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PubMed1994

Incorporation of body components of diverse microorganisms by larval mosquitoes.

A pulse-purge schedule of exposure to labeled microorganisms was used to compare their digestibility by larval mosquitoes. Larvae were placed for an hour in suspensions of diverse axenically grown microorganisms that had been labeled with radioactive carbon (in the form of glucose or glycine). The guts of these mosquitoes were then purged with nonlabeled Sephadex particles for 30 min, and retained radioactivity was measured. Larvae imbibed no dissolved material. Larval mosquitoes differ in their capacity to derive label from algae (sensu lato), and certain algae contribute more label to these mosquitoes than do others. The nature of any algal food, as well as the feeding habits and developmental stage of the larva, influence its capacity to derive label from algae. This pulse-purge method of analysis can assist in the selection of algal "vectors" suitable as vehicles for transgenic larvicide. Although larval mosquitoes fail to assimilate the contents of Palmellacoccus cells with which they are confined, as much as 1/3 of the body contents of a Euglena gracilis cells become incorporated into their bodies. Because larval mosquitoes internalize more material from Euglena than they do from various other algae, these microorganisms provide a promising candidate vehicle for transgenic Bacillus thuringiensis israelensis.

Aedes↗

PubMedDate not supplied

Effects of insulin on urinary calcium excretion by diabetic rats in the absence of blood glucose control.

Male rats weighing approximately 250 g were given streptozotocin (STZ), 60 mg/kg, i.p., on two successive days. Six h after the last STZ injection, some STZ-diabetic rats began receiving daily injections of insulin (NPH, 1.5 U, s.c.). At various times after the above treatments the following studies were performed: control rats, STZ-diabetic rats and NPH-treated STZ-diabetic rats were injected s.c. with calcium-45 and housed individually in metabolic cages. Urine from each rat was collected at 24 h intervals for two weeks. Volumes, osmolalities, glucose concentrations, total calcium and radioactivity due to calcium-45 were measured. The results of the studies showed that: STZ-diabetes caused rapid urinary excretion of the injected 45Ca and increased renal excretion of 40Ca; the increased urinary excretion of calcium was not due to an osmotic effect or to a direct nephrotoxic action of STZ; and insulin therapy instituted early, but insufficient to control blood glucose, significantly reduced the diabetes-induced calcium loss.

Animals↗

PubMed1990

Effects of maternal undernutrition and exercise on glucose kinetics in fetal sheep.

Fetal glucose kinetics were measured using a combination of isotope-dilution and Fick-principle methodology in single-pregnant ewes which were either well-fed throughout, or fed at 0.3-0.4 predicted energy requirement for 7-21 d during late pregnancy. All ewes were studied while standing at rest and then while walking on a treadmill at 0.7 m/s on a 10 degree slope for 60 min. Underfed ewes suffered major decreases in fetal total disposal rate, fetal-placental transfer and umbilical net uptake of glucose, each of which were significantly related to declines in maternal and fetal blood glucose concentrations respectively. In well-fed ewes, fetal endogenous glucose production was negligible, as indicated by the similarity between fetal utilization rate (total glucose disposal rate minus placental uptake of fetal glucose) and umbilical net uptake of glucose, and by nearly identical fetal and maternal arterial blood specific radioactivities of maternally infused D-[2-3H]glucose. By contrast, in underfed ewes, fetal utilization rate greatly exceeded umbilical net uptake of glucose, and the fetal:maternal [3H]glucose specific activity ratio declined significantly, suggesting induction of a substantial rate of fetal endogenous glucogenesis. Exercise caused increases in fetal total glucose disposal rate and glycaemia in fed and underfed ewes. In underfed ewes only, this was accompanied by increased placental uptake of fetal glucose and umbilical net glucose uptake, unchanged fetal glucose utilization and decreased fetal endogenous glucose production. It is concluded that fetal gluconeogenesis makes a major contribution to fetal glucose requirements in undernourished ewes. Increased maternal supply of fetal glucose during exercise substitutes for rather than adds to fetal endogenous glucogenesis.

Animals↗

PubMed2000

Inhibitory action of the anomers of 2-deoxy-D-glucose tetraacetate on the metabolism of D-glucose in rat pancreatic islets.

