Diffusion-weighted MRS of acetate in the rat brain

Acetate has been proposed as an astrocyte-specific energy substrate for metabolic studies in the brain. The determination of the relative contribution of the intracellular and extracellular compartments to the acetate signal using diffusion-weighted magnetic resonance spectroscopy can provide an insight into the cellular environment and distribution volume of acetate in the brain. In the present study, localized (1) H nuclear magnetic resonance (NMR) spectroscopy employing a diffusion-weighted stimulated echo acquisition mode (STEAM) sequence at an ultra-high magnetic field (14.1 T) was used to investigate the diffusivity characteristics of acetate and N-acetylaspartate (NAA) in the rat brain in vivo during prolonged acetate infusion. The persistence of the acetate resonance in (1) H spectra acquired at very large diffusion weighting indicated restricted diffusion of acetate and was attributed to intracellular spaces. However, the significantly greater diffusion of acetate relative to NAA suggests that a substantial fraction of acetate is located in the extracellular space of the brain. Assuming an even distribution for acetate in intracellular and extracellular spaces, the diffusion properties of acetate yielded a smaller volume of distribution for acetate relative to water and glucose in the rat brain.

Proton diffusion spectroscopy and modeling of brain metabolism at 14.1T

Masoumeh Dehghani Moghadam

As a field at the CIBM, Nuclear magnetic resonance (NMR)spectroscopy can be applied non-invasively to explore the metabolic fate of energy fuel substrates, as well as the rate at which they are consumed, using 13C and 1H nuclei. The work of this thesis encompasses both nuclei, and focuses on (1) improving the quantification and modeling of glucose-derived metabolites; and (2) characterizing diffusion-related parameters of the purportedly glial-specific energy substrate, acetate. Both aim to quantitatively explore cerebral energy metabolism, at ultra-high magnetic field, in vivo, in the healthy rat. 13C NMR spectroscopy, as a tool, enables measuring the progressive incorporation of 13C-glucose into brain glucose and then NMR detectable amino acids (glutamate and glutamine); this relies on the infusion of the 13C-labeled energy substrate. The experimentally obtained 13C labelling curves are analyzed using suitable mathematical models to provide an estimation of cerebral metabolic rates. Here, a dynamic model of time-courses of 13C multiplets arising from isotopomers was considered. So beyond the two-compartment neuronal-glial model, we took into account additional data on the dynamics of 13C isotopomers, available from the fine structure multiplets in 13C spectra of glutamate and glutamine, measured under prolonged [1-6,13C] glucose infusion. We concluded that the dynamic analyses of 13C multiplet time courses of glutamate and glutamine resulted in a higher precision for estimating the absolute values of most cerebral metabolic rates. Acetate metabolism is challenging because dynamic metabolic modeling requires prior knowledge of the transport and uptake kinetics of infused acetate. We sought this information by determining the apparent concentration and distribution volume (V_d) of cerebral acetate between the intracellular and the extracellular compartments. Experimentally, the diffusion characteristics of cerebral acetate were measured, relative to that of N-Acetylaspartate (NAA, known to be mainly intracellular) using diffusion-weighted 1H NMR spectroscopy at 14.1T, under prolonged acetate infusion. The detection of an acetate and NAA signal at large diffusion weighting provided direct experimental evidence of intracellular cerebral acetate and NAA, although a substantial fraction of acetate was extracellular. To estimate the apparent concentration of in vivo brain acetate, T1 and T2 relaxation times of acetate were measured. The longer T1 relaxation and shorter T2 relaxation times of acetate compared with NAA provided evidence of its small molecular size, and possibly different chemical environment. Our experimentally determined value of V_dled to cerebral metabolic rates of acetate (CMR acetate) of the same order reported for the glial Krebâs cycle rate, an indication that estimates of CMR acetate are highly dependent on V_d. Finally, in order to pursue metabolic mapping of cerebral acetate uptake in the rat, in vivo, at 14.1 T, the design and construction of a combined transmit-birdcage coil and receive-quadrature pair surface coil was considered. Its performance was compared to a single birdcage coil in the transmit/receive mode. So far, the preliminary results of the 2-coil configuration are promising: homogenous excitation and a gain in sensitivity up to a distance of 5 mm are achievable. Improvements are ongoing for NMR spectroscopic and imaging applications at 14.1 T.

