Succinate Dehydrogenase, Monoamine Oxidase and Glutamine Synthetase in Developing Human Foetal Brain Regions

In human foetal brain ontogeny the cerebral activity of succinate oxidoreductase (EC 1.3.99.1), i.e. succinate dehydrogenase (SDH), is higher than the cerebellar activity. With rise in foetal body weight the activity in all the brain regions gradually declines. SDH in all the brain regions shows two high-activity periods, one at 20-35 g and another at 110-220 g body weight. The enzyme exhibits a craniocaudal pattern of development. At all times of gestation, L-glutamate:ammonia ligase (EC 6.3.1.2), i.e. glutamine synthetase, activity in the spinal cord and medulla is higher than in the other three regions. At 190 g body weight glutamine synthetase shows an activity peak in all brain regions. Monoamine:oxygen oxidoreductase (EC 1.4.3.4). i.e. monoamine oxidase (MAO), is present much before the onset of electrical activity. It develops caudocranially and exhibits a biphasic pattern of development in all the regions. It increases considerably in the medulla and the spinal core towards late gestational periods.     

Changes in Free Amino Acid Levels in Developing Human Foetal Brain Regions

The levels of free amino acids were determined in human foetal brain regions during prenatal development. Variation in the distribution of amino acids and their rate of change in five segments of the CNS at different stages of ontogeny was observed. Striking developmental changes were found in the levels of aspartic acid in medulla-pons and spinal cord, glycine in the spinal cord, gamma-aminobutyric acid in the cerebral cortex, glutamic acid in the cerebral cortex, midbrain, and spinal cord, and taurine in the medulla-pons and spinal cord. At a late gestational period, glutamic acid was found most abundantly over all the brain regions, whereas the level of taurine was highest at an early gestational stage but not in spinal cord.     

ACTIVITY OF 2'',3''-CYCLIC-NUCLEOTIDE 3''-PHOSPHODIESTERASE IN REGIONS OF RAT BRAIN DURING DEVELOPMENT: QUANTITATIVE RELATIONSHIP TO MYELIN BASIC PROTEIN

2′,3′-Cyclic-nucleotide-3′-phosphodiesterase activity was examined in several regions of rat brain during development, namely optic nerve, olfactory bulb, cerebrum, cerebellum, midbrain, brain stem, and spinal cord. From 4 to 120 days the total activity increased in all regions, although the specific activity approached a constant value in adults. The developmental profile of the enzyme appeared to correlate with the onset of myelination and with the levels of myelin basic protein as well as the appearance of galactocerebroside sulfotransferase. A correlation coefficient of 0.91 was found between total basic protein, expressed as the per cent of the adult (120 day) value, and total enzyme activity over 12–42 days of age (P < 0.001) from six different brain regions as well as for whole brain. By increasing the sensitivity of the assay with the use of [³H-8]adenosine 2′,3′-cyclic monophosphate, we were able to detect activity at birth in both whole brain and spinal cord.     

Reevaluation of Taurine Levels and Distribution of Cysteic Acid Decarboxylase in Developing Human Fetal Brain Regions

The possibilities of interference by glycerophosphoryl ethanolamine (GPE) in the estimation of taurine levels in cerebral cortex, midbrain, cerebellum, medullapons, and spinal cord of developing human fetal brain regions were eliminated by hydrolyzing tissue extracts with 6 M HCl. Cysteic acid thus produced was separated from taurine by ion-exchange chromatography using Biorad-AG resin. Fluorescamine was used as fluorogen. Data reveal that the estimation of taurine in human fetal brain regions is affected if GPE is present as a contaminant in the assay system. Cysteic acid decarboxylase activity was measured using cysteic acid as the substrate. Higher enzymic activity was recorded with increased fetal body weight, but the reverse was true for taurine level.     

