Reproducibility of 2D GluCEST in healthy human volunteers at 7 T.

PURPOSE: To investigate the reproducibility of gray and white matter glutamate contrast of a brain slice among a small group of healthy volunteers by using the 2D single-slice glutamate CEST (GluCEST) imaging technique. METHODS: Six healthy volunteers were scanned multiple times for within-day and between-day reproducibility. One more volunteer was scanned for within-day reproducibility at 7T MRI. Glutamate CEST contrast measurements were calculated for within subjects and among the subjects and the coefficient of variations are reported. RESULTS: The GluCEST measurements were highly reproducible in the gray and white matter area of the brain slice, whether it was within-day or between-day with a coefficient of variation of less than 5%. CONCLUSION: This preliminary study in a small group of healthy volunteers shows a high degree of reproducibility of GluCEST MRI in brain and holds promise for implementation in studying age-dependent changes in the brain.

Longitudinal imaging reveals subhippocampal dynamics in glutamate levels associated with histopathologic events in a mouse model of tauopathy and healthy mice.

Tauopathies are neurodegenerative disorders characterized by abnormal intracellular aggregates of tau protein, and include Alzheimer's disease, corticobasal degeneration, frontotemporal dementia, and traumatic brain injury. Glutamate metabolism is altered in neurodegenerative disorders manifesting in higher or lower concentrations of glutamate, its transporters or receptors. Previously, glutamate chemical exchange saturation transfer (GluCEST) magnetic resonance imaging (MRI) demonstrated that glutamate levels are reduced in regions of synapse loss in the hippocampus of a mouse model of late-stage tauopathy. We performed a longitudinal GluCEST imaging experiment paired with a cross-sectional study of histologic markers of tauopathy to determine whether (1) early GluCEST changes are associated with synapse loss before volume loss occurs in the hippocampus, and whether (2) subhippocampal dynamics in GluCEST are associated with histopathologic events related to glutamate alterations in tauopathy. Live imaging of the hippocampus in three serial slices was performed without exogenous contrast agents, and subregions were segmented based on a k-means cluster model. Subregions of the hippocampus were analyzed (cornu ammonis CA1, CA3, dentate gyrus DG, and ventricle) in order to associate local MRI-observable changes in glutamate with histological measures of glial cell proliferation (GFAP), synapse density (synaptophysin, VGlut1) and glutamate receptor (NMDA-NR1) levels. Early differences in GluCEST between healthy and tauopathy mice were measured in the CA1 and DG subregions (30% reduction, P ≤ 0.001). Synapse density was also significantly reduced in every subregion of the hippocampus in tauopathy mice by 6 months. Volume was not significantly reduced in any subregion until 13 months. Further, a gradient in glutamate levels was observed in vivo along hippocampal axes that became polarized as tauopathy progressed. Dynamics in hippocampal glutamate levels throughout lifetime were most closely correlated with combined changes in synaptophysin and GFAP, indicating that GluCEST imaging may be a surrogate marker of glutamate concentration in glial cells and at the synaptic level. © 2016 Wiley Periodicals, Inc.

Muscle oxidative phosphorylation quantitation using creatine chemical exchange saturation transfer (CrCEST) MRI in mitochondrial disorders.

Systemic mitochondrial energy deficiency is implicated in the pathophysiology of many age-related human diseases. Currently available tools to estimate mitochondrial oxidative phosphorylation (OXPHOS) capacity in skeletal muscle in vivo lack high anatomic resolution. Muscle groups vary with respect to their contractile and metabolic properties. Therefore, muscle group-specific estimates of OXPHOS would be advantageous. To address this need, a noninvasive creatine chemical exchange saturation transfer (CrCEST) MRI technique has recently been developed, which provides a measure of free creatine. After exercise, skeletal muscle can be imaged with CrCEST in order to make muscle group-specific measurements of OXPHOS capacity, reflected in the recovery rate (τCr) of free Cr. In this study, we found that individuals with genetic mitochondrial diseases had significantly (P = 0.026) prolonged postexercise τCr in the medial gastrocnemius muscle, suggestive of less OXPHOS capacity. Additionally, we observed that lower resting CrCEST was associated with prolonged τPCr, with a Pearson's correlation coefficient of -0.42 (P = 0.046), consistent with previous hypotheses predicting that resting creatine levels may correlate with 31P magnetic resonance spectroscopy-based estimates of OXPHOS capacity. We conclude that CrCEST can noninvasively detect changes in muscle creatine content and OXPHOS capacity, with high anatomic resolution, in individuals with mitochondrial disorders.

