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Voxel placement for MRS based on anatomy. The high resolution T 2 -weighted fast spin-echo image in axial projection (slice thickness 1 mm) is used as a reference for voxels placement. Hippocampus (left column) and anterior cingulate cortex (right column) voxels positioning in young (PND14, top row) and adult (PND60, bottom raw) rats. The brain levels approximately correspond to levels 7–12 (1.45–3.6 mm anterior of bregma) fo anterior cingulate cortex and levels 27–38 (2.0–5.65 mm posterior of bregma) of the Swanson's anatomical rat brain atlas (Swanson, 2004).
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We utilized proton magnetic resonance spectroscopy to evaluate the metabolic profile of the hippocampus and anterior cingulate cortex of the developing rat brain from postnatal days 14 to 70. Measured metabolite concentrations were modelled using linear, exponential, or logarithmic functions and the time point at which the data reached plateau (i.e...
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Context 1
... in some of these brain metabolites exist even in the same rat species from different breeders and underscores the importance of using appropriate controls in such studies. Voxel placement was particularly difficult to reproduce in the ACC even within this study because of the subjectivity of the effort necessitated by a lack of obvious landmarks (Fig. ...
Context 2
... 2 ). Rat core temperature was maintained throughout the scan at 37.1 70.6°C using warmed circulating water in the animal bed (Bruker BioSpin, Billerica, MA). The spectroscopic voxels were positioned at the left dorsal hippocampus (HC, 4 Â 4 Â 2 mm) and anterior cingulate cortex (ACC, 2.5 Â 2.5 Â 2.5 mm) using a fast spin echo image as a reference (Fig. 1). Voxels were carefully positioned using Swanson's rat brain anatomical atlas as a reference (Swanson, 2004) at approximate levels 7-12 (1.45-3.6 mm anterior of bregma) for ACC, and levels 27-38 (2.00-5.65 mm posterior of bregma) for HC. The magnetic field homogeneity in these voxels was adjusted using FASTMAP to yield a water line ...
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Objective
Despite the serious neurodevelopmental sequelae of epileptic encephalopathy during infancy, the pathomechanisms involved remain unclear. To find potential biomarkers that can reflect the pathogenesis of epileptic encephalopathy, we explored the neurometabolic and microstructural sequelae after infantile spasms using a rat model of infanti...
Citations
... A repeated measures ANOVA revealed the positive effect of the time factor on average T 2 values in all brain regions and the volume of the whole brain, ventricles, and CSF across all subjects. Rats included in this study may still be undergoing developmental changes despite being considered to become mature at approximately 60 days (Sengupta, 2013;Ramu et al., 2016b). However, there is no firm consensus on the age at which rats are considered to be fully developed (Spear, 2000). ...
The assessment of the sensitivity and specificity of any potential biomarker against the gold standard is an important step in the process of its qualification by regulatory authorities. Such qualification is an important step towards incorporating the biomarker into the panel of tools available for drug development. In the current study we analyzed the sensitivity and specificity of T2 MRI relaxometry to detect trimethyltin-induced neurotoxicity in rats. Seventy-five male Sprague-Dawley rats were injected with a single intraperitoneal dose of either TMT (8, 10, 11, or 12 mg/kg) or saline (2 ml/kg) and imaged with 7 T MRI before and 3, 7, 14, and 21 days after injection using a quantitative T2 mapping. Neurohistopathology (the gold standard in the case of neurotoxicity) was performed at the end of the observation and used as an outcome qualifier in receiver-operator characteristic (ROC) curve analysis of T2 changes as a predictor of neurotoxicity. TMT treatment led to a significant increase in T2 values in many brain areas. The biggest changes in T2 values were seen around the lateral ventricles, which was interpreted as ventricular dilation. The area under the ROC curve for the volume of the lateral ventricles was 0.878 with the optimal sensitivity/specificity of 0.805/0.933, respectively. T2 MRI is a promising method for generating a non-invasive biomarkers of neurotoxicity, which shows the dose-response behavior with substantial sensitivity and specificity. While its performance was strong in the TMT model, further characterization of the sensitivity and specificity of T2 MRI with other neurotoxicants is warranted.
... It has been shown that in such stage the cerebral water content drops by about 8-10% [35]. Interestingly, in the studies, where longitudinal brain metabolomics is investigated with the use of MRS, the water content change correction is frequent practice [35][36][37], but some authors explicitly assume that after six weeks of the birth the water content is not changing in healthy rat brain [37], or do not arise this aspect [38]. Based on our findings on R 1 changes, a second, corrected analysis of MRS data was performed by normalizing the metabolites' concentrations to the measured R 1 to diminish the impact of differences in hippocampal water level. ...
