The metabolism of [Me-14C]choline in the brain of the rat in vivo.
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ABSTRACT:
SUBMITTER: Ansell GB
PROVIDER: S-EPMC1187198 | biostudies-other | 1968 Nov
REPOSITORIES: biostudies-other
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Project description:Brain uptake of [(14)C]acetate has been reported to be a useful marker of astrocytic energy metabolism. In addition to uptake values, the rate of radiolabeled acetate washout from the brain appears to reflect CO2 exhaustion and oxygen consumption in astrocytes. We measured the time-radioactivity curves of benzyl [1-(14)C]acetate ([1-(14)C]BA), a lipophilic probe of [1-(14)C]acetate, and compared it with that of benzyl [2-(14)C]acetate ([2-(14)C]BA) in rat brains. The highest brain uptake was observed immediately after injecting either [1-(14)C]BA or [2-(14)C]BA, and both subsequently disappeared from the brain in a single-exponential manner. Estimated [1-(14)C]BA washout rates in the cerebral cortex and cerebellum were higher than those of [2-(14)C]BA. These results suggested that [1-(14)C]BA could be a useful probe for estimating the astrocytic oxidative metabolism. The [1-(14)C]BA washout rate in the cerebral cortex of immature rats was lower than that of mature rats. An autoradiographic study showed that the washout rates of [1-(14)C]BA from the rat brains of a lithium-pilocarpine-induced status epilepticus model were not significantly different from the values in control rat brains except for the medial septal nucleus. These results implied that the enhancement of amino acid turnover rate rather than astrocytic oxidative metabolism was increased in status epilepticus.
| S-EPMC4759666 | biostudies-literature
Project description:1. Acute NH(4) (+) toxicity was studied by using a new apparatus that removes and freezes the brains of conscious rats within 1s. 2. Brains were removed and frozen 5min after intraperitoneal injection of ammonium acetate (2-3min before the onset of convulsions). Arterial [NH(4) (+)] rose from less than 0.01 to 1.74mm at 4-5min. The concentrations of all glycolytic intermediates measured, except glucose 6-phosphate, were increased by the indicated percentage above the control value as follows: glucose (by 41%), fructose 1,6-diphosphate (by 133%), dihydroxyacetone phosphate (by 164%), alpha-glycerophosphate (by 45%), phosphoenolpyruvate (by 67%) and pyruvate (by 26%). 4. Citrate and alpha-oxoglutarate concentrations were unchanged and that of malate was increased (by 17%). 5. Adenine nucleotides and P(i) concentrations were unchanged but the concentration of creatine phosphate decreased slightly (by 6%). 6. Brain [NH(4) (+)] increased from 0.2 to 1.53mm. Net glutamine synthesis occurred at an average rate of 0.33mumol/min per g. 7. The rate of brain glucose utilization was measured in vivo as 0.62mumol/min per g in controls and 0.81mumol/min per g after NH(4) (+) injection. 8. The arteriovenous difference of glucose and O(2) increased by 35%. 9. No significant arteriovenous differences of glutamate or glutamine were detected. Thus, although much NH(4) (+) was incorporated into glutamine the latter was not rapidly released from the brain to the circulation. 10. Plasma [K(+)] increased from 3.3 to 5.4mm. 11. The results indicate that NH(4) (+) stimulates oxidative metabolism but does not interfere with brain energy balance. The increased rate of oxidative metabolism could not be accounted for only on the basis of glutamine synthesis. We suggest that increased extracellular [NH(4) (+)] and [K(+)] decreased the resting transmembrane potential and stimulated Na(+),K(+)-stimulated adenosine triphosphatase activity thus accounting for the increased metabolic rate.
