3-METHOXY-4-HYDROXYPHENYLETHANOL IN THE RAT BRAIN

Abstract— The neutral dopamine metabolite, 3-methoxy4hydroxypenylethanol (MOPET) can be measured in the rat brain by a GLC method using a pentaflouropropionic derivative and electron capture detector. The identity of MOPET is verified by mass spectrographic analyses.
The endogenous level of MOPET of 16.6 ng/g whole rat brain can be raised more than four-fold by intraperitoneal injection of L-DOPA, dopamine or MOPET. In contrast intraventricular injection of dopamine or intraperitoneal injection of L-DOPA plus a peripheral decarboxylase inhibitor (Ro 4-4602)., results in small and insignificant increase bf MOPET in the CNS.
It is concluded that MOPET is probably of low significance to central dopamine metabolism and that MOPET found in the rat brain is predominantly of peripheral origin.     

Determination of kynurenine by a simple gas—liquid chromatographic method applicable to urine,plasma, brain and cerebrospinal fluid

A simple, sensitive and specific method for the determination of kynurenine is described. This is based on alkaline cleavage of kynurenine, followed by solvent extraction, trifluoroacetylation and gas—liquid chromatography with electron capture detection. Using this method kynurenine has been determined in urine and plasma, and for the first time in brain and cerebrospinal fluid. Increases in kynurenine in brain, plasma and urine are demonstrated following tryptophan administration to man and rat.     

FINE STRUCTURE OF THE SURFACE OF THE CEREBRAL CORTEX OF HUMAN BRAIN

Evidence is presented for the existence of arborizing cytoplasmic processes extending from the surface of the cerebral cortex of human brain into the surrounding fluid medium. These originate from subpial fibrous astrocytes and contain the usual cytoplasmic organelles of those cells. They are bordered by basement membrane. Their occurrence is localized and variable over the cortical surface. They are more prevalent in pathological human material than in "normal" human brain and somewhat more prevalent in the latter than in normal rat cortex. Some additional information is presented regarding the relationship of leptomeninges to the cortical surface. The pia mater does not invariably adhere inseparably to the subjacent layer of fibrous astrocytes as generally assumed at present, nor does it always form a continuous layer over the surface of the brain in the material under study. Both collagen and cytoplasmic extensions of astrocytes intervene between these layers. These findings imply that glial elements of the cortex have direct access to the cerebrospinal fluid.     

Urinary Biomarkers of Brain Diseases

Biomarkers are the measurable changes associated with a physiological or pathophysiological process. Unlike blood, urine is not subject to homeostatic mechanisms. Therefore, greater fluctuations could occur in urine than in blood, better reflecting the changes in human body. The roadmap of urine biomarker era was proposed. Although urine analysis has been attempted for clinical diagnosis, and urine has been monitored during the progression of many diseases, particularly urinary system diseases, whether urine can reflect brain disease status remains uncertain. As some biomarkers of brain diseases can be detected in the body fluids such as cerebrospinal fluid and blood,there is a possibility that urine also contain biomarkers of brain diseases. This review summarizes the clues of brain diseases reflected in the urine proteome and metabolome.     

IDENTIFICATION AND QUANTIFICATION OF 3-METHOXY -4-HYDROXYPHENYLLACTIC ACID (VLA) IN CEREBROSPINAL FLUID AND 3-METHOXY-4-HYDROXY PHENYLPYRUVIC ACID (VPA) IN THE URINE OF PARKINSONIAN PATIENTS TREATED WITH L-DOPA

The identification and quantification of 3-methoxy-4-hydroxyphenyllactic acid in cerebrospinal fluid, and 3-methoxy-4-hydroxyphenylpyruvic acid in the urine of Parkinsonian patients on medication with l -3,4-dihydroxyphenylalanine with or without a peripheral decarboxylase inhibitor are described. Data are also given for other catecholamine- and catecholamine precursor metabolites in urine and cerebrospinal fluid. Analyses were performed by means of gas chromatography with flame ionization or mass spectrometric detection. We have concluded that following the administration of l -3.4-dihydroxyphenylalanine + peripheral decarboxylase inhibitor. 3-methoxy-4-hydroxyphenyllactic acid must be considered as a quantitatively important metabolite of L-3.4-dihydroxyphenylalanine in cerebrospinal fluid. In addition, some comment is given on the presence of 3-methoxy-4-hydroxy-phenylpyruvic acid in the urine of these patients.     