... The assumption that water level is constant might lead to uncertainties when comparing metabolites between healthy and diseased cases. Complicating the issue, in longitudinal studies the water content changes due to brain maturation and aging has to be taken into account, what was shown in many MRS studies of brain development [35][36][37]. This aspect, however, is usually not taken into account in MRS studies of depressive disorders. ...
The intestinal microbiome composition and dietary supplementation with psychobiotics can result in neurochemical alterations in the brain, which are possible due to the presence of the brain–gut–microbiome axis. In the present study, magnetic resonance spectroscopy (MRS) and behavioural testing were used to evaluate whether treatment with Lacticaseibacillus rhamnosus JB-1 (JB‑1) bacteria alters brain metabolites’ levels and behaviour during continuous exposure to chronic stress. Twenty Wistar rats were subjected to eight weeks of a chronic unpredictable mild stress protocol. Simultaneously, half of them were fed with JB-1 bacteria, and the second half was given a daily placebo. Animals were examined at three-time points: before starting the stress protocol and after five and eight weeks of stress onset. In the elevated plus maze behavioural test the placebo group displayed increased anxiety expressed by almost complete avoidance of exploration, while the JB-1 dietary supplementation mitigated anxiety which resulted in a longer exploration time. Hippocampal MRS measurements demonstrated a significant decrease in glutamine + glutathione concentration in the placebo group compared to the JB-1 bacteria-supplemented group after five weeks of stress. With the progression of stress the decrease of glutamate, glutathione, taurine, and macromolecular concentrations were observed in the placebo group as compared to baseline. The level of brain metabolites in the JB-1-supplemented rats were stable throughout the experiment, with only the taurine level decreasing between weeks five and eight of stress. These data indicated that the JB-1 bacteria diet might stabilize levels of stress-related neurometabolites in rat brain and could prevent the development of anxiety/depressive-like behaviour.
... 11 Animal and adult human studies have characterized the temporal trajectories of brain metabolites, their regional and sex-based variations, and the influence of mode of delivery; early postnatal studies, however, are currently limited. [12][13][14][15][16] Conventional neonatal 1 H-MR spectroscopy allows measurement of NAA and Cr (markers of neuronal integrity and metabolic activity), Cho (lipid membrane component), and mIns (cytoplasmic osmotic agent). Neonatal brain injury has been associated with lower NAA, 17,18 and simultaneous GABA and glutamate measurements would allow in vivo neurotransmitter-specific interrogation of the developing brain. ...
... We also hypothesized that NAA, Cr, and Glx would increase with advancing age (postmenstrual age [PMA]), whereas Cho (and GABA1) would remain relatively stable. 10,16,19,20 We sought to investigate the influence of postnatal age (weeks of life), sex, and mode of delivery on the neurometabolic profile of the neonatal brain. ...
... 8 We demonstrated that all metabolites except GABA1 positively correlated with advancing PMA, which likely reflects increasing metabolic activity of maturing neurons, dendrites, synapses, and glial cells (Fig 3). The stable GABA1 concentrations observed are consistent with those in previous animal 12, 16 and clinical studies in preterm infants. 7,19 Consistent with previous studies, 16,19 Cr concentrations increased significantly with PMA, whereas Cho remained relatively stable; hence, metabolite/Cho ratios are preferable markers of metabolic changes during early postnatal life. ...
Background and purpose:
Gamma-aminobutyric acid and glutamate system disruptions may underlie neonatal brain injury. However, in vivo investigations are challenged by the need for special 1H-MR spectroscopy sequences for the reliable measurement of the neurotransmitters in this population. We used J-edited 1H-MR spectroscopy (Mescher-Garwood point-resolved spectroscopy) to quantify regional in vivo gamma-aminobutyric acid and glutamate concentrations during the early postnatal period in healthy neonates.
Materials and methods:
We prospectively enrolled healthy neonates and acquired Mescher-Garwood point-resolved spectroscopy spectra on a 3T MR imaging scanner from voxels located in the cerebellum, the right basal ganglia, and the right frontal lobe. CSF-corrected metabolite concentrations were compared for regional variations and cross-sectional temporal trends with advancing age.