BACKGROUND: The tetra-acetate ester of 2-deoxy-D-glucose was recently found to either inhibit or augment insulin secretion, depending on the concentration of the ester. Both the positive and negative insulinotropic actions of the ester display anomeric specificity. METHODS: The effects of the alpha- and beta-anomer of 2-deoxy-D-glucose tetra-acetate (5.0 mM) on the metabolism of D-[5-3H]glucose and D-[U-14C]glucose (8.3 mM) were investigated in isolated rat pancreatic islets. RESULTS: Both the alpha- and beta-anomers of 2-deoxy-D-glucose tetra-acetate inhibited the generation of 3HOH from D-[5-3H]glucose and that of 14CO2, as well as radioactive acidic metabolites and amino acids, from D-[U-14C]glucose. They also lowered the paired ratio between D-[U-14C]glucose oxidation and D-[5-3H]glucose utilization. No significant anomeric difference could be detected, however, in these experiments. CONCLUSIONS: The effects of the alpha- and beta-anomer of 2-deoxy-D-glucose tetra-acetate on the metabolism of D-glucose in isolated rat pancreatic islets reinforce the view that the insulinotropic action of monosaccharide esters involves a dual mode of action, linked to both the metabolic effects of their glucidic moiety and a direct interaction of the esters themselves with a stereospecific receptor system.

Animals↗

PubMed1985

The disposal of an oral glucose load in healthy subjects. A quantitative study.

Although it is an established concept that the liver is important in the disposition of glucose, the quantitative contribution of the splanchnic and peripheral tissues, respectively, to the disposal of an oral glucose load is still controversial. In the present investigation, we have employed the hepatic venous catheter technique in combination with a double-tracer approach (in which the glucose pool is labeled with 3H-glucose and the oral glucose load is labeled with 14C-glucose) to quantitate the four determinants of oral glucose tolerance: rate of oral glucose appearance, splanchnic glucose uptake, peripheral glucose uptake, and suppression of hepatic glucose production. Studies were carried out in 11 normal volunteers in the overnight-fasted state and for 3.5 h after the ingestion of glucose (1 g/kg body wt; range, 55-93 g). In the postabsorptive state, the rate of endogenous (hepatic) glucose production, evaluated from the 3H-glucose infusion, was 2.34 +/- 0.06 mg/min X kg. Glucose ingestion was accompanied by a prompt reduction of endogenous glucose output, which reached a nadir of 0.62 +/- 0.23 mg/min X kg at 45 min and remained suppressed after 3.5 h (0.85 +/- 0.22 mg/min X kg). The average inhibition of hepatic glucose output during the absorptive period was 53 +/- 5%. The appearance of ingested glucose in arterial blood, as derived from the 14C-glucose measurements after correction for recycling 14-C radioactivity, reached a peak after 15-30 min, and 14C-glucose continued to enter the systemic circulation throughout the observation period. The rate of appearance of ingested glucose was 2.47 +/- 0.45 mg/min X kg at 3.5 h. A total of 73 +/- 4% of the oral load was recovered in the systemic circulation within 3.5 h.(ABSTRACT TRUNCATED AT 250 WORDS)

Administration, Oral↗

PubMed1983

Ribose. An oxidation product of glucose 6-phosphate in microsomal fraction.

An oxidative metabolism of glucose 6-phosphate was studied in rat liver microsomal fraction. Although radioactive 14CO2 was formed from [1-14C]glucose 6-phosphate in the microsomal fraction (Hino, Y., and Minakami, S. (1982) J. Biochem. (Tokyo) 92, 547-557), the formation was negligible when [2-14C]glucose 6-phosphate was used as a starting substrate. These results indicated an inability of the microsomal fraction to rearrange [2-14C]glucose 6-phosphate to form [1-14C] glucose 6-phosphate, and it was expected that a certain compound derived from glucose 6-phosphate accumulated as an end-product of the reaction. We, therefore, have tried to identify the product by high performance liquid chromatography, and found that ribose accumulated as the end-product. The formation of ribose was inhibited in the same manner as that of 14CO2 by antibodies against rat liver microsomal hexose-6-phosphate dehydrogenase, and the ratios of ribose to 14CO2 formed in the reaction were 0.5-0.8 on a molar basis. The finding of ribose formation further suggested the involvement of ribose phosphate isomerase and phosphatase activities in the reaction.

