Metabolic Trapping Research Articles

Total-body dynamic PET/CT imaging reveals kinetic distribution of [13N]NH3 in normal organs.

To systematically investigate kinetic metrics and metabolic trapping of [13N]NH3 in organs. Eleven participants performed total-body [13N]NH3 dynamic positron emission tomography (PET). Regions of interest were drawn in organs to obtain time-to-activity curves (TACs), which were fitted with an irreversible two-tissue compartment model (2TC) to investigate constant rates K1, k2 and k3, and to calculate Ki. Additionally, one-tissue compartment model using full data (1TCfull) and the first four minutes of data (1TC4min) were fitted to TAC data. K1 and k2 were compared among different models to assess [13N]NH3 trapping in organs. Kinetic rates of [13N]NH3 varied significantly among organs. The mean K1 ranged from 0.049 mL/cm3/min in the muscle to 2.936 mL/cm3/min in the kidney. The k2 and k3 were lowest in the liver (0.001min- 1) and in the pituitary (0.009min- 1), while highest in the kidney (0.587min- 1) and in the liver (0.800min- 1), respectively. The Ki was largest in the myocardium (0.601 ± 0.259 mL/cm3/min) while smallest in the bone marrow (0.028 ± 0.022 mL/cm3/min). Three groups of organs with similar kinetic characteristics were revealed: (1) the thyroid, the lung, the spleen, the pancreas, and the kidney; (2) the liver and the muscle; and (3) the cortex, the white matter, the cerebellum, the pituitary, the parotid, the submandibular gland, the myocardium, the bone, and the bone marrow. Obvious k3 was identified in multiple organs, and significant changes of K1 in multiple organs and k2 in most organs were found between 2TC and 1TCfull, but both K1 and k2 were comparable between 2TC and 1TC4min. The kinetic rates of [13N]NH3 differed among organs with some have obvious 13N-anmmonia trapping. The normal distribution of kinetic metrics of 13N-anmmonia in organs can serve as a reference for its potential use in tumor imaging.

Application of copper (I) selective ligands for PET imaging of reactive oxygen species through metabolic trapping

IntroductionReactive oxygen species (ROS) are attractive targets for clinical PET imaging. In this study, we hypothesized that PET imaging of ROS would be possible by using chelating ligands (L) that form stable complexes with copper (I) but not with copper (II), based on metabolic trapping. Namely, when [64Cu][CuI(L)2]+ is oxidized by ROS, the oxidized complex will release [64Cu]Cu2+. Then, the released [64Cu]Cu2+ will be trapped inside the cell, resulting in PET signal depending on the redox potential of ROS. To examine the potential of this novel molecular design for ROS imaging, we synthesized copper (I) complexes with bicinchoninic acid (BCA) disodium salt and bathocuproinedisulfonic acid (BCS) disodium salt and evaluated their reactivity with several kinds of ROS. In addition, the cellular uptake of [64Cu][CuI(BCS)2]3− and the stability of [64Cu][CuI(BCS)2]3− in a biological condition were also evaluated. Methods[64Cu]Cu2+ was reduced to [64Cu]Cu+ by ascorbic acid and coordinated with BCA and BCS in the acetate buffer to synthesize [64Cu][CuI(BCA)2]3− and [64Cu][CuI(BCS)2]3−. The radiochemical yields were determined by thin-layer chromatography (TLC). After [64Cu][CuI(BCS)2]3− was incubated with hydroxyl radical, lipid peroxide, superoxide, and hydrogen peroxide, the percentage of released [64Cu]Cu2+ from the parent complex was evaluated by TLC. HT-1080 human fibrosarcoma cells were treated with 0.1 % Dimethyl sulfoxide (control), imidazole ketone erastin (IKE), or IKE + ferrostatin-1 (Fer-1). Then, the uptake of [64Cu][CuI(BCS)2]3− to HT-1080 cells in each group was evaluated as %Dose/mg protein. Lastly, [64Cu][CuI(BCS)2]3− was incubated in human plasma, and its intact ratio was determined by TLC. ResultsThe radiochemical yield of [64Cu][CuI(BCS)2]3− (86 ± 1 %) was higher than that of [64Cu][CuI(BCA)2]3− (44 ± 3 %). [64Cu][CuI(BCA)2]3− was unstable and partially decomposed on TLC. After [64Cu][CuI(BCS)2]3− was reacted with hydroxyl radical, lipid peroxide, and superoxide, 67 ± 2 %, 44 ± 13 %, and 22 ± 3 % of total radioactivity was detected as [64Cu]Cu2+, respectively. On the other hand, the reaction with hydrogen peroxide did not significantly increase the ratio of [64Cu]Cu2+ (4 ± 1 %). These results suggest that [64Cu][CuI(BCS)2]3− could be used for detecting high-redox-potential ROS such as hydroxyl radical and lipid peroxide with high selectivity. The cellular uptake values of [64Cu][CuI(BCS)2]3− in the control, IKE, and Fer-1 group were 42 ± 2, 54 ± 2, and 47 ± 5 %Dose/mg protein (n = 3), respectively, suggesting the ROS specific uptake of [64Cu][CuI(BCS)2]3−. On the other hand, the intact ratio after the incubation of [64Cu][CuI(BCS)2]3− in human plasma was 9 ± 5 %. ConclusionPET imaging of ROS would be possible by using a copper (I) selective ligand, based on metabolic trapping. Although improvement of the membrane permeability and the stability of copper (I) complexes is required, the present results pave the way for the development of novel 64Cu-labeled complexes for PET imaging of ROS.