Results:
Fifty-eight neonates with acceptable spectra acquired at postmenstrual age of 39.1 (SD, 1.3) weeks were included for analysis. Gamma-aminobutyric acid (+ macromolecule) (2.56 [SD, 0.1]) i.u., glutamate (3.80 [SD, 0.2]), Cho, and mIns concentrations were highest in the cerebellum, whereas NAA (6.72 [SD, 0.2]), NAA/Cho, Cr/Cho, and Glx/Cho were highest in the basal ganglia. Frontal gamma-aminobutyric acid (1.63 [SD, 0.1]), Glx (4.33 [SD, 0.3]), Cr (3.64 [SD, 0.2]), and Cho concentrations were the lowest among the ROIs. Glx, NAA, and Cr demonstrated a significant adjusted increase with postmenstrual age (β = 0.2-0.35), whereas gamma-aminobutyric acid and Cho did not.
Conclusions:
We report normative regional variations and temporal trends of in vivo gamma-aminobutyric acid and glutamate concentrations reflecting the functional and maturational status of 3 distinct brain regions of the neonate. These measures will serve as important normative values to allow early detection of subtle neurometabolic alterations in high-risk neonates.
... 56 The changes in brain metabolites become relatively small after PND 35. 57 In our study, the mice were late adolescents (PND [28][29][30][31][32][33][34][35] and therefore the findings may be analogous to adult mice. ...
The study of cerebral metabolites relies heavily on detection methods and sample preparation. Animal experiments in vivo require anesthetic agents that can alter brain metabolism, whereas ex vivo experiments demand appropriate fixation methods to preserve the tissue from rapid postmortem degradation. In this study, the metabolic profiles of mouse hippocampi using proton magnetic resonance spectroscopy (¹H‐MRS) were compared in vivo and in situ with or without focused beam microwave irradiation (FBMI) fixation. Ten major brain metabolites, including lactate (Lac), N‐acetylaspartate (NAA), total choline (tCho), myo‐inositol (mIns), glutamine (Gln), glutamate (Glu), aminobutyric acid (GABA), glutathione (GSH), total creatine (tCr) and taurine (Tau), were analyzed using LCModel. After FBMI fixation, the concentrations of Lac, tCho and mIns were comparable with those obtained in vivo under isoflurane, whereas other metabolites were significantly lower. Except for a decrease in NAA and an increase in Tau, all the other metabolites remained stable over 41 hours in FBMI‐fixed brains. Without FBMI, the concentrations of mIns (before 2 hours), tCho and GABA were close to those measured in vivo. However, higher Lac (P < .01) and lower NAA, Gln, Glu, GSH, tCr and Tau were observed (P < .01). NAA, Gln, Glu, GSH, tCr and Tau exhibited good temporal stability for at least 20 hours in the unfixed brain, whereas a linear increase of tCho, mIns and GABA was observed. Possible mechanisms of postmortem degradation are discussed. Our results indicate that a proper fixation method is required for in situ detection depending on the targeted metabolites of specific interests in the brain.
... GABA concentrations in the developing brain are inherently low (1-2 µmol/g) 19,28,29 and its spectroscopic signals are overlapped by more dominant metabolites in the standard 1 H-MRS spectra 15 . To enhance spectral resolution, animal studies have utilized ultra-high strength magnetic field (7-9 T) scanners 30,31 , while adult human studies on 3 T scanners have used alternative techniques including short-echo time and/or spectral editing techniques like MEGA-PRESS 14,[32][33][34] . Additional challenges in the preterm population including smaller brain volumes limiting voxel size, motion artifacts on non-sedated research scans, and dependence on intensive care during the early postnatal period have further limited in vivo www.nature.com/scientificreports ...
Gamma-aminobutyric acid (GABA) and glutamate are principal neurotransmitters essential for late gestational brain development and may play an important role in prematurity-related brain injury. In vivo investigation of GABA in the preterm infant with standard proton magnetic resonance spectroscopy (¹H-MRS) has been limited due to its low concentrations in the developing brain, and overlap in the spectrum by other dominant metabolites. We describe early postnatal profiles of in vivo GABA and glutamate concentrations in the developing preterm brain measured by using the J-difference editing technique, Mescher-Garwood point resolved spectroscopy. We prospectively enrolled very preterm infants born ≤32 weeks gestational age and non-sedated ¹H-MRS (echo time 68 ms, relaxation time 2000 ms, 256 signal averages) was acquired on a 3 Tesla magnetic resonance imaging scanner from a right frontal lobe voxel. Concentrations of GABA + (with macromolecules) was measured from the J-difference spectra; whereas glutamate and composite glutamate + glutamine (Glx) were measured from the unedited (OFF) spectra and reported in institutional units. We acquired 42 reliable spectra from 38 preterm infants without structural brain injury [median gestational age at birth of 28.0 (IQR 26.0, 28.9) weeks; 19 males (50%)] at a median postmenstrual age of 38.4 (range 33.4 to 46.4) weeks. With advancing post-menstrual age, the concentrations of glutamate OFF increased significantly, adjusted for co-variates (generalized estimating equation β = 0.22, p = 0.02). Advancing postnatal weeks of life at the time of imaging positively correlated with GABA + (β = 0.06, p = 0.02), glutamate OFF (β = 0.11, p = 0.02) and Glx OFF (β = 0.12, p = 0.04). Male infants had higher GABA + (1.66 ± 0.07 vs. 1.33 ± 0.11, p = 0.01) concentrations compared with female infants. For the first time, we report the early ex-utero developmental profile of in vivo GABA and glutamate stratified by age and sex in the developing brain of very preterm infants. This data may provide novel insights into the pathophysiology of neurodevelopmental disabilities reported in preterm infants even in the absence of structural brain injury.
