capillary length, which is canceled by integration along the length of the capillary (6).

The total tissue residue (Qtiss) is the sum of the residues in the various model regions
Q<SUB>tiss</SUB><IT>=</IT>Q<SUB>art</SUB><IT>+</IT>Q<SUB>ven</SUB><IT>+</IT>Q<SUB>c</SUB><IT>+</IT>Q<SUB>e</SUB>
with Qart and Qven representing the residues of 18O-labeled water in the nonexchanging vessels, Qc representing that of the capillary region, and Qe representing that of the extracapillary region. The units for the tissue residues (µmol/g) can be converted into concentrations (µmol/ml) by dividing by the volume of the region (ml/g). To describe intracapillary concentration gradients along the vessel length realistically, the capillary region was represented by 10 segments of tissue exchange units in series. The flow and exchange processes of the capillary region are dealt with in the model by the Lagrangian sliding fluid element approach (3). The individual tissue regions were assumed to consist of parallel independent pathways, each differing from the others only with respect to the local flow. To account for possible effects of flow heterogeneity within tissue samples, 10 pathways of parallel capillary tissue exchange units were simulated (23). Thus each of the 10 parallel pathways contained a capillary region represented by 10 tissue exchange units arranged in series. The flow distribution chosen for the exchanging vessel segment was characterized by a coefficient of variation of 0.3 and a skew of 0.6.

Statistics. Data were processed on a personal computer using SYSTAT 5.0 software (SPSS, Chicago, IL) and are expressed as means ± SE. Correlations between local oxidative metabolism and LMF, coronary venous H218O kinetics and perfusion time, and global PSc value and global myocardial flow were determined by linear least-squares regression analysis. A P value <0.05 was considered to indicate statistical significance.


    RESULTS
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Global MVO2 assessment with analysis of oxidation water. During stable baseline hemodynamics, the global MVO2 in four hearts calculated on the basis of H218O production was >93% of the global MVO2 calculated by the standard Fick method (Table 1).

                              
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  		Table 1.   Global MVO2 from oxidation water versus measurements according to Fick.

Correlation of local H218O residue and LMF. Hemodynamic variables and MVO2 were as follows (n = 8): heart rate, 190 ± 15 beats/min; left ventricular pressure, 84 ± 12 mmHg; aortic flow, 52 ± 8 ml/min; coronary flow, 6.6 ± 0.7 ml · min-1 · g-1; and MVO2, 0.08 ± 0.01 ml · min-1 · g-1.

In 64 tissue samples, LMF ranged between 1.91 (low-flow areas) and 11.24 ml · min-1 · g-1 (high-flow areas). Local H218O residue varied between 0.41 (low-flow areas) and 1.04 µmol/g (high-flow areas). Linear regression analysis between both variables gave the equation H218O = 0.04 LMF + 0.44, with r = 0.62, n = 64, and P < 0.001 (Fig. 2).


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  		Fig. 2.   Linear regression between local myocardial flow (LMF) and local oxidative metabolism (local H218O residue after 240 s of 18O2 perfusion) in 8 isolated, Krebs-Henseleit (KH) buffer-perfused rabbit hearts (r = 0.62, n = 64, [H218O] = 0.04 LMF + 0.44, *P < 0.001).

The coronary venous kinetics of 18O-labeled oxidation water were determined from five rabbit hearts. Ten seconds after the onset of 18O perfusion, 18O-labeled oxidation water appeared in the coronary venous effluent with an increase of the H218O concentration (C) of 0.20 ± 0.12 µmol/ml. The increase became statistically significant after 40 s (0.35 ± 0.09 µmol/ml). Over time, the slope of the curve flattened, indicating that H218O synthesis and H218O removal were approaching equilibrium. At 240 s, the coronary venous H218O concentration was increased by 0.58 ± 0.12 µmol/ml. Fitting the results to a logarithmic function gave the following equation with a significant correlation: C = 0.14 · ln (perfusion time) - 0.16, with r = 0.96, n = 53, and P < 0.05 (Fig. 3).