Synthesis and Biological Evaluation of a Novel Sugar-Conjugated Platinum(II) Complex Having a Tumor-Targeting Effect.

We designed and synthesized a novel platinum complex conjugated with 2-fluorinated 2-deoxyglucoside, named FGC-Pt, to capitalize on the Warburg effect and metabolic trapping properties of [18F]2-deoxy-2-fluoro-d-glucose ([18F]FDG). Then, we conducted comprehensive in vitro and in vivo studies to evaluate the effects of FGC-Pt. In vitro cytotoxicity assays using HeLa cells revealed that FGC-Pt exhibited concentration-dependent cytotoxicity, even though its cytotoxic effect was less pronounced than that of cisplatin. In the evaluation of in vivo biodistribution in mice, platinum concentration in tumors and major organs (muscle, bone, blood, liver, and kidney) and the ratio of platinum concentration in tumors to major organs following the tail vein injection of FGC-Pt and cisplatin suggest that FGC-Pt is more retained in tumors than in other organs and tends to accumulate in tumors more than cisplatin. Furthermore, an in vivo assessment of the antitumor effect conducted in A549 cell-bearing mice demonstrated that FGC-Pt possesses substantial potential as an antitumor agent. It exhibited a tumor growth-inhibitory effect comparable to that of cisplatin while inducing lower toxicity, as evidenced by lower weight loss after administration. Herein, we successfully produced a novel compound with a tumor-growth-inhibitory effect comparable to that of cisplatin and low toxicity.

Synthesis of 4-Deoxy-4-Fluoro-d-Sedoheptulose: A Promising New Sugar to Apply the Principle of Metabolic Trapping.

Fluorinated carbohydrates are important tools for understanding the deregulation of metabolic fluxes and pathways. Fluorinating specific positions within the sugar scaffold can lead to enhanced metabolic stability and subsequent metabolic trapping in cells. This principle has, however, never been applied to study the metabolism of the rare sugars of the pentose phosphate pathway (PPP). In this study, two fluorinated derivatives of d-sedoheptulose were designed and synthesized: 4-deoxy-4-fluoro-d-sedoheptulose (4DFS) and 3-deoxy-3-fluoro-d-sedoheptulose (3DFS). Both sugars are taken up by human fibroblasts but only 4DFS is phosphorylated. Fluorination of d-sedoheptulose at C-4 effectively halts the enzymatic degradation by transaldolase and transketolase. 4DFS thus has a high potential as a new PPP imaging probe based on the principle of metabolic trapping. Therefore, the synthesis of potential radiolabeling precursors for 4DFS for future radiofluorinations with fluorine-18 is presented.

