Assessment of absolute metabolite concentrations in human tissue by 31P MRS in vivo. Part II: Muscle,liver, kidney

Absolute metabolite concentrations were assessed in the muscle, the liver, and the kidney of healthy human volunteers by ³¹P MRS. Fully relaxed in vivo spectra were acquired with a surface coil and were localized with an adiabatic lSlS pulse sequence. The spectra were quantified with a subsequent measurement of a calibration phantom and were processed iteratively in the time domain. The following mean metabolite concentrations (mmollliter) were measured in the resting male calf muscle (n = 9), in the fasting liver (n = 12), and in the orthiotopic kidney (n= 5): [PME] = 2.0 ± 0.6, 3.8 ± 0.7, and 2.6 ± 0.9, [Pi] = 2.9 ± 0.3, 1.8 ± 0.3, and 1.6 ± 0.4, [PDE] = 3.8 ± 0.8, 9.7 ± 1.5, and 4.9 ± 1.1, [PCr] = 22.0 ± 1.2, 0, and 0, [NTP] = 5.7 ± 0.4, 2.9 ± 0.4, and 2.0 ± 0.3, respectively. Several interesting findings are to be emphasized: The concentrations of Pi, PCr, and NTP were 20% lower in the muscle of women than of men. In addition, the pH, was significantly lower in female muscle (6.99 ± 0.03) than in male muscle (7.05 ± 0.03). The pH, in the liver (7.12 ± 0.09) and in the kidney (7.09 ± 0.08) were higher than in the muscle of both genders. The free magnesium concentration (mmollliter) was higher in the lliver (1.40 ± 0.64) than in the kidney (0.79 ± 0.39) and in the muscle (0.52 ± 0.10).     

Comparison of methods for the determination of absolute metabolite concentrations in human muscles by 31P MRS

In order to determine metabolite concentrations in human skeletal muscles by in vivo ³¹P MRS, different quantification methods were analyzed with regard to the accuracy and reproducibility of results and the simplicity of handling. Each quantification method comprised a calibration strategy and a localization technique. Extensive in vivo and in vitro tests showed that homonuclear phantom-based calibration strategies yielded significantly more accurate (lower systematic errors) and more reproducible (lower statistical errors) concentration estimates than heteronuclear strategies using internal water as a concentration standard. Additionally, the former strategies are easier to handle than the latter. Localization with the volume-selective sequence lSlS yielded slightly more reproducible results than localization by surface coil. We conclude that phosphorus metabolite concentrations are determined most accurately with phantom-based calibration strategies in combination with lSlS localization (measurement errors ≈ 5–7%).     

Regional metabolite concentrations in human brain as determined by quantitative localized proton MRS

The regional distribution of brain metabolites was studied in several cortical white and gray matter areas, cerebellum, and thalamus of young adults with use of quantitative single-voxel proton MRS at 2.0 T. Whereas the neuronal compound N-acetylaspartate is distributed homogeneously throughout the brain, N-acetylaspartylglutamate increases caudally and exhibits higher concentrations in white matter than in gray matter. Creatine, myo-inositol, glutamate, and glutamine are less concentrated in cortical white matter than in gray matter. The highest creatine levels are found in cerebellum, parallel to the distribution of creatine kinase and energy-requiring processes in the brain. Also myo-inositol has highest concentrations in the cerebellum. Choline-containing compounds exhibit a marked regional variability with again highest concentrations in cerebellum and lowest levels and a strong caudally decreasing gradient in gray matter. The present findings neither support a metabolic gender difference (except for a 1.3-fold higher myo-inositol level in parietal white matter of female subjects) nor a metabolic hemispheric asymmetry.     