Localized, gradient-reversed ultrafast z-spectroscopy in vivo at 7T.

PURPOSE: To collect ultrafast z-spectra in vivo in situations where voxel homogeneity cannot be assured. THEORY: Saturating in the presence of a gradient encodes the frequency offset spatially across a voxel. This encoding can be resolved by applying a similar gradient during readout. Acquiring additional scans with the gradient polarity reversed effectively mirrors the spatial locations of the frequency offsets so that the same physical location of a positive offset in the original scan will contribute a negative offset in the gradient-reversed scan. METHODS: Gradient-reversed ultrafast z-spectroscopy (GRUFZS) was implemented and tested in a modified, localized PRESS sequence at 7T. Lysine phantoms were scanned at various concentrations and compared with coventionally-acquired z-spectra. Scans were acquired in vivo in human brain from homogeneous and inhomogeneous voxels with the ultrafast direction cycled between read, phase, and slice. Results were compared to those from a similar conventional z-spectroscopy PRESS-based sequence. RESULTS: Asymmetry spectra from GRUFZS are more consistent and reliable than those without gradient reversal and are comparable to those from conventional z-spectroscopy. GRUFZS offers significant acceleration in data acquisition compared to traditional chemical exchange saturation transfer methods with high spectral resolution and showed higher relative SNR effficiency. CONCLUSION: GRUFZS offers a method of collecting ultrafast z-spectra in voxels with the inhomogeneity often found in vivo. Magn Reson Med 76:1039-1046, 2016. © 2016 Wiley Periodicals, Inc.

Lactate Chemical Exchange Saturation Transfer (LATEST) Imaging in vivo A Biomarker for LDH Activity.

Non-invasive imaging of lactate is of enormous significance in cancer and metabolic disorders where glycolysis dominates. Here, for the first time, we describe a chemical exchange saturation transfer (CEST) magnetic resonance imaging (MRI) method (LATEST), based on the exchange between lactate hydroxyl proton and bulk water protons to image lactate with high spatial resolution. We demonstrate the feasibility of imaging lactate with LATEST in lactate phantoms under physiological conditions, in a mouse model of lymphoma tumors, and in skeletal muscle of healthy human subjects pre- and post-exercise. The method is validated by measuring LATEST changes in lymphoma tumors pre- and post-infusion of pyruvate and correlating them with lactate determined from multiple quantum filtered proton magnetic resonance spectroscopy (SEL-MQC (1)H-MRS). Similarly, dynamic LATEST changes in exercising human skeletal muscle are correlated with lactate determined from SEL-MQC (1)H-MRS. The LATEST method does not involve injection of radioactive isotopes or labeled metabolites. It has over two orders of magnitude higher sensitivity compared to conventional (1)H-MRS. It is anticipated that this technique will have a wide range of applications including diagnosis and evaluation of therapeutic response of cancer, diabetes, cardiac, and musculoskeletal diseases. The advantages of LATEST over existing methods and its potential challenges are discussed.

In vivo magnetic resonance imaging of tumor protease activity.