大鼠第三脑室及中脑水管多巴胺能触液神经元的分布

目的观察大鼠第三脑室、中脑水管及中缝背核内多巴胺能触液神经元的分布情况.方法应用CB逆行追踪、TH免疫组织化学和CB/TH免疫荧光双重标记技术,观察多巴胺能触液神经元在间脑及中脑内的分布情况.结果 TH免疫阳性触液神经元分布在第三脑室尾侧部和中脑水管全程的腹侧室管膜上及室管膜内,其胞体呈倒置梨形、圆形或椭圆形、多角形和梭形;在中缝背核内可见少量CB/TH免疫荧光双重标记的远位触液神经元;另在正中隆起部位TH免疫阳性神经末梢含量丰富.结论大鼠第三脑室、中脑水管及中缝背核内存在多巴胺能触液神经元,其在脑-脑脊液之间的信息传递中有着重要的作用.     

The fetal rat binding protein for insulin-like growth factors is expressed in the choroid plexus and cerebrospinal fluid of adult rats

The biological effects of the insulin-like growth factors, IGF-I and IGF-II, on their receptors are modulated by IGF-binding proteins. Recently, we isolated a cDNA clone for one member of the family of IGF-binding proteins, BP-3A, a 30 kilodalton (kDa) protein synthesized by the BRL-3A rat liver cell line. BP-3A is related to but distinct from two other cloned IGF-binding proteins, the human amniotic fluid binding protein and the glycosylated binding subunit of the 150 kDa IGF-binding protein complex in serum. It is expressed in multiple nonneural tissues and in serum in the fetal rat and decreases after birth, similar to the developmental pattern of IGF-II expression. IGF-I, IGF-II, and their receptors are expressed in brain. The present study examines the expression of BP-3A in the rat central nervous system. By Northern blot analysis, BP-3A mRNA is present at high levels in brain stem, cerebral cortex, and hypothalamus from 21-day gestation rats and, like IGF-II mRNA, persists in adult rat brain. The site of BP-3A mRNA synthesis was localized by in situ hybridization to coronal sections of adult rat brain using 35S-labeled oligonucleotides, 48 bases in length, complementary and anticomplementary to the coding region of BP-3A. Specific hybridization of the BP-3A probe was observed exclusively to the choroid plexus extending from the level of the medial preoptic nucleus to the arcuate nucleus of the hypothalamus, similar to the previously reported preferential localization of IGF-II mRNA to the choroid plexus. Synthesis of BP-3A mRNA by choroid plexus suggested that BP-3A might be secreted into the cerebrospinal fluid. A 30 kDa IGF-binding protein was demonstrated in rat cerebrospinal fluid that is recognized by antibodies to BP-3A and, like purified BP-3A, has equal affinity for IGF-I and IGF-II. By analogy with other transport proteins synthesized by the choroid plexus, BP-3A may facilitate the secretion of IGF-II to the cerebrospinal fluid and modulate its biological actions at distant sites within the brain.     

Origin of α-mannosidase activity in CSF

The α-mannosidase activity in human frontal gyrus, cerebrospinal fluid and plasma has been analyzed by DEAE-cellulose chromatography to investigate the origin of the α-mannosidase activity in cerebrospinal fluid (CSF). The profile of α-mannosidase isoenzymes obtained in CSF was similar to that in the frontal gyrus but different from that in human plasma. In particular the two characteristic peaks of lysosomal α-mannosidase, A and B, which have a pH-optimum of 4.5 and are found in human tissues, were present in both the frontal gyrus and CSF. In contrast the majority of α-mannosidase activity in human plasma was due to the so called intermediate form, which has a pH-optimum of 5.5. The results suggest that the intermediate form of α-mannosidase in plasma does not cross the blood–brain barrier and that the α-mannosidase activity present in the cerebrospinal fluid is of lysosomal type and of brain origin. Thus the α-mannosidase activity in cerebrospinal fluid might mirror the brain pathological changes linked to neurodegenerative disorders such as Parkinson's disease.     