Regional Development of Glutamate Dehydrogenase in the at Brain

The development of glutamate dehydrogenase enzyme activity in rat brain regions has been followed from the late foetal stage to the adult and through to the aged (greater than 2 years) adult. In the adult brain the enzyme activity was greatest in the medulla oblongata and pons greater than midbrain = hypothalamus greater than cerebellum = striatum = cortex. In the aged adult brain, glutamate dehydrogenase activity was significantly lower in the medulla oblongata and pons when compared to the 90-day-old adult value, but not in other regions. The enzyme-specific activity of nonsynaptic (free) mitochondria purified from the medulla oblongata and pons of 90-day-old animals was about twice that of mitochondria purified from the striatum and the cortex. The specific activity of the enzyme in synaptic mitochondria purified from the above three brain regions, however, remained almost constant.     

Regional and Subcellular Distribution in Mammalian Brain of the Enzymes Producing Adenosine

The activities of 5'-nucleotidase, 2'-nucleotidase, alkaline phosphatase, and acid phosphatase were measured in rat and autopsied human brains. The four phosphatases in the rat brain showed little change in activity after death. The activities of adenosine-producing enzymes were compared in various parts of rat and human brains. When phosphatase activity was measured at pH 7.5, 5'-nucleotidase showed the highest activity in the most parts of the brain. The activity of 2'-nucleotidase and that of nonspecific phosphatase were almost the same at pH 7.5. However, higher phosphatase activity was observed in all parts of the brain when nonspecific phosphatase activity was measured at pH 10.0 or 5.5. High specific activity of 5'-nucleotidase in the brain was detected in the membranous components, especially in the synaptic membranes. The activity of 2'-nucleotidase was distributed in the soluble and synaptosomal fractions. The highest activity of both alkaline and acid phosphatases was recovered in the crude mitochondrial fraction, with the highest specific activity in the microsomal fraction. Phosphatase activity was distributed widely in the rat brain. The activity of 5'-nucleotidase was high in the medulla oblongata, thalamus, and hippocampus, but low in the peripheral nerve, spinal cord, and occipital lobe. The activity of 2'-nucleotidase was high in the vermis and frontal lobe. The highest acid and alkaline phosphatase activities were detected in the frontal lobe and in the olfactory bulb, respectively. The distribution of the four phosphatases in the autopsied human brain was similar to that in the rat brain. The highest 5'-nucleotidase activity was observed in the temporal lobe and thalamus.(ABSTRACT TRUNCATED AT 250 WORDS)     

S-Adenosyl-l-homocysteine in brain

Administration of methionine sulfoximine (MSO) to rats and mice significantly decreased cerebral levels ofS-adenosyl-l-homocysteine (AdoHcy). Concurrent administration of methionine prevented this decrease and, when methionine was given alone, significantly elevated AdoHcy levels resulted in both species. Regionally, AdoHcy levels varied from 20 nmol/g in rat cerebellum and spinal cord to about 60 nmol/g in hypothalamus and midbrain. MSO decreased AdoHcy in all regions tested except striatum, midbrain, and spinal cord. AdoMet/AdoHcy ratios (methylation index) varied from 0.48 in hypothalamus to 2.4 in cerebellum, and MSO administration decreased these ratios in all regions except hypothalamus. AdoHcy hydrolase activity was lowest in hypothalamus, highest in brainstem and, generally, varied inversely with regional AdoHcy levels. MSO decreased AdoHcy hydrolase activity in all regions except hypothalamus and spinal cord. Cycloleucine administration resulted in significantly decreased levels of mouse brain AdoHcy, whereas the administration of dihydroxyphenylalanine (DOPA) failed to affect AdoHcy levels. It is concluded that (a) cerebral AdoHcy levels are more tightly regulated than are those of AdoMet after MSO administration, (b) slight fluctuations of AdoHcy levels may be important in regulating AdoHcy hydrolase activity and hence AdoHcy catabolism in vivo, (c) the AdoMet/AdoHcy ratio reflects the absolute AdoMet concentration rather than the transmethylation flux, (d) the decreased AdoMet levels in midbrain, cortex, and striatum after MSO with no corresponding decrease in AdoHcy suggest an enhanced AdoMet utilization, hence an increased transmethylation in the MSO preconvulsant state.Supported by USPHS, NINCDS grant NS-06294.     