... The levels of metabolites observed in the hippocampus are in agreement with previous works which analyzed the rat hippocampus during early infant development (from p7 to p28) (Tkáč et al., 2003) or across the entire developmental period (from p10 to p70) (Ramu et al., 2016). To date, no one has investigated on the difference in the metabolism during the specific transitions (the juvenile and the adolescence one) occurring in this period. ...
Adolescence is an age of transition when most brain structures undergo drastic modifications, becoming progressively more interconnected and undergoing several changes from a metabolic and structural viewpoint. In the present study, three MR techniques are used in rats to investigate how metabolites, structures and patterns of connectivity do change. We focused in particular on areas belonging to the limbic system, across three post-weaning developmental stages: from “early” (PND 21–25) to “mid” (i.e., a juvenile transition, PND 28–32) and then to “late” (i.e., the adolescent transition, PND 35–39). The rs-fMRI data, with comparison between early and mid (juvenile transition) age-stage rats, highlights patterns of enhanced connectivity from both Striata to both Hippocampi and from there to (left-sided) Nucleus accumbens (NAcc) and Orbitofrontal Cortex (OFC). Also, during this week there is a maturation of pathways from right Striatum to ipsilateral NAcc, from right OFC to ipsilateral NAcc and vice versa, from left Prefrontal Cortex to ipsilateral OFC and eventually from left Striatum, NAcc and Prefrontal Cortex to contralateral OFC. After only 1 week, in late age-stage rats entering into adolescence, the first pathway mentioned above keeps on growing while other patterns appear: both NAcc are reached from contralateral Striatum, right Hippocampus from both Amygdalae, and left NAcc -further- from right Hippocampus. It's interesting to notice the fact that, independently from the age when these connections develop, Striata of both hemispheres send axons to both Hippocampi and both NAcc sides, both Hippocampi reach left NAcc and OFC and finally both NAcc sides reach right OFC. Intriguingly, the Striatum only indirectly reaches the OFC by passing through Hippocampus and NAcc. Data obtained with DTI highlight how adolescents' neurite density may be affected within sub-cortical gray matter, especially for NAcc and OFC at “late” age-stage (adolescence). Finally, levels of metabolites were investigated by 1H-MRS in the anterior part of the hippocampus: we put into evidence an increase in myo-inositol during juvenile transition and a taurine reduction plus a total choline increase during adolescent transition. In this paper, the aforementioned pattern guides the formulation of hypotheses concerning the correlation between the establishment of novel brain connections and the emergence of behavioral traits that are typical of adolescence.
... Metabolic changes of the developing brain have been investigated non-invasively by in vivo MR spectroscopy in animal models and also in humans [15][16][17][18][19][20][21]. For ex vivo applications, nuclear magnetic resonance (NMR) spectroscopy has been established in recent years, besides mass spectrometry, as important tool for metabolic profiling in pathology and toxicology [22,23]. ...
... Metabolic changes of the developing brain have also been investigated non-invasively by in vivo MR spectroscopy in animal models and also in humans [15][16][17][18][19][20][21]. In vivo NMR measurements commonly report clear NAA and Cre increases during brain development, which is in line with our findings in the 3D rat brain cell aggregates. ...
... However, mIno was found to increase during rat brain development in another study [21] in accordance with our results. A strong decrease was noted for taurine in the developing rat brain [16], in line with our finding of a strong decrease following a very early rise. Developmental differences between studies were attributed to differences in animal species or to regional differences in the brain [16]. ...
