bluuebunny/crossref_metadata_2025 huggingface dataset:
In Vivo Biosynthesis of 35S‐Substance P from [35S]Methionine in the Rat Striatum and Its Transport to the Substantia Nigra
Authors: Torrens Y. , Michelot R. , Beaujouan J. C. , Glowinski J. , Bockaert J.
Published: October 2006

Abstract: Rats with a push‐pull cannula implanted in the right striatum were used to study the biosynthesis of 35S‐substance P (SP) from [35S]methionine and its transport to the ipsilateral substantia nigra. [35S]Methionine was delivered for 2, 3 or 5 h to the push‐pull cannula. 35S‐SP in striatal and nigral tissues was estimated after immunoadsorption and HPLC. Higher levels of 35‐SP in striatal homogenates were found after a 5‐h labelling period. 35S‐SP biosynthesis was inhibited when cycloheximide was superfused together with [35S]methionine. The identity of 35S‐SP was further checked by its conversion into 35S‐SP sulphoxide. After a 5‐h labelling period, 35S‐SP was also recovered in the substantia nigra. This was not the case after hemisection of striato‐nigral fibers. When rats were killed 15 or 24 h after the 5‐h labelling period, 35S‐SP levels in the substantia nigra were higher than those found just after the 5‐h labelling period, while the reverse was observed in the striatum.
Calmodulin Affinity for Brain Coated Vesicle Proteins
Authors: Moskowitz Nathan , Schook William , Lisanti Michael , Hua Elaine , Puszkin Saul
Published: October 2006

Abstract: A systematic characterization of the affinity of calmodulin for brain coated vesicles was undertaken. Binding of [125I‐labeled calmodulin to coated vesicles was saturable and competed with unlabeled calmodulin, but not with troponin‐C. Scatchard analysis revealed one high‐affinity, low‐capacity binding site, KD= 3.9 ± 0.6 nM, Bmax= 16.3 ± 2.4 pmol/mg, and one low‐affinity, high‐capacity binding site, KD= 102 ± 15.0 nM, Bmax= 151 ± 23.0 pmol/mg. Radioimmunoassay revealed that coated vesicles contain 1.05 μg calmodulin/mg protein. Because this value remained constant even after removal of clathrin, the major coat protein, from the coated vesicle, it is apparent that calmodulin is associated with the vesicle per se rather than with its clathrin lattice. When a Triton X‐100‐treated extract of coated vesicles was passed through a Sepharose 4B‐calmodulin affinity column, polypeptides with Mrs (molecular weights) of 100,000, 55,000, and 30,000 bound in a Ca2+‐dependent manner. A 30,000 Mr, protein doublet purified from coated vesicles was completely eluted by EGTA from the calmodulin affinity column, confirming that this protein doublet represents one of the coated vesicle calmodulin binding sites. Because calmodulin stimulated [Ca2+‐Mg2+]‐ATPase activity as well as Ca2+ uptake in coated vesicles, it is postulated that the 100,000 and 55,000 Mr calmodulin binding proteins represent the [Ca+2‐Mg2+]‐ATPase complex, the other coated vesicle calmodulin binding site.
Cysteine Sulfinic Acid in the Central Nervous System: Uptake and Release of Cysteine Sulfinic Acid by a Rat Brain Preparation
Authors: Iwata Heitaroh , Yamagami Satoru , Mizuo Hiroyuki , Baba Akemichi
Published: October 2006

Abstract: Uptake and release of cysteine sulfinic acid by synaptosomal fractions (P2) and slices of rat cerebral cortex were investigated. The P2 fraction had a Na+‐dependent high‐affinity uptake system for cysteine sulfinic acid (Km, 12μM), which was restricted to the synaptosomes. High‐affinity uptake of cysteine sulfinic acid was competitively inhibited by glutamate, aspartate, and cysteic acid. None of the various centrally acting drugs tested specifically inhibited this transport system. Release of [14C]cysteine sulfinic acid from preloaded cortical slices or P2 fractions was examined by a superfusion method, which avoided reuptake of released [14C]cysteine sulfinic acid. High K+ (56 mM) and veratridine (10μM) stimulated the release of cysteine sulfinic acid from slices and the P2 fraction in a partly Ca2+‐dependent manner. Diazepam at concentrations of 10 and 100 μM markedly inhibited the stimulated release, but not the spontaneous release, by cortical slices. On the contrary, it had no effect on the stimulated release of cysteine sulfinic acid from the P2 fraction.
Polypeptide Elongation Factors of the Developing Chick Brain
Authors: Murakami K. , Miyamoto K.
Published: October 2006

Abstract: The polypeptide elongation factors (EF‐1L, EF‐1H, and EF‐2) of the developing chick brain were separated and purified by means of a combination of gel chromatographic methods. The molecular weight of EF‐1H of the chick brain ranged from 5 to 10 × 105, and was different from that of the chick liver (about 7 × 105). The molecular weight of other purified factors was about 5 × 104 for EF‐1L. and 9.4 × 104 for EF‐2. High activities of polyphenylalanine (poly‐Phe) synthesis per mg protein in the developing chick brain were observed between the 3rd embryonic week and the 1st post‐hatch week and declined afterwards. On the other hand, the levels of both EF‐1 and EF‐2 per mg protein in the brain were observed to be high in an early embryonic stage, gradually declining afterwards to the adult level. The brain EF‐1L was a major component of EF‐1 in an early embryonic stage, while EF‐1H became recognizable in the 3rd embryonic week. Moreover, the EF‐1H activities were found to be more than double with regard to the binding reaction and to be more than 10‐fold as active in respect to poly‐Phe synthesis in comparison with the activities of EF‐1L. It is proposed that the brain EF‐1H could be due to aggregates consisting of EF‐1L, a stimulatory factor, and other components.
Regional Amino Acid Distribution in Relation to Function in Insulin Hypoglycaemia
Authors: Butterworth Roger F. , Merkel Andrea D. , Landreville France
Published: October 2006

