Effects of Changes in Brain Metabolism on the Levels of Citric Acid Cycle Intermediates

Abstract

The levels of all the free intermediates of the Krebs cycle except succinyl coenzyme A were measured in cerebral tissue of mice after periods of ischemia; anesthesia by Amytal, phenobarbital, and ether; hyperthermia; fluoroacetate poisoning; and insulin hypoglycemia. The levels of glucose, glucose 6-phosphate, pyruvate, phosphocreatine, adenosine triphosphate, adenosine diphosphate, and adenylic acid were also determined. During 5 sec of complete ischemia (decapitation) the levels of citrate, α-ketoglutarate, and oxalacetate fell 15 to 60% whereas the concentrations of succinate and fumarate rose 40% over a 30-sec interval. These changes presumably represent the conversion of a steady state system into an equilibrium system. Succinate accumulation is postulated to result from pyruvate carboxylation and fumarate reduction rather than from α-ketoglutarate oxidation. Decreasing the metabolic flux with anesthetic agents resulted in each case in marked decreases in α-ketoglutarate, fumarate, and malate, whereas increasing the flux by hyperthermia resulted in increases in these same three substrates. Neither anesthesia nor hyperthemia caused significant change in the levels of citrate, isocitrate, succinate, or oxalacetate. Fluoroacetate at first increased citrate levels without much effect on other substrates. Subsequently, when citrate had increased more than 3-fold, changes occurred in levels of the other Krebs cycle members. The accompanying rise in glucose 6-phosphate and fall in pyruvate suggested that inhibition of glycolysis at the phosphofructokinase step had taken place. Insulin hypoglycemia resulted in a general lowering of the levels of all Krebs cycle intermediates, as well as of glucose 6-phosphate and pyruvate. The changes associated with increasing, decreasing, or blocking flux in the Krebs cycle by the various procedures are used to assess the kinetic behavior of individual steps and to attempt to identify control points.