EPFL2016

Biophysical Basis of the Diffusion-Weighted Magnetic Resonance Signal in the Rat Brain

Nicolas Kunz

Nuclear magnetic resonance (NMR) can be used in-vivo in a vast array of applications, such as anatomical imaging (magnetic resonance imaging, MRI), localized chemical composition characterization (magnetic resonance spectroscopy, MRS), cellular structure assessment (diffusion tensor imaging, DTI) and cerebral activity mapping (functional imaging, fMRI) for the most important one. This thesis focused on the development of diffusion NMR spectroscopy and imaging methods at ultra-high magnetic field, with the aim of a better characterization of the diffusion mechanism in-vivo. DTI measures the water molecule displacement due to the thermal agitation in the sample. In cellular tissue, the molecules are restrained in compartments delimited by the cell membranes, which mean that DTI can provide information on the cerebral cellular microstructure. DTI is thus widely used to investigate cerebral disorders such as brain ischemia, trauma, and tumors, as well as the structural changes occurring during brain development and normal aging. However, the presence of water molecules in both the intra- and extra-cellular compartments may bias the assessed structural information. On the other hand, the metabolites, measured by 1H-MRS, are mostly localized in the intracellular compartment, which may provide a more precise insight of the cellular structure. With the availability of high magnetic field systems, 1H-MRS allows the measurement of 15 – 20 metabolites in-vivo in localized region in the order of &icro;l in the rodent brain. These metabolites are involved in cell energy management, neurotransmission, neuro-protection, neuronal development and membrane growth, and thus provide a rather complete status of the brain neurochemical profile. In this context, a diffusion tensor spectroscopy pulse sequence (DT-MRS) was implemented and used to investigate the metabolite diffusion properties in specific brain regions in-vivo at ultra-high magnetic field 14.1 T to benefit from the increased sensitivity and spectral resolution at high B0. The diffusion tensor of 5 metabolites were reconstructed in the corpus-callosum, CC, (NAA, Tau, Glu, Ins and Mac) and 8 in the cortex, Cx, (with in addition Cr, PCr, GABA and GSH). Metabolites apparent diffusion coefficients were significantly higher in the CC (0.115 – 0.153 µm2/ms) than in the Cx (0.087 – 0.107 µm2/ms) and so was the fractional anisotropy (0.51 – 0.62 and 0.34 – 0.51 respectively). In addition, the metabolite preferential diffusion directions showed an excellent agreement with the known rat brain structure. The aforementioned DW-MRS pulse sequence was then combined with an inversion recovery module, allowing direct assessment of the macromolecule (MM) signal, which overlaps with metabolites resonances. An inaccurate estimation of the MM baseline may result in a significant bias during the quantification process. At ultra-high magnetic field, the MM spectral shape becomes more complex, and therefore, the previously reported experimental techniques would require post-processing to remove residual metabolite signals. In this study, several metabolite contaminations remaining in the metabolite nulled spectrum were successfully identified and attributed to creatine, myo-inositol, taurine, choline, N-acetylaspartate, glutamine and glutamate. The macromolecule baseline assessed by DW-MRS resulted in a twofold reduction of the metabolite residuals with minimal effect on the macromolecule signal (i.e. < 8% signal reduction). Although at high magnetic field spectral resolution and sensitivity are improved, metabolite diffusion measurements remain very time consuming and challenging due to the low metabolite concentrations in-vivo (1 – 15 mM) compared to water (44 M). Therefore, a fast and robust EPI-based DTI pulse sequence was developed, implemented and validated to investigate the cerebral architecture in-vivo at 14.1 T through water diffusion. Combined with the use of a purpose-built quadrature surface coil and adiabatic RF-pulses, it resulted in images with a full coverage of the rat brain and a total acquisition time of one hour. The rapid signal loss arising from the use of a surface coil for the MRI acquisitions, which was previously in this thesis overcome experimentally, was also addressed in collaboration with the Signal Processing Core of CIBM, by a direct correction applied on reconstructed magnitude images. Based on a low-pass frequency mode; the developed method showed better accuracy in enhancing image intensity compared to standard approaches. In view of a future application of the presented diffusion NMR techniques, investigating structural changes during brain development, a study was performed to characterize different intra-uterine growth retardation (IUGR) models by 1H-MRS. Dexamethasone-treated animals (100 µg/kg/day induced from the day 15 of gestation) showed more severe alterations in cerebral energy metabolism and neurotransmitter action at postnatal day 7 and 21 than the reference IUGR model, caloric restriction (30% food intake of ad libitum fed dams from day 1 of gestation). In a continuation of the IUGR study, the exposure to the estrogen-like molecule (i.e. bisphenol-A, BPA), was investigated. Here, 1H-MRS showed a significant increase of the Glu/Asp ratio in the hippocampus of the BPA-treated group at 20 days old, indicating a possible deviation in the malate-aspartate shuttle. Finally, the diverse NMR techniques used and developed during this thesis have required the implementation of several data processing tools, which were centralized in a framework written in Matlab (Mathworks, Inc). Two principal modules were build: 1. to reconstruct DTI experiments data and 2. to facilitate and standardize the 1H-MRS spectra quantification by LCModel. The proposed NMR data processing framework was used to process most of the data presented in this thesis and has demonstrated good performance. In addition, this framework was tested and used by five external users to analyze for data-analysis in ongoing studies.

