bluuebunny/crossref_metadata_2025 huggingface dataset:
THE SUBCELLULAR DISTRIBUTION OF [14C]GABA AND [3H]DOPAMINE IN THE RETINA
Authors: Attlrwill C. K. , Neal M. J.
Published: December 2006

Abstract— Rabbit retinae were homogenized in isotonic sucrose and subjected to differential and density gradient centrifugation. Preliminary electron microscopic examination of some of the fractions indicated that in addition to the subcellular particles usually observed in brain homogenates, the photoreceptor cells gave rise to several characteristic fragments. These included fragmented outer limbs, aggregations of mitochondria from the inner segments, and photoreceptor terminals. Unlike the synaptosomes formed from the conventional type of synapses in the retina, these photoreceptor terminals appeared to sediment mainly in the low speed crude nuclear pellet (P1).

Retinae were incubated with low concentrations of [14C]GABA and/or [3H]dopamine prior to subcellular fractionation and in these experiments the P2 pellet was further fractionated on sucrose density gradients. Analysis of the radioactivity in the fractions showed that labelled GABA was accumulated by osmotically sensitive particles which had the sedimentation characteristics of synaptosomes. The panicles accumulating [3H]dopamine appeared to belong to a different, slightly lighter, population than those accumulating [14C]GABA. It is tentatively suggested that the particles accumulating labelled GABA were synaptosomes because the fractions containing these particles also possessed most of the GAD activity of the gradient. In contrast, GABA‐T and MAO activity was found in the dense fractions of the gradients usually associated with mitochondria.

When retinae were incubated with a high concentration of labelled GABA a‘lighter’population of particles seemed to accumulate the amino acid than when a low external GABA concentration was used. These results suggest that the high and low affinity uptake processes for GABA in the retina may have different cellular sites.
DEVELOPMENTAL AND AGING CHANGES IN PROTEIN CONCENTRATION AND 2′,3′‐CYCLIC NUCLEOSIDEMONOPHOSPHATE PHOSPHODIESTERASE ACTIVITY (EC 3.1.4.16) IN HUMAN CEREBRAL WHITE AND GRAY MATTER AND SPINAL CORD
Authors: Toews A. D. , HORROCKS L. A.
Published: December 2006

Abstract— The concentration of protein as assayed by the Lowry method and the specific activity of 2′.3’‐cyclic nucleosidemonophosphate phosphodiesterase (CNP), an enzyme characteristic of the myelin sheath, were determined in human CNS tissues obtained at autopsy from subjects ranging in age from 26 weeks gestation to 83 y.

CNP activity in cerebral white matter samples was very low until approx 2 months of age when it increased rapidly, reaching near‐adult levels by 2 y of age. CNP activity in adult (15–60 y) cerebral white matter was 8.1 ± 1.0 μmol/min/mg protein (mean ±s.d.). The protein concentration of cerebral white matter increased from 64 mg/g wet tissue at 26 weeks gestation to adult levels (118.5 ± 10.0 mg/g wet tissue) by 16–18 months. CNP activity in cerebral gray matter was initially very low and showed only a small increase during development to adult values of approx 1.4 μmol/min/mg protein. In spinal cord, adult values (3.7 ± 0.56 μmol/min/mg protein) were found shortly after birth. The increase in CNP activity to near‐adult values occurred earlier in cross‐sections of cervical spinal cord than in cerebral white matter. The increase in spinal cord protein concentration showed a similar trend (adult values = 103.1 ± 9.5 mg/g wet tissue).

The white matter protein concentration decreased significantly with age over the 15–83 y interval examined but the CNP specific activity in white matter did not. The protein concentration of the 61–83 y group was 8% lower than that of the 15–60 y group. The spinal cord protein concentration decreased significantly and the spinal cord CNP specific acitivity increased significantly with increasing time between death and sample freezing. The sex of the individual had no significant effect on any of the variables examined.

The developmental curves obtained for these tissues are consistent with the hypothesis that CNP is an intrinsic myelin component in human CNS myelin. The marked increase in CNP activity in white matter coincides with the period of rapid myelin deposition as determined by other parameters. CNP activity may be useful as an index of myelination in human CNS tissues.
METABOLIC RELATIONSHIPS BETWEEN MYELIN SUBFRACTIONS: ENTRY OF GALACTOLIPIDS AND PHOSPHOLIPIDS 1
Authors: Benjamins Joyce A. , Miller S. L. , Morell P.
Published: December 2006

Abstract— Partially purified myelin from the brains of 17‐day‐old rats was separated into 4 subfractions on a three‐step sucrose gradient by virtue of heterogeneity in density and particle size. Precursor‐product relationships between different membrane fractions were investigated by determining the specific radioactivity of individual lipids in each subcellular fraction 15 min after intracranial injection of an appropriate precursor. Rats were injected with [2‐3H]glycerol. myelin subfractions prepared, and individual lipids separated by TLC. For choline and ethanolamine phospholipids, specific radioactivity was highest in the densest fraction (D), intermediate in the next densest fraction (C), and lowest in the lighter fractions (B and A). Similar results were observed for cerebroside and sulphatide when [3H]galactose was the precursor. These data are consistent with (but do not prove) a precursor‐product relationship for individual lipids from the densest to the lightest subfraction.