Fatigue in Patients on Chronic Hemodialysis: The Role of Indoleamine 2,3-Dioxygenase (IDO) Activity, Interleukin-6, and Muscularity.

Fatigue is a frequent symptom in hemodialysis (HD), and the indolamine-2,3-dioxygenase (IDO) metabolic trap has been hypothesized in the pathogenesis of fatigue. The association between IDO activity according to fatigue and its relationship with muscle mass and function in HD patients was verified. Chronic HD patients were considered, and fatigue was assessed. The plasma kynurenines and tryptophan ratio (Kyn/Trp), as surrogate of IDO activity, and interleukin (IL)-6 were measured. Muscularity was assessed by BIA and muscle strength by hand-grip dynamometer. 50 HD patients were enrolled, and fatigue was present in 24% of the cohort. Patients with fatigue showed higher Kyn/Trp (p = 0.005), were older (p = 0.007), and IL-6 levels resulted higher than in non-fatigue patients (p < 0.001). HD patients with fatigue showed lower intracellular water (surrogate of muscle mass) (p < 0.001), as well as lower hand grip strength (p = 0.02). The Kyn/Trp ratio positively correlated with IL-6 and ECW/ICW (p = 0.004 and p = 0.014). By logistic regression analysis, higher ICW/h2 was associated with lower odds of fatigue (OR, 0.10; 95% CI, 0.01 to 0.73). In conclusion, our cohort fatigue was associated with a higher Kyn/Trp ratio, indicating a modulation of IDO activity. The Kyn/Trp ratio correlated with IL-6, suggesting a potential role of IDO and inflammation in inducing fatigue and changes in muscularity.

It's Not What You Take Up, It's What You Keep: How Discoveries from Diverse Disciplines Directed the Development of the FDG PET/CT Scan.

Although imaging glucose metabolism with positron emission tomography combined with X-ray CT (FDG-PET/CT) has become a standard diagnostic modality for the discovery and surveillance of malignant tumors and inflammatory processes, its origins extend back to more than a century of notable discoveries in the fields of inorganic and organic chemistry, nuclear physics, mathematics, biochemistry, solute transport physiology, metabolism, and imaging, accomplished by pioneering and driven investigators, of whom at least ten were recipients of the Nobel Prize. These tangled and diverse roots eventually coalesced into the FDG-PET/CT method, that through its many favorable characteristics inherent in the isotope used (18F), the accurate imaging derived from coincidence detection of positron annihilation radiation combined with computed tomography, and the metabolic trapping of 2-deoxy-2-[18F]fluoro-D-glucose (FDG) in tissues, provides safety, sensitivity, and specificity for tumor and inflammation detection. The authors hope that this article will increase the appreciation among its readers of the insight, creativity, persistence, and drive of the many investigators who made this technique possible. This article is followed by a review of the many applications of FDG-PET/CT to the gastrointestinal tract and hepatobiliary system (Mandelkern in Dig Dis Sci 2022).

Advancing Cancer Treatment by Targeting Glutamine Metabolism-A Roadmap.

