Emilio Bossi

Brain metabolite composition during early human brain development as measured by quantitative in vivo 1H magnetic resonance spectroscopy

Biochemical maturation of the brain can be studied noninvasively by 1H magnetic resonance spectroscopy (MRS) in human infants. Detailed time courses of cerebral tissue contents are known for the most abundant metabolites only, and whether or not premature birth affects biochemical maturation of the brain is disputed. Hence, the last trimester of gestation was observed in infants born prematurely, and their cerebral metabolite contents at birth and at expected term were compared with those of fullterm infants. Successful quantitative short‐TE 1H MRS was performed in three cerebral locations in 21 infants in 28 sessions (gestational age 32–43 weeks). The spectra were analyzed with linear combination model fitting, considerably extending the range of observable metabolites to include acetate, alanine, aspartate, cholines, creatines, γ‐aminobutyrate, glucose, glutamine, glutamate, glutathione, glycine, lactate, myo‐inositol, macromolecular contributions, N‐acetylaspartate, N‐acetylaspartylglutamate, o‐phosphoethanolamine, scyllo‐inositol, taurine, and threonine. Significant effects of age and location were found for many metabolites, including the previously observed neuronal maturation reflected by an increase in N‐acetylaspartate. Absolute brain metabolite content in premature infants at term was not considerably different from that in fullterm infants, indicating that prematurity did not affect biochemical brain maturation substantially in the studied population, which did not include infants of extremely low birthweight. Magn Reson Med 48:949–958, 2002.

Structural and neurobehavioral delay in postnatal brain development of preterm infants.

Postnatal brain development of healthy prematurely born infants was assessed to study possible influence of premature birth and early extrauterine environment on structural, biochemical, and functional brain development. Myelination and differentiation of gray and white matter were studied byin vivo magnetic resonance (MR) imaging (MRI), changes in cerebral metabolism by 1H MR spectroscopy (MRS), and changes in early human neurobehavior by the assessment of preterm infants behavior (APIB). The stage of intrauterine and extrauterine brain development in prematurely born infants at term was compared with the stage of mainly intrauterine brain development in a group of full-term infants. Eighteen preterm infants unremarkable with respect to neurologic and medical status were studied at approximately 2 wk of postnatal age [gestational age (GA) 1: 32.5 ± 1.2 wk] and again at term(GA 2: 40.0 ± 1.1 wk). For comparison a group of 13 full-term born infants (GA T: 40.6 ± 2.1 wk) were studied by MR and six by APIB. When GA 2 to GA 1 was compared, significant maturational changes were found with MRI in gray and white matter and myelination, with 1H MRS in the concentration of N-acetylaspartate and with all scores of APIB. In preterm infants at term (GA 2) compared with full-term infants (GA T) significantly less gray and white matter differentiation and myelination was observed as well as significantly poorer performance in four neurobehavioral parameters (autonomic reactivity, motoric reactivity, state organization, attentional availability). We conclude that MRI and 1H MRS can be used to study postnatal brain development in preterm infants. Structural and biochemical maturation is accompanied by functional maturation as shown with the neurobehavior assessment. Preterm infants at term compared with full-term infants show a structural as well as a functional delay in brain development assessed at 40 wk of postconceptional age.

Magnetic resonance in preterm and term newborns: 1H-spectroscopy in developing human brain.

ABSTRACT: Localized proton magnetic resonance spectra were recorded from human cerebellum in vivo with a 1.5-T magnet. The spectra from healthy adults and preterm and term babies showed resonances from N-acetylaspartate, creatine and phosphocreatine, choline-containing compounds such as phosphocholine and glycerophosphocholine, taurine, and inositol. The age-dependent changes of in vivo molar concentrations of N-acetylaspartate, choline, taurine, and inositol were estimated in preterm babies, babies at term, and adults. The range of postconceptional age in the studied babies was 31 to 45 wk. Taking the biochemically measured creatine concentrations in age-corresponding autopsy material as an internal standard, the in vivo concentrations of the other metabolites were calculated from the proton spectra. N-acetylaspartate showed an increase from 1.9 mM in preterm babies to 3.1 mM in term babies and to 6.5 mM in adult brain. Taurine was noted to increase from 1.1 mM in preterm infants to 2.3 mM in term infants and did not decrease significantly in adult brain. Choline and inositol concentrations did not change significantly throughout the studied age groups. These new data on in vivo, localized 1H-spectroscopy show that it is a sensitive method for studying early metabolic brain development in humans.