Another experimental design involving time staggered injections of [3H] and [14C] precursors was developed which enables a more definitive result with regard to precursor‐product relationships to be obtained. A precursor‐product relationship between a given lipid in a dense myelin membrane fraction, and the same lipid in a lighter subfraction, would be indicated by a change in isotope ratio. If there is no precursor‐product relationship. Ihe isotope ratio should be constant. Such experiments were done with [3H] and [14C]glycerol. The data indicated that phosphatidyl ethanolamine and its plasmalogen analog were added first to the densest subfraction and then in turn to the lighter subfractions. In contrast, phosphatidyl choline and its plasmalogen analog were added “simultaneously” (i.e. with delays of much less than 15min) to each of the subfractions. Similar experiments with [3H] and [14C]galactose showed that cerebroside, sulphatide and galactosyl diglyceride also entered the subfractions simultaneously rather than in sequential order. Thus the assembly of the myelin sheath involves an obligate order of addition of certain lipids. while other lipids are probably added in a random order.
DEVELOPMENTAL CHANGES IN THE BRAIN LEVELS OF NEUROTRANSMITTERS AS INFLUENCED BY MATERNAL ETHANOL CONSUMPTION IN THE RAT1
Authors: Rawat A. K.
Published: December 2006

Abstract— Effects of prolonged maternal ethanol consumption by pregnant and lactating rats were investigated on the cerebral contents of certain neurotransmitters and the activities of their enzymes in fetal and neonatal brains. Ethanol consumption by the pregnant rats resulted in a significant decrease in the fetal brain contents of acetylcholine and an increase in γ‐aminobutyric acid and glutamate. The cerebral levels of 5‐hydroxytryptamine (5‐HT), norepinephrine of choline in the fetal brains from the ethanol group were not significantly different from those observed in the control group. With the increase in gestational age, an increase in the levels of 5‐HT, norepinephrine, acetylcholine, γ‐amino‐butyric acid (GABA) and glutamate was observed in the fetal brains from both ethanol and control groups. The brains of neonates suckling on ethanol‐fed mothers showed a significant decrease in acetylcholine levels and an increase in GABA and glutamate levels as compared to the corresponding controls. The activities of L‐glutamate decarboxylase and GABA‐aminotransferase in the fetal as well as neonatal brains were decreased in the ethanol‐fed group as compared to the controls. A single injection of acetaldehyde resulted in a decrease in acetylcholine, GABA and glutamate levels in both neonatal and adult brains.
SEPARATION OF NEUROTOXINS FROM HORNET (VESPA INSULARIS) VENOM AND THEIR ACTIONS ON CRUSTACEAN NEUROMUSCULAR TRANSMISSION
Authors: Hori S. , Kawai N. , Niwa A. , Ohotani S.
Published: December 2006

Abstract— The venom of homet (Vespa insularis) was separated into three components by gel filtration on Sephadex G‐50, Sephadex G‐25 and Sephadex G‐10. The effect of each component on crustacean neuromuscular junctions was studied electrophysiologically using intracellular recordings. Two components (Fractions D and E) suppressed the postsynaptic potentials; Fraction D preferentially depressed excitatory postsynaptic potentials (epsps) without affecting inhibitory postsynaptic potentials (ipsps), and fraction E depressed both epsps and ipsps with concurrent decrease in membrane resistance. Another component (fraction F) augmented both epsps and ipsps. Fractions D and F were possibly acting presynaptically while fraction E was acting on the postsynaptic membrane. The active substances in fraction D, E and F were separated on TLC and the active substance in fraction F was confirmed to be serotonin. The active substances in fractions D and E are assumed to be peptides from the results of peptidase digestion. The effect of fraction D was degraded by treatment with trypsin but not by chymotrypsin nor carboxypeptidase A and B. On the other hand, the effect of fraction E was degraded by chymotrypsin and carboxypeptidase B but not by trypsin nor carboxypeptidase A.
GLUTAMINE UPTAKE AND METABOLISM BY THE ISOLATED TOAD BRAIN: EVIDENCE PERTAINING TO ITS PROPOSED ROLE AS A TRANSMITTER PRECURSOR
Authors: Shank R. P. , Aprison M. H.
Published: December 2006

Abstract— Hemisections of toad brains, when incubated in a physiological medium containing no glutamine. released considerable amounts of this amino acid into the medium. When glutamine was included in the medium at a concentration of 0.2 mm the net efflux from the tissue was reduced but not totally prevented. Although there was no net uptake of glutamine, the tissue did accumulate [U‐14C]glu‐tamine and some of this labelled glutamine was rapidly metabolized to glutamate, GABA and aspartate. The precursor‐product relationship for the metabolism of glutamine to glutamate differed from the classic single compartment model in that the specific radioactivity of glutamate rose very quickly to approx one‐tenth that of glutamine, but increased slowly thereafter. These data suggest that the [14C]glutamine was taken up into two metabolically distinct compartments and/or that some of the [14C]glutamine was converted to [14C]glutamate during the uptake process.

The uptake of [14C]glutamine was diminished when the tissue was incubated in a non‐oxygenated medium or when Na+ was omitted (substituted with sucrose) and K+ was concomitantly elevated. However, on a relative basis, the incorporation of radioactivity into glutamate and GABA was increased by these incubation conditions. The metabolism of glutamine to aspartate was greatly depressed when the tissue was not oxygenated. The glutamate formed from [U‐14C]glutamine taken up by the tissue was converted to GABA at a faster rate than was glutamate derived from [U‐14C]glucose. [U‐14C]gly‐cerol or exogenous [U‐14C]glutamate. This suggests that glutamine was metabolized to GABA selectively; i.e. on a relative basis, glutamine served as a better source of carbon for the synthesis of GABA than did glucose, glycerol or exogenous glutamate.

When the brain hemisections were incubated in the normal physiological medium with or without glutamine. there was very little efflux of glutamate, GABA or aspartate from the tissue. However when NaCl was omitted from the medium (substituted with sucrose) and K+ was elevated to 29 miu. a marked efflux of these three amino acids into the medium did occur, and over a period of 160min, the content of each amino acid in the tissue was depleted considerably. When glutamine (0.2 mm) was included in the Na+ deficient‐high K.+ medium, the average amount of glutamate, GABA and aspartate in the tissue plus the medium was greater than when glutamine was not included in the medium. Such data indicate that CNS tissues can utilize glutamine for a net synthesis of glutamate, GABA and aspartate.