Lithium: Effect on [3H]spiperone binding,ionic content,and amino acid levels in the brain of rats

After prolonged treatment of rats with lithium (pellets, 0.21% lithium carbonate, or 0.5 mg/ml lithium chloride in drinking water) for three months, the level of lithium in plasma was 0.87 meq/liter; in several brain regions, between 1.06–1.39 eq/g wet weight. The content of sodium and potassium in the plasma was normal. The level of potassium in the brain regions tested increased by 13–30% and that of sodium by about 10%. Glycine levels increased significantly in all the regions (cerebral cortex, midbrain, cerebellum, and spinal cord). In the cerebellum GABA was also increased, while glutamine was decreased. In midbrain, apart from increases in glycine levels, alamine, valine, GABA and lysine were also increased. In the spinal cord, glutamic acid was also increased. Changes were largely in the putative neurotransmitters. Long-term treatment with lithium also influenced the high-affinity binding of [³H]spiperone in the cerebral cortex and corpus striatum. Two specific binding sites were found in both brain regions; the main change was the reduction in the lower affinity binding site (B _max2).     

Regional and Subcellular Distribution of Cyanide Metabolizing Enzymes in the Central Nervous System

Activities of cyanide metabolizing enzymes were measured in various subcellular fractions and regions in the central nervous system. Brain rhodanese and liver beta-mercaptopyruvate sulfurtransferase showed a slight decrease in activity after death. The activity of beta-mercaptopyruvate sulfurtransferase was negligible in the rat brain, compared with that of rhodanese. A small amount of thiocyanate was produced from cyanide and beta-mercaptopyruvate in the human brain, probably due to contamination with red blood cells. Rhodanese activity was widely distributed in all the areas of nervous tissue examined. In the rat the olfactory bulb showed the highest rhodanese activity, and high activity was also observed served in the thalamus, septum, hippocampus, and dorsal part of the midbrain. Rhodanese activity was low in various parts of the cerebral cortex. The distribution pattern of rhodanese in post-mortem human brain was essentially similar to that in rat brain. The thalamus, amygdala, centrum semiovale, colliculus superior, and cerebellar cortex showed high rhodanese activity in the human brain. Rhodanese activity was detected in the spinal cord. Anterior horn showed the highest rhodanese activity in the cervical, thoracic, and lumbar cord. Most rhodanese activity in the rat brain was recovered in the mitochondrial fraction with the highest specific activity. Rhodanese activity was lower in spinal cords obtained from autopsied cases with amyotrophic lateral sclerosis than in those of control subjects. A significant decrease in rhodanese was observed in the posterior column of the cervical or thoracic cord, but the activity in the anterior horn did not differ significantly between the two groups.     

Identification with a Monoclonal Antibody of a Phylogenetically Conserved Brain-Specific Determinant on a 130,000 Molecular Weight Glycoprotein of Human Brain

Human brain glycoproteins depleted of Thy-1 antigen were used to immunise Balb/c mice for monoclonal antibody production. The F3-87-8 antibody described in this paper interacts with a determinant present in large amounts on all human brain subregions studied (cerebral cortical grey matter, white matter, caudate, thalamus, dentate nucleus, putamen, cerebellar cortex) but absent from all other tissues examined (liver, heart, kidney, spleen, thymus, lymph node, erythrocyte, adrenal gland, and peripheral nerve). The determinant is conserved in mammalian evolution, as the brains of the rat and dog have amounts equal to that found in human brain. Balb/c mouse brain has approximately one-third as much antigen activity as these other mammalian brains, whereas brains of the frog and chicken have no detectable antigenic activity. Developmental studies showed that 16-week human foetal brain and neonatal dog brain had little or no antigen activity, indicating a dramatic increase in the amount of the determinant with brain maturation. Biochemical studies showed that the F3-87-8-bearing molecule was a major sialoglycoprotein of human brain with an apparent molecular weight of 130,000. It was shown by immunofluorescence to be particularly localised in what appeared to be fibre tracts in the thalamus and basal ganglia, and in the dentate nucleus, although all regions including grey matter were stained.     

