Hongxia Lei

Effect of Deep Pentobarbital Anesthesia on Neurotransmitter Metabolism in Vivo: On the Correlation of Total Glucose Consumption with Glutamatergic Action

The effect of deep barbiturate anesthesia on brain glucose transport, TCA cycle flux, and aspartate, glutamate, and glutamine metabolism was assessed in the rat brain in vivo using 13C nuclear magnetic resonance spectroscopy at 9.4 T in conjunction with [1-13C] glucose infusions. Brain glucose concentrations were elevated, consistent with a twofold reduced cerebral metabolic rate for glucose (CMRglc) compared with light α-chloralose anesthesia. Using a mathematical model of neurotransmitter metabolism, several metabolic reaction rates were extracted from the rate of label incorporation. Total oxidative glucose metabolism, CMRglc(ox), was 0.33 ± 0.03 μmol·g−1 · min−1. The neuronal TCA cycle rate was similar to that in the glia, 0.35 ± 0.03 μmol · g−1 · min−1 and 0.26 ± 0.06 μmol · g−1 · min−1, respectively, suggesting that neuronal energy metabolism was mainly affected. The rate of pyruvate carboxylation was 0.03 ± 0.01 μmol·g−1 · min−1. The exchange rate between cytosolic glutamate and mitochondrial 2-oxoglutarate, Vx, was equal to the rate of neuronal pyruvate dehydrogenase flux. This indicates that Vx is coupled to CMRglc(ox), implying that the malate-aspartate shuttle is the major mechanism that facilitates label exchange across the inner mitochondrial membrane. The apparent rate of glutamatergic neurotransmission, VNT, was 0.04 ± 0.01 μmol·g−1 · min−1, consistent with strong reductions in electrical activity. However, the rates of cerebral oxidative glucose metabolism and glutamatergic neurotransmission, CMRglc(ox)/VNT, did not correlate with a 1:1 stoichiometry.

Neuroprotective role of lactate after cerebral ischemia

It is well established that lactate can be used as an energy substrate by the brain by conversion to pyruvate and a subsequent oxidation in the mitochondria. Knowing the need for readily metabolizable substrates directly after ischemia and the protective effect of lactate after excitotoxicity, the aim of this study was to investigate whether lactate administration directly after ischemia could be neuroprotective. In vitro, the addition of 4 mmol/L l-lactate to the medium of rat organotypic hippocampal slices, directly after oxygen and glucose deprivation (OGD), protected against neuronal death, whereas a higher dose of 20 mmol/L was toxic. In vivo, after middle cerebral artery occlusion in the mouse, an intracerebroventricular injection of 2 μL of 100 mmol/L l-lactate, immediately after reperfusion, led to a significant decrease in lesion size, which was more pronounced in the striatum, and an improvement in neurologic outcome. A later injection 1 h after reperfusion did not reduce lesion size, but significantly improved neurologic outcome, which is an important point in the context of a potential clinical application. Therefore, a moderate increase in lactate after ischemia may be a therapeutic tool.

Evolution of the neurochemical profile after transient focal cerebral ischemia in the mouse brain

Evolution of the neurochemical profile consisting of 19 metabolites after 30 mins of middle cerebral artery occlusion was longitudinally assessed at 3, 8 and 24 h in 6 to 8 µL volumes in the striatum using localized 1H-magnetic resonance spectroscopy at 14.1 T. Profound changes were detected as early as 3 h after ischemia, which include elevated lactate levels in the presence of significant glucose concentrations, decreases in glutamate and a transient twofold glutamine increase, likely to be linked to the excitotoxic release of glutamate and conversion into glial glutamine. Interestingly, decreases in N-acetyl-aspartate (NAA), as well as in taurine, exceeded those in neuronal glutamate, suggesting that the putative neuronal marker NAA is rather a sensitive marker of neuronal viability. With further ischemia evolution, additional, more profound concentration decreases were detected, reflecting a disruption of cellular functions. We conclude that early changes in markers of energy metabolism, glutamate excitotoxicity and neuronal viability can be detected with high precision noninvasively in mice after stroke. Such investigations should lead to a better understanding and insight into the sequential early changes in the brain parenchyma after ischemia, which could be used for identifying new targets for neuroprotection.