Animals↗

PubMed1983

Precursor of carbon atom five and hydroxymethyl carbon atom of the pyrimidine moiety of thiamin in Escherichia coli.

The pyrimidine moiety of thiamin can be synthesized in bacteria from 5-aminoimidazole ribotide (AIR), an intermediate in purine biosynthesis. To transform the imidazole ring of AIR to the pyrimidine, the bond between C-4 and C-5 of the imidazole ring should be ruptured and a two-carbon unit should be inserted. We investigated a precursor of the two-carbon unit which should be inserted to form the pyrimidine. To examine the possibility of the two-carbon compound as a precursor which is formed from a three-carbon intermediate in the glycolytic pathway and its three-carbon derivative by decarboxylation in cells, we studied the incorporation of [2-14C]glycerol, [2-14C]pyruvate, [U-14C]alanine and [3-14C]serine into the pyrimidine by Escherichia coli. Glycerol, pyruvate and alanine were not incorporated into the pyrimidine. Radioactive carbon of [3-14C]serine was incorporated into C-2 of the pyrimidine via one-carbon units pool. Then, we investigated the possibility of a ribose moiety of AIR being a precursor of the two-carbon unit. As ribose has low permeability into E. coli cells, the ribose moiety of AIR was labeled with [14C]glucose which was added to the growth medium. The results show that two radioactive carbons of [U-14C]glucose were incorporated into the pyrimidine and radioactive carbon of [6-14C]glucose was incorporated into hydroxymethyl carbon attached to C-5 of the pyrimidine. The dilution rates of the specific radioactivity of the pyrimidine from [U-14C]glucose and [6-14C]glucose well coincide with those of the ribose moiety of the nucleotide (AMP). Radioactive carbon of [1-14C]glucose was not incorporated into the pyrimidine and nucleotide. It is concluded that the two-carbon fragment derived from C-4' and C-5' of the ribose moiety of AIR can be incorporated into the pyrimidine moiety of thiamin as the precursor of C-5 and hydroxymethyl carbon.

Alanine↗

PubMed1993

Reverse iontophoresis: development of a noninvasive approach for glucose monitoring.

Solvent flow generated during iontophoresis can be used to convect neutral molecules through the skin, thereby greatly enhancing their flux. This concept was exploited to realize noninvasive glucose measurement by its iontophoretic extraction from the subcutaneous tissue. The hypothesis was tested in vitro using hairless mouse skin. The dermal surface was bathed with a glucose solution; chambers on the epidermal surface housed the current delivery electrodes. Iontophoresis (at 0.36 mA/cm2) was performed for 2 hr, at the end of which the solutions in contact with the electrodes were analyzed. The amount extracted was proportional to the glucose solution concentration bathing the dermis. Higher radioactivity levels were found at the anode than at the cathode, possibly because of glucose metabolism during its outward transport across the skin. Glucose biotransformation results in negatively charged metabolites which migrate to the anode. Two sensitive glucose sensors were developed; one was selective for glucose, the other for glucose and related compounds. Both sensors indicated the presence of glucose at the cathode but an abnormally high value was also recorded at the anode. This signal, however, was not due to glucose but rather to electroactive ascorbate withdrawn from the skin. Finally, a system has been developed with which glucose can be extracted noninvasively from the subcutaneous tissue and unambiguously measured. Whether iontophoretic glucose sampling in vivo will be equally successful remains to be answered.

Animals↗

PubMed1978

Evidence for catabolite degradation in the glucose-dependent inactivation of yeast cytoplasmic malate dehydrogenase.

The cytoplasmic malate dehydrogenase of Saccharomyces cerevisiae was radioactively labeled during its synthesis on a glucose-free derepression medium. After purification a sensitive radio-immunoassay for this enzyme could be developed. The assay showed that after the physiological, glucose-dependent 'catabolite inactivation' of cytoplasmic malate dehydrogenase an inactive enzyme protein is immunologically not detectable. Together with the irreversibility of this reaction in vivo this finding strongly suggest a proteolytic mechanism of enzyme inactivation. For this process the term 'catabolite degradation' is used.