Evaluation of 31P metabolite differences in human cerebral gray and white matter

³¹P NMR is commonly used to study brain energetics in health and disease. Due to sensitivity constraints, the NMR measurements are typically made in volumes that do not contain pure gray or white matter. For accurate evaluation of abnormalities in brain metabolite levels, it is necessary to consider the differences in normal levels of ³¹P metabolites in gray and white matter. In this study, voxels from a three-dimensional spectroscopic image acquisition were analyzed for their dependence on tissue type to assess differences in metabolite levels between gray and white matter. Specifically, gray matter was found to have significantly higher ratios of phosphocreatine (PCr) to γ-ATP and PCr to the total ³¹P metabolite signal, whereas pH and the ratio of PCr to inorganic phosphate (P₁) were found to differ insignificantly between gray and white matter. Thus, tissue type can be an important factor to consider for alterations in bioenergetics by ³¹P NMR spectroscopic studies of the brain.     

Use of phased array coils for a determination of absolute metabolite concentrations.

This work describes the use of phased array coils for a quantification of absolute metabolite concentrations. The method is demonstrated for single-voxel localized proton MRS of human brain with an eight-element receive-only head coil. It is based on the transmitter reference amplitude of the body coil used for RF transmission. A relative sensitivity of every element of the phased array coil is derived from a combination of two reference scans without water suppression that correspond to either the body coil in transmit-receive mode or the phased array coil in conjunction with body coil excitation. Experimental results were obtained at 2.9 T for both phantoms and 12 human subjects in different locations of gray and white matter. The data demonstrate that the procedure is technically robust and without a penalty in measuring time. Moreover, it takes full advantage of the signal-to-noise gain for quantitative proton MRS and may be extended to other phased array coils without the need for a recalibration.     

Localized proton MRS of cerebral metabolite profiles in different mouse strains.

Localized proton MR spectroscopy (MRS) was used to quantify cerebral metabolite concentrations in NMRI (n = 8), BALB/c (n = 7), and C57BL/6 (n = 8) mice in vivo and 1 hr after global irreversible ischemia (2.35 T, STEAM, TR/TE/TM = 6000/20/10 ms, 4 x 3 x 4 mm(3) volume, corrections for cerebrospinal fluid). Anatomical MRI and proton MRS revealed significant differences of the C57BL/6 strain in comparison with both BALB/c and NMRI mice. While MRI volumetry yielded larger ventricular spaces of the C57BL/6 strain, proton MRS resulted in elevated concentrations of N-acetylaspartate (tNAA), creatine and phosphocreatine (tCr), choline-containing compounds (Cho), glucose (Glc), and lactate (Lac) relative to BALB/c mice and elevated Glc relative to NMRI mice. Apart from the expected decrease of Glc and increase of Lac 1 hr post mortem, C57BL/6 mice presented with significant reductions of tNAA, tCr, and Cho, whereas these metabolites remained unchanged in BALB/c and NMRI mice. The results support the hypothesis that the more pronounced vulnerability of C57BL/6 mice to brain ischemia is linked to strain-dependent differences of the cerebral energy metabolism.     

Reproducibility of in vivo metabolite quantification with proton magnetic resonance spectroscopic imaging

PURPOSE: To investigate intra- and interscanner in vivo reproducibility of brain metabolite quantification using 1H magnetic resonance spectroscopic imaging (1H-MRSI) (PRESS localization, TE = 30 msec, voxel volume = 2.3 mL) and the linear combination model (LCModel). MATERIALS and METHODS: One subject had a total of nine scans on three occasions at a single site, and three subjects had single scans at two sites. Coefficients of variation (CVs) were estimated using different statistical models applied to intra- and interscanner data; therefore, only qualitative comparisons may be made between results. RESULTS: CV (intra-/interscanner) for metabolite quantifications were choline, 12.3%/10.1%; creatine, 9.9%/10.6%; glutamate + glutamine, 15.8%/13.6%; myo-inositol, 18.5%/14.7%; and N-acetyl-aspartate + N-acetyl-aspartyl-glutamate, 6.1%/7.0%. Overall, total intra- and intersubject variability was greater than intra- and interscanner variability. CONCLUSION: When quantifying metabolic concentrations using the methods employed in this study, biological factors contribute a greater proportion to measurement variability than measurement errors. Using this technique, intra- and intersite measurement errors are of the same order.     