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  		Fig. 3.   Coronary venous H218O kinetics in 5 isolated rabbit hearts perfused with 18O2-equilibrated KH buffer solution for 240 s. After 40 s, the increase of coronary venous H218O concentration (C) attained statistical significance (+0.35 ± 0.09 µmol/ml) and was in steady state after ~120 s (+0.54 ± 0.10 µmol/ml). At the end of perfusion (t = 240 s), the increase in C averaged 0.58 ± 0.12 µmol/ml [r = 0.96, n = 53, C = 0.14 ln (perfusion time) - 0.16, *P < 0.05].

Model analysis. Experimental measurements of global myocardial flow and arterial 18O2 concentration from nine hearts were used as model input parameter values. The experimental switch to perfusion with 18O2 was simulated by an inflow step increase of the 18O2 concentration from 0 to 1.19 ± 0.20 µmol/ml, which is equal to twice the concentration of molecular 18O2 in water at a partial pressure of 600 mmHg and a temperature of 38°C. Thus 1 mol of atomic oxygen yielded 1 mol of oxidation water. The model output was constrained by using the experimental value of oxygen consumption according to Fick. The PSc for water was used as a free parameter. Satisfactory model solutions were obtained for individual data sets by choosing a PSc between 2 and 10 ml · min-1 · g-1. PSc linearly correlated with the global myocardial flow (GMF) according to the equation PSc = 2.23GMF - 4.24, with r = 0.70 and P < 0.05 (Fig. 4). The average PSc value was 5.11 ± 2.56 ml · min-1 · g-1, and the median was 5 ml · min-1 · g-1. In each experiment, a single PSc value could be used for fitting the residue and outflow data. The agreement of experimental results and model solutions is summarized in Table 2.


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  		Fig. 4.   Linear regression between global myocardial flow (GMF) and permeability-surface area product for water (PSc) (n = 9, r = 0.70, PSc = 2.23GMF - 4.24, *P < 0.05).


                              
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  		Table 2.   Agreement between experiment and model

After calibration of this model with regard to its PSc value, it was used to calculate the local metabolic rate of oxygen. With a flow-proportional PSc value (Fig. 4), local flow differences between 3 and 10 ml · min-1 · g-1 corresponded to a 6.45-fold difference in the metabolic rate of oxygen.


    DISCUSSION
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Recent studies in anesthetized dogs indicate that local myocardial substrate metabolism is as heterogeneous as LMF (9, 11). However, substrate consumption depends strongly on experimental conditions and cannot be reliably extrapolated to oxidative metabolism.

Our technique of evaluating local myocardial oxidative metabolism by labeling oxidation water with 18O was intended to circumvent this problem (31, 32). Under physiological, normoxic conditions as used in this study, the myocardium incorporates about 98% of its oxygen uptake into oxidation water, whereas oxidant generation is negligible and amounts to only 1-2% (7, 34). Thus our technique is well suited for direct measurements of myocardial oxidative phosphorylation.

The application of oxygen isotopes to the assessment of oxidative metabolism has a long history. As early as 1960, 18O-labeled water was used to determine the turnover of energy-rich phosphates (ATP, ADP) in isolated muscle fibers (16). We have replaced 18O-labeled water with 18O-oxygenated buffer and extended this technique to use in whole hearts. Similarly, 15O2 was used in previous experiments on isolated, blood-perfused rabbit hearts (12), but only an extensive mathematical model analysis of tracer kinetics made it possible to distinguish between the produced oxidation water and nonmetabolized oxygen using this technique. In contrast, the present technique permits the analytic separation of the 18O-labeled water from the 18O-labeled oxygen. In another study in anesthetized dogs, 18O-labeled erythrocytes were injected into the coronary circulation, and the 18O tracer transient in the coronary venous outflow was analyzed (29). Because of sensitivity limitations, 18O-labeled water as reaction product could not be determined in the coronary venous effluent and, furthermore, the local tissue tracer residue was not analyzed. On the other hand, measurements of the tissue residue of 17O2 using nuclear magnetic resonance, 15O2 using positron-emission tomography, or 18O2 using mass spectrometry require mathematical model analysis to correct the time-dependent changes of the water tissue residues for the effects of water transport.