ON THE ELIMINATION OF CENTRALLY FORMED 5-HYDROXYINDOLEACETIC ACID BY CEREBROSPINAL FLUID AND URINE

Abstract— In rats, the release of centrally formed 5-hydroxyindoleacetic acid (5-HIAA) into brain and spinal cord perfusates and urine was measured. Data from spinal cord perfusion of anaesthetized rats indicate that more than about 36% of the spinal production (122ng/h) of 5-HIAA is eliminated via the cerebrospinal fluid (CSF). More than 30% of cerebrally formed 5-HIAA (265.0 ng/h) was calculated to be released into ventricular-cisternal perfusates. Of the total amount of 5-HIAA found in the urine we estimated that about 8% originates in the central nervous system (CNS).
In probenecid treated animals there was a substantial increase in the outflow of 5-HIAA in both perfusion systems. In the combined perfusion experiments no proportional increase of the cerebral contribution to the cisternal outflow was found after probenecid. Our data indicate that a significant proportion of centrally formed 5-HIAA is eliminated by the CSF. No evidence was found for an increased contribution of cerebral 5-HIAA to lumbar CSF after application of the probenecid test. Urinary levels of 5-HIAA do not reflect quantitatively central 5-hydroxytryptamine metabolism.     

MAXIMUM ACTIVITIES, PROPERTIES AND DISTRIBUTION OF 5''NUCLEOTIDASE, ADENOSINE KINASE AND ADENOSINE DEAMINASE IN RAT AND HUMAN BRAIN

Abstract— The maximum activities of 5'nucleotidase, adenosine kinase and adenosine deaminase have been measured in several areas of rat and human brain. There is no major difference between the activities of nucleotidase and kinase between rat and human brain, but the activity of deaminase is considerably higher in human brain. The activities of all these enzymes are similar in three areas of rat brain and nine areas of human brain, except for hind brain of the human, which has a low activity of adenosine deaminase. This variation may indicate the existence of different steady-state concentrations of adenosine in certain areas of the brain.
Subcellular fractionation of different areas of rat brain showed that, whereas adenosine kinase and deaminase activities were located mainly in the soluble fractions, 5'nucleotidase was present in all subcellular fractions (i.e. membrane, synaptosomal, mitochondrial and soluble). In particular, there was no major localisation within the synaptosomal fraction. Thus it is unlikely that the regulation of the activities of these enzymes is dependent upon changes within a specific compartment (e.g. synaptosomes) in the brain.     

CONTROL OF PYRUVATE AND β-HYDROXYBUTYRATE UTILIZATION IN RAT BRAIN MITOCHONDRIA AND ITS RELEVANCE TO PHENYLKETONURIA AND MAPLE SYRUP URINE DISEASE

—The effects of the amino acids (phenylalanine, valine, leucine and isoleucine) which accumulate in phenylketonuria (PKU) and maple syrup urine disease (MSUD), and their analogue α-keto acids (phenylpyruvate, α-keto isovalerate, α-keto isocaproate, α-keto-β-Me valerate) have been studied on rat brain mitochondrial respiration. Both phenylpyruvate and α-keto isocaproate specifically inhibited the oxidation of pyruvate plus malate and β-hydroxybutyrate plus malate by rat brain mitochondria in the presence of ADP. However, no inhibitory effects of similar concentrations of phenylpyruvate or α-keto isocaproate were observed on the isolated semipurified pyruvate or β-hydroxybutyrate dehydrogenases from rat brain mitochondria. The transport of pyruvate and β-hydroxybutyrate across the brain mitochondrial membrane was studied by both uptake and exchange of radioactively labelled substrates. Both these processes were inhibited by phenylpyruvate and α-ketoisocaproate. The results are interpreted as providing evidence for both pyruvate and β-hydroxybutyrate translocases across the brain mitochondrial membrane, and that the inhibition of these systems by phenylpyruvate and α-keto isocaproate may be important lesions in phenylketonuria and maple syrup urine disease respectively.     