Simple SummaryDysregulated glutamine metabolism is one of the metabolic features evident in cancer cells when compared to normal cells. Cancer cells utilize glutamine for energy generation as well as the synthesis of other molecules that are critical for cancer growth and progression. Therefore, drugs targeting glutamine metabolism have been extensively investigated. However, inhibition of glutamine metabolism in cancer cells results in the activation of other metabolic pathways enabling cancer cells to survive. In this review, we summarize and discuss the targets in glutamine metabolism, which has been probed in the development of anticancer drugs in preclinical and clinical studies. We further discuss pathways activated in response to glutamine metabolism inhibition, enabling cancer cells to survive the challenge. Finally, we put into perspective combined treatment strategies targeting glutamine metabolism along with other pathways as potential treatment options.Tumor growth and metastasis strongly depend on adapted cell metabolism. Cancer cells adjust their metabolic program to their specific energy needs and in response to an often challenging tumor microenvironment. Glutamine metabolism is one of the metabolic pathways that can be successfully targeted in cancer treatment. The dependence of many hematological and solid tumors on glutamine is associated with mitochondrial glutaminase (GLS) activity that enables channeling of glutamine into the tricarboxylic acid (TCA) cycle, generation of ATP and NADPH, and regulation of glutathione homeostasis and reactive oxygen species (ROS). Small molecules that target glutamine metabolism through inhibition of GLS therefore simultaneously limit energy availability and increase oxidative stress. However, some cancers can reprogram their metabolism to evade this metabolic trap. Therefore, the effectiveness of treatment strategies that rely solely on glutamine inhibition is limited. In this review, we discuss the metabolic and molecular pathways that are linked to dysregulated glutamine metabolism in multiple cancer types. We further summarize and review current clinical trials of glutaminolysis inhibition in cancer patients. Finally, we put into perspective strategies that deploy a combined treatment targeting glutamine metabolism along with other molecular or metabolic pathways and discuss their potential for clinical applications.

Late metabolic trapping of 13N-ammonia is a significantly better predictor of functional improvement after revascularization when compared to absolute myocardial blood flow

Late metabolic trapping of 13N-ammonia is a significantly better predictor of functional improvement after revascularization when compared to absolute myocardial blood flow

Gallagher's Principle of Metabolic Trapping.

The term metabolic trapping means intracellular enzymatic conversion of a radiotracer into a metabolic product that is trapped and concentrated in key tissues. In a classic paper selected by the editors of JNM for the 60th anniversary edition, Brian Gallagher et al. reported on the dominant

The IDO Metabolic Trap Hypothesis for the Etiology of ME/CFS.

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a debilitating noncommunicable disease brandishing an enormous worldwide disease burden with some evidence of inherited genetic risk. Absence of measurable changes in patients’ standard blood work has necessitated ad hoc symptom-driven therapies and a dearth of mechanistic hypotheses regarding its etiology and possible cure. A new hypothesis, the indolamine-2,3-dioxygenase (IDO) metabolic trap, was developed and formulated as a mathematical model. The historical occurrence of ME/CFS outbreaks is a singular feature of the disease and implies that any predisposing genetic mutation must be common. A database search for common damaging mutations in human enzymes produces 208 hits, including IDO2 with four such mutations. Non-functional IDO2, combined with well-established substrate inhibition of IDO1 and kinetic asymmetry of the large neutral amino acid transporter, LAT1, yielded a mathematical model of tryptophan metabolism that displays both physiological and pathological steady-states. Escape from the pathological one requires an exogenous perturbation. This model also identifies a critical point in cytosolic tryptophan abundance beyond which descent into the pathological steady-state is inevitable. If, however, means can be discovered to return cytosolic tryptophan below the critical point, return to the normal physiological steady-state is assured. Testing this hypothesis for any cell type requires only labelled tryptophan, a means to measure cytosolic tryptophan and kynurenine, and the standard tools of tracer kinetics.

Deuterium Kinetic Isotope Effect Studies of a Potential in Vivo Metabolic Trapping Agent for Monoamine Oxidase B