Abstract: The amino acids glutamate, aspartate, γ‐aminobutyric acid (GABA), and glutamine were measured as their dansyl derivatives in whole brain and specific brain regions by a sensitive double‐labelling technique at various times during the development of hypoglycaemic encephalopathy. Hypoglycaemia was induced by administration of insulin (100 i.u./kg) to 24‐h fasted rats. No significant changes in glutamate, GABA, or glutamine were detected in whole brain at any time up to and including the onset of hypoglycaemic convulsions. In cerebral cortex, however, GABA levels were reduced to 65% of normal prior to the appearance of neurological symptoms of hypoglycaemia. Onset of symptoms (severe catalepsy and loss of righting reflex, but before the onset of convulsions) was accompanied by marked decreases of glutamate and glutamine in striatum and hippocampus. These regions, in addition to cerebral cortex, show the greatest vulnerability to hypoglycaemic insult, according to previous anatomical studies. Aspartate levels were significantly increased (p < 0.01) in the cerebral cortex of convulsing animals, confirming a previous report. No changes were detectable in any of the amino acids studied in medulla‐pons at any time during the progression of hypoglycaemia. Cerebral cortex and striatum showed a selective net loss of amino acids (2.2 and 3.5 μmol/g, respectively) prior to the onset of insulin‐hypoglycaemic convulsions.
Soluble Spiroperidol Binding Factors from Bovine Caudate Nucleus
Authors: Winkler M. H. , Berl S.
Published: October 2006

Abstract: Several properties of soluble spiroperidol binding factors separated from bovine caudate nucleus have been investigated by a previously unreported procedure. Data consistent with high particle weight and rapid binding equilibration are reported for high‐affinity (+)butaclamol‐sensitive components of a digitonin extract. A slower sedimenting component is found that also exhibits high affinity for spiroperidol but is not sensitive to (+)butaclamol. Centrifugation of a caudate nucleus homogenate yields a supernatant that appears to contain a component that exhibits spiroperidol binding that is more sensitive to displacement by (‐) than by (+)butaclamol. The procedure used effects rapid separation of bound from unbound tritiated ligand on short columns of Sephadex G‐15 followed by extrusion and sectioning of the Sephadex. The radioactivity remaining with each section is determined. The procedure is very rapid; the addition of active phases or the changing of the ionic environment, which may disturb the equilibrium, is avoided; and recovery of the protein free of bound ligand is easily affected.
4‐Aminobutyraldehyde Dehydrogenase Activity in Rat Brain
Authors: Tago Kozue , Kurioka Susumu , Matsuda Makoto
Published: October 2006

Abstract: An enzyme with NAD+‐dependent 4‐aminobutyraldehyde dehydrogenase activity was purified about 360‐fold from rat brain extract. AMP‐Sepharose chromatography was effective in separating the enzyme from other NAD+‐dependent aldehyde dehydrogenases included in the extract. The Kms for the substrates NAD+ and 4‐aminobutyraldehyde were 4.8 × 10−4 and 8.3 × 10−5M, respectively. The pH optimum for the enzyme was about 8.0. The ratio of activities toward 4‐aminobutyraldehyde, propionaldehyde, succinate semialdehyde, and benzaldehyde was 1.00:0.17:0.24:0.09:0.03 when the activity toward 4‐aminobutyraldehyde was set equal to 1.00. The enzyme activity in subcellular fractions of rat brain was localized in cytosol.
Composition and Metabolism of Gangliosides in Rat Peripheral Nervous System During Development
Authors: Chou K. H. , Nolan C. E. , Jungalwala F. B.
Published: October 2006

Abstract: Ganglioside composition of rat trigeminal nerve was studied during development in order to understand the changes that occur as a result of cellular differentiation in the nerve. The ganglioside composition of the trigeminal nerve was entirely different from that of brain. The major gangliosides in adult trigeminal nerve were GM3, GD3, and LM1 (sialosyl‐lactoneotetraosylceramide or sialosylparagloboside). The structure of LM1 and other gangliosides was established by enzymatic degradation and by analysis of the products of acid hydrolysis. At 2 days after birth, when the Schwann cells were immature, GM3 and GD3 were the major gangliosides in the nerve, 50 and 18 mol %, respectively. As the nerve developed and Schwann cells proliferated and myelinated the axons, the mol % of GM3 and GD3 reduced and that of LM1 steadily increased. Polysialogangliosides did not change drastically with nerve development. The rate of deposition of LM1 in the nerve with age was very similar to that of myelin marker lipids, cerebrosides, and sulfatides; thus, deposition appears to be localized mainly in the rat nerve myelin. LM1 also had long‐chain fatty acids 22:0 and 24:0, which are not usually found in CNS gangliosides. The ganglioside pattern of the rat trigeminal nerve was very similar to that of rat sciatic nerve, but was different from that of rabbit and chicken sciatic nerve. The activity of the two key enzymes involved in the metabolism of GM3, viz., CMP‐N‐acetylneuraminic acid:lactosylceramide sialyltransferase and UDP‐N‐acetylgalactosamine:GM3‐N‐acetylgalactosaminyltransferase, was also studied during development of the nerve and brain. The developmental profiles of both enzymes were consistent with the amounts of GM3 present in the nerve.
Peptide Mapping Studies of the Chromogranins and of Two Chromaffin Granule Proteoglycans
Authors: Banerjee Sila , Margolis Richard U.
Published: October 2006

