Pyruvate Dehydrogenase Activity in the Rat Cerebral Cortex following Cerebral Ischemia

Abstract

Summary: The effect of cerebral ischemia on the activity of pyruvate dehydrogenase (PDH) enzyme complex (PDHC) was investigated in homogenates of frozen rat cerebral cortex following 15 min of bilateral common ca rotid occlusion ischemia and following 15 min, 60 min, and 6 h of recirculation after 15 min of ischemia. In frozen cortical tissue from the same animals, the levels of labile phosphate compounds, glucose, glycogen, lactate, and pyruvate were determined. In cortex from control ani mals, the rate of [l_14C]pyruvate decarboxylation was 9.6 ± 0.5 nmol CO2/(min-mg protein) or 40% of the total PDHC activity. This fraction increased to 89% at the end of 15 min of ischemia. At 15 min of recirculation following 15 min of ischemia, the PDHC activity decreased to 50% of control levels and was depressed for up to 6 h post ischemia. This decrease in activity was not due to a de crease in total PDHC activity. Apart from a reduction in ATP levels, the acute changes in the levels of energy metabolites were essentially normalized at 6 h of recov ery. Dichloroacetate (DCA), an inhibitor of PDH kinase, given to rats at 250 mg/kg i.p. four times over 2 h, signifThe pyruvate dehydrogenase (POH) complex (POHC) is a mitochondrial multienzyme complex composed of three enzyme activities, participating in the decarboxylation of pyruvate and formation of acetyl-CoA (Reed, 1981). It is a central enzyme in the metabolism of glucose and in acetylcholine syn thesis (Tucek and Cheng, 1974; Gibson et aI. , 1975), but has also been proposed to be important in neu ronal plasticity and accommodative processes via effects on the cellular calcium ion homeostasis (Morgan and Routtenberg, 1981; Browning et aI. , 1981;Baudry and Lynch, 1985 350icantly decreased blood glucose levels from 7.4 ± 0.6 to 5.1 ± 0.3 mmoUL and fully activated PDHC. In animals in which the plasma glucose level was maintained at control levels of 8. 3 ± 0.5 IJ-mol/g by intravenous infusion of glu cose, the active portion of PDHC increased to 95 ± 4%. In contrast, the depressed PDHC activity at 15 min fol lowing ischemia was not affected by the DCA treatment. In both DCA + glucose-treated control and recovery groups, the pyruvate levels decreased by 50%. No signif icant difference in the lactate levels was seen. We con clude that the depressed postischemic PDHC activity is not due to loss of enzyme protein nor to an increased PDH kinase activity, but is probably due to a decreased activity of PDH phosphatase. This could in turn be sec ondary to a change in the cellular levels of PDH phos phatase regulators, most probably a decreased intramito chondrial concentration of calcium. The postischemic de crease in PDH activity may be related to the postischemic metabolic depression. Key Words: Brain-Dichloro acetate-Ischemia-Pyruvate dehydrogenase-Rat.Phosphorylation-dephosphorylation of the a-subunit protein of POHC regulates the enzyme activity via reactions catalyzed by a PDH phos phatase that activates, and a PDH kinase that inac tivates, the enzyme (for review see Reed,...

Preischemic Hyperglycemia and Postischemic Alteration of Rat Brain Pyruvate Dehydrogenase Activity

“…This study confirms the previously reported post ischemic depression in PDHC activity in normogly cemic animals (Cardell et al, 1989) and extends the observation to 18 h of recovery. In addition, our present results show that PDHC activity was re-…”

Postischemic hyperoxia reduces hippocampal pyruvate dehydrogenase activity

“…However, given our in vitro experiments that show that "active" purified PDHC is targeted and inactivated by peroxynitrite, the most likely mechanism of enzyme inhibition is that PDHC in its active, dephosphorylated form is susceptible to attack and subsequent inactivation by peroxynitrite. In addition, loss of PDHC activity after cerebral ischemia and reperfusion in rat arterial occlusion models is the result of impaired maximal activity rather than covalent inhibition caused by increased protein phosphorylation [21,54].…”

“…However, given our in vitro experiments that show that "active" purified PDHC is targeted and inactivated by peroxynitrite, the most likely mechanism of enzyme inhibition is that PDHC in its active, dephosphorylated form is susceptible to attack and subsequent inactivation by peroxynitrite. In addition, loss of PDHC activity after cerebral ischemia and reperfusion in rat arterial occlusion models is the result of impaired maximal activity rather than covalent inhibition caused by increased protein phosphorylation [21,54].The results obtained in vivo suggesting that peroxynitrite mediates postischemic PDHC enzyme inhibition are supported by in vitro experimentation in which incubation of purified PDHC with SIN-1 is associated with decreased PDHC activity and increased 3-nitrotyrosine immunoreactivity The inactivation of PDHC by peroxynitrite, in vitro, is primarily due to tyrosine nitration, although the small protection afforded by DTT indicates that S-nitrosation also contributes to enzyme inhibition (Fig. 6).…”

NADH hyperoxidation correlates with enhanced susceptibility of aged rats to hypoxia

“…The inhibition of this enzyme is suggested to occur as a result of free radical mediated protein oxidation and may be potentiated by increased calcium concentrations (Bogaert et al, 1994). Inhibition of the PDHC (to 50% of control levels) takes place as early as 15 min following reperfusion (Cardell et al, 1989) consistent with the occurrence of NADH hyperoxidation following hypoxia (as shown in this study) and ischemia (Feng et al, 1998). Inhibition may, as a result, restrict the supply of reducing equivalents and substrates to mitochondria thereby decreasing aerobic metabolism.…”