Absolute quantification of human liver metabolite concentrations by localized in vivo 31P NMR spectroscopy in diffuse liver disease

Phosphorus-31 NMR spectroscopy using slice selection (DRESS) was used to investigate the absolute concentrations of metabolites in the human liver. Absolute concentrations provide more specific biochemical information compared to spectrum integral ratios. Nine patients with histopathologically proven diffuse liver disease and 12 healthy individuals were examined in a 1.5-T MR scanner (GE Signa LX Echospeed plus). The metabolite concentration quantification procedures included: (1) determination of optimal depth for the in vivo measurements, (2) mapping the detection coil characteristics, (3) calculation of selected slice and liver volume ratios using simple segmentation procedures and (4) spectral analysis in the time domain. The patients had significantly lower concentrations of phosphodiesters (PDE), 6.3±3.9 mM, and ATP-, 3.6±1.1 mM, (P<0.05) compared with the control group (10.0±4.2 mM and 4.2±0.3 mM, respectively). The concentrations of phosphomonoesters (PME) were higher in the patient group, although this was not significant. Constructing an anabolic charge (AC) based on absolute concentrations, [PME]/([PME] + [PDE]), the patients had a significantly larger AC than the control subjects, 0.29 vs. 0.16 (P<0.005). Absolute concentration measurements of phosphorus metabolites in the liver are feasible using a slice selective sequence, and the technique demonstrates significant differences between patients and healthy subjects.     

Measurement of glycine in gray and white matter in the human brain in vivo by 1H MRS at 7.0 T

The concentration of glycine (Gly) was measured in gray matter (GM) and white matter (WM) in the human brain using single‐voxel localized ¹H MRS at 7 T. A point‐resolved spectroscopy sequence with echo time = 150 ms was used for measuring Gly levels in various regions of the frontal and occipital lobes in 11 healthy volunteers and one subject with a glioblastoma. The point‐resolved spectroscopy spectra were analyzed with LCModel using basis functions generated from density matrix simulations that included the effects of volume localized radio‐frequency and gradient pulses. The fraction of GM and white matter within the voxels was obtained from T₁‐weighted image segmentation. The metabolite concentrations within the voxels, estimated with respect to the GM + WM water concentrations, were fitted to a linear function of fractional GM content. The Gly concentrations in pure GM and white matter were estimated to be 1.1 and 0.1 mM, with 95% confidence intervals 1.0–1.2 and 0.0–0.2, respectively. Magn Reson Med, 2012. © 2012 Wiley Periodicals, Inc.     

A critical assessment of noise-induced errors in 31P MRS: Application to the measurement of free intracellular magnesium in vivo

Phosphorus magnetic resonance spectroscopy (³¹P MRS) is a noninvasive technique that has been used to estimate free intracellular magnesium concentration (free [Mg²⁺]). Free [Mg²⁺] is computed from the chemical shift separation between the α- and β-phosphate resonances of ATP. The current study was undertaken to critically assess the influence of noise effects in estimating free [Mg²⁺] in rat brain subjected to moderate parasagittal fluid percussion-induced injury. We show that contrary to published data, free [Mg²⁺] does not significantly change for up to 4 h after moderate trauma in different rat strains and using different surface coils. Before injury, free [Mg²⁺] = 0.56 ± 0.11 (mean ± SD, n = 36) and 4 h post-trauma, free [Mg²⁺] = 0.56 ± 0.28. Our results suggest that explanations for this discrepancy comprise errors of chemical shift assignments accompanying low signal-to-noise ratios and the method of analysis employed. Indeed, the authors propose that spectra of β-ATP signal-to-noise ratio less than 5:1 will produce significant noise-induced errors. We conclude that without knowledge of the inherent errors in ³¹P MRS spectroscopy and appropriate statistical analysis, caution should be exercised in calculating free [Mg²⁺] and using these changes as a basis for proposing pharmacotherapeutic interventions.     

Assessment of glutamate and glutamine contribution to in vivo N-acetylaspartate quantification in human brain by (1)H-magnetic resonance spectroscopy.