The results of this study provide further evidence in support of the concept that the functional (transmitter) pools of glutamate and GABA are maintained and regulated in part via biosynthesis from glutamine. One specific mechanism instrumental in regulating the content of glutamate in nerve terminals may be a process of glutamine uptake coupled to deamidation.
COMPARISON OF THE BINDING’ OF β‐ALANINE AND y‐AMINOBUTYRIC ACID IN SYNAPTOSOMAL‐MITOCHONDRIAL FRACTIONS OF RAT BRAIN1
Authors: Somoza E. , Pugnaire Michele P. , Orensanz Munoz L. M. , Portal Clara Gonzalez , Ibanez A. Elena , DeFeudis F. V.
Published: December 2006

Abstract— Na+‐dependent ‘binding’ of β‐alanine and GABA was examined with synaptosomal‐mitochondrial fractions of rat brain incubated for 10 min at 0°C. GABA was bound to a much greater extent than β‐alanine to particles of cerebral cortex, whole cerebellum and brain stem. For cerebral cortex, the binding capacity (Bmax) for GABA was about 18 limes greater than that for β‐alanine. and the affinity of the particles for GABA was about 2′ times greater than for β‐alanine. The order of potency of GABA binding to brain regions was cerebral cortex > cerebellum > brain stem, whereas that for β‐alanine was the reverse. If the binding of β‐alanine is taken to indicate the glial component of the Na+‐dependent binding process for GABA, then most of the GABA was bound to neuronal elements under the conditions employed.
CALCIUM ION DEPENDENCE OF THE 2‐SITE IMMUNORADIOMETRIC ASSAY OF MOORE'S S‐100 PROTEIN
Authors: Miles L. E. M. , Lee Y. L. , Eng L. F.
Published: December 2006

Abstract— The two‐site immunoradiometric assay (two‐site IRMA) for a specific protein of the nervous system, S‐100, is carried out by reaction of the S‐100 protein solution with a solid‐phase anti(S‐100) followed by a second reaction in which the insoluble product is incubated with purified, radioactive anti(S‐100). Unreacted labeled antibodies remain in solution and are washed away. As the amount of S‐100 increases, the radioactivity in the solid‐phase increases. The most significant assay variable is the effect of calcium on the assay dose‐response. 0.1 mM‐EDTA causes a total inhibition of the dose‐response curve which is reversed by increasing the concentration of calcium ions. The dose‐response reaches a maximum at 1.0mM‐Ca2+. then becomes progressively inhibited as the Ca2+ concentration is increased further. Previous immunochemical studies of S‐100 which did not allow for this effect must now be interpreted with caution.
EFFECT OF OESTRADIOL ON TURNOVER OF TYPE A MONOAMINE OXIDASE IN BRAIN
Authors: Luine Victoria N. , McEwen B. S.
Published: December 2006

Abstract— Administration of oestrogen (oestradiol‐17β or oestradiol‐17β‐benzoate) to ovariectomized (OVX) rats for 1–4 weeks results in an approx 30% decrease in the activity of monoamine oxidase (MAO) in the basomedial‐hypothalamus (BM‐Hyp) and corticomedial‐amygdala (CM‐Amy) but not in cerebral cortex. Further investigation shows that (1) decreased MAO activity in the BM‐Hyp and CM‐Amy occurs only in Type A MAO (serotonin as substrate) and does not occur in Type B MAO (phenylethylamine as substrate); (2) decreased MAO activity does not occur when a single large dose of oestrogen is given i. v. or when homogenates from oestrogen treated rats are mixed with homogenates from OVX rats suggesting that direct enzyme inhibition is not responsible for the change in activity; (3) oestrogen administration to OVX rats increases the rate constant of degradation for MAO in BM‐Hyp and CM‐Amy but not in cerebral cortex as determined in turnover studies using pargyline, an irreversible inhibitor of MAO. The increased rate of degradation results in shorter half lives (t 1/2) for MAO in the BM‐Hyp and CM‐Amy of oestrogen treated rats. In OVX rats the t 1/2 is 9.8 days in BM‐Hyp and 12.7 days in CM‐Amy. Oestrogen administration results in a t 1/2 of 7.6 days in BM‐Hyp and 7.8 days in CM‐Amy. The possible relationship between oestrogen dependent decreased MAO activity and estrogen dependent lordosis behavior is discussed.
IN VIVO SYNTHESIS OF STABLE RNA WITHIN THE RAT NODOSE GANGLIA FOLLOWING VAGOTOMY
Authors: Kaye P. L. , Gunning P. W. , Austin L.
Published: December 2006

Abstract— We have investigated the in vivo synthesis of stable RNA in the rat nodose ganglia following unilateral vagotomy. Incorporation of precursor into rRNA, and possibly tRNA, in the injured ganglion increases rapidly after vagotomy, reaching twice the normal value at 2 days post‐crush. In contrast to this, incorporation within the uninjured colateral ganglion decreases to about 65% of normal values after 4 days. But, by 7 days, both ganglia appear to be returning to their normal levels. One phase of DNA synthesis is detected in the injured ganglion, at days 1 and 2. while two phases are found in the contralateral ganglion, at days 2 and 34.
IN VIVO SYNTHESIS OF RAPIDLY‐LABELLED RNA WITHIN THE RAT NODOSE GANGLIA FOLLOWING VAGOTOMY
Authors: Gunning P. W. , Kaye P. L. , Austin L.
Published: December 2006

Abstract— The synthesis of rapidly‐labelled RNA was studied in the nodose ganglia following unilateral vagotomy. Early changes were detected in the electrophoretic patterns of RNA from both ganglia following a crush of the right vagus. The right ganglionic response is characterized by two phases, the first involving rapid processing of rRNA with associated changes in ‘message‐like’ RNA (mlRNA). A second change, detected by 14 days, appears to involve an increase in the heterodispersity of mlRNA and decreased processing of rRNA. The left ganglion follows a very similar response to that of the right with the exception of the changes in rRNA. However, the left response lags behind that of the right by at least 1 day. We conclude that extensive changes in gene expression are occurring in both ganglia. The similarity of the responses of both ganglia suggests that axon regeneration (in the right) and collateral sprouting (in the left) may produce analogous changes in gene expression, but not in rRNA synthesis.
IMMUNOLOGICAL QUANTITATION OF GLUTAMIC ACID DECARBOXYLASE IN DEVELOPING MOUSE BRAIN1
Authors: Hadjian R. A. , Stewart J. A.
Published: December 2006