Capillary permeability-surface area product. The use of 18O-labeled water tissue residues in the present study provided physiologically significant data after the measurements were subjected to a rather simple mathematical model analysis. Assuming that one of the major model parameters influencing the results, namely, the PSc for water, was proportional to flow, the model estimated a 6.45-fold difference in local H218O production rate for a 3.33-fold difference in LMF.

Because the extraction fraction of oxygen can be increased by only about 50% in the myocardium, changes in flow are expected to be of similar magnitude to changes in oxidative metabolism. Thus we expected a difference smaller than 6.45-fold for the H218O production rate as predicted by the model. To obtain an estimate of the lower possible limit of the local H218O production rate, a constant PSc was also calculated from our data; this value (5.5 ml · min-1 · g-1) gave a 2.6-fold difference in local H218O production rate.

In the present study, the PSc for water was estimated by modeling the kinetics of intracellularly produced water. Previous studies have used intracoronary bolus injections of 3H-labeled water and analyzed the kinetics of the resulting coronary effluent concentration (1, 30). The key difference between these approaches is that, with parenchymal production of labeled water, there is only net outward diffusion into the capillary region, whereas there is a time-dependent net inward and net outward diffusion of labeled water in classic indicator dilution experiments. In one such previous study, the capillary wall PSc was estimated to be 2.8 ml · min-1 · g-1 (1), whereas in another, capillary and parenchymal cell membrane PSc was 18.0 and 3.8 ml · min-1 · g-1, respectively (30). These estimates refer to blood flow ranges between 0.8 and 1.5 ml · min-1 · g-1.

Our PSc values varied between 2 and 10 ml · min-1 · g-1 (mean = 5.11 ± 2.56 ml · min-1 · g-1) and were used to fit the kinetics of intracellularly produced water. To reduce the number of degrees of freedom in the model analysis, it was assumed that one lumped PSc value was sufficient to characterize the water-exchanging processes between parenchymal cells and the capillary region. Because parenchymal cell membranes and capillary walls represent two barriers in series, PSc may be calculated as PStot = 1/[(1/PS1) + (1/PS2)], where PStot represents the lumped PSc and PS1 and PS2 represent the individual permeability-surface area products. This simplification seems justified for the conditions of the present study, because the labeled water is produced in the extracapillary region, from which it slowly escapes into the capillary region. In agreement, the PSc value found in another study [3.8 ml · min-1 · g-1 (30)] falls within the range of lumped PSc values calculated in the present study.

One important limitation of the present modeling is the uncertainty of the estimated local PSc values and, hence, local MVO2. Previous studies conducted on the global heart have provided evidence for a significant direct relationship between flow rate and PSc (1, 19). Because both variables were correlated in the present experiments, a flow-proportional PSc was used for the analysis of local MVO2. However, mean coronary flow varied between 3.10 and 5.25 ml · min-1 · g-1 in the experiments that we used to calibrate the model with respect to the PSc, i.e., mean coronary flow did not cover a wide range. In addition, only a single estimate of PSc was obtained for each heart, ranging from 2 to 10 ml · min-1 · g-1.

On the other hand, we are confident that the PSc values used in the present study were reliable, because in contrast to most previous analyses, they were constrained by independent measurements of coronary outflow concentrations and tissue residues. There is only a narrow range in which the estimated PSc parameter fits both measures equally well. It is reassuring that the analysis of classic indicator dilution curves after intracoronary bolus injection of tracer water (1, 30) yielded results similar to those of the present study.

The relatively high 6.45-fold difference of local H218O can only be explained with difficulty. Using a constant PSc gives a value that is certainly too low, once a flow-proportional PSc is accepted (1, 19). An increase in the extraction fraction of oxygen would also increase local H218O. Thus we must attribute the uncertainty in estimating PSc values to an unknown rest.