Visualizing the in vivo activity of monoamine oxidase B (MAO-B) is a valuable tool in the ongoing investigation of astrogliosis in neurodegeneration. Existing strategies for imaging changes in MAO enzyme expression or activity have utilized the irreversible suicide inhibitors or high-affinity reversibly binding inhibitors as positron emission tomography (PET) ligands. As an alternative approach, we developed 4-methyl-7-[(1-[11C]methyl-1,2,3,6-tetrahydropyridin-4-yl)oxy]-2 H-chromen-2-one ([11C]Cou) as a metabolic trapping agent for MAO-B. Trapping of [11C]Cou in rhesus monkey brain demonstrated MAO-B selectivity. In this work, we have attempted to improve on the in vivo pharmacokinetics of [11C]Cou by using the deuterium kinetic isotope effect (KIE) to slow the MAO-B-mediated oxidation step and thus reduce the rate of trapping in brain tissues. However, in vitro assays of enzyme kinetics and in vivo PET imaging of pharmacokinetics in primate brain showed no effects of deuterium substitution on the tetrahydropyridine ring of [11C]Cou. The results are possibly due to masking of the KIE by a second step in the overall metabolism of the new imaging agent.

PnuT uses a facilitated diffusion mechanism for thiamine uptake.

Membrane transporters of the bacterial pyridine nucleotide uptake (Pnu) family mediate the uptake of various B-type vitamins. For example, the PnuT transporters have specificity for vitamin B1 (thiamine). It has been hypothesized that Pnu transporters are facilitators that allow passive transport of the vitamin substrate across the membrane. Metabolic trapping by phosphorylation would then lead to accumulation of the transported substrates in the cytoplasm. However, experimental evidence for such a transport mechanism is lacking. Here, to determine the mechanism of thiamine transport, we purify PnuTSw from Shewanella woodyi and reconstitute it in liposomes to determine substrate binding and transport properties. We show that the electrochemical gradient of thiamine solely determines the direction of transport, consistent with a facilitated diffusion mechanism. Further, PnuTSw can bind and transport thiamine as well as the thiamine analogues pyrithiamine and oxythiamine, but does not recognize the phosphorylated derivatives thiamine monophosphate and thiamine pyrophosphate as substrates, consistent with a metabolic trapping mechanism. Guided by the crystal structure of the homologous nicotinamide riboside transporter PnuC, we perform mutagenesis experiments, which reveal residues involved in substrate binding and gating. The facilitated diffusion mechanism of transport used by PnuTSw contrasts sharply with the active transport mechanisms used by other bacterial thiamine transporters.

Molecular Imaging of GLUT1 and GLUT5 in Breast Cancer: A Multitracer Positron Emission Tomography Imaging Study in Mice.

Use of [18F]FDG-positron emission tomography (PET) in clinical breast cancer (BC) imaging is limited mainly by insufficient expression levels of facilitative glucose transporter (GLUT)1 in up to 50% of all patients. Fructose-specific facilitative hexose transporter GLUT5 represents an alternative biomarker for PET imaging of hexose metabolism in BC. The goal of the present study was to compare the uptake characteristics of selected hexose-based PET radiotracers in murine BC model EMT6. Uptake of 1-deoxy-1-[18F]fluoro-d-fructose (1-[18F]FDF), 6-deoxy-6-[18F]fluoro-d-fructose (6-[18F]FDF), 1-deoxy-1-[18F]fluoro-2,5-anhydro-mannitol (1-[18F]FDAM), 2-deoxy-2-[18F]fluoro-d-glucose (2-[18F]FDG), and 6-deoxy-6-[18F]fluoro-d-glucose (6-[18F]FDG) was studied in EMT6 cells, tumors, and muscle and correlated to GLUT1 and GLUT5 expression levels. Fructose-derivative 6-[18F]FDF revealed greater tumor uptake than did structural analog 1-[18F]FDF, whereas 1-[18F]FDAM with locked anomeric configuration showed similar low tumor uptake to that of 1-[18F]FDF. Glucose-derivative 6-[18F]FDG reached maximum tumor uptake at 20 minutes, with no further accumulation over time. Uptake of 2-[18F]FDG was greatest and continuously increasing owing to metabolic trapping through phosphorylation by hexokinase II. In EMT6 tumors, GLUT5 mRNA expression was 20,000-fold lower compared with GLUT1. Whereas the latter was much greater in tumor than in muscle tissue (GLUT1 50:1), the opposite was found for GLUT5 mRNA expression (GLUT5 1:6). GLUT5 protein levels were higher in tumor versus muscle tissue as determined by Western blot and immunohistochemistry. Our data suggest that tumor uptake of fructose metabolism-targeting radiotracers 1-[18F]FDF, 6-[18F]FDF, and 1-[18F]FDAM does not correlate with GLUT5 mRNA levels but is linked to GLUT5 protein levels. In conclusion, our results highlight the importance of detailed biochemical studies on GLUT protein expression levels in combination with PET imaging studies for functional characterization of GLUTs in BC.

