Exogenously administered fructose 1,6-bisphosphate reportedly protects ischemic or hypoxic tissue and facilitates metabolic recovery. The mechanism of action of exogenous fructose 1,6-bisphosphate has been an issue of considerable debate, since there is a lack of direct evidence that fructose 1,6-bisphosphate can cross the cell membrane and act as an intermediate in glycolysis. We synthesized [1,6-13C]fructose 1,6-bisphosphate and directly examined its cellular metabolism in hog carotid artery segments using 13C-nuclear magnetic resonance (NMR) spectroscopy. [1,6-13C]fructose 1,6-bisphosphate (2.1 mM) was metabolized by hog carotid artery during normoxia and hypoxia with a major metabolic product being [3-13C]lactate. The production of [3-13C]lactate was greater during hypoxia than during normoxia, indicating that fructose 1,6-bisphosphate metabolism responded to the energetic state of the tissue. We found that exogenously added fructose 1,6-bisphosphate at 2.1 mM did not significantly improve the ability of hypoxic hog carotid artery to maintain isometric force, whereas 20 mM fructose 1,6-bisphosphate did significantly, although modestly, improve isometric force maintenance. These results indicate that exogenously added fructose 1,6-bisphosphate is capable of entering cells and serving as a glycolytic intermediate.

Vascular smooth muscle glycogen stores are traditionally thought to be small compared to other glycogen-containing tissues such as striated muscle or liver. However, glycogen has been thought to be an important carbon substrate for oxidative metabolism in support of contraction in vascular smooth muscle. We examined the synthesis and degradation of glycogen in isometrically mounted hog carotid artery using ¹³C-NMR spectroscopy. The rate of net glycogen synthesis from 1-¹³C-glucose was found to be constant during the first 8 h of incubation of carotid arteries with 10 mM glucose at 37 °C and then decreased towards a rate of zero by 14 h of incubation. During 8 h of incubation in the presence of 5 mM glucose, the content of glycogen increased from 1.5 to 8.1 µmol/g blot weight in the absence of insulin and to 11.4 µmol/g blot weight in the presence of 0.5 U/ml insulin. During prolonged glycogen loading, there was a simultaneous degradation of previously synthesized 6-¹³C-glycogen during synthesis of 1-¹³C-glycogen from 1-¹³C-glucose indicating substrate cycling of glycogen metabolism. This substrate cycling results in a pattern of glycogen utilization in which the most recently synthesized glucosyl units of glycogen are utilized only slightly more readily than the previously synthesized glucosyl units of glycogen. We conclude that glycogen stores are larger and more dynamic than previously thought in vascular smooth muscle consistent with an important role for glycogen as a carbon source for smooth muscle energy metabolism.

We examined the metabolism of exogenously added 13C-labeled fructose 1,6-bisphosphate (either labeled at the first and sixth carbons or labeled at the first carbon only) and of [2-13C]glucose in well-oxygenated and well-superfused hog carotid artery segments. Exogenously added fructose 1,6-bisphosphate was utilized by hog carotid artery and primarily participated in gluconeogenesis while the production of [3-13C]lactate was not significantly different from zero. When [1,6-13C]fructose 1,6-bisphosphate or [1-13C]fructose 1,6-bisphosphate was utilized individually, gluconeogenic flux occurred without metabolism through aldolase and triosephosphate isomerase resulting in formation of [1,6-13C]-glucose and [1-13C]glucose respectively. When [2-13C]glucose was the sole exogenous substrate, it was utilized and exclusively participated in glycolytic flux with production of [3-13C]lactate and no gluconeogenic flux from the trioses to [5-13C]glucose. When both glucose and fructose 1,6-bisphosphate were provided together as exogenous substrates, glucose still participated exclusively in glycolytic flux with no trioses participating in gluconeogenesis while fructose 1,6-bisphosphate participated in glycolytic flux with [3-13C]lactate production approximately being approximately half of the [1,6-13C]glucose production from [1,6-13C]fructose 1,6-bisphosphate. In the presence of glucose, [1-13C]fructose 1,6-bisphosphate also participated in glycolytic flux and gluconeogenic flux simultaneously. However in the presence of [2-13C]glucose, [1-13C]fructose 1,6-bisphosphate underwent isomerization through the trioses prior to gluconeogenesis since [6-13C]glucose was produced.(ABSTRACT TRUNCATED AT 250 WORDS)

The extent to and the mechanism by which fructose-1,6-bisphosphate (FDP) crosses cell membranes are unknown. We hypothesized that its transport is either via band 3 or a dicarboxylate transporter. The question was addressed in isolated Langendorff rat hearts perfused under normoxic conditions. Groups of hearts received the following metabolic substrates (in mM): 5 FDP; 5 FDP + either 5, 10, or 20 fumarate; 10 FDP and either 5, 10, or 20 fumarate; or 5 FDP + 2 4,4'-dinitrostilbene-2,2'-disulfonate (DNDS), a band 3 inhibitor. FDP uptake and metabolism were measured as production of [(13)C]lactate from [(13)C]FDP or (14)CO(2) and [(14)C]lactate from uniformly labeled [(14)C]FDP in sample perfusates. During 30 min of perfusion, FDP metabolism was 12.4 +/- 2.6 and 31.2 +/- 3.0 micromol for 5 and 10 mM FDP, respectively. Addition of 20 mM fumarate reduced FDP metabolism over a 30-min perfusion period to 3.1 +/- 0.6 and 6.3 +/- 0.5 micromol for 5 and 10 mM FDP groups, respectively. DNDS did not affect FDP utilization. These data are consistent with transport of FDP by a dicarboxylate transport system.

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