Appearance of a brain specific antigen (th S-100 protein) during human foetal development

—The concentration of a protein specific to brain, the S-100 protein, was measured in various regions of the human foetal brain at gestational ages ranging from 10 weeks until term. The relative increase in concentrations of the S-100 protein during development of the human foetal brain proceeded in a caudal-rostral fashion. This observation is emphasized by the delayed appearance of the S-100 protein in the frontal cerebral cortex until the 30th week of gestation.     

A reassessment of the phosphoglycerate bypass enzymes in human erythrocytes.

Dissociation of the human erythrocyte into cytoplasmic and membranous components, shows that all of the cell's intrinsic 2,3-diphosphoglycerate phosphatase activity is associated with the soluble component. Further fractionaction of the cytoplasm on DEAE cellulose illustrates that both 1,3-diphosphoglycerate mutase and 2,3-diphosphoglycerate phosphatase activities occur coincidently within one peak. Thermal denaturation of the peak proteins at 60° results in a parallel loss in phosphatase and mutase activity. The identical phenomenon is observed in the presence of the 2,3-diphosphoglycerate phosphatase activator, 2-phosphoglycolate. Homogeneous 1,3-diphosphoglycerate mutase, which quantitatively accounts for all of the intrinsic 2,3-diphosphoglycerate phosphatase within the red cell, also exhibits thermal instability at 60°. These findings suggest that the phosphoglycerate bypass in erythrocytes is under the control of a single, bifunctional enzyme.     

Comparative study on the activity, pH optimum and thermal stability of some glycosidases and acid phosphatase from pig and mule leukocytes.

1. alpha-D-mannosidase, beta-D-galactosidase, alpha-L-fucosidase, beta-N-acetylgalactosaminidase, alpha-D-glucosidase and acid phosphatase activities were studied in circulating blood leukocytes from Sus scropha var. domestica L. (pig) and Equus asinus x caballus (mule) by spectrophotometric procedures using p-nitrophenyl derivatives as substrates and three different buffer solutions. 2. The highest specific activity corresponds to acid phosphatase. The specific activities of the glycosidases, all relatively close together in all cases, were low in comparison with that of phosphatase. 3. Generally, each of the above-mentioned enzymes shows one common peak for the pH optimum between 3.5 and 6.0, except alpha-D-glucosidase, which shows two peaks. 4. The pH optima values are generally similar in three buffer solutions employed. 5. Specific activities of the studied enzymes show a parallelism in leukocytes from both pig and mule. Also, this parallelism is observed in their pH optima values. 6. Thermal stability of alpha-D-mannosidase is high whereas that of acid phosphatase is low, in both materials. For other enzymes, differences in the thermal stability was observed according to their source.     

CHANGES OCCURRING IN THE CHEMICAL COMPOSITION OF THE CENTRAL NERVOUS SYSTEM DURING FOETAL AND POST-NATAL DEVELOPMENT OF THE SHEEP