Early predictive biomarkers for lesion after transient cerebral ischemia.

Background and Purpose— Despite the improving imaging techniques, it remains challenging to predict the outcome early after transient cerebral ischemia. The aim of this study was thus to identify early metabolic biomarkers for outcome prediction. Methods— We modeled transient ischemic attacks and strokes in mice. Using high-field MR spectroscopy, we correlated early changes in the neurochemical profile of the ischemic striatum with histopathologic alterations at a later time point. Results— A significant increase in glutamine was measured between 3 hours and 8 hours after all ischemic events followed by reperfusion independently of the outcome and can thus be considered as an indicator of recent transient ischemia. On the other hand, a reduction of the score obtained by summing the concentrations of N-acetyl aspartate, glutamate, and taurine was a good predictor of an irreversible lesion as early as 3 hours after ischemia. Conclusions— We identified biomarkers of reversible and irreversible ischemic damage, which can be used in an early predictive evaluation of stroke outcome.

Neurochemical profile of the mouse hypothalamus using in vivo1H MRS at 14.1T

The hypothalamus plays an essential role in the central nervous system of mammals by among others regulating glucose homeostasis, food intake, temperature, and to some extent blood pressure. Assessments of hypothalamic metabolism using, e.g. 1H MRS in mouse models can provide important insights into its function. To date, direct in vivo 1H MRS measurements of hypothalamus have not been reported. Here, we report that in vivo single voxel measurements of mouse hypothalamus are feasible using 1H MRS at 14.1T. Localized 1H MR spectra from hypothalamus were obtained unilaterally (2–2.2 µL, VOI) and bilaterally (4–4.4 µL) with a quality comparable to that of hippocampus (3–3.5 µL). Using LCModel, a neurochemical profile consisting of 21 metabolites was quantified for both hypothalamus and hippocampus with most of the Cramér‐Rao lower bounds within 20%. Relative to the hippocampus, the hypothalamus was characterized by high γ‐aminobutryric acid and myo‐inositol, and low taurine concentrations. When studying transgenic mice with no glucose transporter isoform 8 expressed, small metabolic changes were observed, yet glucose homeostasis was well maintained. We conclude that a specific neurochemical profile of mouse hypothalamus can be measured by 1H MRS which will allow identifying and following metabolic alterations longitudinally in the hypothalamus of genetic modified models. Copyright

Minimization of Nyquist ghosting for echo-planar imaging at ultra-high fields based on a "negative readout gradient" strategy

To improve the traditional Nyquist ghost correction approach in echo planar imaging (EPI) at high fields, via schemes based on the reversal of the EPI readout gradient polarity for every other volume throughout a functional magnetic resonance imaging (fMRI) acquisition train.

Proton and Phosphorus Magnetic Resonance Spectroscopy of a Mouse Model of Alzheimer's Disease

The development of new diagnostic criteria for Alzheimers disease (AD) requires new in vivo markers reflecting early pathological changes in the brain of patients. Magnetic resonance (MR) spectroscopy has been shown to provide useful information about the biochemical changes occurring in AD brain in vivo. The development of numerous transgenic mouse models of AD has facilitated the evaluation of early biomarkers, allowing researchers to perform longitudinal studies starting before the onset of the pathology. In addition, the recent development of high-field animal scanners enables the measurement of brain metabolites that cannot be reliably quantified at lower magnetic fields. In this report, we studied a new transgenic mouse model of AD, the 5xFAD model, by in vivo proton and phosphorus MR spectroscopy. This model, which is characterized by an early-onset and a robust amyloid pathology, developed changes in the neurochemical profile, which are typical in the human disease, i.e., an increase in myo-inositol and a decrease in N-acetylaspartate concentrations, as early as in the 40th week of age. In addition, a significant decrease in the γ-aminobutyrate concentration was observed in transgenic mice at this age compared to controls. The pseudo-first-order rate constant of the creatine kinase reaction as well as relative concentrations of phosphorus-containing metabolites were not changed significantly in the 36 and 72-week old transgenic mice. Overall, these results suggest that mitochondrial activity in the 5 × FAD mice is not substantially affected but that the model is relevant for studying early biomarkers of AD.

