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Plasma concentrations of amino acids are frequently elevated in insulin-resistant states, and a proteinenriched diet can impair glucose metabolism. This study examined effects of short-term plasma amino acid (AA) elevation on whole-body glucose disposal and cellular insulin action in skeletal muscle. Seven healthy men were studied for 5.5 h during euglycemic (5.5 mmol/l), hyperinsulinemic (430 pmol/l), fasting glucagon (65 ng/ l), and growth hormone (0.4 g/l) somatostatin clamp tests in the presence of low (ϳ1.6 mmol/l) and increased (ϳ4.6 mmol/l) plasma AA concentrations. Glucose turnover was measured with D-[6,6-2 H 2 ]glucose. Intramuscular concentrations of glycogen and glucose-6-phosphate (G6P) were monitored using 13 C and 31 P nuclear magnetic resonance spectroscopy, respectively. A ϳ2.1-fold elevation of plasma AAs reduced whole-body glucose disposal by 25% (P < 0.01). Rates of muscle glycogen synthesis decreased by 64% (180 -315 min, 24 ؎ 3; control, 67 ؎ 10 mol ⅐ l ؊1 ⅐ min ؊1 ; P < 0.01), which was accompanied by a reduction in G6P starting at 130 min (⌬G6P 260 -300 min , 18 ؎ 19; control, 103 ؎ 33 mol/l; P < 0.05). In conclusion, plasma amino acid elevation induces skeletal muscle insulin resistance in humans by inhibition of glucose transport/phosphorylation, resulting in marked reduction of glycogen synthesis. Diabetes 51:599 -605, 2002 P lasma concentrations of alanine and particularly branched-chain amino acids (AAs) are elevated in insulin-resistant states such as obesity (1,2), and high dietary protein intake impairs glucose metabolism mainly by changing the utilization of gluconeogenic precursors (3-6).The mechanisms by which AAs could reduce skeletal muscle glucose uptake are as yet unclear. At the cellular level, availability of substrates for energy production, such as AAs and free fatty acids (FFAs), may play an important role in modulating the response to insulin (7). In vitro studies demonstrated that AAs may inhibit glucose utilization in skeletal muscle at various levels. AAs could decrease glucose oxidation by substrate competition with glucose (8,9) and/or reduce glucose uptake (10) by interaction with early steps of insulin signaling (11). Studies in humans, however, revealed controversial results. Infusion of AAs decreased forearm and whole-body glucose disposal in some (12-15), but not all (16,17), studies. Moreover, endogenous release of insulin (18) and glucagon (19) induced by plasma AA elevation might have obscured possible direct effects of AAs in those studies. Taking together all these factors, it is uncertain whether AAs directly induce skeletal muscle insulin resistance in vivo and if so, which mechanism (glucose uptake versus substrate competition) is responsible for such an effect.This study was therefore designed to examine effects of plasma AA elevation on skeletal muscle glucose metabolism by combining isotope dilution technique with in vivo nuclear magnetic resonance (NMR) spectroscopy of gastrocnemius muscle from healthy young humans. In vivo [ 13 C]NMR spectroscop...

31P MRS offers a unique view of muscle metabolism in vivo, but correct quantification is important. Inter-study correlation of estimates of [Pi] and [phosphocreatine (PCr)] in a number of published studies suggest that the main technical problem in calibrated 31P MRS studies is the measurement of PCr and Pi signal intensities, rather than absolute quantification of [ATP]. For comparison, we discuss the few published biopsy studies of calf muscle and a selection of the many studies of quadriceps muscle. The ATP concentration is close to the value that we obtained in calf muscle in our own study, presented here, on four healthy subjects, by localised 31P MRS using a surface coil incorporating an internal reference and calibrated using an external phantom. However, the freeze-clamp biopsy PCr concentration is approximately 20% lower than the value obtained by 31P MRS, consistent with PCr breakdown by creatine kinase during freezing. Finally, we illustrate some consequences of uncertainty in resting [PCr] for analysis of mitochondrial function from PCr kinetics using a published 31P MRS study of exercise and recovery: the lower the assumed resting [PCr], the lower the absolute rate of oxidative ATP synthesis estimated from the PCr resynthesis rate; in addition, the lower the assumed resting [PCr], or the higher the assumed [total creatine], the higher the apparent resting [ADP], and therefore the more sigmoid the relationship between the rate of oxidative ATP synthesis and [ADP]. Correct quantification of resting metabolite concentrations is crucially important for this sort of analysis. Our own results ([PCr] = 33 +/- 2 mM, [Pi] = 4.5 +/- 0.2 mM, and [ATP] = 8.2 +/- 0.4 mM; mean +/- SEM) are close to the overall mean values of the 10 published studies on calf muscle by 'calibrated' 31P MRS (as in the present work), and of [PCr] and [Pi] in a representative selection of 'uncalibrated' 31P MRS studies (i.e. from measured PCr/ATP and Pi/ATP ratios, assuming a literature value for [ATP]).