Abstract— l‐Glutamic acid decarboxylase (GAD) was isolated from bovine cerebellum and purified approx 32‐fold by a combination of DEAE‐Sephadex chromatography and gel filtration. This preparation was purified electrophoretically. Rabbit antiserum against the electrophoretically purified bovine GAD was found to react with the decarboxylase of bovine cerebellum and mouse brain. Examination of GAD enzyme specific activity at various postnatal ages of developing mouse brain showed that an initial rise in GAD activity occurs at 6 days postnatally. followed by a rapid increase in enzymatic activity which reaches a maximum at 28 days postnatally. Quantitative immunoprecipitation of mouse GAD by rabbit anti‐GAD antisera indicated that the amount of GAD per brain increases 10‐fold over the period between 1 and 28 days postnatally. This increase coincides closely with the GAD enzyme activity profile. Therefore, the increase in GAD enzyme specific activity during the postnatal development of mouse brain represents an increase in the absolute amount of GAD enzyme protein.
RELATED CHANGES IN AMOUNTS OF ACh AND ATP IN RESTING AND ACTIVE TORPEDO NERVE ELECTROPLAQUE SYNAPSES
Authors: Israel M. , Lesbats B. , Manaranche R. , Marsal J. , Mastour‐Frachon P. , Meunier F. M.
Published: December 2006

Abstract— Closely related changes in the levels of acetylcholine (ACh) and adenosine triphosphate (ATP) in the electric organ of Torpedo exist during rest and synaptic activity. The present work clarifies these relations by showing:

That both substances are involved in an oscillatory process induced by nerve stimulation.

Both substances are present in synaptic vesicles; the size of the bound pool of ACh is Ca2‐dependent and is large when Ca2+ is low. Free ACh and transmission are restored when Ca2+ is present in the incubation medium.

The amount of ATP in the tissue is also Ca2+‐dependent but is low when Ca2+ is omitted. The addition of Ca2+ to the physiological medium restores the amount of ATP in the tissue.

There is a postsynaptic release of ATP triggered by transmitter depolarization. This release was measured after single nerve impulses.

When added to the incubation medium, nucleotides strongly inhibit transmitter release. It is suggested that the postsynaptic release of ATP regulates transmitter release.
MODULATION OF CYCLIC AMP LEVELS IN THE BOVINE SUPERIOR CERVICAL GANGLION BY PROSTAGLANDIN E, AND DOPAMINE
Authors: Tomasi V. , Biondi Carla , Trevisani A. , Martini Marta , Perri V.
Published: December 2006

Abstract— Dopamine, norepinephrine, carbamylcholine and PGE1 (prostaglandin E1). increased cyclic AMP concentrations in slices of bovine superior cervical ganglia. PGF1α was less effective and neither PGE2 nor PGF2α had any effect. Dopamine and PGE, alone or in combination, did not modify low Km cyclic AMP phosphodiesterase activity. Combinations of dopamine and PGE, showed a marked synergistic effect, increasing ganglionic cyclic AMP to a much greater extent than that observed when the two compounds were tested alone. Norepinephrine (10 μM), which increased cyclic AMP as much as 10 μm‐dopamine, showed no synergistic effect when tested in the presence of PGE1 or other PGs. Phentolamine, fluphenazine and triflupromazine blocked the dopamine effect without suppressing its synergism with PGE1 Adenylate cyclase of synaptosomes isolated from the ganglia under a variety of experimental conditions appeared to be as responsive to PGE1 as the slices, but it was poorly stimulated by dopamine and was not synergistically modulated by dopamine in the presence of PGE1

These and other data are interpreted as indicating the presence of both a PGE1‐sensitive and a PGE1‐modulated dopamine‐sensitive adenylate cyclase in the cervical ganglion. These adenylate cyclases are tentatively assigned to pre‐ and post‐synaptic structures respectively.
ON THE MECHANISM OF ELECTROSHOCK‐INDUCED INHIBITION OF PROTEIN SYNTHESIS IN RABBIT CEREBRAL CORTEX
Authors: Metafora. S. , Persico Maria , Felsani A. , Ferraiuolo Rosa , Giuditta A.
Published: December 2006

Abstract— The mechanism of electroshock (ES)‐induced inhibition of protein synthesis in rabbit cerebral cortex has been investigated by using a cell‐free system. The protein biosynthetic activity of the post‐mitochondrial supernatant (PMS) obtained from the cerebral cortex of ES‐treated animals was found to be markedly lower than in controls (C). This inhibition was accompanied by a decrease of polysomes and an increase of monomers. In addition, a relative increase in light polysomes was evident at short intervals after ES treatment. No difference was found in the total soluble activity and in the activity of the elongation factors and ribonuclease present in the cell sap of C and ES animals. The biosynthetic activity of ES‐total. free and membrane‐bound ribosomes was approx 45% lower than that of the corresponding C fractions: polysome/monomer ratios were similarly reduced. The total content of cortical ribosomes was not affected by ES. Following ES treatment there was no change in the ribo‐somal ability to elongate, terminate and release polypeptide chains, nor a decrease in the polysomal content of poly(A)‐containing mRNA. These data strongly suggest that the ES‐induced inhibition of protein synthesis results from a defect in the initiation process. The possible mechanisms mediating this defect have been discussed.
NUCLEAR RIBONUCLEASE ACTIVITIES OF RAT BRAIN DURING POSTNATAL DEVELOPMENT
Authors: Ittel M. E. , Mandel P.
Published: December 2006