Non-invasive quantification of brain glycogen absolute concentration

The only currently available method to measure brain glycogen in vivo is 13C NMR spectroscopy. Incorporation of 13C‐labeled glucose (Glc) is necessary to allow glycogen measurement, but might be affected by turnover changes. Our aim was to measure glycogen absolute concentration in the rat brain by eliminating label turnover as variable. The approach is based on establishing an increased, constant 13C isotopic enrichment (IE). 13C‐Glc infusion is then performed at the IE of brain glycogen. As glycogen IE cannot be assessed in vivo, we validated that it can be inferred from that of N‐acetyl‐aspartate IE in vivo: After [1‐13C]‐Glc ingestion, glycogen IE was 2.2 ± 0.1 fold that of N‐acetyl‐aspartate (n = 11, R2 = 0.77). After subsequent Glc infusion, glycogen IE equaled brain Glc IE (n = 6, paired t‐test, p = 0.37), implying isotopic steady‐state achievement and complete turnover of the glycogen molecule. Glycogen concentration measured in vivo by 13C NMR (mean ± SD: 5.8 ± 0.7 μmol/g) was in excellent agreement with that in vitro (6.4 ± 0.6 μmol/g, n = 5). When insulin was administered, the stability of glycogen concentration was analogous to previous biochemical measurements implying that glycogen turnover is activated by insulin. We conclude that the entire glycogen molecule is turned over and that insulin activates glycogen turnover.

High-resolution spatial mapping of changes in the neurochemical profile after focal ischemia in mice

After ischemic stroke, the ischemic damage to brain tissue evolves over time and with an uneven spatial distribution. Early irreversible changes occur in the ischemic core, whereas, in the penumbra, which receives more collateral blood flow, the damage is more mild and delayed. A better characterization of the penumbra, irreversibly damaged and healthy tissues is needed to understand the mechanisms involved in tissue death. MRSI is a powerful tool for this task if the scan time can be decreased whilst maintaining high sensitivity. Therefore, we made improvements to a 1H MRSI protocol to study middle cerebral artery occlusion in mice. The spatial distribution of changes in the neurochemical profile was investigated, with an effective spatial resolution of 1.4 μL, applying the protocol on a 14.1‐T magnet. The acquired maps included the difficult‐to‐separate glutamate and glutamine resonances and, to our knowledge, the first mapping of metabolites γ‐aminobutyric acid and glutathione in vivo, within a metabolite measurement time of 45 min. The maps were in excellent agreement with findings from single‐voxel spectroscopy and offer spatial information at a scan time acceptable for most animal models. The metabolites measured differed with respect to the temporal evolution of their concentrations and the localization of these changes. Specifically, lactate and N‐acetylaspartate concentration changes largely overlapped with the T2‐hyperintense region visualized with MRI, whereas changes in cholines and glutathione affected the entire middle cerebral artery territory. Glutamine maps showed elevated levels in the ischemic striatum until 8 h after reperfusion, and until 24 h in cortical tissue, indicating differences in excitotoxic effects and secondary energy failure in these tissue types. Copyright

Deep thiopental anesthesia alters steady-state glucose homeostasis but not the neurochemical profile of rat cortex

Barbiturates are regularly used as an anesthetic for animal experimentation and clinical procedures and are frequently provided with solubilizing compounds, such as ethanol and propylene glycol, which have been reported to affect brain function and, in the case of 1H NMR experiments, originate undesired resonances in spectra affecting the quantification. As an alternative, thiopental can be administrated without any solubilizing agents. The aim of the study was to investigate the effect of deep thiopental anesthesia on the neurochemical profile consisting of 19 metabolites and on glucose transport kinetics in vivo in rat cortex compared with α‐chloralose using localized 1H NMR spectroscopy. Thiopental was devoid of effects on the neurochemical profile, except for the elevated glucose at a given plasma glucose level resulting from thiopental‐induced depression of glucose consumption at isoelectrical condition. Over the entire range of plasma glucose levels, steady‐state glucose concentrations were increased on average by 48% ± 8%, implying that an effect of deep thiopental anesthesia on the transport rate relative to cerebral glucose consumption ratio was increased by 47% ± 8% compared with light α‐chloralose‐anesthetized rats. We conclude that the thiopental‐induced isoelectrical condition in rat cortex significantly affected glucose contents by depressing brain metabolism, which remained substantial at isoelectricity.