N-Acetylaspartate (NAA) is one of the most important metabolites detectable by brain (1)H-MRS being considered an index of neuronal integrity. At the low magnetic field used in most clinical settings beta,gamma-glutamate/glutamine (Glx) resonances are very close and partially overlap the methyl-NAA resonance interfering with NAA quantification especially at low TE and in the presence of increased Glx signals. NAA overestimation due to Glx on a set of model solutions containing NAA, glutamate, and glutamine in variable amounts was evaluated and the result tested in vivo in six healthy controls and five age- and sex-matched patients with hepatic encephalopathy (HE), the latter having an increased Glx content. A method to assess in vivo the NAA overestimation caused by Glx is proposed. A perfect match was obtained between the assessment of Glx contamination on the NAA of healthy controls and that obtained on the model solutions. However, a substantial difference in NAA overestimation was found between controls and HE patients that cannot be explained by our model. An interpretative hypothesis is provided.     

Quantitative (31)P spectroscopic imaging of human brain at 4 Tesla: assessment of gray and white matter differences of phosphocreatine and ATP.

This report describes the implementation and application of a multicompartment analysis of (31)P spectroscopic imaging data to determine the tissue-specific heterogeneities in metabolite content in the human brain and surrounding tissue. Using this information and a multicompartment regression analysis the phosphocreatine and ATP content of "pure" cerebral gray and white matter, the cerebellum, and skeletal muscle was determined in a group of 10 healthy volunteers. The data were converted to mM units using previously reported values for the T(1)s of phosphocreatine and ATP at 4 T, the water content of human brain, and an external reference for absolute quantification. The phosphocreatine concentration in cerebral gray and white matter, the cerebellum, and skeletal muscle was 3.53 +/- 0.33, 3.33 +/- 0.37, 3.75 +/- 0.66, and 25.8 +/- 2.3 mM, respectively. The ATP concentration in cerebral gray and white matter, the cerebellum, and skeletal muscle was 2.19 +/- 0.33, 3.41 +/- 0.33, 1.75 +/- 0.58, and 8.5 +/- 1.9 mM, respectively. Magn Reson Med 45:46-52, 2001.     

In vivo 31P magnetic resonance spectroscopy of human brain at 7 T: an initial experience.

In vivo (31)P spectra were acquired from the human primary visual cortex at 7 T. The relaxation times of the cerebral metabolites, intracellular pH, rate constant (k(f)) of the creatine kinase (CK) reaction, and nuclear Overhauser enhancement (NOE) on the detected phosphorus moieties from irradiation of the water spins were measured from normal subjects. With a 5-cm-diameter surface coil, 3D (31)P chemical shift imaging was performed with a spatial resolution of 7.5 ml and an acquisition resolution of 8 min, resulting in a signal-to-noise ratio (SNR) for phosphocreatine (PCr) resonance of 32. The apparent T(1) and T(2) of PCr measured at 7 T were 3.37 +/- 0.29 s and 132.0 +/- 12.8 ms, respectively, which were considerably longer than those of adenosine triphosphate (ATP) (T(1): 1.02-1.27 s; T(2): 25-26 ms). The NOE measured in this study was 24.3% +/- 1.6% for PCr, and 10% for ATP. The k(f) measured in the human primary visual cortex was 0.24 +/- 0.03 s(-1). The results from this study suggest that ultra-high-field strength is advantageous for performing in vivo (31)P magnetic resonance spectroscopy (MRS) in the human brain.     

Differentiation of metabolic concentrations between gray matter and white matter of human brain by in vivo 1H magnetic resonance spectroscopy

Differentiation of absolute metabolite concentrations between gray and white matter in the occipital region of normal human brain was performed by in vivo localized single-voxel ¹H magnetic resonance spectroscopy at 1.5 Testa with long echo time (136 ms). With the combination of image segmentation between white and gray matter and cerebrospinal fluid, signal compensation of T, and T₂ effects, tissue water signal as the internal concentration reference, as well as compensation by different water contents in gray and white matters, it was determined that the levels of N-acetylaspartate (NAA), creatine and/or phosphocreatine (Cr), and choline-containing compounds (Cho) in gray matter were significantly higher than in white matter. The averaged NAA, Cr, and Cho concentrations in gray matter were 11.0, 9.7, and 1.9 mM/liter, respectively, in comparison with 7.5, 5.2, and 1.6 mM/liter in white matter. These results suggest that precise composition of white and gray matter and cerebrospinal fluid is necessary to avoid partial voluming effect in a single voxel and to accurately quantify the metabolite concentrations.     