EPFL2010

Ultra high magnetic field for glial contribution into brain metabolism studied by MR spectroscopy and CEST methods for molecular imaging of glycogen

Elise Marie Catherine Vinckenbosch

Magnetic resonance at ultra-high field increases signal and spectral dispersion. In this thesis, I use those characteristics to investigate three different subjects in 13C spectroscopy, hyperpolarized methods and chemical exchange saturation transfer (CEST) used for molecular imaging. In the brain, metabolism is driven by glia-neuron interactions and is composed of at least two compartments with distinct TCA cycle kinetics. The first study examines cerebral metabolic adaptation after an induced glial TCA-cycle impairment using 1H and 13C spectroscopy. We treated rats with fluoroacetate, which specifically blocks glial aconitase and measured changes in neurochemical profile at euglycemic and hyperglycemic condi-tions by in vivo 1H MR spectroscopy. In addition, we evaluated the brain-compartmentalized metabolic adap-tation by following 13C-incorporation into brain amino acids by modern dynamic 13C MRS methods during infu-sion of [1,6-13C2]glucose. By following [1,6-13C2]glucose incorporation, we successfully observed an augmen-tation of pyruvate carboxylase and glutamine synthetase fluxes and a 20% decrease of glial TCA cycle flux. Neuronal metabolism was also affected. In the second study, we focused on hyperpolarized methods to increase acetate 13C polarization allowing us to follow the kinetics of its metabolic products in the TCA cycle. Indeed, infusion of hyperpolarized [1-13C]acetate, an astrocyte-specific precursor, and the in vivo detection of 2-oxo[5-13C]glutarate (2OG), a TCA intermediate, enables the direct analysis of glial Krebs cycle activity. We examined the effect of hyperpolar-ized acetate concentration on its cerebral metabolism and calculate the production rate of 2OG. A healthy group was compared to rodents treated with fluoroacetate. The conversion rates of acetate to 2OG were calculated and were found to depend on the substrate dose. We successfully estimated 2OG production rates. In partially inhibited TCA cycle conditions, the production rate of 2OG was reduced by a quarter. This level of reduction is agrees with the percentage of inhibition calculated in the first study. Finally, we investigated glycogen imaging in skeletal muscle using the CEST strategy. It has been demonstrat-ed in perfused liver that glycogen can be detected indirectly through the water signal using CEST. Glycogen detection is, due to its resonance close to water, strongly sensitive to spectral dispersion and improved by a higher static magnetic field. In contrast to liver, which contains ~300 mmol/g of glycogen and enjoys good glycogen specificity, myocellular content is also rich in creatine, which produces a spectral signal on the same order of amplitude as its glycogen content of ~80-30 mmol/g. In this study, we allowed glycogen to be de-tectable in vivo and distinguishable from the neighboring creatine at 14.1T. We propose to use creatine as an internal reference for muscular glycoCEST analysis. Scanning muscle after physical exercise, resulted in a 50% reduction of glycoCEST compared to muscle at resting state. This result supportis the specificity of the CEST method and is consistent with literature. This last study demonstrates for the first time that glycoCEST imag-ing is feasible in vivo.

EPFL2018