Abstract— The activities of alkaline and acidic RNAses were determined in soluble and insoluble fractions from nuclei of brain hemispheres of rats, aged from 1 day to adult. The activities increased rapidly and reached a maximum, at 30 days, of about 10 times (alkaline RNAsel or 5 times (acidic RNAse) that seen at day 1.
THE EFFECT OF PHOSPHOLIPASE C, PHOSPHOLIPASE A2 AND NEURAMINIDASE ON THE UPTAKE OF [3H]NOREPINEPHRINE AND [3H]SEROTONIN
Authors: Rotman A.
Published: December 2006

Abstract— The uptake of [3H]norepinephrine ([3H]NE) and [3H]serotonin ([3H]5‐HT) by rat brain synaptosomes is reduced as a result of pretreatment of the synaptosomes with phospholipase C (EC 3.1.4.3) or phospholipase A2 (EC 3.1.1.4). This effect is not due to inhibition of the Na+‐K+‐ATPase but rather is caused by hydrolysis of neuronal membrane phospholipids, mainly phosphatidylcholine, which seem to be important to the uptake.
A SUBCELLULAR POOL OF HYPO‐OSMOTICALLY RESISTANT PARTICLES CONTAINING THYROTROPIN RELEASING HORMONE, α‐MELANOCYTE STIMULATING HORMONE, AND LUTEINIZING HORMONE RELEASING HORMONE IN THE RAT HYPOTHALAMUS
Authors: Barnea Ayalla , Neaves William B. , Cho Gloria , Porter John C.
Published: December 2006

Abstract— A 10% homogenate of male rat hypothalami, prepared in 0.32M‐sucrose‐10μM‐CaCl2, was diluted either with one volume of 0.32M‐sucrose‐10μM‐CaCl2 (iso‐osmotic) or with 10μM‐CaCl2 (hypo‐osmotic). A 900 g supernatant fluid fraction (0.9 K‐S) was prepared from the diluted homogenates and fractionated on continuous sucrose density gradients under non‐equilibrium and equilibrium conditions. In the iso‐osmotic 0.9 K‐S thyrotropin releasing hormone (TRH), α‐melanocyte stimulating hormone (α‐MSH), and luteinizing hormone releasing hormone (LHRH) were each found to be sequestered in two populations of particles which were different in size but similar in density. In the hypo‐osmotic 0.9 K‐S, TRH, α‐MSH, and LHRH were each found to be sequestered in a single population of particles. In their sedimentation properties (as judged by differential, non‐equilibrium, and equilibrium density centrifugations), the hypo‐osmotically resistant particles and the small particles present in the iso‐osmotic 0.9 K‐S were identical. However, in their peptide content, the two sets of particles differed from each other. If the total quantity of particle‐bound peptides recovered after gradient centrifugation of the iso‐osmotic 0.9 K‐S is taken as 100%, one finds that the amount of TRH, α‐MSH. and LHRH recovered in the small particles is 39%, 50%, and 39%. respectively, whereas the amount of TRH, a‐MSH, and LHRH recovered in the hypo‐osmotically resistant particles is 42%, 68%, and 67%, respectively. This increase in the quantity of peptides sequestered in the small particles occurred concomitantly with the disappearance of peptides from the large synaptosome‐like particles. It is estimated that within the large synaptosome‐like particles 7% of the TRH, 35% of the a‐MSH, and 45% of the LHRH are associated with hypo‐osmotically resistant particles. Ultrastructural analysis of purified hypo‐osmotically resistant particles containing TRH. α‐MSH. or LHRH revealed a predominance of membrane‐bounded packets of electron‐dense material.
TRYPTIC PEPTIDES FROM BOVINE WHITE MATTER PROTEOLIPIDS
Authors: Chan David S. , Lees Marjorie B.
Published: December 2006

Abstract— The amino acid composition of the fractions obtained after tryptic digestion of performic acid oxidized and non‐oxidized white matter proteolipids was studied. The acid‐soluble fraction from the tryptic digest represented between 25 and 30% of the starting material and was relatively enriched in hydrophilic amino acids and deficient in hydrophobic amino acids. The acid‐soluble peptides were separated by high voltage paper electrophoresis, and the amino acid compositions of 16 peptides were determined; three additional peptides were obtained from the acid‐soluble digest of the oxidized proteolipid. The sequence of 7 peptides including the N‐ and C‐terminal peptides is reported. The results suggest that the protein is segregated into hydrophilic and hydrophobic regions and that small hydrophilic regions are separated by large hydrophobic areas.
ISOLATION AND CHARACTERIZATION OF MYELIN‐RELATED MEMBRANES
Authors: McIntyre R. J. , Quarles R. H. , Webster H. deF. , Brady R. O.
Published: December 2006

Abstract— Myelin related membrane fractions from rat brain and spinal cord were isolated from material normally discarded during standard myelin isolation procedures. A fraction which floated on 0.32 M‐sucrose (F) and the material released after subjecting the myelin fraction to osmotic shock at two stages in the purification (W1 and W2) were characterized. These fractions were subjected to subfractionation on three step discontinuous sucrose gradients. Morphologically, the heavier subfrac‐tions of W1 and W2 were shown to consist mainly of single membranes and vesicles. Sodium dodecyl sulphate (SDS) polyacrylamide gel electrophoresis showed that, relative to myelin, proteolipid and basic protein were reduced in all subfractions, while the high molecular weight proteins were increased. The specific activity of 2′,3′‐cyclic nucleotide 3′‐phosphohydrolase (CNP) was up to 2‐fold higher than that of myelin in the heavier subfractions of W1 and W2. The major myelin‐associated glycoprotein was also increased in these subfractions as determined by periodic acid‐Schiff staining. Differential centrifugation of the initial tissue homogenate to remove microsomes prior to myelin isolation gave rise to W1 and W2 subfractions with a CNP specific activity 3–4 times that of myelin. The high molecular weight proteins and glycoproteins were enriched in these microsome‐depleted subfractions, but were qualitatively similar to those of myelin. Some of the membranes in these fractions may be derived from the continuum between the plasma membrane of the oligodendrocyte and compact myelin. Fraction F consisted of small membrane fragments and many vesicles, and was particularly deficient in proteolipid. The specific activity of CNP in fraction F was about the same as myelin, while the major myelin associated glycoprotein could not be detected. Fraction F from normal CNS tissue appears to be similar to the floating fractions previously isolated in larger amounts from pathological brain undergoing edematous demyelination.
THE EFFECT OF INTRAOCULAR INJECTION OF CORDYCEPIN ON RETINAL RNA SYNTHESIS AND ON RNA AXONALLY TRANSPORTED DURING REGENERATION OF THE OPTIC NERVES OF GOLDFISH
Authors: Ingoglia Nicholas A.
Published: December 2006