Abstract: The chromogranins, a family of related acidic glycoproteins, and two chondroitin sulfate/dermatan sulfate proteoglycans were isolated from the soluble contents of bovine adrenal chromaffin granules by chromatography on DEAE‐cellulose. These chromaffin granule matrix glycoconjugates were treated with trypsin, and the resulting peptides were fractionated by HPLC. The two proteoglycans, which differ in their concentration of glycosaminogly‐cans and glycoprotein oligosaccharides, yielded almost identical peptide patterns and would both appear to have the same protein moiety. The peptide profile of the proteoglycans differs, however, from that of the chromogranins, which they closely resemble in terms of amino acid composition. The various chromogranin fractions obtained by gel filtration were also found to have significant differences in the chromatographic patterns of their tryptic peptides.
Combined Effects of a Metabolic Inhibitor (Gabaculine) and an Uptake Inhibitor (Ketamine) on the γ‐Aminobutyrate System in Mouse Brain
Authors: Wood J. D. , Geddes J. W. , Tsui S.‐K. , Kurylo E.
Published: October 2006

Abstract: The intramuscular administration of a γ‐aminobutyrate‐α‐oxoglutarate aminotransferase (GABA‐T) inhibitor, gabaculine, to mice resulted in significant increases in GABA content and decreases in the content of aspartate, glutamate, and glutamine in the nerve endings (synaptosomes). These effects were ameliorated by the concurrent administration of the GABA uptake inhibitor ketamine. A major cause of these effects was the gabaculine‐induced inhibition of GABA‐T activity and the lessening of this inhibition by ketamine. The latter phenomenon was not due to a direct action of ketamine on the enzyme, nor to an interaction between gabaculine and ketamine. Rather, it appeared that ketamine might be interfering with the transport of gabaculine into the cellular structures. The anticonvulsant action of the GABA‐T inhibitor and the GABA uptake inhibitor together was little different from that of the GABA‐T inhibitor alone.
Comparison of the Kinetics of [3H]Diazepam and [3H]Flunitrazepam Binding to Cortical Synaptosomal Membranes
Authors: Chiu Ted H. , Rosenberg Howard C.
Published: October 2006

Abstract: [3H]Diazepam and [3H]flunitrazepam ([3H]FNP) binding to washed and frozen synaptosomal membranes from rat cerebral cortex were compared. In Tris‐citrate buffer, γ‐aminobutyric acid (GABA) and NaCl both increased [3H]diazepam binding more than [3H]FNP binding. GABA and pentobarbital both enhanced this effect of NaCl. Because of the extremely rapid dissociation of [3H]diazepam in the absence of NaCl and GABA, the Bmax (maximal binding capacity) was smaller by the filtration assay than by the centrifugation assay. [3H]FNP, which dissociates more slowly, had the same Bmax in both assays. [3H]Diazepam association had two components, and was faster than [3H]FNP association. [3H]Diazepam dissociation, which also had two components, was faster than that of [3H]FNP, and also had a greater fraction of rapidly dissociating species. [3H]FNP dissociation was similar when initiated by diazepam, flunitrazepam, clonazepam, or Ro15‐1788, which is a benzodiazepine antagonist. [3H]Diazepam dissociation with Ro15‐1788, flunitrazepam, or clonazepam was slower than with diazepam. GABA and NaCl, but not pentobarbital, increased the percentage of slowly dissociating species. This effect of NaCl was potentiated by GABA and pentobarbital. The results support the cyclic model of benzodiazepine receptors existing in two interconvertible conformations, and suggest that, distinct from their binding affinity, some ligands (like flunitrazepam) are better than others (like diazepam) in inducing the conversion of the receptor to the higher‐affinity state.
Comparative Studies of Fresh and Postmortem Isolated Rat Cortical Synaptosomes: Morphology, K+ Transport, and Respiration
Authors: Kálmán Mihály , Hajós Ferenc
Published: October 2006

Abstract: Synaptosomal fractions were isolated from rat cerebral cortex immediately and at 1, 2, 3, and 4 h postmortem. The capability to undergo function‐dependent configurational changes in relation to ouabain‐ and protoveratrine‐sensitive K+ uptake, and the rate of O2 consumption were compared between fresh and postmortem synaptosomal fractions. Our results suggest that a 2 h postmortem storage at room temperature does not decrease substantially the functional performance of synaptosomes, while longer storage periods interfere with their capacity to maintain intracellular cation levels. The findings appear to be relevant to the use of human autopsy material for the preparation of synaptosomes
An Improved Filtration Procedure for Measuring Opiate Receptors in Small Regions of Rat Brain
Authors: Bardo M. T. , Bhatnagar R. K. , Gebhart G. F.
Published: October 2006

Abstract: A modified filtration method for in vitro receptor binding was used to determine specific binding of [3H]naloxone to small regions of adult rat brain. Reliable determinations of ligand binding were quantified with about 50 μg of protein per assay tube. Large differences in [3H]naloxone binding were obtained between various brain nuclei, and these differences were consistent with prior determinations of opiate receptor densities in various rat brain nuclei using autoradiographic techniques.
Identification, Development, and Regional Distribution of Thymidylate Synthetase in Adult Rabbit Brain
Authors: Suleiman Suleiman A. , Spector Reynold
Published: October 2006

Abstract: The development and regional distribution of thymidylate synthetase (TS) (EC 2.1.1.45) in rabbit brain were determined. After optimization of the assay for brain, TS activity in brain was measured by a nonspecific (3H2O release) and specific method. The specific method involved the conversion of [6‐3H]deoxyuridine monophosphate (dUMP) to [3H]thymidine phosphate and the subsequent identification of [3H]thymidine. The specific activity of the enzyme in whole brain of newborn rabbits declined from 10.35 ± 1.17 units/mg protein to 0.71 ± 0.09 units/mg protein at 10–12 weeks of age. Two‐year‐old rabbits had 0.81 ± 0.04 units/mg protein. The decline in specific activity with age was not due to an inhibitor of TS activity or a change in the Km for dUMP. The Km for dUMP of the unpurified enzyme in the brains of both 10‐day‐old and young adult rabbits was 0.8 μm. In young adult rabbits (3 months) the specific activity of TS was similar in the various regions of the brain tested except for the cerebellum, which had 40% higher specific activity than the whole brain. The results show that TS is widely distributed in adult rabbit brain, and, although the activity declines with age, it stabilizes at adult levels at 3 months of age.
Topographic Distribution of Enzymes Synthesizing Phosphatidylcholine and Phosphatidylethanolamine in Chicken Brain Microsomes
Authors: Freysz L. , Harth S. , Dreyfus H.
Published: October 2006