Antibody Specificity↗

PubMed1993

Localization of four new markers to pig chromosomes 1, 6, and 14 by radioactive in situ hybridization.

The genes coding for glucose regulated protein, 78kDal (GRP78), hormone-sensitive lipase (LIPE), plasminogen activator or urokinase (PLAU), and D-amino acid oxidase (DAO) were localized in the pig by radioactive in situ hybridization. GRP78 was mapped to 1q2.10-->q2.13 and LIPE was localized to chromosome 6cen-->q1.2. The genes for PLAU and DAO were both assigned to chromosome 14, in the region q2.4-->q2.6 and q2.1-->q2.3, respectively. The results are compared to mapping data in other mammalian species.

Animals↗

PubMed1978

Association between 5-hydroxytryptamine release and insulin secretion.

The efflux of radioactivity after loading with trace amounts of tritiated 5-hydroxytryptamine ([3H]5-HT) or 5-hydroxytryptophan ([3H]5-HTP) was studied in perifused beta-cell-rich pancreatic islets from ob/ob mice. Analysis of the effluent revealed that more than 90% of the radioactivity was released as [3H]5-HT after loading with [3H]5-HTP. Increasing the concentration of glucose in the perifusion medium from 3 to 20 mmol/l enhanced the efflux when islets from fed mice were used and this effect was potentiated by the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (IBMX). Whereas 20 mM-glucose alone did not stimulate the efflux of 5-HT from islets isolated from mice starved for 3 days, a stimulatory effect was observed in the presence of IBMX. Stimulation of the efflux of radioactivity by glucose was inhibited if calcium was omitted from or adrenaline added to the medium. The results are consistent with the concept of exocytotic release of 5-HT occurring in response to stimulation of insulin secretion, although basal non-exocytotic transport must also be occurring across the beta-cell membrane.

5-Hydroxytryptophan↗

PubMed1972

The biosynthesis in vitro of keratan sulphate in bovine cornea.

1. Bovine corneas were incubated in vitro with [U-(14)C]glucose. 2. The glycosaminoglycans of corneal stroma were isolated and fractionated on cetylpyridinium chloride-cellulose columns. The major components were keratan sulphate (71%), chondroitin 4-sulphate (17%) and chondroitin 6-sulphate (4%). 3. The acid-soluble nucleotides and intermediates of glycosaminoglycan biosynthesis of corneal stroma were separated on Dowex 1 (formate form) and the tissue content and cellular concentrations were determined. 4. The rates of synthesis of the intermediates of glycosaminoglycan biosynthesis in corneal stroma were determined. 5. The incorporation of radioactivity from [U-(14)C]glucose into the uronic acid and hexosamine components of the glycosaminoglycans present in corneal stroma were measured and the turnover rates of these polymers were calculated. It was found that keratan sulphate was turning over in about 723h and chondroitin 6-sulphate in 251h.

Animals↗

PubMed2003

Assessment of treatment response by autoradiography with (14)C-aminocyclopentane carboxylic acid, (67)Ga-DTPA, and (18)F-FDG in a herpes simplex virus thymidine kinase/ganciclovir brain tumor model.

