Effects of aminooxyacetate on glutamate compartmentation and TCA cycle kinetics in rat hearts

The nonspecific transaminase inhibitor aminooxyacetate (AOA) has multiple influences on the dynamics of C-13 appearance in glutamate in rat hearts as measured by C-13 nuclear magnetic resonance (NMR) without altering O-2 consumption or tricarboxylic acid (TCA) cycle flux. These include the following: 1) a reduced rate of C-13 enrichment at glutamate C3 and C4; 2) a near coalescence of the C3 and C4 fractional enrichment curves; 3) a dramatic alteration in the time-dependent evolution of the glutamate C4 multiplets, C4S and C4D34; and 4) a decrease in the NMR visibility of glutamate. A fit of the C-13 fractional enrichment curves of glutamate C4 and C3 in the absence of inhibitor to a kinetic model of the TCA cycle gave values for transaminase flux of 7.5 mu mol.min(-1).g dry wt(-1) and TCA cycle flux of 7.5 mu mol.min(-1).g dry wt-l, thereby confirming reports by others that the kinetics of C-13 enrichment of glutamate C3 and C4 in heart tissue is significantly affected by flux through reactions other than TCA cycle. The C-13 fractional enrichment data collected in the presence of 0.5 mM AOA could not be fitted using this same kinetic model. However, kinetic simulations demonstrated that the time-dependent changes in C4S and C4D34 are only consistent with a 10-fold reduction in the size of intermediate pools undergoing rapid turnover in the TCA cycle. We conclude that inhibition of glutamic-oxalacetic transaminase by AOA effectively reduces the size of the alpha-ketoglutarate pool in rapid exchange with the TCA cycle. Our data indicate that changes in glutamate multiplet areas in the C-13 NMR spectra of heart (as demonstrated by glutamate C4S and C4D34) are more sensitive to alterations in metabolic pool sizes in exchange with the TCA cycle than are measurements of C-13 fractional enrichment at glutamate C3 and C4.