Abstract— The presence of relatively large amounts of RNA has been demonstrated in regenerating axons of the goldfish optic nerve. Previous experiments have suggested that this R NA may be composed of only small molecular weight 4S RNA. The present experiments were performed in order to see if inhibiting RNA transport by intraocular injections of cordycepin causes a selective depletion of 4S RNA arriving in the contralateral optic tectum, and thus add further evidence that 4S RNA is axonally transported.

Optic nerves were crushed in a group of goldfish and 18 days later 10.0 /tg of cordycepin was injected into the right eye followed 3 h later by injections of [3H]uridine into the same eye. Six days later the amount of axonally transported [3H]RNA was decreased by 89% compared with non‐cordycepin treated controls. The effect of cordycepin on retinal RNA synthesis was shown by autoradiography to be primarily on retinal ganglion cell RNA synthesis with lesser effects on other cellular elements of the retina. SDS polyacrylamide gel electrophoresis at both 1 and 6 days after intraocular injections of cordycepin and [3H]uridine, showed that cordycepin blocks the retinal synthesis of ribosomal RNAs but appeared to have little effect on the synthesis of 4S RNA. When transported RNA in the tectum was fractionated by gel electrophoresis 6 days after injection, it was found that the amount of ribosomal RNA was decreased by approx 70% as a result of cordycepin pretreatment. This correlated well with the effect of cordycepin on the transport of available RNA precursors (also decreased by approx 70%) and is consistent with the contention that in these experiments ribosomal RNA is synthesized in the tectum itself and is not axonal. The amount of [3H] 4S RNA arriving in the tectum, however, was decreased by greater than 90% suggesting that its presence in the tectum was not entirely dependent on the availability of 3H precursors for local synthesis in the tectum.

These results are consistent with data suggesting that 4S RNA is the predominant, if not the only, RNA species axonally transported during regeneration of goldfish optic nerves.
SUBCELLULAR DISTRIBUTION AND TRANSPORT OF ACETYLCHOLINE IN CAT VENTRAL ROOTS AND SCIATIC NERVE
Authors: Waldenlind L. , Anderson P.
Published: December 2006

Abstract— The axonal transport of radioactive ACh, which was labelled by injecting radioactive choline into the ventral horn of the cat, was studied. Radioactivity analysed as ACh was transported in the nerves at a rate of 20–25 cm/24 h from the place of injection. Morphological and biochemical analysis after density gradient centrifugation revealed that endogenous ACh was principally distributed in three fractions‐(a) a soluble fraction (provided homogenization was carried out in the presence of eserine), (b) a vesicular fraction (diameter 600–1600 A) in the 0.4M‐sucrose layer of the density gradient, and (c) a fraction containing very small structures (size 90–100 A) in the 0.8–1.0 M‐sucrose interphase. The radioactive ACh, however, was exclusively found in the soluble fraction (supernatant) after density gradient centrifugation. Analysis of ACh metabolites showed that radioactive ACh may have been formed locally in the nerves after transport of its precursor. Thus the morphologic and metabolic results do not support the hypothesis that ACh is transported in vesicles.
A FLUOROMETRIC ASSAY FOR CHOLINE ACETYLTRANSFERASE and ITS USE IN THE PURIFICATION OF THE ENZYME FROM HUMAN PLACENTA
Authors: Hersh Louis B. , Coe Barbara , Casey Lynette
Published: December 2006

Abstract— A fluorometric assay for choline acetyltransferase has been developed. This assay is based on coupling the choline acetyltransferase dependent formation of acetyl‐CoA from acetylcholine and coenzyme A, to the reactions catalyzed by the enzymes citrate synthase and malic dehydrogenase. Although this assay is not as sensitive as previously described radiometric assays, it can be conveniently used during enzyme purification.

Employing this assay method, choline acetyltransferase has been purified from human placenta to a specific activity of 92.7 μmol acetylcholine formed/min/mg protein.
Ca2+ UPTAKE BY SYNAPTOSOMES AND ITS EFFECT ON THE INHIBITION OF ACETYLCHOLINE RELEASE BY BOTULINUM TOXIN
Authors: Wonnacott S. , Marchbanks R. M. , Fiol C.
Published: December 2006

Abstract— 45Ca2+ uptake by cerebral cortex synaptosomes was determined by gel filtration, glass fibre disc filtration under suction and by centrifugation with EGTA present. The filtration methods gave comparable results which were higher than values obtained by the centrifugation method. Uptake was increased by 25mM‐K+ at all times investigated. The accumulated 45Ca2+ was bound within the synaptosome. 45Ca2+‐ionophore A23187 stimulated uptake only during the first min; levels of intra‐synaptosomal 45Ca2+ then returned to control values. A23187 also increased intra‐synaptosomal Na+ and Cl− contents. Botulinum toxin inhibits the K.+‐stimulated release of [14C]ACh from synaptosomes but the ionophore released [14C]ACh from both normal and botulinum‐treated preparations in a Ca2+‐dependent manner. However, it also elicited Ca2+‐dependent release of [choline. Increased extracellular Ca2+ (10 mM and 20 mM) released [14C]ACh (but not [14C]choline) from both normal and botulinum‐treated synaptosomes. It is concluded that botulinum toxin interferes with the provision of Ca2+ essential for the mechanism of ACh release.
CEREBRAL, CEREBELLAR, AND BRAIN STEM GANGLIOSIDES IN MICE SUSCEPTIBLE TO AUDIOGENIC SEIZURES
Authors: Seyfried Thomas N. , Glaser Gilbert H. , Yu Robert K.
Published: December 2006