(1) The chemical composition of the CNS (separated into cerebrum, cerebellum, brain stem and spinal cord) was determined in sheep during foetal and post-natal development and in adults. (2) The spinal cord differed from the remainder of the CNS in growing more after the period studied (50-day-old foetuses to 5-week-old lambs) than before it. This was largely attributable to lipid accumulation. (3) Chemical growth (accumulation of DNA, protein and lipid) proceeded linearly in spinal cord, logarithmically in cerebrum and cerebellum while in brain stem growth was described by a sigmoid function. (4) Fat-free dry matter, protein, total lipid, cholesterol and phospholipid concentrations increased progressively in all parts of the CNS but DNA concentrations changed little. In the cerebrum alone there was an increase in DNA concentration during maturation suggesting an increased cell population. Cholesterol was present predominantly in the free form but esters were detected in foetal tissues from 70 up to 120 days gestation. (5) Cerebroside, the characteristic lipid of myelin, increased in concentration soon after 85 days of gestation, up to which point very low values were recorded, the rate varying according to the region of the CNS examined. Rates of increase in total regional cerebroside content were used to identify periods of myelination and the results suggest that there are two periods of peak activity, one about 20 days before birth and the other at 10-20 days after birth. (6) The composition of lipids added during the two phases of myelination and during maturation were characteristically different. In the spinal cord, lipid analyses best reflect changes in myelin composition.     

100Hz电针刺受时大鼠脑和脊髓k受体结构特性的改变

西方采用放射配体结合实验研究了１００ＨＺ电针耐受发生发展过程中大鼠脑和脊髓Ｋ受体结构特性的变化。大鼠每天给予１００ＨＺ电针１次,连续７ｄ。分别在电针的第１、３、５、７天取不同脑区进行观察。     

Distribution of Branch Point Prenyltransferases in Regions of Bovine Brain

Abstract: Bovine brains contain large amounts of isoprenoid compounds and the enzymes involved in their biosynthesis were investigated. Ten different regions were dissected from fresh bovine brains and, in addition, fractions from cerebellum, spinal cord, and hypophysis were obtained. The cholesterol concentration was found to be ∼8 mg/g in the cortex regions and three times higher in the pons, medulla oblongata, and white matter. Dolichol concentration varied between 8 and 40 µg/g in the different tissues, and ubiquinone was found at a lower level, which varied between 3 and 25 µg/g. Farnesyl-pyrophosphate synthase activity in cytosolic fractions from various regions exhibited only a twofold variation, whereas geranylgeranyl pyrophosphate synthase displayed larger differences, being particularly rich in the pons, medulla oblongata, white matter, and spinal cord. Squalene synthase activity was lowest in the thalamus and threefold higher in the pons. Determination of specific activity based on cholesterol content revealed that enzyme activities in various regions are not related to the actual lipid amount present. Both cis - and trans -prenyltransferases exhibited similarities in their regional distribution showing up to 20-fold differences in activity. Thus, it appears that the mevalonate pathway lipids and the various branch point enzymes involved in their syntheses vary greatly in different brain regions and are subjected to separate regulation.     

Human brain natriuretic peptide-like immunoreactivity in human brain.

The presence of immunoreactive human brain natriuretic peptide in the human brain was studied with a specific radioimmunoassay for human brain natriuretic peptide-32. This assay showed no significant cross-reaction with human alpha atrial natriuretic peptide, porcine brain natriuretic peptide or rat brain natriuretic peptide. Immunoreactive human brain natriuretic peptide was found in all 5 regions of human brain examined (cerebral cortex, thalamus, cerebellum, pons and hypothalamus) (0.6-6.7 pmol/g wet weight, n = 3). These values were comparable to the concentrations of immunoreactive alpha atrial natriuretic peptide in human brain (0.5-10.1 pmol/g wet weight). However, Sephadex G-50 column chromatography showed that the immunoreactive human brain natriuretic peptide in the human brain eluted earlier than synthetic human brain natriuretic peptide-32. These findings suggest that human brain natriuretic peptide is present in the human brain mainly as larger molecular weight forms.     