H218O tissue residues and LMF. The 2.5-fold difference in the measured local H218O tissue residues represents a considerable underestimate of the true disparity of local oxidative metabolism between low- and high-flow areas. The relatively small difference of H218O residues between low- and high-flow areas (see Fig. 2) is due to the constant drain of 18O-labeled water with the coronary effluent, which is influenced by the following two factors: 1) the metabolic rate turns out to be higher in high-flow areas, which gives rise to a higher extravascular 18O-labeled water concentration in these areas; and 2) because of the higher flow rate, washout of vascular 18O-labeled water occurs more rapidly, thus lowering the concentration in the capillary region. These two factors result in a greater concentration difference between extracapillary and capillary regions in high-flow areas.

Errors in assessing LMF might also account for the flat slope of the relation between the H218O tissue residues and LMF, but they are likely to be small, because tissue sample mass averaged 100 ± 27 mg, which is close to the optimal size (80 mg) for local flow determinations in rabbit hearts using microspheres (5). By estimating the precision (V) of flow measurements according to V approx  1.96 · <RAD><RCD>1</RCD></RAD>/n (14, 28), with n = 600-4,000 microspheres per sample of 100 ± 27 mg, the methodological error ranged between 3 and 9% and therefore seems not to obscure the observed large flow heterogeneity. Potential temporal flow fluctuations, on the other hand, are unlikely to occur during 240 s and, if anything, would tend to underestimate LMF. Thus the 2.5-fold difference of local H218O residues as measured in this study must be regarded as a minimum difference associated with the observed 5.8-fold flow range.

In conclusion, we have quantitatively characterized the relationship between LMF and the production of oxidation water. Oxidative metabolism and hence MVO2 differ between low- and high-flow areas, and our data thus confirm previous studies (35, 36) using other techniques such as microbiopsies and spectrophotometry. In context with recent results on the lower flow threshold that is associated with an increase in local cytosolic adenosine concentration in myocardial low-flow areas (27), the results of the present study indicate that low-flow samples do not represent hypoperfused areas. In turn, high-flow samples do not represent regions of luxury perfusion but, rather, regions in which aerobic metabolism is above average (27). Because no local histological differences have so far been demonstrated and, in particular, because the mitochondria are uniformly distributed within myocardial tissue, we propose that oxidative metabolism is controlled by cellular mechanisms as yet unknown. Thus the present study provides the first experimental evidence that, in structurally homogeneous myocardium (13, 33), both MVO2 and flow are heterogeneous but correlated to each other. Whether low oxidative metabolism and low flow are secondary to low contractile function on the same spatial level, potentially representing physiological hibernation, remains unknown at present (20).


    ACKNOWLEDGEMENTS

We thank Prof. J. Schrader, who initiated these experiments many years ago. We gratefully acknowledge the thorough reading and correcting of the manuscript by Prof. S. Cleveland. We greatly appreciate the opportunity to use the model of oxygen transport and metabolism developed at the National Simulation Resource at the University of Washington (Dr. J. B. Bassingthwaighte, Director).


    FOOTNOTES

A. Deussen was a Heisenberg Fellow of the German Research Foundation, and U. Schwanke was supported by a fellowship from the German Cardiac Society. This study was supported by Deutsche Forschungsgemeinschaft Grants Schi 201/6-1 and He 1320/10-1.

Address for reprint requests and other correspondence: G. Heusch, Dept. of Pathophysiology, Univ. of Essen Medical School, Hufelandstrasse 55, D-45122 Essen, Germany (E-mail: gerd.heusch{at}uni-essen.de).

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Received 22 November 1999; accepted in final form 17 March 2000.


    REFERENCES
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

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Am J Physiol Heart Circ Physiol 279(3):H1029-H1035
0363-6135/00 $5.00 Copyright © 2000 the American Physiological Society


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