Acyl CoA synthetase-1 links facilitated long chain fatty acid uptake to intracellular metabolic trafficking differently in hearts of male versus female mice

RationaleAcyl CoA synthetase-1 (ACSL1) is localized at intracellular membranes, notably the mitochondrial membrane. ACSL1 and female sex are suggested to indirectly facilitate lipid availability to the heart and other organs. However, such mechanisms in intact, functioning myocardium remain unexplored, and roles of ACSL1 and sex in the uptake and trafficking of fats are poorly understood. ObjectiveTo determine the potential for ACSL1 and sex-dependent differences in metabolic trapping and trafficking effects of long-chain fatty acids (LCFA) within cardiomyocytes of intact hearts. Methods and results13C NMR of intact, beating mouse hearts, supplied 13C palmitate, revealed 44% faster trans-sarcolemmal uptake of LCFA in male hearts overexpressing ACSL1 (MHC-ACSL1) than in non-transgenic (NTG) males (p<0.05). Acyl CoA content was elevated by ACSL1 overexpression, 404% in males and 164% in female, relative to NTG. Despite similar ACSL1 content, NTG females displayed faster LCFA uptake kinetics compared to NTG males, which was reversed by ovariectomy. NTG female LCFA uptake rates were similar to those in ACSL1 males and ACSL1 females. ACSL1 and female sex hormones both accelerated LCFA uptake without affecting triglyceride content or turnover. ACSL1 hearts contained elevated ceramide, particularly C22 ceramide in both sexes and specifically, C24 in males. ACSL1 also induced lower content of fatty acid transporter-6 (FATP6) indicating cooperative regulation with ACSL1. Surprisingly, ACSL1 overexpression did not increase mitochondrial oxidation of exogenous palmitate, which actually dropped in female ACSL1 hearts. ConclusionsACSL1-mediated metabolic trapping of exogenous LCFA accelerates LCFA uptake rates, albeit to a lesser extent in females, which distinctly affects LCFA trafficking to acyl intermediates but not triglyceride storage or mitochondrial oxidation and is affected by female sex hormones.

Measurement of Long-Chain Fatty Acyl-CoA Synthetase Activity.

Long-chain fatty acyl-CoA synthetases (ACS) are a family of essential enzymes of lipid metabolism, activating fatty acids by thioesterification with coenzyme A. Fatty acyl-CoA molecules are then readily utilized for the biosynthesis of storage and membrane lipids, or for the generation of energy by ß-oxidation. Acyl-CoAs also function as transcriptional activators, allosteric inhibitors, or precursors for inflammatory mediators. Recent work suggests that ACS enzymes may drive cellular fatty acid uptake by metabolic trapping, and may also regulate the channeling of fatty acids towards specific metabolic pathways. The implication of ACS enzymes in widespread lipid associated diseases like type 2 diabetes has rekindled interest in this protein family. Here, we describe in detail how to measure long-chain fatty acyl-CoA synthetase activity by a straightforward radiometric assay. Cell lysates are incubated with ATP, coenzyme A, Mg(2+), and radiolabeled fatty acid bound to BSA. Differential phase partitioning of fatty acids and acyl-CoAs is exploited to quantify the amount of generated acyl-CoA by scintillation counting. The high sensitivity of this assay also allows the analysis of small samples like patient biopsies.

State-of-the-art of bone marrow imaging in multiple myeloma.