| S-EPMC1177908 | biostudies-other
Project description:Biologic scaffold materials are used for repair and reconstruction of injured or missing tissues. Such materials are often composed of allogeneic or xenogeneic extracellular matrix (ECM) manufactured by decellularization of source tissue, such as dermis. Dermal ECM (D-ECM) has been observed to degrade and remodel in vivo more slowly than other biologic scaffold materials, such as small intestinal submucosa (SIS-ECM). Histologic examination is a common method for evaluating material degradation, but it lacks sensitivity and is subject to observer bias. Utilization of (14)C-proline labeled ECM is a quantitative alternative for measuring degradation of ECM scaffolds. Using both methods, the amount of degradation of D-ECM and SIS-ECM was determined at 2, 4, and 24 weeks post-implantation in a rodent model. Results utilizing (14)C liquid scintillation counting (LSC) analysis showed distinct differences in degradation at the three time points. D-ECM material in situ stayed the same at 76% remaining from 2 to 4 weeks post-implantation, and then decreased to 44% remaining at 24 weeks. In the same time period, implanted SIS-ECM material decreased from 72% to 13% to 0%. Visual examination of device degradation by histology overestimated degradation at 2 weeks and underestimated device degradation at 24 weeks, compared to the (14)C method.
| S-EPMC5372693 | biostudies-literature
Project description:1. Adult rat hepatocytes were isolated by collagenase perfusion and were maintained in monolayer culture for 24h. 2. Choline metabolism and phosphatidylcholine biosynthesis were studied in these cells by performing pulse-chase studies at physiological concentrations (1-40 microM) of (Me-3H)-labelled or unlabelled choline in the culture medium. 3. During the 15 min pulse incubation, choline entering the cells was rapidly phosphorylated to phosphocholine or oxidized to betaine. Low concentrations of choline in the medium decreased the relative amount of choline oxidized. 4. During the 3 h chase period, the radioactivity in the phosphocholine pool was transferred to phosphatidylcholine. Very little radioactivity was associated with CDP-choline. These results provide good evidence that the rate-limiting step for phosphatidylcholine biosynthesis in these cultured hepatocytes is the conversion of phosphocholine into CDP-choline. Similar results were obtained for all concentrations of choline in the culture medium. 5. Cellular concentrations of phosphocholine were unaffected by the concentration of choline (1-40 microM) in the medium. 6. The majority of the label associated with betaine was secreted into the culture medium during the chase incubation. 7. From the pulse-chase studies, and the cellular phosphocholine concentrations, it was possible to estimate the rate of phosphatidylcholine biosynthesis (2.2, 2.8, 3.1 and 3.7 nmol/min per g wet weight of cells cultured in 1, 5, 10 and 40 microM-choline respectively for up to 4.25 h).
| S-EPMC1162990 | biostudies-other
Project description:Phospholipases D (PLDs) catalyze hydrolysis of the diester bond of phospholipids to generate phosphatidic acid and the free lipid headgroup. In mammals, PLD enzymes comprise the intracellular enzymes PLD1 and PLD2 and possibly the proteins encoded by related genes, as well as a class of cell surface and secreted enzymes with structural homology to ectonucleotide phosphatases/phosphodiesterases as typified by autotaxin (ENPP2) that have lysoPLD activities. Genetic and pharmacological loss-of-function approaches implicate these enzymes in intra- and intercellular signaling mediated by the lipid products phosphatidic acid, lysophosphatidic acid, and their metabolites, while the possibility that the water-soluble product of their reactions is biologically relevant has received far less attention. PLD1 and PLD2 are highly selective for phosphatidylcholine (PC), whereas autotaxin has broader substrate specificity for lysophospholipids but by virtue of the high abundance of lysophosphatidylcholine (LPC) in extracellular fluids predominantly hydrolyses this substrate. In all cases, the water-soluble product of these PLD activities is choline. Although choline can be formed de novo by methylation of phosphatidylethanolamine, this activity is absent in most tissues, so mammals are effectively auxotrophic for choline. Dietary consumption of choline in both free and esterified forms is substantial. Choline is necessary for synthesis of the neurotransmitter acetylcholine and of the choline-containing phospholipids PC and sphingomyelin (SM) and also plays a recently appreciated important role as a methyl donor in the pathways of "one-carbon (1C)" metabolism. This review discusses emerging evidence that some of the biological functions of these intra- and extracellular PLD enzymes involve generation of choline with a particular focus on the possibility that these choline and PLD dependent processes are dysregulated in cancer.
