Abstract
Metabolic control analyses of glucose utilization were performed for four groups of working rat hearts perfused with Krebs-Henseleit buffer containing 10 mM glucose only, or with the addition of 4 mM D-beta-hydroxybutyrate/1 mM acetoacetate, 100 nM insulin (0.05 unit/ml), or both. Net glycogen breakdown occurred in the glucose group only and was converted to net glycogen synthesis in the presence of all additions. The flux of [2-3H]glucose through P-glucoisomerase (EC 5.3.1.9) was reduced with ketones, elevated with insulin, and unchanged with the combination. Net glycolytic flux was reduced in the presence of ketones and the combination. The flux control coefficients were determined for the portion of the pathway involving glucose transport to the branches of glycogen synthesis and glycolysis. Major control was divided between the glucose transporter and hexokinase (EC 2.7.1.1) in the glucose group. The distribution of the control was slightly shifted to hexokinase with ketones, and control at the glucose transport step was abolished in the presence of insulin. Analysis of the pathway from 3-P-glycerate to pyruvate determined that the major control was shared by enolase (EC 4.2.1.1) and pyruvate kinase (EC 2.7.1.40) in the glucose group. Addition of ketones, insulin, or the combination shifted the control to P-glycerate mutase (EC 5.4.2.1) and pyruvate kinase. These results illustrate that the control of the metabolic flux in glucose metabolism of rat heart is not exerted by a single enzyme but variably distributed among enzymes depending upon substrate availability, hormonal stimulation, or other changes of conditions.
Highlights
Metabolic control analyses of glucose utilization were pmol/min/g to 12pmol/min/g while at the same time decreasing performed for four groups of working rat hearts per- the K, in the reverse direction from7 to 3 mM [4]
The flux control coefficients weredetermined for the study it was observed that [Glc,] was about 1.5 mM when hearts were perfused in the working mode, provided that extracellular glucose was elevated to the mildly hyperglycemic concentration of 16 mM a n d that the atrial pressure was maintained below 5 cm H,O to limit the work-related requirement portion of the pathway involving glucose transport to for substrate
Ysis of the pathwayfrom3-P-glycerate to pyruvate In contrast to the view that glucose transport is the “ratedetermined that the major control was shared by eno- limiting” or “pacemaker” reaction of glucose utilization, an allase (EC 4.2.1.1) and pyruvate kinase (EC 2.7.1.40) in the ternative view has been expressed that many, if not all, of t h e glucose group
Summary
The flux control coefficient can be determined indirectly by first calculating theelasticity (E), the fractional t o give maximal insulin responsein vitro [38]; or 4) 10 mM glucose plus the combination of ketone bodies and insulin During this period, a number of parameters of cardiac function were measuredas previously described [36]: aortic flow, coronary flow, left ventricular dPldt (Gould G4615-71, Valley View, OH), systolic aorticpressure(Spectramed P23X1, Oxnard CA), diastolic aortic pressure, mean aortic pressure, change in the net rate of a reaction catalyzed by a particular enzyme brought aboutby infinitesimal fractional changein the concentration of its substrate or product (Equation 6). The differences between top-down and bottom-up analyses are: 1) top-down analysis applies t o the whole of glucose metabolism, i.e. the effects of reactions beyond the block analyzed are included; 2) because differbelnotck elasticities were derivedby taking different pairsof the experimental sets, there is a single result from the top-down method, which is anapproximate average over the states considered, Fuller explanaatsiomnsodels are given below
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