Multiple myeloma is a common hematological malignancy arising in the bone marrow. Bone lesions were initially depicted with conventional radiography, although recently F-FDG PET/CT and MRI are recognized as having a clear role in the initial workup and in the evaluation of therapy response. Tumor development produces osteolysis and expansive lesions. Although tumor burden and extent are key prognostic factors, different cancer hallmarks can also be evaluated in vivo through noninvasive imaging. This imaging-based virtual biopsy approach might be useful to define several relevant prognostic markers, such as angiogenesis, cellularity, metabolic trapping and bone morphology and elasticity, both before and during treatment, to predict tumor behavior and the early effect of therapy.

A perspective on the future role of brain pet imaging in exercise science

Positron Emission Tomography (PET) bears a unique potential for examining the effects of physical exercise (acute or chronic) within the central nervous system in vivo, including cerebral metabolism, neuroreceptor occupancy, and neurotransmission. However, application of Neuro-PET in human exercise science is as yet surprisingly sparse. To date the field has been dominated by non-invasive neuroelectrical techniques (EEG, MEG) and structural/functional magnetic resonance imaging (sMRI/fMRI). Despite PET having certain inherent disadvantages, in particular radiation exposure and high costs limiting applicability at large scale, certain research questions in human exercise science can exclusively be addressed with PET: The “metabolic trapping” properties of 18F-FDG PET as the most commonly used PET-tracer allow examining the neuronal mechanisms underlying various forms of acute exercise in a rather unconstrained manner, i.e. under realistic training scenarios outside the scanner environment. Beyond acute effects, 18F-FDG PET measurements under resting conditions have a strong prospective for unraveling the influence of regular physical activity on neuronal integrity and potentially neuroprotective mechanisms in vivo, which is of special interest for aging and dementia research. Quantification of cerebral glucose metabolism may allow determining the metabolic effects of exercise interventions in the entire human brain and relating the regional cerebral rate of glucose metabolism (rCMRglc) with behavioral, neuropsychological, and physiological measures. Apart from FDG-PET, particularly interesting applications comprise PET ligand studies that focus on dopaminergic and opioidergic neurotransmission, both key transmitter systems for exercise-related psychophysiological effects, including mood changes, reward processing, antinociception, and in its most extreme form ‘exercise dependence’. PET ligand displacement approaches even allow quantifying specific endogenous neurotransmitter release under acute exercise interventions, to which modern PET/MR hybrid technology will be additionally fruitful. Experimental studies exploiting the unprecedented multimodal imaging capacities of PET/MR in human exercise sciences are as yet pending.

In Vivo Metabolic Trapping Radiotracers for Imaging Monoamine Oxidase-A and -B Enzymatic Activity.

The isozymes of monoamine oxidase (MAO-A and MAO-B) are important enzymes involved in the metabolism of numerous biogenic amines, including the neurotransmitters serotonin, dopamine, and norepinephrine. Recently, changes in concentrations of MAO-B have been proposed to be an in vivo marker of neuroinflammation associated with Alzheimer's disease. Previous developments of in vivo radiotracers for imaging changes in MAO enzyme expression or activity have utilized the irreversible propargylamine-based suicide inhibitors or high-affinity reversibly binding inhibitors. As an alternative approach, we have investigated 1-[(11)C]methyl-4-aryloxy-1,2,3,6-tetrahydropyridines as metabolic trapping agents for the monoamine oxidases. MAO-mediated oxidation and spontaneous hydrolysis yield 1-[(11)C]methyl-2,3-dihydro-4-pyridinone as a hydrophilic metabolite that is trapped within brain tissues. Radiotracers with phenyl, biphenyl, and 7-coumarinyl ethers were evaluated using microPET imaging in rat and primate brains. No isozyme selectivity for radiotracer trapping was observed in the rat brain for any compound, but in the monkey brain, the phenyl ether demonstrated MAO-A selectivity and the coumarinyl ether showed MAO-B selectivity. These are lead compounds for further development of 1-[(11)C]methyl-4-aryloxy-1,2,3,6-tetrahydropyridines with optimized brain pharmacokinetics and isozyme selectivity.