Effect of fractions of the cerebrospinal fluid from patients with drug dependence on basic electrophysiological characteristics of the rat olfactory cortex slices

Heroin addicts at the initial stage of abstinence syndrome were subjected to detoxication by liquorosorption technique. The fractions of their cerebrospinal fluid obtained by the thin layer chromatography technique were analyzed. The substances extracted from the cerebrospinal fluid of drug addicts, presumably peptides, negatively affected the conductive function and synaptic transmission in surviving slices of the olfactory cortex of rats. The conclusion was drawn about a possibility of application of surviving rat brain slices as test object for estimation of the extent of purification of the cerebrospinal fluid from toxic endogenous substances after the liquorosorption.     

FREE AMINO ACID LEVELS IN THE CEREBROSPINAL FLUID OF NORMAL HUMANS AND THEIR VARIATION IN CASES OF EPILEPSY AND SPIELMEYER-VOGT-BATTEN DISEASE

Abstract— The concentrations of free amino acids in the plasma and cerebrospinal fluid from control subjects and from patients suffering from epilepsy and Spielmeyer-Vogt-Batten disease were determined using an automatic amino acid analyser. It was found that the plasma/cerebrospinal fluid ratio of amino acid concentrations showed the variation in the amounts of free amino acids in epilepsy more clearly than the cerebrospinal fluid levels alone.     

DEVELOPMENTAL PROFILES OF GANGLIOSIDES IN HUMAN AND RAT BRAIN

Abstract— The developmental profiles of individual gangliosides of human brain were compared with those of rat brain. Interest was focused mainly on the pre- and early postnatal development. Human frontal lobe cortex covering the period from 10 foetal weeks to adult age and the cerebrum of rat from birth to 21 days were analysed. Lipid-NANA and lipid-P were followed; in the rat, also protein and brain weight. A limited number of samples of human cerebral white matter and cerebellar cortex were also studied. The following major results were obtained:

• 1 The ganglioside concentration increased approximately three-fold within a short period: in rat cerebrum, from birth to the 17th day; in human cerebral cortex, from the 15th foetal week to the age of about 6 months. The largest increase in the rat brain occurred by the 11th to the 13th day; in human brain by term. The relative increase of gangliosides during this period was more rapid than that of phospholipids.
• 2 A hitherto unknown distinct early period of ganglioside and phospholipid formation in rat occurred by the second to fourth day.
• 3 The changes in brain ganglioside pattern, characteristic of the developmental stages of the rat, were found to be equally pronounced in the human brain.
• 4 Regional developmental differences in the ganglioside pattern were demonstrated in human brain. A characteristic white matter pattern, rich in monosialogangliosides, had developed by the age of 1 year. The increase in ganglioside concentration and the formation of the definitive ganglioside pattern of cerebellar cortex occurred later than in cerebral cortex. This cerebellar pattern was characterized by a very large trisialoganglioside fraction.
• 5 The two periods of rapid ganglioside metabolism in rat brain preceded the two periods of rapid protein biosynthesis.

     

Regional Distribution of Neurotensin in Human Brain

Abstract: Neurotensin (NT) is an endogenous neuropeptide that is active in many preclinical screening tests for neuroleptic drugs. Using a radioimmunoassay, we have studied the regional distribution of NT in postmortem human brain and in cerebrospinal fluid. Highest levels were present in the hypothalamus, substantia nigra, and limbic areas, whereas much lower amounts were found in the cortex and striatum. The chromatographic properties of hypothalamic immunoreactivity on ion-exchange and high pressure liquid chromatography were similar to those of the synthetic tridecapeptide. We conclude that NT is present in human brain with a distribution resembling that seen in other species, such as rat and monkey.     