Abstract— The quantitative and qualitative distribution of gangliosides was investigated in the cerebrum, cerebellum and brain stem of audiogenic seizure resistant (C57BL/6J) and susceptible (DBA/2J) mice at 21 days of age. The concentration of gangliosides (μg/unit weight) was higher in the DBA cerebrum and brain stem, but lower in the DBA cerebellum compared to the concentration in C57 mice. In general, the brain water content was lower in DBA mice than in C57 mice. The distributions of a number of gangliosides were found to be different between the two strains and the differences were often in the same direction across the three brain regions. The most consistant and significant difference in ganglioside pattern observed between the strains was the higher concentration of GM1 in all three regions of the DBA brain. These results suggest that DBA mice have a more heavily myelinated CNS than C57 mice. The relationship of these observations to inherent audiogenic seizure susceptibility is discussed.
PERINATAL METHADONE EXPOSURE AND BRAIN DEVELOPMENT: A BIOCHEMICAL STUDY1
Authors: Zagon Ian S. , McLaughlin Patricia J.
Published: December 2006

Abstract— The neurochemical effect of maternally administered methadone (5 mg/kg, DL‐methadone‐HCI) on the brain (including the olfactory bulbs, cerebellum, and brain stem) and cerebellum of offspring exposed during gestation and/or lactation was studied in 10‐, 21‐, and 60‐day old rats. Brain weights were significantly reduced in all methadone‐exposed groups at 10 days of age, while only those rats subjected to methadone during gestation or lactation had deficits in brain weights at day 21; no differences were found at 60 days. Brain DNA content was significantly reduced in all opiate‐exposed offspring at every age examined, but RNA/DNA and protein/DNA ratios were only consistently increased in rats of the gestation group. Cerebellar weight was reduced at 10 days in the gestation‐lactation pups, at 21 days in rats of the gestation and lactation groups, and at 60 days in animals of the gestation and gestation‐lactation groups. Cerebellar DNA content was significantly decreased in pups of the gestation group at every age investigated, but only reduced at 21 days in the lactation group and at 60 days in the gestation‐lactation group. Rats in the lactation group had the greatest number of alterations in terms of RNA and protein, with the most noticeable being decreases in mean cellular RNA content on days 21 and 60 and a reduction in the mean cellular protein content on day 60. These data suggest that prenatal and/or postnatal methadone treatment affects the biochemical maturation of the central nervous system; deficits in neurons and/or glia, as well as a reduction in myelination, might be reflected in these changes.
UPTAKE OF [14C]ASPARTIC ACID AND [14C]GLUTAMIC ACID BY RETINAL SYNAPTOSOMAL FRACTIONS
Authors: Thomas Thomas N. , Redburn Dianna A.
Published: December 2006

Abstract— Uptake systems for [14C]aspartate and [14C]glutamate were characterized in two distinct synaptosomal fractions solated from rabbit retina. The P, synaptosomal fraction was highly enriched in large photoreceptor cell synaptosomes but contained very few conventional sized synaptosomes from amacrine, horizontal or bipolar cells. In contrast, the P2 synaptosomal fraction contained numerous conventional sized synaptosomes and was virtually free of photoreceptor cell synaptosomes. Both synaptosomal fractions took up [14C]aspartate and [14C]glutamate with high affinity [Km= 1–2μM). Uptake characteristics were similar to those described for high affinity uptake systems in brain synaptosomes, i.e. saturation kinetics; temperature and Na+ dependence. Although the presence of a high affinity uptake system is not a definitive criterion for demonstration of functional neurotransmitter systems, it is an important and necessary prerequisite and can thus be considered as supportive evidence for the involvement of asparate and glutamate in neurotransmission in rabbit retina.
STEADY STATE KINETICS OF RAT BRAIN PYRUVATE DEHYDROGENASE MULTIENZYME COMPLEX
Authors: Ngo T. T. , Barbeau A.
Published: December 2006

Abstract— The overall steady state kinetic mechanism of pyruvate dehydrogenase multienzyme complex purified from rat brain has been investigated. Initial rate patterns were a series of parallel lines regardless of which substrate was varied at several fixed concentrations of other substrates. Product inhibition patterns showed that acetyl CoA is competitive vs CoA, that NADH is competitive vs NAD, and that both acetyl CoA and NADH are uncompetitive vs pyruvate. Both acetyl CoA and NADH are noncompetitive vs NAD and CoASH, respectively. These results are inconsistent with classical ‘hexa uni’ ping‐pong mechanisms, but are consistent with a non‐classical 3‐site ping‐pong mechanism.
METHYLATION OF E. COLI TRANSFER RIBONUCLEIC ACIDS BY A tRNA ADENINE‐l‐METHYLTRANSFERASE FROM RAT BRAIN CORTEX AND BULK‐ISOLATED NEURONS
Authors: Salas Carlos E. , Sellinger Otto Z.
Published: December 2006

Abstract— Brain cortices or bulk‐isolated neuronal cell bodies prepared from cortices of 8‐day old male rats were used as the source of a l‐methyl adenine‐specific tRNA methyltransferase (tRNA‐AMT). Ammonium sulfate fractionation and chromatography on spheroidal hydroxylapatite and Sephadex G‐200 yielded an 80‐fold purified enzyme, as determined by using E. coli bulk tRNA as substrate. The kinetic parameters of tRNA‐AMT for the substrate S‐adenosyl‐l‐methionine (SAM) (Km= 6 μM) and the inhibitor, S‐adenosyl‐l‐homocysteine (SAH) (Ki= 3.4 μm) were determined and several SAH analogs tested as inhibitors. S‐Adenosyl‐l‐cysteine (SAC) (10‐4m) and S‐adenosyl‐d‐homocysteine (SADH) (10‐4m) produced a 35 and a 21% reduction in enzyme activity, respectively. The effects of Mg2+, NH4+ acetate and of the polyamines spermine, putrescine and spermidine on the brain tRNA‐AMT mimicked the effects of these agents on hepatic tRNA‐AMT (Glicket al, 1975).