| S-EPMC7768751 | biostudies-literature
Project description:1. [2-(14)C]Indole has been synthesized from [(14)C]formate and o-toluidine via N[(14)C]-formyltoluidine. 2. When fed to rats, the (14)C of [(14)C]indole (dose 70-80mg./kg. body wt.) is fairly rapidly excreted, and in 2 days an average of 81% appears in the urine, 11% in the faeces and 2.4% as carbon dioxide in the expired air. 3. Radioactivity is excreted in the urine as indoxyl sulphate (50% of the dose), indoxyl glucuronide (11%), oxindole (1.4%), isatin (5.8%), 5-hydroxyoxindole conjugates (3.1%), N-formylanthranilic acid (0.5%) and unchanged indole (0.07%). The faeces contain indoxyl sulphate (0.4% of the dose) and indole (0.2%), but the major metabolites have not been identified. 4. Fed to rats with biliary cannulae an average of 5.6% of a dose of [(14)C]indole (20-60mg./kg. body wt.) is excreted in the bile in 2 days. Radioactivity is present as indoxyl sulphate (0.8% dose) and 5-hydroxyoxindole conjugates (0.6%). 5. Rats further metabolize indoxyl into N-formylanthranilic acid and anthranilic acid, and oxindole into 5-hydroxyoxindole. 6. With rat-liver microsomes plus supernatant under aerobic conditions, indole gives indoxyl, oxindole, possibly isatin, N-formylanthranilic acid and anthranilic acid, but under anaerobic conditions gives only oxindole. Similarly, under aerobic conditions, oxindole gives 5-hydroxyoxindole, anthranilic acid and o-aminophenylacetic acid. 7. Indole is metabolized by two pathways, one via indoxyl to isatin, N-formylanthranilic acid and anthranilic acid, and the other via oxindole to 5-hydroxyoxindole and possibly to o-aminophenylacetic and anthranilic acid. 8. The following new compounds are described: 4-hydroxy-2-nitrophenylacetic acid, 3-, 4- and 5-benzyloxy-2-nitrophenylacetic acid, 5- and 7-hydroxyoxindole and 5-aminoacridine indoxyl sulphate.
| S-EPMC1264825 | biostudies-other
Project description:Determination of oxidative metabolism in the brain using in vivo ¹³C NMR spectroscopy (¹³C MRS) typically requires repeated blood sampling throughout the study to measure blood glucose concentration and fractional enrichment (input function). However, drawing blood from small animals, such as young rats, placed deep inside the magnet is technically difficult due to their small total blood volume. In the present study, a custom-built animal holder enabled temporary removal of the animal from the magnet for blood collection, followed by accurate repositioning in the exact presampling position without degradation of B? shimming. ¹³C label incorporation into glutamate C4 and C3 positions during a 120 min [1,6-¹³C?] glucose infusion was determined in 28-day-old rats (n = 4) under ?-chloralose sedation using localized, direct-detected in vivo ¹³C MRS at 9.4T. The tricarboxylic acid cycle activity rate (V(TCA)) determined using a one-compartment metabolic modeling was 0.67 ± 0.13 ?mol/g/min, a value comparable to previous ex vivo studies. This methodology opens the avenue for in vivo measurements of brain metabolic rates using ¹³C MRS in small animals.