Abstract: The localization of phosphatidylethanolamine and phosphatidylcholine biosynthetic enzymes within the transverse plane of chicken brain microsomes was investigated by using proteases (trypsin and pronase) and neuraminidase. Treatment of intact microsomes with the proteases inactivated the phosphocholine transferase completely and the ethanolamine phosphotransferase only slightly. This latter enzyme was, however, completely inactivated when deoxycholate‐treated microsomes were exposed to proteases. Treatment of intact microsomes with neuraminidase had no effect on both phosphotransferases, although 65% of the sialic acid of sialoglycoproteins and 37% of that of gangliosides were removed. With deoxycholate‐disrupted microsomes nearly all sialic acid from the sialoglycoproteins and about 70% of that of gangliosides were released. In parallel, the phosphoethanolamine transferase was 90% inactivated. It is suggested that phosphocholine transferase is localized on the outer face of the microsomal vesicle, whereas the phosphoethanolamine transferase could be a sialoglycoprotein, possibly situated on the inner face of the vesicle, or perhaps a transmembrane protein.
Effects of Neonatal Undernutrition on the Lipid Composition of Gray Matter and White Matter in Rat Brain
Authors: Reddy T. Sanjeeva , Horrocks Lloyd A.
Published: October 2006

Abstract: Separate analyses were made of gray matter and white matter from rat brain after neonatal undernutrition. Newborn rats were redistributed into control, large‐litter, and protein‐deficient groups. Large litters had 16 rather than 8 pups with a dam. Protein‐deficient dams were fed a 4%, instead of a 24%, casein diet. For controls at 21 days of age, the 2′,3′‐cyclic nucleotide‐3′‐phosphohydrolase activity was more than fivefold greater in white matter than in gray matter. Severe undernutrition (protein‐deficient) gave 2′,3′‐cyclic nucleotide‐3′‐phosphohydrolase activities that were 36% lower in gray matter and 56% lower in white matter. Lipid galactose concentrations were 17% less than control in both gray matter and white matter. In protein‐deficient white matter, phospholipid concentrations were 15% lower than control. Ethanolamine plasmalogens and phosphatidyl serine were affected most. Moderate undernutrition (large litter) had no effect on 2′,3′‐cyclic nucleotide‐3′‐phosphohydrolase activity. A 14% deficit of galactolipids was the only difference from controls in large‐litter white matter. In large‐litter gray matter, phospholipid concentrations were 16% higher than controls. Nearly all glycerophos‐pholipids, including plasmalogens, were affected. With the exception of the myelination markers, 2′,3′‐cyclic nucleotide‐3′‐phosphohydrolase and lipid galactose, the development of lipids in gray matter is almost completely spared from the effects of undernutrition. The primary effect of undernutrition is on myelination, especially in white matter.
The Distribution of Enzymes Which Synthesize Nicotinamide Adenine Dinucleotide and Nicotinamide Adenine Dinucleotide Phosphate in Monkey, Rabbit, and Ground Squirrel Retinas
Authors: Berger Sosamma J. , DeVries Gerald W.
Published: October 2006

Abstract: The activity of adenosinetriphosphate:nicotinamide adenylyl‐transferase (EC 2.7.7.1) was measured in all the layers of monkey, rabbit, and ground squirrel retinas. Nicotinamide adenine dinucleotide (NAD) kinase (EC 2.7.1.23) distribution was measured in monkey and rabbit retinas. An attempt was made to measure NAD synthetase (EC 6.3.5.1), but the activities in the retinal layers were too low to produce a reliable increment in the levels of endogenous NAD+. In monkey retina the adenylyl transferase was highest by far in the outer and inner nuclear layers, lower and variable in ganglion cell and fiber layers, and almost absent elsewhere. Rabbit retina differed in that activity was nearly absent in the outer nuclear layer, whereas in the ground squirrel outer nuclear layer activity was double that of the inner nuclear layer. The species differences suggest that adenylyl transferase is almost absent from cone cell nuclei and high in rod cell nuclei. NAD kinase distribution in monkey retina was almost the mirror image of that of adenylyl transferase.
Blocked and Methylated 5′‐Terminal Cap Structures of Rat Brain Messenger Ribonucleic Acids
Authors: Ven Murthy M. R.
Published: October 2006

Abstract: The presence and identity of 5′‐terminal cap structures in rat brain polysomal mRNA were investigated by radiolabeling the mRNA by periodate oxidation and [3H]sodium borohydride reduction or by β‐elimination of 5′‐terminal nucleoside and incorporation of 32P in the presence of polynucleotide kinase. The labeled mRNAs were digested with nucleases and the cap structures were isolated and identified by chromatographic and electrophoretic procedures. The results showed that rat brain mRNAs contained cap 1 and cap 2 structures and no caps of the zero type. The proportion of cap 2 was higher than that of cap 1. Both caps had 7‐methylguanosine (m7G) as the 5′‐terminal nucleoside, which was linked to the next nucleoside by an inverted triphosphate bridge, as in other eukaryotic mRNAs. The most prominent nucleoside in the 5′‐penultimate position was 6‐methyl‐2′‐O‐methyiadenosine [m6A(m)] followed by 2′‐O‐methyladenosine [A(m)], which together contributed to nearly 70% of both cap 1 and cap 2 structures. 2′‐O‐Methylguanosine [G(m)] accounted for approximately 18%, the rest being made up of 2′‐O‐methyl‐cytidine [C(m)] and 2′‐O‐methyluridine [U(m)].
Effects of Bovine Growth Hormone on the Retarded Cerebral Development Induced by Neonatal Hydrocortisone Intoxication
Authors: Noguchi Tetsuya , Sugisaki Tetsuro , Watanabe Masayuki , Kohsaka Shinichi , Tsukada Yasuzo
Published: October 2006