Increased expression of cathepsins has diagnostic as well as prognostic value in several types of cancer. Here, we demonstrate a novel magnetic resonance imaging (MRI) method, which uses poly-L-glutamate (PLG) as an MRI probe to map cathepsin expression in vivo, in a rat brain tumor model. This noninvasive, high-resolution and non-radioactive method exploits the differences in the CEST signals of PLG in the native form and cathepsin mediated cleaved form. The method was validated in phantoms with known physiological concentrations, in tumor cells and in an animal model of brain tumor along with immunohistochemical analysis. Potential applications in tumor diagnosis and evaluation of therapeutic response are outlined.

In vivo measurement of glutamate loss is associated with synapse loss in a mouse model of tauopathy.

Glutamate is the primary excitatory neurotransmitter in the brain, and is implicated in neurodegenerative diseases such as Alzheimer's disease (AD) and several other tauopathies. The current method for measuring glutamate in vivo is proton magnetic resonance spectroscopy ((1)H MRS), although it has poor spatial resolution and weak sensitivity to glutamate changes. In this study, we sought to measure the effect of tau pathology on glutamate levels throughout the brain of a mouse model of tauopathy using a novel magnetic resonance imaging (MRI) technique. We employed glutamate chemical exchange saturation transfer (GluCEST) imaging, which has been previously validated as a complimentary method for measuring glutamate levels with several important advantages over conventional (1)H MRS. We hypothesized that the regional changes in glutamate levels would correlate with histological measurements of pathology including pathological tau, synapse and neuron loss. Imaging and spectroscopy were carried out on tau transgenic mice with the P301S mutation (PS19, n=9) and their wild-type littermates (WT, n=8), followed by immunohistochemistry of their brain tissue. GluCEST imaging resolution allowed for sub-hippocampal analysis of glutamate. Glutamate was significantly decreased by 29% in the CA sub-region of the PS19 hippocampus, and by 15% in the thalamus, where synapse loss was also measured. Glutamate levels and synapse density remained high in the dentate gyrus sub-region of the hippocampus, where neurogenesis is known to occur. The further development of GluCEST imaging for preclinical applications will be valuable, as therapies are being tested in mouse models of tauopathy.

In vivo chemical exchange saturation transfer imaging of creatine (CrCEST) in skeletal muscle at 3T.

PURPOSE: To characterize the chemical exchange saturation transfer (CEST)-based technique to measure free creatine (Cr), a key component of muscle energy metabolism, distribution in skeletal muscle with high spatial resolution before and after exercise at 3T. MATERIALS AND METHODS: CrCEST saturation parameters were empirically optimized for 3T. CEST, T2 , magnetization transfer ratio (MTR), and (31) P magnetic resonance spectroscopy (MRS) acquisitions of the lower leg were performed before and after mild plantar flexion exercise on a 3T whole-body MR scanner on six healthy volunteers. RESULTS: The feasibility of imaging Cr changes in skeletal muscle following plantar flexion exercise using CrCEST was demonstrated at 3T. This technique exhibited good spatial resolution and was able to differentiate differences in muscle use among subjects. The CrCEST results were compared with (31) P MRS results, showing good agreement in the Cr and PCr recovery kinetics. A relationship of 0.45% CrCESTasym /mM Cr was observed across all subjects. CONCLUSION: It is demonstrated that the CrCEST technique could be applied at 3T to measure dynamic changes in creatine in muscle in vivo. The widespread availability and clinical applicability of 3T scanners has the potential to clinically advance this method.

Glutaminase catalyzes reaction of glutamate to GABA.

Here, for the first time, we report an NMR spectroscopy study of l-Glutamine (Gln) conversion by Glutaminase (Glnase), which shows that the reaction involves two distinct steps. In the first step, Glnase rapidly hydrolyzes Gln to Glutamate (Glu) (∼16.87 µmol of Gln/min/mg of Glnase) and in the second step, Glu generated in the first step is decarboxylated into gamma-amino butyric acid (GABA) with a much slower rate (∼0.185 µmol/min/mg). When Glnase was added to the sample containing l-Glu alone, it was also converted to GABA, at a similar rate as in the second step mentioned above. The rate of Glu decarboxylation into GABA by Glnase is about an order of magnitude lower than that by commonly known enzyme, Glutamate decarboxylase. Potential impact of these findings, on the mechanistic aspects of Gln-Glu shuttle in neuroscience and glutaminolysis in tumors, is discussed.