The aim of the present study was to establish the developmental profile of metabolic changes of 3D aggregating brain cell cultures by 1H high-resolution magic angle spinning (HR-MAS) NMR spectroscopy. The histotypic 3D brain aggregate, containing all brain cell types, is an excellent model for mechanistic studies including OMICS analysis; however, their metabolic profile has not been yet fully investigated. Chemometric analysis revealed a clear separation of samples from the different maturation time points. Metabolite concentration evolutions could be followed and revealed strong and various metabolic alterations. The strong metabolite evolution emphasizes the brain modeling complexity during maturation, possibly reflecting physiological processes of brain tissue development. The small observed intra- and inter-experimental variabilities show the robustness of the combination of 1H-HR-MAS NMR and 3D brain aggregates, making it useful to investigate mechanisms of toxicity that will ultimately contribute to improve predictive neurotoxicology.
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Metabolites play important roles in brain development and their levels change rapidly in the prenatal period and during infancy. Metabolite levels are thought to stabilize during childhood, but the development of neurochemistry across early-middle childhood remains understudied. We examined the developmental changes of key metabolites (total N-acetylaspartate, tNAA; total choline, tCho; total creatine, tCr; glutamate+glutamine, Glx; and myo-inositol, mI) using short echo-time magnetic resonance spectroscopy (MRS) in the anterior cingulate cortex (ACC) and the left temporo-parietal cortex (LTP) using a mixed cross-sectional/longitudinal design in children aged 2-11 years (ACC: N=101 children, 112 observations; LTP: N=95 children, 318 observations). We found age-related effects for all metabolites. tNAA increased with age in both regions, while tCho decreased with age in both regions. tCr increased with age in the LTP only, and mI decreased with age in the ACC only. Glx did not show linear age effects in either region, but a follow-up analysis in only participants with ≥3 datapoints in the LTP revealed a quadratic effect of age following an inverted U-shape. These substantial changes in neurochemistry throughout childhood likely underlie various processes of structural and functional brain development.
Gamma-aminobutyric acid (GABA) and glutamatergic system perturbations following premature birth may explain neurodevelopmental deficits in the absence of structural brain injury. Using GABA-edited spectroscopy (MEscher-GArwood Point Resolved Spectroscopy [MEGA-PRESS] on 3 T MRI), we have described in-vivo brain GABA+ (+macromolecules) and Glx (glutamate + glutamine) concentrations in term-born infants. We report previously unavailable comparative data on in-vivo GABA+ and Glx concentrations in the cerebellum, the right basal ganglia, and the right frontal lobe of preterm-born infants without structural brain injury. Seventy-five preterm-born (gestational age 27.8 ± 2.9 weeks) and 48 term-born (39.6 ± 0.9 weeks) infants yielded reliable MEGA-PRESS spectra acquired at post-menstrual age (PMA) of 40.2 ± 2.3 and 43.0 ± 2 weeks, respectively. GABA+ (median 2.44 institutional units [i.u.]) concentrations were highest in the cerebellum and Glx higher in the cerebellum (5.73 i.u.) and basal ganglia (5.16 i.u.), with lowest concentrations in the frontal lobe. Metabolite concentrations correlated positively with advancing PMA and postnatal age at MRI (Spearman's rho 0.2-0.6). Basal ganglia Glx and NAA, and frontal GABA+ and NAA concentrations were lower in preterm compared with term infants. Moderate preterm infants had lower metabolite concentrations than term and extreme preterm infants. Our findings emphasize the impact of premature extra-uterine stimuli on GABA-glutamate system development and may serve as early biomarkers of neurodevelopmental deficits.
Cognitive and behavioral disabilities in preterm infants, even without obvious brain injury on conventional neuroimaging, underscores a critical need to identify the subtle underlying microstructural and biochemical derangements. The gamma-aminobutyric acid (GABA) and glutamatergic neurotransmitter systems undergo rapid maturation during the crucial late gestation and early postnatal life, and are at-risk of disruption after preterm birth. Animal and human autopsy studies provide the bulk of current understanding since non-invasive specialized proton magnetic resonance spectroscopy (¹H-MRS) to measure GABA and glutamate are not routinely available for this vulnerable population due to logistical and technical challenges. We review the specialized ¹H-MRS techniques including MEscher-GArwood Point Resolved Spectroscopy (MEGA-PRESS), special challenges and considerations needed for interpretation of acquired data from the developing brain of preterm infants. We summarize the limited in-vivo preterm data, highlighted the gaps in knowledge and future directions for optimal integration of available in-vivo approaches to understand the intricate role of GABA and glutamate on neurodevelopmental outcomes after preterm birth.