Abstract: In comparison with normal controls, hydrocortisone‐intoxicated rats (HC rats) had smaller cerebra, lowered 2′,3′‐cyclic nucleotide 3′‐phosphohydrolase (CNPase) activity, and greatly reduced learning ability. The reduction in cerebral weight and DNA content was considered to be caused by a decrease in the number of proliferating glial cells, because the usual postnatal elevation of thymidine kinase (TK) activity was found to be suppressed in the cerebra from the HC rats. Electron microscopic observation of the pituitary gland revealed that the 5‐day‐old HC rat contained growth hormone (GH) secretory cells which were fully packed with GH granules, suggesting a disorder in the system which releases GH. In an attempt to promote cerebral development in the HC rats, we administered bovine GH (bGH) to some of the HC rats daily from the day of birth until weaning (HC + bGH rats). In the HC + bGH rats, the cerebral DNA was restored to normal levels and a concomitant increase in TK and CNPase activity was noted. Furthermore, in the brightness discrimination test, whereas the HC + bGH rats attained the learning ability of the normal controls after only 10 sessions, the HC rats were unable to reach an equivalent level even after 25 sessions.
Endogenous Peptides That Inhibit Brain Cyclic AMP Phosphodiesterase
Authors: Collier H. O. J. , Butt N. M. , Saeed S. A.
Published: October 2006

Abstract: Peptide extracts of rat brain powerfully inhibited the cyclic AMP phosphodiesterase activity of rat brain homogenate. Similar extracts of ox brain showed comparable although less potent activity. Preliminary investigation of the physicochemical properties of brain extracts indicated that the rat brain extract contained an active peptide of low molecular weight (about 1400), whereas ox brain contained two such peptides (about 1400 and 900). These studies indicate that endogenous oligopeptides that inhibit cyclic AMP phosphodiesterase are present in brain. Experiments on several pure peptides known to be present in the CNS showed that the majority were inactive against brain phosphodiesterase, but ACTH(1.24), somatostatin, substance P and Lys8‐vasopressin, in descending order of potency, were active. To help distinguish the peptides found in rat and ox brain extracts from known peptides, preliminary analyses of amino acid composition were performed. These suggested that the peptides found in brain extracts were distinct from known peptides having the ability to inhibit cyclic AMP phosphodiesterase.
Biochemical and Histochemical Evidence of 16S Acetylcholinesterase in Salivary Glands
Authors: Kása P. , Rakonczay Z.
Published: October 2006

Abstract: Three forms of acetylcholinesterase (AChE) have been reported: the 4S, 10S, and 16S forms. It was suggested previously that 16S AChE is characteristic of the end‐plate region; subsequently its presence has also been demonstrated in sciatic nerve, vagus nerve, several nerve trunks, cardiac atria, and distal ileum of rat. The purpose of the present study was to investigate further the occurrence of 16S AChE. We found that not only is it present in motor axons but that it may also occur in the secretory postganglionic parasympathetic fibers in the synapse‐free parotid and submandibular glands.
Distribution of Angiotensin‐Converting Enzyme Activity in Specific Areas of the Rat Brain Stem
Authors: Chevillard Claude , Saavedra Juan M.
Published: October 2006

Abstract: Angiotensin‐converting enzyme (ACE) activity was measured by a radiochemical assay in 30 specific areas of the rat brain stem. ACE activity is unevenly distributed, with a 60‐fold difference between the lowest and the highest activity. The area postrema exhibits the highest activity. The substantia nigra (pars reticulata), the locus coeruleus, the areas A1 and A2, the nuclei commissuralis, and tractus solitarii have a substantial ACE activity, whereas the lowest activity is found in the raphe nuclei and the nuclei of the reticular formation.
Contribution of Sulfate Conjugation, Deamination, and O‐Methylation to Metabolism of Dopamine and Norepinephrine in Human Brain
Authors: Rivett A. Jennifer , Eddy Barbara J. , Roth Jerome A.
Published: October 2006

Abstract: The kinetic constants were determined for dopamine (DA) and norepinephrine (NE) metabolism by phenolsulfotransferase (PST), type A and B monoamine oxidase (MAO), and membrane‐bound and soluble catechol‐O‐ methyltransferase (COMT) in frontal lobe preparations of human brain. PST and membrane‐bound COMT were found to have the lowest Km, values for both catecholamines. By means of the appropriate rate equations and the calculated kinetic constants for each enzyme, the activity of each enzymatic pathway was determined at varying concentrations of DA and NE. Results indicate that deamination by MAO is the principal pathway for the enzymatic inactivation of DA whereas NE is largely metabolized by MAO type A and membrane‐bound COMT under the in vitro assay conditions used. At concentrations less than 100 μM, soluble COMT’contributes less than 5% to the total catabolism of either catecholamine. PST can contribute up to 15% of the total DA metabolism and 7% of NE metabolism.
Cytochrome Reductase Activities in Rat Brain Microsomes During Development
Authors: Takeshita Masazumi , Miki Mayumi , Yubisui Toshitsugu
Published: October 2006

Abstract: Postnatal developmental alterations of microsomal NADH‐cyto‐chrome b5 reductase and NADPH‐cytochrome c reductase activities were determined in the brain of rats. The reductase activities increased from a low level in the immature brain to a maximum level at 23 to 30 days of age, and then decreased slightly to a plateau. The periods of the activity increments were in accord with those of the enhancement of microsomal fatty acid elongation. The specific activities of these reductases were high in cerebral hemispheres and medulla oblongata, intermediate in midbrain, and lowest in cerebellum of the four regions of 20‐day‐old rat brain.
Effects of Monensin on Assembly of Po Protein into Peripheral Nerve Myelin
Authors: Rapaport Raymond N. , Benjamins Joyce A. , Skoff Robert P.
Published: October 2006