Assessment of metabolite quantitation reproducibility in serial 3D-(1)H-MR spectroscopic imaging of human brain using stereotactic repositioning.

Intrasubject reproducibility of metabolite quantitation in three-dimensional proton magnetic resonance spectroscopic imaging (3D-MRSI) was investigated in 10 healthy volunteers over five separate sessions using two echo times (TEs): 144 and 30 ms. The use of a Gill-Thomas-Cosman (GTC) stereotactic head frame enabled precise subject repositioning and immobilization. Metabolite levels from each voxel in the volume of interest (VOI) were quantified using the Linear Combination of Model spectra (LCModel) analysis algorithm, and coefficients of variation (CVs) were calculated. Standard error estimates (%SD or Cramer-Rao lower bounds) generated by LCModel were used as a confidence filter. The 95% confidence interval (CI) was found for each metabolite, providing an indication of the normal fluctuation expected for 3D-MRSI. In vivo, median CVs at the %SD < or = 20 level were found to be (%CV for TE = 144 and 30 ms, respectively): N-acetyl-aspartate plus N-acetyl-aspartyl-glutamate (NAA): 10.2% and 13.5%; creatine plus phosphocreatine (Cr), 14.4% and 21.7%; and choline-containing compounds (Cho), 15.2% and 18.4%. Relaxing the statistical filtering criteria to %SD < or = 30 increased median CVs by less than 5% and permitted in vivo quantitation reproducibility to be evaluated for glutamine plus glutamate (Glx) and myoinositol (Ins) for TE = 30 ms, yielding CVs of 24.0% and 21.0%, respectively.     

Experimental design of 31P MRS assessment of human forearm muscle function: Restrictions imposed by functional anatomy

The restrictions imposed by the functional anatomy of the finger flexor muscles on the experimental design of ³¹P MRS assessment of human forearm muscle function employing surface coil localization and voluntary exercise were investigated. It was found that ³¹P MRS metabolic data of finger flexor muscle should be correlated with mechanical data of combined flexion of only the ring and little fingers, rather than all four fingers as has been commonly the case in previously reported studies.     

Comparative study of cerebral white matter in autism and attention-deficit/hyperactivity disorder by means of magnetic resonance spectroscopy

RATIONALE AND OBJECTIVES: Autism and attention-deficit/hyperactivity disorder (ADHD) are neurodevelopmental disorders whose pathophysiology is mostly unknown. As far as the symptoms are different and, in some aspects, opposed, we hypothesize that there must be biochemical differences in the brain of the afflicted children. The aim of the study is to analyze comparatively the metabolite concentration of the cerebral white matter in autism, in ADHD, and in a control group of healthy children to test the hypothesis that N-acetyl aspartate (NAA) is decreased in autism and increased in ADHD. PATIENTS AND METHODS: We included 21 autistic children according to DSM-IV criteria, 8 children with ADHD meeting the respective criteria of DSM-IV, and 12 healthy controls of similar age. Single-voxel proton magnetic resonance spectroscopy was performed on all of them with an echo time of 30 milliseconds and a repetition time of 2500 milliseconds. The voxel was placed in the left centrum semiovale. Metabolite ratios relative to creatine were reported for NAA, choline, and myoinositol. RESULTS: Although we did not observe differences between autistic children and controls, we found a mean higher concentration of NAA in the left centrum semiovale of ADHD children (2.2; SD, 0.21) than that found in autistic children (1.88; SD, 0.18) and controls (1.91; SD, 0.01), which was significant (P = .01 in parametric and in nonparametric test). CONCLUSION: We conclude that white matter of autistic children does not present alterations on MRS. We hypothesize that the higher concentration of NAA in the white matter of ADHD points to mitochondrial hypermetabolism. This may constitute a new substrate in the pathophysiology and merits further research.