Comparing the ability of cerebral tRNA‐AMT to methylate E. coli tRNAglu2, tRNAval, tRNAphe and bulk tRNA revealed tRNAglu2 as the best and tRNAphe as the least effective substrate.

tRNA‐AMT prepared from neuronal cell bodies showed closely similar characteristics to the cortical enzyme. A comparison of the activities of tRNA‐AMT in neurons and glial cells revealed higher values in the former.
STUDIES ON THE LOW‐MOLECULAR WEIGHT GLYCOPROTEIN GP‐350: MOLECULAR AND IMMUNOLOGICAL HETEROGENEITY
Authors: Simonian S. , Heijlman J. , Hooghwinkel G. J. M.
Published: December 2006

Abstract— GP‐350 was isolated from the water soluble cell fraction of bovine brain and liver. The isolated protein preparations were electrophoresed in the presence of SDS in 19% polyacrylamide gels and in the absence or presence of Triton X‐100 and urea in 7.5% polyacrylamide gels. These experiments show that the GP‐350 protein fraction from the different tissues behaves as a class of low‐molecular weight proteins with different intrinsic charges. The majority of the protein bands which were resolved in the presence or absence of Triton X‐100 and urea in 7.5% polyacrylamide gels were not reactive with the antiserum directed against the total GP‐350 protein fraction.

Moreover, on gel chromatography in Sephadex G‐50, GP‐350 was fractionated into several peaks. The reactivity with the GP‐350 antiserum in double immunodiffusion was present primarily in the major peak with a molecular weight between 9500 and 11,500; this peak gave three precipitin lines. Furthermore, lipid analysis of GP‐350 has shown that GP‐350 protein preparations from brain contained about 17% (w/w) choloroform‐methanol (2:Insoluble lipids. The lipids were for the major part of neutral type and only trace amounts of glycolipids were detectable. The lipid‐free GP‐350 protein was immunologically identical to the total GP‐350 fraction.

On the basis of this heterogeneity in charge, molecular composition and immunological properties we conclude that GP‐350 is a mixture of low‐molecular weight protein and lipid constituents.
RIBOSOMAL ACTIVITY IN PRENATAL MOUSE BRAIN
Authors: Gilbert Brian E.
Published: December 2006

Abstract— Regulation of protein synthesis is important for the proper growth and development of the brain. Our previous work on the regulation of protein synthetic activity in fetal mouse brain cell suspensions showed that the rate of protein synthesis decreased during the prenatal period. In the present study, ribosomal activity of cell‐free homogenates and purified ribosomes obtained from fetal neural tissue was measured. The post‐mitochondrial supernatant (PMS) fraction actively incorporated amino acids into polypeptides using either endogenous mRNA or polyuridylic acid as template. The protein synthetic activity was dependent upon the age of the fetus. Ribosomes purified from this fraction were also active in protein synthesis. Incorporation of phenylalanine was linear for 20 min, and dependent upon the concentration of ribosomes and the pH 5 enzyme fraction. The age dependent decrease in protein synthetic activity observed with the post‐mitochondrial supernatant fractions was not found when these purified ribosomes were employed. Ribosomes obtained from fetal, newborn or adult neural tissue were compared and found equally active in their protein synthetic capacity.
EFFECTS OF TETRODOTOXIN, CALCIUM AND MAGNESIUM ON THE RELEASE OF AMINO ACIDS FROM SLICES OF GUINEA‐PIG CEREBRAL CORTEX
Authors: Potashner S. J.
Published: December 2006

Abstract— Tetrodotoxin, Ca2+‐deprivation and high‐Mg2+ were used in an effort to identify the portion of the evoked release of endogenous amino acids, labelled via metabolism of [14C]‐glucose, and several exogenous labelled amino acids, that came from nerve terminals when slices of guinea pig cerebral cortex were superfused with glucose‐free solutions and stimulated electrically.

With some exceptions, spontaneous release of labelled amino acids was decreased by 2 μm‐tetrodotoxin but increased in Ca2+‐free medium and in solutions containing an extra 24 mm‐MgCl2. Tetrodotoxin suppressed 85–90% of the stimulated release of almost all labelled amino acids, but had a smaller effect on the release of endogenous 14C‐labelled threonine‐serine‐glutamine (unseparated). In Ca2+‐free solution, the stimulated release of endogenous 14C‐labelled glutamate, aspartate and GABA was suppressed by 80–90%, but that of endogenous 14C‐labelled threonine‐serine‐glutamine was unaffected as was most of the release of the other labelled amino acids. In medium containing an extra 24mM‐MgCl2, the stimulated release of endogenous 14C‐labelled glutamate, aspartate and GABA was suppressed by 75‐85%, that of exogenous labelled aspartate and GABA by 50–65%, but the release of the other labelled amino acids was unaffected. The control stimulated releases of endogenous 14C‐labelled glutamate, aspartate and GABA were much larger than those of other labelled amino acids but were reduced by tetrodotoxin, Ca2+‐deprivation and high‐Mg2+ to a level similar to that of the control stimulated releases of the other labelled amino acids.

These results suggest that almost all of the stimulated release of endogenous 14C‐labelled glutamate, aspartate and GABA came from nerve terminals while those of the other labelled amino acids came from other tissue elements. In addition, they are in accord with a transmitter role for glutamate, aspartate and GABA in cerebral cortex.