Abstract: The ionophore monensin has been used in a variety of systems to block secretion of glycoproteins or assembly of glycoproteins into membranes. We examined the effects of monensin on assembly of the Po glycoprotein into PNS myelin, and compared this agent with the glycosylation inhibitor tunicamycin in our system. Sciatic nerves from 9‐day‐old rat pups were sliced and incubated in vitro. Electron microscopy of the Schwann cells in slices incubated with monensin revealed extensive swelling of the Golgi complex. Incubation with 10−7M monensin inhibited total protein synthesis by about 20% and fucose incorporation into protein about 35%. Following isolation of myelin, proteins were separated by sodium dodecyl sulfate gel electrophoresis. Monensin inhibited the appearance of Po in myelin, while causing its accumulation in a denser membrane fraction. In addition, a slightly faster‐migrating species of Po labeled with both [3H]fucose and [14C]glycine was observed in all fractions. Assembly of basic proteins into myelin was not affected. Preincubation with 10 μg/ml tunicamycin for 30 min prior to incubation with [3H]fucose and [14C]glycine for 2 h resulted in a 65% decrease in [3H]fucose incorporation into Po, and the appearance of a new [14C]glycine‐labeled peak that migrated in the region of the 23K protein reported by Smith and Sternberger. [3H]Fucose incorporation was inhibited earlier, and to a greater extent, than protein synthesis. Our results show that processing of the Po glycoprotein is sensitive to both monensin and tunicamycin, and that monensin partially blocks assembly of Po into myelin.
Effects of Monovalent and Divalent Cations on Ca2+ Fluxes Across Chromaffin Secretory Membrane Vesicles
Authors: Krieger‐Brauer Heidemarie I. , Gratzl Manfred
Published: October 2006

Abstract: Bovine chromaffin secretory vesicle ghosts loaded with Na+ were found to take up Ca2+ when incubated in K+ media or in sucrose media containing micromolar concentrations of free Ca2+. Li+‐ or choline+loaded ghosts did not take up Ca2+. The Ca2+ accumulated by Na+‐loaded ghosts could be released by the Ca2+ ionophore A23187, but not by EGTA. Ca2+ uptake was inhibited by external Sr2+, Na +, Li +, or choline +. All the 45Ca2+ accumulated by Na+‐dependent Ca2+ uptake could be released by external Na +, indicating that both Ca2+ influx and efflux occur in a Na+‐dependent manner. Na + ‐dependent Ca2+ uptake and release were only slightly inhibited by Mg2+. In the presence of the Na+ ionophore Monensin the Ca2+ uptake by Na +‐loaded ghosts was reduced. Ca2+ sequestered by the Na+‐dependent mechanism could also be released by external Ca2+ or Sr2+ but not by Mg2+, indicating the presence of a Ca2+/Ca2+ exchange activity in secretory membrane vesicles. This Ca2+/Ca2+ exchange system is inhibited by Mg2+, but not by Sr2+. The Na + ‐dependent Ca2+ uptake system in the presence of Mg2+ is a saturable process with an apparent Km of 0.28 μM and a Vmax= 14.5 nmol min−1 mg protein−1. Ruthenium red inhibited neither the Na+/Ca2+ nor the Ca2+/Ca2+ exchange, even at high concentrations.
Lack of Stereoselectivity in Ability of Nicotine to Release Dopamine from Rat Synaptosomal Preparations
Authors: Connelly Michael S. , Littleton John M.
Published: October 2006

Abstract: Both the naturally occurring (—)‐isomer and the synthetic (+)‐isomer of nicotine caused release of 3H from a crude synaptosomal fraction of rat brain prein‐cubated with [3H]dopamine. The isomers were equipotent in producing this response, which was concentration‐dependent, a significant effect on the fractional release of dopamine being observed at 10−4M nicotine. The effect did not appear to be the result of synaptosomal damage, as levels of the intrasynaptosomal marker lactate dehy‐drogenase did not increase in the supernatant. Nicotine‐induced release was inhibited by removal of external Ca2+ and by the presence in vitro of pempidine (230 μ.M). Neither hexamethonium (500 μ.M) in vitro nor the chronic administration of (‐)‐nicotine in vivo had any effect on the nicotine‐induced release of [3H]dopamine. It is concluded that nicotine exerts this effect via a presynaptic nicotinic receptor of the “ganglionic” type, but that this receptor differs from that in the periphery by showing a relative lack of stereospecificity. There is no evidence of a functional “down regulation” in this receptor on chronic exposure to nicotine in vivo.
Metabolic Processes in Alzheimer's Disease: Adenine Nucleotide Content and Production of 14CO2 from [U‐14C]Glucose In Vitro in Human Neocortex
Authors: Sims N. R. , Bowen D. M. , Neary D. , Davison A. N.
Published: October 2006

Abstract: Samples of neocortex removed at diagnostic craniotomy from patients with Alzheimer's disease and incubated in vitro showed an increased production of 14CO2 from [U‐14C]glucose compared with neurosurgical controls. This was a feature of incubations in the presence of both 5 mM K+ (142% control) and 31 mM K + (126%). Specific labelling of the amino acid pool was unaltered, suggesting that the apparent increase of CO2 production was not merely a reflection of changes in dilution of the radiolabel from glucose. The content of adenine nucleotides was significantly less than control values in the tissue from patients with Alzheimer's disease after in vitro incubations but the adenylate energy charge was unchanged, indicating that normal energy metabolism was not grossly impaired in these preparations.
Comparison of Voltage‐Dependent 45Ca2+ Uptake Rates by Synaptosomes Isolated from Rat Brain Regions
Authors: Leslie S. W. , Barr E. , Chandler L. J.
Published: October 2006