Regional Metabolic Assessment of Human Brain during Development by Proton Magnetic Resonance Spectroscopy In Vivo and by High-Performance Liquid Chromatography/Gas Chromatography in Autopsy Tissue

To study the course of regional metabolite concentrations during early brain development, we measured in vivo metabolites [N-acetyl-aspartate (NAA), choline-containing compounds, and myo-inositol (M-Ino)] in the precentral area of the cerebrum by short echo-time single volume proton magnetic resonance spectroscopy and compared in vivo established spectroscopic data with classic chromatographic data (HPLC) on age-corresponding autopsy tissue in different regions of the brain. In autopsy tissue, regional (frontal lobe, precentral area, basal ganglia, thalamus) and age-dependent differences of the concentration of creatine, NAA, and M-Ino were determined. In vivo measurement of NAA by proton magnetic resonance spectroscopy shows a significant increase of NAA by increasing postconceptional age. M-Ino shows a weak correlation and a nonsignificant decrease with increasing postconceptional age. Choline shows no age-dependent changes. Creatine concentrations measured by HPLC in different regions of the developing brain at autopsy showed an age-dependent increase that was identical for the left and right side and similar for the precentral area and frontal lobe and more pronounced for the basal ganglia and thalamus. Comparison of the results obtained by the two methods shows agreement for the age-dependent changes and the absolute concentration of M-Ino. NAA determined in autopsy tissue by HPLC is significantly lower than that measured in vivo by proton magnetic resonance spectroscopy. A comparison of the concentrations measured by HPLC in frontal lobe, basal ganglia, and thalamus with the results obtained from the precentral area showed significant regional differences in all measured metabolites. These results define important age-dependent changes detected with both methods and further indicate limitations of both methods that have to be considered when presenting absolute concentration values.

Sources of blood glucose in the rat fetus.

Extract: Pregnant Sprague-Dawley rats from the 18th to the 21st day of gestation were fasted for 16 hr and anesthetized, and a constant infusion of uniformly labeled glucose-14C and of glucose-2-3H was started. The purpose was to see if the regulation of fetal blood glucose concentration differed from the maternal during fasting. The results are given in Figures 2 and 3. During the course of the infusions, the specific activity of glucose was, at all points measured, two to four times lower in the fetal than in the maternal serum. The higher fetal than maternal supply of blood glucose could be derived either from gluconeogenesis or from glycogenolysis in the fetus.In vitro studies indicate that the fetal rat liver is unable to perform gluconecgenesis. Therefore, the placenta was investigated for the presence of a gluconeogenetic pathway. Placenta slices showed no ability to convert pyruvate into glucose (Fig. 8). No phosphoenolpyruvate carboxykinase (EC. 4.1.1.32) activity could be found in placenta homogenates.The possibility of the presence of gluconeogenesis in the rat fetus has been investigated in vivo. Intraperitoneal injections of pyruvate-2-14C and glycerol-1, 3-14C were given to 1-day-old newborns, prematures (fetuses which were delivered 1 day prior to term), and fetuses in situ. The 1-day-old newborns do convert both substrates to glucose, with averages of 50 and 70%, respectively, of serum radioactivity being due to glucose-14C, thus proving the validity of this way of investigating gluconeogenesis in vivo. The prematures did not synthesize glucose from pyruvate, but glucose synthesis occurred from glycerol, an average of 70% of serum radioactivity being due to glucose-14C. The fetuses which were injected in situ did not synthesize glucose from either one of the two substrates. These experiments would seem to exclude fetal gluconeogenesis as the source of fetal blood glucose in the constant infusion experiments mentioned above.Fetal hepatic glycogenolysis can account only in part for the higher dilution of the specific activity of glucose in the fetus after a 16-hr fast. The data in Table I show that at the time when the constant infusions were started (after a 16-hr fast) the fetal liver glycogen concentration had diminished from 111 ± 5 to 82 ± 10 mg/g wet weight, as compared to the fed state. This decrease is too small to account for the higher fetal than maternal dilution of blood glucose specific activity, or for the maintenance of unchanged blood glucose levels in the fetus after a 16-hr fast (86 ± 8 and 92 ± 11 mg/100 ml, respectively).An unknown mechanism leading to the privileged situation of the fetus with regard to blood glucose levels during a fasting state must be postulated.Speculation: Gluconeogenesis is inactive in the rat fetus, as investigated by in vivo and in vitro methods. Fetal hepatic glycogenolysis cannot be the only source of fetal blood glucose during fasting.

Utilization of D-β-Hydroxybutyrate and Oleate as Alternate Energy Fuels in Brain Cell Cultures of Newborn Mice after Hypoxia at Different Glucose Concentrations

ABSTRACT: In dissociated whole brain cell cultures from newborn mice, we have previously shown that during glucose deprivation under normoxia, D-β-hydroxybutyrate and oleic acid are increasingly used for energy production. We now asked whether this glucose dependency of the utilization of D-β-hydroxybutyrate and oleic acid as alternate energy fuels is also present after a hypoxic phase. 3- Hydroxy[3-14C]butyrate or [U-14C]oleic acid were added to 7- and 14-d-old cultures and 14CO2-production compared after hypoxia in normal and glucose-deprived conditions. After hypoxia, the ability of the cells 7 d in culture to increase D-β-hydroxybutyrate consumption in response to glucose deprivation is diminished, 14-d-old cells lose this ability. In contrast, after hypoxia, both 7- and 14-d-old cultures maintain or even improve the ability to increase oleate consumption, when glucose is lacking.