UNLABELLED: Assessments of herpes simplex virus 1 thymidine kinase (HSV-tk)/ganciclovir (GCV) treatment response, early in the course of therapy, are important in the evaluation and clinical management of patients. This study addresses whether imaging amino acid transport, glucose utilization, and passive vascular permeability provides an early indication of treatment response and can predict long-term outcome. METHODS: Fischer 344 rats with intracerebral HSV-tk transduced RG2TK+ xenografts were studied. GCV-treated (50 mg/kg twice daily) and saline-treated control animals were compared; triple-label quantitative autoradiography was performed 3 d after initiating treatment, and long-term survival was determined. Autoradiograms of (18)F-FDG, (67)Ga-diethylenetriaminepentaacetic acid ((67)Ga-DTPA), and (14)C-aminocyclopentane carboxylic acid ((14)C-ACPC) were obtained; measurements of (14)C-ACPC and (67)Ga-DTPA plasma clearance (K(1)), (14)C-ACPC transport ( partial differential K(1)), relative glucose utililization (R), and normalized radioactivity (% dose/g) were obtained in tumor and brain tissues. Adjacent sections were stained to detect apoptotic cells, microvessels, and type L neutral amino acid transporter in tumor and normal brain. RESULTS: GCV treatment reduced partial differential K(1) and % dose/g of (14)C-ACPC in RG2TK+ xenografts to approximately 30% of that in nontreated animals (from 34 +/- 9 [mean +/- SD] to 9.5 +/- 2.7 microL/min/g and from 0.28 +/- 0.09 to 0.11 +/- 0.04 % dose/g, respectively). GCV had a significant but substantially smaller effect than toxicity on glucose utilization and little or no effect on passive vascular permeability of RG2TK+ xenografts. These differences could not be explained by differences in plasma amino acid or glucose concentration at the time of the study. Histology revealed a large fraction of dead tumor cells and only a sparse distribution of apoptotic cells in GCV-treated tumors. Many CD34-positive endothelial cells in GCV-treated tumors showed only weak or marginal LAT1 staining, whereas CD98 staining remained unchanged. Survival was significantly increased by GCV treatment from 18 +/- 4 to 56 +/- 17 d. CONCLUSION: (14)C-ACPC influx, K(1)(ACPC), facilitated transport, partial differential K(1)(ACPC), and % dose/g (ACPC) are good indicators of early treatment response after HSV-tk/GCV gene therapy. The parametric images and changes in K(1)(ACPC), partial differential K(1)(ACPC), and % dose/g (ACPC) are substantial and are better than the corresponding measures obtained in the same animals and in the same tissue (tumor) regions with (67)Ga-DTPA and (18)F-FDG. Amino acid transport imaging may be a good surrogate paradigm to monitor treatment response of brain tumors.

Animals↗

PubMed1995

Synthesis of fluoroacetate from fluoride, glycerol, and beta-hydroxypyruvate by Streptomyces cattleya.

Streptomyces cattleya produces fluoroacetate and 4-fluorothreonine from inorganic fluoride added to the culture broth. We have shown by 19F nuclear magnetic resonance (NMR) spectrometry that fluoroacetate is accumulated first in the culture broth and that accumulation of 4-fluorothreonine is next. To show precursors of the carbon skeleton of fluoroacetate, we carried out tracer experiments with various 14C- and 13C-labeled compounds. Radioactivity of [U-14C]glucose, [U-14C]glycerol, [U-14C]serine, and [U-14C]beta-hydroxypyruvate was incorporated into fluoroacetate to an extent of 0.2 to 0.4%, whereas [3-14C]pyruvate, [2,3-14C]succinate, and [U-14C]aspartate were less efficiently incorporated (0.04 to 0.08%). The addition of [2-13C]glycerol to the mycelium suspension of Streptomyces cattleya caused exclusive enrichment of the carboxyl carbon of fluoroacetate with 13C; about 40% of carboxyl carbon of fluoroacetate was labeled with 13C. We studied the radioactivity incorporation of [3-14C]-, [U-14C]-, and [1-14C]beta-hydroxypyruvates to show that C-2 and C-3 of beta-hydroxypyruvate are exclusively converted to the carbon skeleton of fluoroacetate. These results suggest that the carbon skeleton of fluoroacetate derives from C-1 and C-2 of glycerol through beta-hydroxypyruvate, whose hydroxyl group is eventually replaced by fluoride.

Carbon Isotopes↗

PubMedDate not supplied

Effects of fructose on D-[6-3H]-glucose uptake and sorbitol metabolism of bovine retina in vitro.

