Increase In Glycolytic Flux Research Articles

Inhibition of Muscle Pyruvate Kinase by Creatine Phosphate

Abstract At neutral pH, concentrations of creatine phosphate comparable to those found in skeletal muscle and heart are highly inhibitory toward rabbit skeletal muscle pyruvate kinase. Creatine phosphate appears to be competitive with phosphoenolpyruvate with a Ki of 2.0 mm. That it is not a simple competition, however, is indicated by the following evidence. The inhibition is increased in the presence of high concentrations of ADP. ATP, a known inhibitor of pyruvate kinase, acts synergistically with creatine phosphate; that is, in the presence of both ATP and creatine phosphate the inhibition is much greater than the sum of the actions of the inhibitors separately. No synergism is seen between ATP and the inhibitory substrate analogue, 2-phosphoglyceric acid. The characteristics of the creatine phosphate effect are quite distinct from the inhibition noted previously with phenylalanine. Phenylalanine inhibition is maximal above pH 8.0 whereas creatine phosphate is more potent at neutral pH. Alanine, which reverses phenylalanine inhibition, has no effect on creatine phosphate inhibition. Phenylalanine does not inhibit the Mn2+-activated enzyme whereas creatine phosphate inhibition is more potent in the presence of manganous ion. Rabbit erythrocyte pyruvate kinase is only slightly inhibited by creatine phosphate. In the presence of the activator, fructose 1,6-diphosphate, the inhibition is much more pronounced. The results are discussed in relation to the regulation of muscle glycolysis and it is suggested that falling creatine phosphate levels may be one of the primary factors contributing to the increase in glycolytic flux that accompanies muscle contraction.

The effects of calcium ions on the activities of trehalase, hexokinase, phosphofructokinase, fructose diphosphatase and pyruvate kinase from various muscles.

1. The effects of Ca(2+) on the activities and regulatory properties of trehalase, hexokinase, phosphofructokinase, fructose diphosphatase and pyruvate kinase from vertebrate red and white muscle and insect fibrillar and non-fibrillar muscle have been investigated. These muscles were selected because of the possible difference in the role of glycolysis in energy production in the vertebrate muscles, and the possible difference in the role of Ca(2+) in the control of contraction in the two types of insect muscle. An increase in Ca(2+) concentration from 0.001mum to 10mum did not modify the activities nor did it modify the regulatory properties of these enzymes from these various muscles. 2. Concentrations of Ca(2+) above 0.1mm inhibited the activities of hexokinase and phosphofructokinase from the different muscles. It has been suggested that this inhibition may provide the basis for a theory of regulation of glycolysis (Margreth et al., 1967). If phosphofructokinase is located within the sarcoplasmic reticulum, its activity will be inhibited when the muscle is at rest, but the release of Ca(2+) from the reticulum during contraction will lead to a stimulation of its activity and hence an increase in glycolytic flux. The distribution of hexokinase and phosphofructokinase in the various cell fractions of these muscles was very variable. In particular, both enzymes were present almost exclusively in the 100000g supernatant fraction in the extracts of insect flight muscles. Thus there is no correlation between the properties of the enzymes and their distribution in muscle. 3. It is concluded that Ca(2+) does not control the activities of the important regulatory enzymes of glycolysis in muscle. It is suggested that in some muscles the sensitivity of the control mechanism at the level of phosphofructokinase to changes in the concentration of AMP may be increased by a process known as ;substrate-cycling'.

The effect of audiogenic seizures on regional CNS energy reserves, glycolysis and citric acid cycle flux.

AbstractSelected energy reserves, glycolytic intermediates and citric acid cycle intermediates were measured in the cerebral cortex, thalamus, brain stem, cerebellum and spinal cord of susceptible mice during audiogenic seizures. Changes in energy reserves (ATP, phosphocreatine and glucose) differed strikingly in extent and temporal pattern from region to region. The audiogenic seizure produced a transient, large decrease in thalamic energy reserves during the early, pretonic phase of the seizure. Less extensive decreases were observed in brain stem and spinal cord; but in these latter regions the changes persisted throughout the pretonic and tonic phases of the seizures. In cerebellum there was a biphasic decrease in energy reserves; a small decrease was observed immediately after the sound stimulus and a second much greater decrease was observed during the tonic phase of the seizure. No change in energy reserves was observed in cerebral cortex. Changes in glycolytic intermediates (glucose 6‐phosphate, fructose diphosphate, pyruvate and lactate) also varied from region to region in response to the decreases in energy reserves. In contrast, changes in the two citric acid cycle intermediates, α‐oxoglutarate and malate, were essentially the same in all regions studied. α‐Oxoglutarate decreased during the tonic phase of the seizure and rose during recovery. Malate remained at control levels throughout the seizure and then slowly increased.These findings are interpreted as indicating regional variations in nueronal activity during audiogenic seizures. During the period when clinical seizure activity is apparent neuronal activity increases in the subcortical regions. This is reflected by an increase in energy utilization and an increase in glycolytic flux in these areas. However, a concomitant increase in citric acid cycle flux does not seem to occur during this period. Citric acid cycle flux does appear to increase after the seizure is over.