Effects of diethylmaleate on DNA damage and repair in the mouse brain

The enzyme 8-oxoguanine DNA glycosylase 1 participates in the repair of damaged DNA by excising the oxidized base 8-hydroxy-2'-deoxyguanosine. We have previously demonstrated that enzymatic activity of this enzyme is inversely related to the levels of the damaged base in specific brain regions. We now report that the activity of 8-oxoguanine DNA glycosylase 1 is increased in a region-specific manner following treatment with diethylmaleate, a compound that reduces glutathione levels in the cell. A single treatment with diethylmaleate elicited a significant increase ( approximately 2-fold) in the activity of 8-oxoguanine DNA glycosylase 1 in three brain regions with low basal levels of activity (cerebellum, cortex, and pons/medulla). There was no change in the activity of 8-oxoguanine DNA glycosylase 1 in those regions with high basal levels of activity (hippocampus, caudate/putamen, and midbrain). This is the first report to demonstrate that DNA repair capacity can be upregulated in the CNS, and the increased repair activity correlates with a reduction in the levels of DNA damage. The brain region-specific capacity to deal with increased oxidative damage to DNA may be responsible, in part, for the vulnerability of specific neuronal populations with aging, sources of oxidative stress, and neurodegenerative diseases.     

A major phosphotyrosyl-protein phosphatase from bovine heart is associated with a low-molecular-weight acid phosphatase

The phosphotyrosyl [Tyr(P)]-immunoglobulin G (IgG) phosphatase activity in the extracts of bovine heart, bovine brain, human kidney, and rabbit liver can be separated by DEAE-cellulose at neutral pH into two fractions. The unbound fraction exhibits a higher activity at acidic than neutral pH while the reverse is true for the bound fraction. Of all tissues examined, the Tyr(P)-IgG phosphatase activity in the unbound fraction measured at pH 5.0 is higher than that in the bound fraction measured at pH 7.2. The acid Tyr(P)-IgG phosphatase activity has been extensively purified from bovine heart. It copurified with an acid phosphatase activity (p-nitrophenyl phosphate (PNPP) as a substrate) throughout the purification procedure. These two activities coelute from various ion-exchange and gel filtration chromatographies and comigrate on polyacrylamide gel electrophoresis, indicating that they reside on the same protein molecule. The phosphatase has a Mr = 15,000 by gel filtration and exhibits an optimum between pH 5.0 and 6.0 when either Tyr(P)-IgG-casein or PNPP is the substrate. It is highly specific for Tyr(P)-protein with little activities toward phosphoseryl [Ser(P)]- or phosphothreonyl [Thr(P)]-protein. The enzyme activities toward Tyr(P)-casein and PNPP are strongly inhibited by microM molybdate and vanadate but insensitive to inhibition by L(+)-tartrate, NaF, or Zn2+. The molecular and catalytic properties of the acid Tyr(P)-protein phosphatase purified from bovine heart are very similar to those of the low-molecular-weight acid phosphatases of Mr = 14,000 previously identified and purified from the cytosolic fraction of human liver, placenta, and other animal tissues.     

Co-identity of brain angiotensin converting enzyme with a membrane bound dipeptidyl carboxypeptidase inactivating Met - enkephalin.

The distribution in rat brain of angiotensin converting enzyme (EC3.4.15.1) using hippuryl-His-Leu as substrate was identical to a dipeptidyl carboxypeptidase present in membranes assayed with Met-enkephalin as substrate. Highest activity occurred in pituitary, followed by cerebellum, corpus striatum, midbrain, pons-medulla, hypothalamus, cerebral cortex and spinal cord. The ratio of products His-Leu/Tyr-Gly-Gly was identical for all regions but differed from His-Leu/Tyr. Angiotensin converting enzyme purified by immunoaffinity chromatography gave a K_m for hippuryl-His-Leu of 0.5mM and for Met-enkephalin of 0.1 mM. In the presence of the specific inhibitor of angiotensin converting enzyme, SQ 14,225, the Ki value was 10^?7M. Present data point to the co-identity of brain angiotensin converting enzyme with the dipeptidyl carboxypeptidase inactivating enkephalin.