| S-EPMC3184325 | biostudies-literature
Project description:Betaine critically contributes to the control of hepatocellular hydration and provides protection of the liver from different kinds of stress. This study investigates to what extent hepatocellular hydration changes affect the expression levels of enzymes involved in the metabolism of betaine and related organic osmolytes by using qRT-PCR gene expression studies in rat hepatoma cells as well as metabolic and gene expression profiling in 5,10 - methylene tetrahydrofolate reductase (MTHFR) deficient primary hepatocytes. The results demonstrate a coordinated regulation of betaine degradation and synthesis under anisoosmotic conditions. Expression of betaine degrading enzymes is downregulated by hyperosmolarity and strongly induced by hypoosmolarity. In contrast, synthesis of glycerophosphocholine from phosphoethanolamine and conversion of choline to betaine are both induced by hyperosmolarity but decreased under hypoosmotic conditions. In addition we evaluated the flux of choline and its derivates in liver and plasma of methylene tetrahydrofolate reductase knockout (Mthfr-/-) mice by tandem mass spectrometry. Analyses of system-wide alterations of osmolyte metabolism with microarray studies revealed expression changes similar to those after hypoosmotic exposure in this betaine depletion model. In conclusion, regulation of betaine synthesis and degradation and concomitant changes in intracellular osmolyte concentrations contribute to long-term adaptation to anisoosmotic exposure of the liver. Expression of 280 genes were analyzed in wild type and mthr-/- mice (n=7) with spotted oligonucleotides.
2013-04-01 | E-GEOD-42275 | biostudies-arrayexpress
Project description:OBJECTIVE:To characterize the absorption, distribution, metabolism, and excretion of naloxegol, a PEGylated derivative of the µ-opioid antagonist naloxone, in healthy male subjects. MATERIALS AND METHODS:[14C]-Labeled naloxegol (27 mg, 3.43 MBq) was administered as an oral solution to 6 fasted subjects. Blood, fecal, and urine samples were collected predose and at various intervals postdose. Naloxegol and its metabolites were quantified or identified by liquid chromatography with radiometric or mass spectrometric detection. Pharmacokinetic parameters were calculated for each subject, and metabolite identification was performed by liquid chromatography with parallel radioactivity measurement and mass spectrometry. RESULTS:Naloxegol was rapidly absorbed, with a maximum plasma concentration (geometric mean) of 51 ng/mL reached before 2 hours after dosing. A second peak in the observed naloxegol and [14C] plasma concentration-time profiles was observed at ~3 hours and was likely due to enterohepatic recycling of parent naloxegol. Distribution to red blood cells was negligible. Metabolism of [14C]-naloxegol was rapid and extensive and occurred via demethylation and oxidation, dealkylation, and shortening of the polyethylene glycol chain. Mean cumulative recovery of radioactivity was 84.2% of the total dose, with ~68.9% recovered within 96 hours of dosing. Fecal excretion was the predominant route of elimination, with mean recoveries of total radioactivity in feces and urine of 67.7% and 16.0%, respectively. Unchanged naloxegol accounted for ~1/4 of the radioactivity recovered in feces. CONCLUSIONS:Naloxegol was rapidly absorbed and cleared via metabolism, with predominantly fecal excretion of parent and metabolites.
| S-EPMC4564822 | biostudies-literature
Project description:The concentration of unesterified choline in the plasma in the jugular vein of the rat (0.85 nmol/ml) was found to be three times that of the arterial supply to the brain (0.25 nmol/ml), indicating a higher efflux than uptake of unesterified choline by the brain. No such difference was found for the rabbit and no arterio-venous difference for phosphatidylcholine or lysophosphatidylcholine was observed in either species. No arterio-venous difference was found for choline in blood cells. The infusion of [Me-3H]choline into the circulation of the rat or rabbit indicated an uptake of radioactive choline by the brain and an efflux of non-radioactive choline. In the rabbit such an infusion produced a steady rise in the labelling of phosphatidylcholine and lysophosphatidylcholine in the plasma. When [14C2]ethanolamine was injected intraperitoneally into the rat there was a labelling of phosphatidylcholine, lysophosphatidylcholine and sphingomyelin in the plasma and cells of blood from the jugular vein and the arterial supply, as well as in the brain tissue. However, no labelling of unesterified choline in these tissues could be detected. Unesterified choline was shown to be liberated into the plasma when whole blood from the rat or man, but not the rabbit, was incubated for short periods at 30 degrees C.
| S-EPMC1172684 | biostudies-other