A technique for in vivo mapping of myocardial creatine kinase metabolism.

ATP derived from the conversion of phosphocreatine to creatine by creatine kinase provides an essential chemical energy source that governs myocardial contraction. Here, we demonstrate that the exchange of amine protons from creatine with protons in bulk water can be exploited to image creatine through chemical exchange saturation transfer (CrEST) in myocardial tissue. We show that CrEST provides about two orders of magnitude higher sensitivity compared to (1)H magnetic resonance spectroscopy. Results of CrEST studies from ex vivo myocardial tissue strongly correlate with results from (1)H and (31)P magnetic resonance spectroscopy and biochemical analysis. We demonstrate the feasibility of CrEST measurement in healthy and infarcted myocardium in animal models in vivo on a 3-T clinical scanner. As proof of principle, we show the conversion of phosphocreatine to creatine by spatiotemporal mapping of creatine changes in the exercised human calf muscle. We also discuss the potential utility of CrEST in studying myocardial disorders.

Method for high-resolution imaging of creatine in vivo using chemical exchange saturation transfer.

PURPOSE: To develop a chemical exchange saturation transfer (CEST)-based technique to measure free creatine (Cr) and to validate the technique by measuring the distribution of Cr in muscle with high spatial resolution before and after exercise. METHODS: Phantom studies were performed to determine contributions from other Cr kinase metabolites to the CEST effect from Cr (CrCEST). CEST, T2 , magnetization transfer ratio and (31) P magnetic resonance spectroscopy acquisitions of the lower leg were performed before and after plantar flexion exercise on a 7T whole-body magnetic resonance scanner on healthy volunteers. RESULTS: Phantom studies demonstrated that while Cr exhibited significant CEST effect there were no appreciable contributions from other metabolites. In healthy human subjects, following mild plantar flexion exercise, increases in the CEST effect from Cr were observed, which recovered exponentially back to baseline. This technique exhibited good spatial resolution and was able to differentiate differences in muscle utilization among subjects. The CEST effect from Cr results were compared with (31) P magnetic resonance spectroscopy results showing good agreement in the Cr and phosphocreatine recovery kinetics. CONCLUSION: Demonstrated a CEST-based technique to measure free Cr changes in in vivo muscle. The CEST effect from Cr imaging can spatially map changes in Cr concentration in muscle following mild exercise. This may serve as a tool for the diagnosis and treatment of various disorders affecting muscle.

Imaging of glutamate in the spinal cord using GluCEST.

Glutamate (Glu) is the most abundant excitatory neurotransmitter in the brain and spinal cord. The concentration of Glu is altered in a range of neurologic disorders that affect the spinal cord including multiple sclerosis (MS), amyotrophic lateral sclerosis (ALS) and spinal cord injury. Currently available magnetic resonance spectroscopy (MRS) methods for measuring Glu are limited to low spatial resolution, which makes it difficult to measure differences in gray and white matter glutamate. Recently, it has been shown that Glu exhibits a concentration dependent chemical exchange saturation transfer (CEST) effect between its amine (-NH2) group protons and bulk water protons (GluCEST). Here, we demonstrate the feasibility of imaging glutamate in the spinal cord at 7T using the GluCEST technique. Results from healthy human volunteers (N=7) showed a significantly higher (p<0.001) GluCESTasym from gray matter (6.6±0.3%) compared to white matter (4.8±0.4%). Potential overlap of CEST signals from other spinal cord metabolites with the observed GluCESTasym is discussed. This noninvasive approach potentially opens the way to image Glu in vivo in the spinal cord and to monitor its alteration in many disease conditions.