Some neurochemical aspects of pentamethylenetetrazole convulsive activity in rat brain.

Abstract— The concentrations of several metabolites, including glucose, glycogen, hexose phosphates, adenine nucleotides, amino acids and some tricarboxylic cycle intermediates were estimated in cerebral tissue of rats killed at various stages of pentamethylenetetrazole convulsions. Prior to and during the seizure there was an increase in glycolytic flux and depletion of energy reserves. An accumulation of alanine concomitant with a decreased citrate level observed before the convulsion indicated that a possible interference of pyruvate entry into the tricarboxylic cycle had occurred. At the tonic extensor phase of the seizure and during post‐convulsive sedation a small but significant rise in GABA and an attempt to restore some of the labile constituents to more normal values was apparent. The significance of these results in relation to cerebral metabolic control and possible mechanisms of initiation and arrest of the seizure is discussed.

Regulation of Glycolysis in Brain, in Situ, during Convulsions

Abstract The concentrations of metabolites, including glucose, glycogen, glycolytic intermediates, adenine nucleotides, phosphates, amino acids, and citrate, in mouse brain in situ were measured from 15 to 180 sec after exposure of mice to the convulsant, Indoklon (bis(2,2,2-trifluoroethyl) ether). An increase in glycolytic flux of at least 3 times the basal rate was estimated for the brain in the convulsive state. During the period of hyperactivity, new steady state levels were established for several metabolites. In the transition, the concentrations of glucose and hexose monophosphates in the brain decreased. Coincident with these decreases, the concentration of fructose 1,6-diphosphate increased rapidly but transiently. The concentration of cerebral glycogen was unchanged during the first 30 sec, decreased about 20% during the next min, and then remained constant. Lactate accumulated throughout the period of convulsion. At the initiation of convulsion, the concentration of creatine phosphate fell precipitously to half its original value. The decrease in adenosine triphosphate was relatively small. Correspondingly, the concentrations of ADP, AMP, and inorganic phosphate in the brain increased rapidly with the induction of hyperactivity. The concentrations of citrate, glutamate, and aspartate did not change significantly. The level of proline was lowered during the convulsion. With the induction of convulsions and concomitant with the increased rate of glycolysis, the enzymes phosphofructokinase and hexokinase were coordinately activated. These two enzymatic sites represent the major control points of glycolysis in cerebral tissue in vivo. The ability of the brain to adjust to a new steady state following an increase in glycolytic rate, as well as its ability to maintain adequate supplies of energy reserves and oxygen, serves to distinguish the convulsive from the ischemic condition.

Regulation of Metabolism in Working Muscle in Vivo: I. CONCENTRATIONS OF SOME GLYCOLYTIC, TRICARBOXYLIC ACID CYCLE, AND AMINO ACID INTERMEDIATES IN INSECT FLIGHT MUSCLE DURING FLIGHT

The concentrations of 28 metabolites, including glycogen, trehalose, components of the glycolytic pathway and the Krebs cycle, and amino acids in flight muscle of the blowfly, Phormia regina, were measured concurrently after periods of flight ranging from 5 sec to 1 hour. At the onset of contraction, there was a rapid utilization of trehalose followed by catabolism of thoracic glycogen and then of the polysaccharide stored in the abdomen. Coincident with the decrease in the concentration of trehalose within the muscle was a rapid and transient increase in that of glucose. The concentrations of the hexose monophosphates were constant or declined. The concentration of fructose-1,6-diphosphate rose with the initiation of flight and then fell. The concentrations of triose phosphates, including α-glycerophosphate, increased slightly, but transiently, or were steady during continuous flight. The concentration of pyruvate increased strikingly at the start of exercise. Initially, pyruvate was largely converted to alanine. There were no accumulations of lactate or citrate. The amino group made available for the enormous increase in the concentration of alanine within the muscle after commencement of flight seems to have been derived largely from proline. The concentration of glutamate was lowered slightly during flight; no significant changes in the concentrations of ammonia, aspartate, glutamine, and asparagine within the muscle were found after initiation of flight. The concentration of malate increased sharply at the onset of active contraction, declined slightly for 2 min, and then remained high throughout the entire flight. The concentrations of α-ketoglutarate and oxaloacetate were apparently not altered. Measurements of coincident changes in metabolites concomitant with the 100-fold increase in glycolytic flux attained on initiation of flight provided an unique system to determine the steps that control glycolysis in vivo, in the transition from a tissue at rest to one performing strenuous work. Three sites of regulation—the phosphorylation of fructose 6-phosphate, the cleavage of trehalose, and the phosphorolysis of glycogen—were established in this system in vivo. Possible additional controls at the oxidations of α-glycerophosphate and of proline are suggested.