Abstract 233: Metabolic Trapping Effects of ACS1 for Fatty Acid Uptake and Intracellular Trafficking are Different in Hearts of Males and Females

Disruption of cardiomyocyte lipid metabolism has been observed in the responses to both pathogenic and nutrient stresses in human patients and animal models, with distinct sex differences. This investigation examined cardiac acyl CoA synthetase-1 (ACS)-mediated, LCFA uptake and trafficking with hearts of male and female mice. Isolated mouse hearts with cardiac specific overexpression of ACS1 (MHC-ACS) and non-transgenic littermates (NTG) were perfused with buffer containing 13C palmitate, 10 mM glucose, and 1 mM lactate. Dynamic 13C NMR elucidated the two phases of LCFA incorporation into triacylglycerol (TAG). The time constant (τ) of the initial exponential phase, corresponding to LCFA trans-sarcolemmal uptake, was lower, indicating faster LCFA uptake and esterification in male ACS hearts compared to NTG males (ACS male τ = 1.59±0.67 min vs NTG male τ = 2.85±0.90 min, P&lt;0.05). Consistent with greater metabolic trapping, acyl CoA content was 470% higher in male ACS hearts (NTG male 57.5±9.4 vs ACS male 327.4±42 pmol/mg protein. P&lt;0.001). Despite no differences in ACS content, NTG females displayed faster uptake than NTG males at a rate similar to ACS males, which was reversed by ovariectomy (OVX) indicating an estrogen dependent response (NTG female τ = 1.56±0.46 min, P&lt;0.05 vs NTG OVX female τ = 3.02±0.77 min, P&lt;0.05). Despite prior reports of ACS localization distal from the sarcolemma, ACS level and female gender both independently accelerated LCFA uptake without affecting TAG content or turnover. The data are consistent with metabolic trapping to facilitate LCFA uptake due to ACS activity. ACS also leads to higher total ceramide in hearts, with specific elevation of C18 and C22 in both genders, C24 in males, and C20 in females (male P&lt;0.05; female P&lt;0.01). ACS overexpression induced lower content of the cardiac fatty acid transporter (FATP) isoform, FATP6, indicating cooperative regulation of LCFA uptake between membrane transport proteins and intracellular acyl chain activation (P&lt;0.05 male; P&lt;0.001 female and female OVX). These results demonstrate that perturbation of LCFA metabolism though facilitated metabolic trapping by ACS1 affects sarcolemma transporter expression and induces the conversion of the CoA activated LCFA to ceramide.

18F]-FDG positron emission tomography--an established clinical tool opening a new window into exercise physiology.

Positron emission tomography (PET) with [(18)F]-fluorodeoxyglucose (FDG) is an established clinical tool primarily used to diagnose and evaluate disease status in patients with cancer. PET imaging using FDG can be a highly valuable tool to investigate normal human physiology by providing a noninvasive, quantitative measure of glucose uptake into various cell types. Over the past years it has also been increasingly used in exercise physiology studies to identify changes in glucose uptake, metabolism, and muscle activity during different exercise modalities. Metabolically active cells transport FDG, an (18)fluorine-labeled glucose analog tracer, from the blood into the cells where it is then phosphorylated but not further metabolized. This metabolic trapping process forms the basis of this method's use during exercise. The tracer is given to a participant during an exercise task, and the actual PET imaging is performed immediately after the exercise. Provided the uptake period is of sufficient duration, and the imaging is performed shortly after the exercise; the captured image strongly reflects the metabolic activity of the cells used during the task. When combined with repeated blood sampling to determine tracer blood concentration over time, also known as the input function, glucose uptake rate of the tissues can be quantitatively calculated. This synthesis provides an accounting of studies using FDG-PET to measure acute exercise-induced skeletal muscle activity, describes the advantages and limitations of this imaging technique, and discusses its applications to the field of exercise physiology.