In order to study the influence of fructose on sorbitol formation, bovine retinal tissue was incubated with different concentrations of glucose and fructose, and supplemented with tracer amounts of D-[6-3H]-glucose. Combining high-performance liquid chromatography (HPLC) with radioactivity determinations allowed detection of sorbitol and fructose derived from glucose in the incubation medium. In addition, the total amount of sorbitol was measured with a sensitive bioluminescence method. In this way, it was possible to distinguish between sorbitol formation from glucose and fructose. High concentrations of glucose in the medium increased the formation of sorbitol and fructose from glucose. Addition of fructose to the incubation medium diminished the sorbitol and fructose formation from glucose although the total amount of sorbitol increased significantly. Incubating retinal tissue with an aldose reductase inhibitor decreased sorbitol formation from glucose but did not influence the formation of sorbitol from fructose. Thus, the present findings clearly demonstrate the important influence exerted by fructose on sorbitol formation. The possible significance of the present finding is discussed with respect to diabetes retinopathy and retinal osmoregulation.

Animals↗

PubMed1998

Lactate is released and taken up by isolated rabbit vagus nerve during aerobic metabolism.

To determine if lactate is produced during aerobic metabolism in peripheral nerve, we incubated pieces of rabbit vagus nerve in oxygenated solution containing D-[U-14C]glucose while stimulating electrically. After 30 min, nearly all the radioactivity in metabolites in the nerve was in lactate, glucose 6-phosphate, glutamate, and aspartate. Much lactate was released to the bath: 8.2 pmol (microg dry wt)(-1) from the exogenous glucose and 14.2 pmol (microg dry wt)(-1) from endogenous substrates. Lactate release was not increased when bath PO2 was decreased, indicating that it did not come from anoxic tissue. When the bath contained [U-14C]lactate at a total concentration of 2.13 mM and 1 mM glucose, 14C was incorporated in CO2 and glutamate. The initial rate of formation of CO2 from bath lactate was more rapid than its formation from bath glucose. The results are most readily explained by the hypothesis that has been proposed for brain tissue in which glial cells supply lactate to neurons.

Aerobiosis↗

PubMed1995

Kinetic analysis of thyroid hormone action on glucose metabolism in man.

Thyroid hormone action on insulin's effect on glucose kinetics was investigated with the use of a physiological three compartment model. In six healthy volunteers before and after 14 days of thyroxine treatment (300 micrograms/day), a bolus of [3-H3]glucose was injected and the time course of plasma radioactivity was followed closely for 150 min. Then a hyperinsulinemic (1 mU.min-1.kg-1) and euglycemic clamp was started, and euglycemia was maintained for another 250 min. A second bolus of the tracer was then given at 240 min, and the plasma radioactivity was followed for 160 min. Insulin stimulated basal plasma glucose clearance fourfold (p < 0.001) and completely suppressed basal hepatic glucose production (p < 0.001). Concomitantly, the total distribution volume of glucose was increased by 19% (p < 0.05); this change was accompanied by about 50% expansion of the slowly exchanging glucose pool (putatively representing the insulin-dependent compartment). Thyroxine treatment increased plasma triiodothyronine by about 20% (0.1 > p > 0.05) but did not affect basal glucose turnover, insulin-stimulated plasma glucose clearance or the insulin-induced suppression of endogenous glucose output. However, thyroxine treatment blunted the insulin-induced increases in total distribution volume and the slowly exchanging pool of glucose (p = NS vs the basal state). We conclude that minor changes in plasma triiodothyronine (such as occur during overfeeding) do not interfere with the ability of insulin to stimulate the rate of disappearance of glucose or suppress endogenous glucose release; however, our data suggest that they induce finer changes in glucose kinetics, possibly reflecting acceleration or intracellular glucose degradation.

Adult↗

PubMed1977

UDP-glucose: Glucan Synthetase in Developing Cotton Fibers: II. Structure of the Reaction Product.

The solubility properties, composition, and structure of the radioactive product synthesized from UDP-[(14)C]glucose by a highly active cotton fiber glucan synthetase have been determined. Product obtained under the following three different conditions was analyzed: at high and low substrate concentrations by detached fibers, and at high substrate concentrations with an isolated particulate preparation. The results of acetic and nitric acid digestion, enzyme digestion, total acid hydrolyses, periodate oxidation, partial acid hydrolyses, and methylation analyses all support the conclusion that the product of the glucan synthetase produced under all three assay conditions is a linear beta-(1-->3)-glucan.

Journal Article↗