Pathways of Study and Methodological Approaches on Glucose Metabolism

(A) The Glycolytic / Gluconeogenic Pathways

The different components of the glycolytic / gluconeogenic pathways are depicted by the reactions shown in box A of Figure 1.

I) Level of Glucose-6-P

The control of blood glucose is the end result of many regulatory steps. The most immediate control is the balance between hepatic glucose phosphorylation versus hepatic glucose release (Nordlie, 1985). The former process depends on blood glucose concentration and the activity of glucokinase, and the latter depends on the glucose-6-P concentration and glucose-6-Pase. The glucose-6-P concentration is also regulated by interconnecting pathways of glycogen synthesis and release, and the isomerization of G-6-P to fructose-6-P by phosphoglucose isomerase.

There are several established tracer methods for the study of the balance of these reactions:

i) Direct and indirect pathways of glycogen synthesis 每 When specifically labeled glucose is given to a whole animal, it is well known that labeled liver glycogen molecules are of two kinds, those of the original label and those with the label rearranged indicating separate pathways of glycogen synthesis. When labeled glucose is incorporate directly, the label will remain intact and this process is presumed to represent the contribution of the activity of glucokinase to glycogen synthesis. Other labeled or unlabeled molecules are derived from pathways ending at the level of fructose-6-P. Direct and indirect pathways of glycogen synthesis represent the relative contribution of glucokinase and gluconeogenesis or recycling. There are a number of glucose isotopes that have been used for the study of direct and indirect pathway of glycogen synthesis. These are 1-¹³C-glucose (Roden et al., 1996),U¹³C-glucose (Katz and Lee, 1991; Katz et al, 1991), 1, 2-¹³C-glucose. The quantitation of direct and indirect pathway relies on the comparison of enrichment of enrichment in blood and glycogen glucose, or the ability to determine the enrichment of different positional isotopomers resulting from recycling of ¹³C carbons. When glycogen glucose is not accessible as in in vivo studies, acetaminophen-glucuronide can be used as a surrogate of glycogen glucose.

ii) Phosphoglucose isomerase:- The isomerization of G-6-P and F-6-P is measured using 2-²H-glucose. The conversion of aldose to ketose will cause an exchange of the hydrogen atom in the 2 position of glucose and the loss of the deuterium label. When 1-¹³C-glucose and 2-²H-glucose are given to an animal, the recovery of 1-¹³C-glucose and 2-²H-glucose in glycogen (or a glucuronide) can be used to determine the relative flux of glucokinase and the isomerase. (Landau et al., 1964)

iii) G-6-P futile cycle: The process by which glucose is phosphorylated in the liver and then dephosphorylated and released is known as the futile cycle. It is a measure of the balance between glucokinase and G-6-Pase. This process is estimated by the simultaneous measurement of glucose turnover using 1-¹³C-glucose and 2-²H-glucose. (Efendic et al, 1988) Since the isomerization between G6-P and F-6-P is rapid, the turnover of 2-²H-glucose will be greater than that measured by 1-¹³C-glucose. The difference is due to the recycling of 1-¹³C-glucose or the futile cycle.