Pressure distribution in the cranial cavity

Values of interstitial fluid pressure in surface layers of the human brain and of the pressure of the cerebrospinal fluid "bathing" the brain are presented. The results show a difference in these pressures. The analysis with a spherical model shows that intratissue structural (biomechanical) connections, and those between the brain, its membranes and cranium are extremely important in significantly weakening the transmission of the cerebrospinal fluid pressure to the cortex and surface layers of the brain.     

Conjugated 3,4 dihydroxy phenyl acetic acid (DOPAC) in human and monkey cerebrospinal fluid and rat brain and the effects of probenecid treatment

Gas chromatography-mass spectrometry (GC-MS) was used to measure 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in cerebrospinal fluid from humans and monkeys and in rat caudate nuclei. DOPAC was found to be present mainly in conjugated form. In human lumbar CSF the average concentration of total DOPAC before probenecid treatment was 1.48 ± 0.31 ng/ml; after probenecid it increased to 15.06 ± 3.17 ng/ml. This increase was mainly due to conjugated DOPAC but increases in free DOPAC also occurred. There is a relatively greater accumulation of DOPAC than of HVA, suggesting that in human CSF conjugated DOPAC may have a faster turnover rate than HVA. In monkey, ventricular CSF contained higher concentrations of DOPAC and HVA than did lumbar CSF.In rat brain, treatment with probenecid caused increases in DOPAC, HVA and their conjugates.These results suggest that DOPAC is conjugated in brain and that both compounds are removed from brain and CSF by a probenecid-sensitive acid transport system in the same manner as is HVA.     

大鼠脑内远位触液神经元p38丝裂原活化蛋白激酶的分布及在噪声应激时的表达

目的：观察脑内远位触液神经元内p-p38丝裂原活化蛋白激酶（MAPK）的分布及其在噪声应激时的表达。方法：用霍乱毒素亚单位B与辣根过氧化物酶复合物（CB-HRP）标记和免疫组织化学相结合的双重标记技术．观察SD大鼠脑实质内远位触液神经元中p-p38MAPK的分布：进一步制作噪声应激动物模型,观察噪声应激后该类神经元中p-p38MAPK的表达变化。结果：在脑干的特定部位恒定出现被CB-HRP标记的两组神经细胞簇,其他脑区未见CB-HRP标记神经细胞簇。不予应激刺激,该细胞簇内仅有个别神经元见有CB-HRP／p—p38MAPK;噪声应激刺激1d时,上述特定部位细胞簇的CB-HRP／p-p38MAPK双重标记神经元数目没有明显变化;噪音应激刺激5d时,CB-HRP／p—p38MAPK双重标记神经元数目较对照组显著增多（P〈0．05）;噪音应激刺激10d时CB-HRP／p—p38MAPK双重标记神经元数目较对照组显著增多（P〈0．05）;噪音应激刺激20d时,CB-HRP／p—p38MAPK双重标记神经元数目较对照组显著增多（P〈0．01）：结论：在脑干特定部位恒定存在的两组被CBHRP标记的细胞团为远位触液神经元,其中少数触液神经元有p-p38MAPK表达,且当给予动物噪声应激刺激时,p-p38MAPK免疫阳性神经元和CB-HRP／p—p38MAPK双重标记神经元数量显著增加,提示脑实质内的这种远位触液神经元中的P—p38MAPK可能参与了机体对噪声应激的信息传递或调控,其作用随应激天数增加而日趋增强．     

Aldehyde dehydrogenase activity in the barrier brain structures

The histochemical method was used to study the aldehyde dehydrogenase (EC 1.2.1.3.; ALDH) activity in capillaries and glial structures of different regions in the rat central nervous system (CNS). The occurrence of three metabolic barriers for aldehydes on systemic level in the CNS has been shown. They are: the barrier between blood and the nervous tissue (represented by capillary endothelium and surrounding astrocytes ALDH), that between blood and cerebrospinal fluid (ALDH in ependymocytes of vascular plexus), and that between cerebrospinal fluid and nervous tissue (ALDH of ependymocytes covering brain cavities). On the single microregions level a similar barrier is between interstitial fluid and neurons (ALDH of satellite oligodendrocytes).