The translation of MRS to clinical practice has been impeded by the lack of technical standardization. There are multiple methods of acquisition, post-processing, and analysis whose details greatly impact the interpretation of the results. These details are often not fully reported, making it difficult to assess MRS studies on a standardized basis. This hampers the reviewing of manuscripts, limits the reproducibility of study results, and complicates meta-analysis of the literature. In this paper a consensus group of MRS experts provides minimum guidelines for the reporting of MRS methods and results, including the standardized description of MRS hardware, data acquisition, analysis, and quality assessment. This consensus statement describes each of these requirements in detail and includes a checklist to assist authors and journal reviewers and to provide a practical way for journal editors to ensure that MRS studies are reported in full.

With a 40-year history of use for in vivo studies, the terminology used to describe the methodology and results of magnetic resonance spectroscopy (MRS) has grown substantially and is not consistent in many aspects. Given the platform offered by this special issue on advanced MRS methodology, the authors decided to describe many of the implicated terms, to pinpoint differences in their meanings and to suggest

By improving spatial and anatomical specificity, localized spectroscopy can enhance the power and accuracy of the quantitative analysis of cellular metabolism and bioenergetics. Localized and nonlocalized dynamic 31P magnetic resonance spectroscopy using a surface coil was compared during aerobic exercise and recovery of human calf muscle. For localization, a short echo time single-voxel magnetic resonance spectroscopy sequence with adiabatic refocusing (semi-LASER) was applied, enabling the quantification of phosphocreatine, inorganic phosphate, and pH value in a single muscle (medial gastrocnemius) in single shots (TR = 6 s). All measurements were performed in a 7 T whole body scanner with a nonmagnetic ergometer. From a series of equal exercise bouts we conclude that: (a) with localization, measured phosphocreatine declines in exercise to a lower value (79 ± 7% cf. 53 ± 10%, P = 0.002), (b) phosphocreatine recovery shows shorter half time (t1/2 = 34 ± 7 s cf. t1/2 = 42 ± 7 s, nonsignificant) and initial postexercise phosphocreatine resynthesis rate is significantly higher (32 ± 5 mM/min cf. 17 ± 4 mM/min, P = 0.001) and (c) in contrast to nonlocalized 31P magnetic resonance spectroscopy, no splitting of the inorganic phosphate peak is observed during exercise or recovery, just an increase in line width during exercise. This confirms the absence of contaminating signals originating from weaker-exercising muscle, while an observed inorganic phosphate line broadening most probably reflects variations across fibers in a single muscle. Magn Reson Med, 2012. © 2012 Wiley Periodicals, Inc.

A novel method based on interleaved localized 31 P-and 1 H MRS is presented, by which lactate accumulation and the accompanying changes in high energy phosphates in human skeletal muscle can be monitored simultaneously during exercise and recovery. Lactate is quantified using a localized double quantum filter suppressing the abundant lipid signals while taking into account orientation dependent signal modulations. Lactate concentration after ischemic exercise directly quantified by DQF 1 H spectroscopy was 24 ± 3 mmol/L cell water, while 22 ± 3 mmol/L was expected on the basis of 31 major disadvantage of 31 P MRS compared to needle biopsy has been the inability to quantify lactate directly. This has necessitated indirect 31 P MRS approaches (4) to important [and still much debated (5-7)] issues in the control of glycolysis (5,8) and cellular acid handling (4,7,9-11) in vivo. There are several technical obstacles to quantifying lactate in vivo by 1 H MRS, particularly overlapping lipid resonances (at 0.5-1.5 ppm) and orientation-dependent dipolar coupling effects which reduce lactate visibility. These necessitate techniques to selectively acquire the lactate resonances while suppressing the strong background signals. Here, we describe an interleaved 31 P-and 1 H MRS method by which lactate accumulation and the accompanying changes in phosphorus metabolites can be monitored simultaneously in a well-defined gradient localized VOI positioned in a muscle during exercise and recovery (12). Also interleaved is a standard 1 H STEAM pulse sequence (13), to monitor extra-and intramyocellular lipid (which becomes depleted in exercise of longer duration than that described here), TMA, and total creatine. The results advance present knowledge of muscle cellular acidbase metabolism and open the way for much more detailed studies of this and the control of glycolysis. METHODS SubjectsHealthy subjects (n = 7) executed ischemic plantar-flexion on a pedal ergometer (13) at 50% MVC (measured prior to MRS, under aerobic conditions) until fatigue, 2 min 20 sec, on the average. Written informed consent to the protocol, which was approved by the local ethics committee, was obtained from all volunteers. Ischemia in the lower leg was induced by inflating a pneumatic cuff. Control of the cuff pressure was given over to the volunteers themselves, to enhance compliance. To ensure fully ischemic exercise, subjects were instructed to inflate the cuff to 230 mm Hg (i.e., well above the systolic blood pressure) 2 min before commencing plantar flexion exercise, to maintain constant cuff pressure, and to deflate the cuff after an ischemic postexercise period as long as the subjects would tolerate. On average, postexercise ischemia lasted 4 min 20 sec. Subjects were instructed to confine exercise to periods without RF excitation and reception, i.e., not to exert force on the pedal during and shortly after gradient noise, in order to reduce calf motion during acquisition. Force on the pedal and its angle were measured using sensors and were recor...

We demonstrate the feasibility of dynamic perfusion quantification in skeletal muscle at 7 Tesla using PASL. This may help to better investigate the physiological processes in the skeletal muscle and also in diseases such as diabetes mellitus and peripheral arterial disease.