The Flux of Pyruvate in Perfused Rat Heart

Abstract

1.It was possible to find a steady-state lactate concentration in a closed-circuit heart perfusion containing added glucose. Insulin but not growth hormone (4 pglml) increased the equilibrium concentration in perfusate by 200 Ole. Intracellular lactate also remained constant and was about twice that in perfusate: this distribution is difficult to explain whether ionic or undissociated lactate is the permeating species.2. Pyruvate accumulated linearly in perfusate, reaching a peak after 30 min perfusion and then declined non-linearly. Insulin increased the peak concentration and growth hormone slowed the decline after the peak. The intracellular pyruvate concentration stayed constant and, except in the first few minutes, was lower than the perfusate concentration.3. Pyruvate addition a t zero time could inhibit pyruvate accumulation but altered neither the non-linear decline, the intracellular pyruvate level nor the lactate equilibrium. The [NAD]/[NADH]ratio, after 10 min of pre-perfusion on open circuit, was 23. During 2 h of perfusion the ratio rose slowly and then declined. If either insulin or growth hormone were present in the medium, the ratio fell to about 11 during the 6rst few minutes of perfusion and remained low for 90 min. These values could not easily be correlated with either the varying extracellular, or the constant intracellular [lactate]/[pyruvate] ratios.5. The glucose and oxygen consumption of the hearts during perfusion was measured, and the change in glycogen concentration in the tissue. It was then possible to draw up a balance sheet which indicated that the control hearts oxidied little other than carbohydrate for the 60 min of observation, but both insulin and growth hormone-treated hearts oxidised significant quantities of fatty acid during the first 15 min. It appears that growth hormone, like insulin, has an immediate effect on heart metabolism. The addition of pyruvate a t zero time appeared to stimulate carbohydrate oxidation in hearts perfused with insulin, even though the hormone had been present in the pre-perfusion fluid.6. Consideration of the variation in the extracellular [lactate]/[pyruvate] ratio with length of perfusion, and its lack of relationship to the intracellular [lactate]/[pyruvate] ratio, together with similar reports of other workers, leads us to conclude that neither lactate nor pyruvate establish a concentration gradient across the heart cell membrane which is based on passive diffusion. The discrepancy is particularly marked with respect to pyruvate. Our results strongly suggest that the extracellular [lactate]/[pyruvate] ratio cannot be used as an index of the cytoplasmic redox potential in perfused heart. 7.A tracer study of pyruvate metabolism in the reproducible non-steady-state conditions described can offer an explanation for the behaviour of the observed metabolic parameters : this study is reported in the following papers.The experiments which are reported in this and succeeding papers were designed to produce data that could be used to test a new metho...

The Computation of Parameters in Isotopic Tracer Systems not in a Steady‐State

“…It was therefore necessary to include in all optimisation runs, two parallel seta of equations of the type of Equation (a), one describing the behilviour of the total radioactivity (q) in each pool, and one set that of the total chemical Computing Methods The preliminary analyses, using the few mean specific activities of the perfusate lactate and pyruvate that were available, were carried out on a Solartron twin TR15 analog computer, using a Bryant x -y plotter to record the outputs. I n spite of the fact that so few separate time points were available for curve fitting (only 3, including zero time, for pyruvate), we found as discussed in detail earlier [8] that we were able to discard a number of unsuitable models without difficulty. This convinces us of the value of having a few really reliable points taken from, in our case, the mean of eight experiments, rather than a large number of points on a single curve which must be fitted by "curvepeeling".…”

“…or by applying the arbitrary restriction that R, = R-, and so on throughout the system. Similarly, it was found possible, when analysing the data described in the accompanying papers [7,8] and using only the radioactivity measurements (which did involve knowledge of the perfusate pyruvate pooI size), to get a perfect fit to the data which did not simulate the behaviour of the chemical pools a t all (Fig.1). It was therefore necessary to include in all optimisation runs, two parallel seta of equations of the type of Equation (a), one describing the behilviour of the total radioactivity (q) in each pool, and one set that of the total chemical Computing Methods The preliminary analyses, using the few mean specific activities of the perfusate lactate and pyruvate that were available, were carried out on a Solartron twin TR15 analog computer, using a Bryant x -y plotter to record the outputs.…”

“…Since in the work with which this analysis is associated ( [7,8] and this paper) a t least one of the chemical pools was varying in size during the tracer experiment, it is with this form of the equation that we shall be concerned. Q Q+ ...…”

“…This convinces us of the value of having a few really reliable points taken from, in our case, the mean of eight experiments, rather than a large number of points on a single curve which must be fitted by "curvepeeling". When it came to deciding which of the possible sources of perfusate pyruvate shown in Models C , D and E [7,8] best fitted the data, we found that stray currents in the quarter-squares multiplier prevented us from getting enough accuracy in the simulation when the intracellular pyruvate pool (1000 times smaller than the lactate pool) was to be multiplied by the function R, e-". From this point onwards the analysis was carried out by digital computer.…”

The effect of insulin on pyruvate dehydrogenase interconversion in heart muscle of alloxan-diabetic rats

“…However, a number of additional factors, such as pyruvate concentration, the ratios of NADH/ NAD and of acetyl-CoA/CoA, have been demonstrated in studies with isolated rat heart mitochondria to act on PDH interconversion [21,[27][28][29][30]. Pertaining to pyruvate, insulin neither causes an accunmlation in the perfusion medium (Table 1) nor in the heart tissue [31,32]. Also with respect to ATP and ADP insulin did not change the total cellular contents of perfused hearts from normal and diabetic animals [34,35].…”