Abstract: 45Ca2+ uptake by synaptosomes isolated from cerebral cortex, cerebellum, midbrain, and brain stem of male Sprague‐Dawley rats was measured at 1‐, 3‐, 5‐, 15‐, 30‐, and 60‐s time periods. The fastest rate of depolarization‐dependent calcium uptake occurred in each brain region between 0 and 1 s. Uptake rates dropped off quickly with 3–5‐s rates at approximately 15–20% of those observed at 0–1 s in cerebral cortex, cerebellum, and midbrain. Uptake rates at the 1–3‐s interval were maintained at a relatively high rate in these three brain regions suggesting mixed fast‐ and slow‐phase processes. The magnitude and rate of 45Ca2+ uptake were similar in synaptosomes from cerebral cortex, cerebellum, and midbrain but were significantly less in brain stem synaptosomes. These results suggest a fast and a slow component to voltage‐dependent 45Ca2+ uptake by presynaptic nerve terminals from various brain regions.
Further Studies on 4‐Hydroxy‐3‐Methoxyphenylglycol Oxidation in Humans: Effect of Pool Expansion and Stereochemistry
Authors: Mådh Göran
Published: October 2006

Abstract: The in vivo oxidation of the norepinephrine metabolite 4‐hydroxy‐3‐methoxyphenylglycol (HMPG) to 4‐hydroxy‐3‐methoxymandelic acid was studied in man with two different doses of deuterium‐labeled HMPG and a tracer dose of [14C]HMPG. HMPG oxidation appeared to be dose‐dependent with an oxidation of 62–70% for doses below or equal to 2.2 μmol. With the use of a capillary column coated with an optically active phase (Chirasil‐Val) and gas chromatography mass‐spectrometry the human urinary excretions of the two stereoisomers of deuterium‐labelled HMPG (free + conjugates) were found to be equal.
Binding of [3H]DMCM, a Convulsive Benzodiazepine Ligand, to Rat Brain Membranes: Preliminary Studies
Authors: Braestrup C. , Nielsen M. , Honoré T.
Published: October 2006

Abstract: DMCM (methyl 6,7‐dimethoxy‐4‐ethyl‐β‐carbo‐line‐3‐carboxylate) produces convulsions in mice and rats, probably by interacting with benzodiazepine (BZ) receptors. Investigation of specific binding of [3H]DMCM to rat hippocampus and cortex revealed polyphasic saturation curves, indicating a high‐affinity site (KD= 0.5–0.8 nM) and a site with lower affinity (KD= 3–6 nM). BZ receptor ligands of various chemical classes, but not other agents, displace [3H]DMCM from specific binding sites—indicating that [3H]DMCM binds to BZ receptors in rat brain. The regional distribution of [3H]DMCM binding is complementary to that of the BZ1‐‐selective radioligand [3H]PrCC. Specific binding of [3H]DMCM (0.1 nM) was reduced by γ‐aminobutyric acid (GABA) receptor agonist to ∼20% of the control value at 37°C in chloride‐containing buffers; the reduction was bicuculline methiodideand RU 5135‐sensitive. The effective concentrations of 10 GABA analogues in reducing [3H]DMCM binding correlated closely to published values for their GABA receptor affinity. Specific binding of [3H]DMCM is regulated by unknown factors; e.g., enhanced binding was found by Ag+ treatment of membranes, in the presence of picrotoxinin, or by exposure to ultraviolet light in the presence of flunitrazepam. In conclusion, [3H]DMCM appears to bind to high‐affinity brain BZ receptors, although the binding properties are different from those of [3H]flunitrazepam and [3H]PrCC. These differences might relate in part to subclass selectivity and in part to differences in efficacy of DMCM at BZ receptors.
Choline and Acetylcholine Metabolism in Rat Neostriatal Slices
Authors: Weiler Molly H. , Il Bak Jin , Jenden Donald J.
Published: October 2006

Abstract: Choline (Ch) uptake and release and acetylcholine (ACh) synthesis and release have been studied by gas chromatography mass spectrometry (GCMS) in slices of rat neostriatum in vitro to assess the effects of depolarization by 25 mM K+ and the influence of elevated concentrations of Ch in the incubation medium. During the first 60 min after preparation, 25 mM K+ increased ACh release by 182% and reduced ACh levels by 40%. The rate of ACh synthesis was unchanged. After a 1‐h equilibration period, the rate of ACh synthesis was considerably less (2.41 nmol mg−1 h−1, compared to 9.78 nmol mg−1 h−1). Exposure to 25 mM K+ during the second hour increased the rate to 6.47 nmol mg−1 h−1. During the first 10 min of exposure to 25 mM K+, ACh synthesis was reduced, regardless of incubation. Increasing concentrations of external [2H4]Ch apparently favored initial rates of net ACh synthesis, since the rank order of initial net ACh synthesis rates is the same as the rank order of external [2H4]Ch concentration under both normal and depolarized conditions. However, the only significant effect of external [2H4]Ch on ACh metabolism was that it increased ACh release during the initial 10 min, when the preparation was depolarized with K+. The efflux of endogenous [2H0]Ch was increased initially (10 min) and slowed over a 60‐min period by 25 mM K+, and increased when [2H4]Ch in the medium was increased. Changes in ACh synthesis and release were dependent upon the time exposure of slices to high K+, and the results suggest that Ch favors initial rates of ACh synthesis, but that Ch influences ACh release primarily under conditions of stress (i.e., depolarization).
Changes in Hexokinase Isoenzymes in Regions of Rat Brain During Insulin‐Induced Hypoglycemia
Authors: Kaur Gurcharan , Singh Rameshwar , Baquer Najma Zaheer
Published: October 2006