TAURINE CONCENTRATIONS IN BRAIN OF PRETERM AND TERM NEWBORNS MEASURED BY IN VIVO 1 H-MRS AND CHROMATOGRAPHY IN AUTOPSY TISSUE

Our study presents data on in vivo measurements of regional Tau concentrations in different stages of early human brain development. We used the method of short echotime 1H-spectroscopy in single volumes of 3.3cm3 for the in vivo assessment. 27 preterm (≥37wk) and 22 term newborns(≥38wk), all fed maternal milk, were examined by 1H-MRS either in cerebellum or cerebrum. In comparison age-corresponding autopsy tissue was examined by high performance liquid chromatography (HPLC).Concentrations of Tau measured in vivo by 1H-MRS and in autopsy tissue by HPLC are comparable. In cerebellum Tau concentration increases from preterm to term newborns.Cerebellar Tau concentrations measured in autopsy tissue of term newborns are significantly higher than cerebral Tau concentrations (p<0.05). The same tendency is observed in the in vivo 1H-MRS data.

46 BRAIN DEVELOPMENT IN PRETERM INFANTS studied by MRI, MRS, APIB, Dubowitz scale and BAEP

We designed a longitudinal study to investigate brain maturation in preterm infants, using the Assessment of Preterm Infants Behavior(APIB), MRI(GW;M), MRS(NAA/Cho), Dubowitz scale(D) for neurologic assessment and brainstem auditory evoked potentials(BAEP:V). Age-dependent maturational changes are presented in the first six patients, studied at a mean gestational age(GA) of 32.7±1.4 weeks and again at due date(mean GA:40.1±1.3w).Conclusion: First results indicate, that grey-while matter differentiation and myelination in MRI clearly discriminates maturational stages in brain development followed by MRS, behavioral studies assessed in autonomic-, motoric organization, self-regulation and examiner facilitation, neurologic assessment and BAEP.

176 LUNG MECHANICS AND GAS EXAWNCE DURING ARTIFICIAL SUPFACTANT |[lpar]|EXOSURE=E|[rpar]| ADMINISTRATION FOR HYALINE MEMBRANE DISEASE |[lpar]|HMD|[rpar]|

Studies with animal surfactant preparations have shown dramatic improvements in oxygenation without concomitant changes in dynamic compliance. The present study deals with the effects of a purely synthetic surfactant (E) upon arterial blood gases (ABG) and compliance (Crs)/resistance (Rrs), determined by the recently introduced single breath occlusion technique. Six premature babies with HMD, enrolled in the European OSIRIS trial were studied. Crs/Rrs and ABG were determined 30 min. before and after E administration and repeated at 24 h intervals.Results: Pre- and post values (means) are given for the 1st and 2nd dose. Measurements 3 and 4 are follow-ups. *p<0.05 vs. previous value.Conclusion: E fails to induce an acute amelioration of lung mechanics and gas exchange, later improvements in oxygenation parallel those seen in Crs. The lack of immediate effect of E may be due to the absence of surfactant-associated proteins in this preparation.

47 ABNORMAL LONG FUNCTION 8 YEARS AFTER VENTILATION FOR HYALINE MEMBRANE DISEASE (HMD) WITHOUT BRONCHOPULMONARY DYSPLASIA (BPD)

Ventilation for HMD can lead to BPD resulting in long lasting functional lung impairment. Whether this also occurs when BPD does not develop, is not known. Therefore, we tested lung function in 18 children (median age 7 7/12 years) who had been ventilated for HMD without BPD, in 20 non-ventilated siblings (9 10/12) and in 20 controls matched for gestational age, birth weight, and sex (6 2/12).Method: Lung function was measured by whole-body plethysmography. Airway reactivity was assessed by carbachol provocation. Abnormal lung function was defined when hyperinflation (TGV>130 % predicted) and/or small airway obstruction (maximal expiratory flow at 50 % VC<80 %) was present.Results: 10/18 ventilated children, 1/20 siblings (p<0.001) and 3/20 matched controls (p vs. ventilated<0.01) have abnormal lung function. The occurence of airway hyperreactivity was not different between the groups (5/18, 4/20 and 6/20).Conclusion: Ventilation for HMD not resulting in BPD is associated with hyperinflation and/or collapse of the small bronchioli at a median age of 7 7/12 years. The abnormal lung function is probably not due to genetic predisposition (sig. less abnormal lung function in siblings, no difference between siblings and controls, no difference in airway hyperreactivity) nor to prematurity itself (sig. less abnormalities in gestational age- and birth weight-matched controls).