Abstract: Activities of hexokinase isoenzymes were determined during insulin‐induced hypoglycemia in soluble and total particulate fractions from three regions of rat brain. Type I hexokinase isoenzyme activity showed a small decrease in both soluble and particulate fractions from the cerebral hemispheres. In cerebellum and brain stem, however, Type I isoenzyme showed a decrease only in the soluble fraction. A significant increase was observed in hexokinase Type II isoenzyme from both the fractions, in all the three brain regions 1 h after insulin administration.
In Vivo Release from Cerebral Cortex of [14C]Glutamate Synthesized from [U‐14C]Glutamine
Authors: Thanki C. M. , Sugden D. , Thomas A. J. , Bradford H. F.
Published: October 2006

Abstract: Awake, unrestrained, and behaviourally normal animals with superfusion cannulae implanted over the sensorimotor cortex were used in a study of the capacity of infused [U‐14C]glutamine for labelling glutamate and other amino acids released by depolarising stimuli. A spontaneous background release of [14C]glutamate was detected. This was increased by tityustoxin (1 μM). The specific radioactivity of glutamate increased eightfold during the evoked‐release period. [14C]Aspartate was also detected and showed increased release, but not increased specific labelling, in response to depolarisation. Evoked γ‐aminobutyric acid (GABA) release occurred but only small amounts of [14C]GABA were detected. Glutamine showed increased rates of uptake to the sensorimotor cortex during stimulation periods, suggesting an accelerated breakdown via glutaminase.
Relative Importance of 3‐Methoxy‐4‐Hydroxyphenylglycol and 3,4‐Dihydroxyphenylglycol as Norepinephrine Metabolites in Rat, Monkey, and Humans
Authors: Elsworth J. D. , Roth R. H. , Redmond D. E.
Published: October 2006

Abstract: A gas chromatographic‐mass spectrometric assay, which allowed simultaneous measurement of 3‐methoxy‐4‐hydroxyphenylglycol (MHPG) and 3,4‐dihydroxyphenylglycol (DHPG), was used to show that the concentration of MHPG in primate CNS far exceeded that of DHPG and that both metabolites were mainly in the unconjugated form. In rat brain, DHPG concentration was generally higher than that of MHPG, and both existed predominantly as conjugates. Rat and primate plasma contained more MHPG than DHPG. In plasma of primates but not of rats, higher proportions of the metabolites were conjugated, compared to those in brain. Significant correlations existed between MHPG and DHPG in rat brain, monkey brain, human plasma, and both monkey CSF and plasma. In monkeys, a significant CSF‐plasma correlation was found for MHPG, but not for DHPG. Acute administration of piperoxane raised rat brain MHPG and DHPG concentration; desipramine prevented this rise in DHPG, but not in MHPG. Desipramine alone decreased DHPG, but not MHPG, concentration. Piperoxane increased monkey brain MHPG, but not DHPG, concentration. These data suggest that DHPG is a valuable metabolite to measure when assessing norepinephrine metabolism in the rat. Under certain conditions, measurement of rat brain MHPG and DHPG may provide information concerning the site of norepinephrine metabolism. However, in primates the importance of monitoring DHPG, in addition to MHPG, is uncertain.
Postnatal Changes in Cathepsin D in Rat Neural Tissue
Authors: Snyder D. Stephen , Whitaker John N.
Published: October 2006

Cathepsin D, an aspartyl endopeptidase, was analyzed in cortex from forebrain and cerebellum, spinal cord, and optic and sciatic nerves, and in the liver of rats from 1 to 120 days of age. Cathepsin D was quantitated in tissue extracts by measurement of enzyme specific activity on a substrate of [methyl‐14C]‐methylated hemoglobin and by radioimmunoassay. Immunocytochemistry was used to ascertain the identity of the mixed cell types that contributed to the cathepsin D detected. As quantitated by radioimmunoassay, immunoreactive cathepsin D varied between 0.2 and 1 ng/μg of total protein. Maximum activity occurred at approximately the 15th postnatal day; the least amount of immunoreactive cathepsin D was found at 30 or 60 days of age. A subsequent increase of varying magnitude occurred at postnatal day 120. There was good correspondence between immunoreactive enzyme and enzyme specific activity, which ranged from 1 to 4 ng/μg of total protein, and the activities determined by the two methods provided similar, but not identical, developmental profiles. Cathepsin D was demonstrated by immunocytochemistry to be present in most neurons, in all choroid plexus epithelium, and in certain oligodendrocytes from the first postnatal day. Cathepsin D was present in oligodendrocytes in cord lateral funiculi and optic nerve by the first postnatal day, and by the sixth postnatal day many oligodendrocytes were abundantly stained. In contrast, oligodendrocytes in the corpus callosum and in the cerebellar white matter did not contain demonstrable cathepsin D until postnatal days 10 and 15, respectively. These results indicate a role for cathepsin D during the postnatal development of rat CNS and suggest that this proteinase may be involved in the steps of myelination.
Amino Acids in the Substantia Nigra of Rats with Striatal Lesions Produced by Kainic Acid
Authors: Korf Jakob , Venema Kor
Published: October 2006

In an attempt to estimate the pool size of glutamate and other amino acids in γ‐aminobutyric acid (GABA)‐containing neurons, we determined the content of 12 amino acids in the bilateral substantia nigra of rats, in which unilateral striatal lesions had been made with kainic acid two weeks earlier. The assay of the amino acids (including glutamate, aspartate, glutamine, asparagine, glycine, and GABA) and ethanolamine was based on HPLC and fluorimetric detection after precolumn derivatization with o‐phthaldialdehyde. The levels of all measured amino acids (except those of tyrosine, threonine, and ethanolamine) were decreased in the affected striatum, but only the levels of aspartate, taurine, and GABA were lowered in the ipsilateral substantia nigra. These results indicate that the pool size of the various amino acids in the striatonigral GABAergic pathway is small compared to their nigral content, and that in addition to GABA a significant fraction of aspartate and taurine may be confined to nerve terminals in the substantia nigra.