bluuebunny/crossref_metadata_2025 huggingface dataset:
SYNAPTOSOMAL PLASMA MEMBRANES. ACYL GROUP COMPOSITION OF PHOSPHOGLYCERIDES and (Na++ K+)‐ATPase ACTIVITY DURING FATTY ACID DEFICIENCY
Authors: Sun G. Y. , Sun A. Y.
Published: December 2006

Abstract— Essential fatty acid deficiency was induced in mice after feeding a fatty acid deficient diet for 6 months. Activity of the (Na++ K+)‐ATPase in the total brain homogenates and in isolated synaptosomal plasma membranes was significantly higher (P & lt; 0 05) in the deficient mice than the controls. Analysis of the acyl group composition of phosphoglycerides in brain as well as in the synaptosomal plasma membranes showed that mice fed the deficient diet had increased levels of 20:3(n‐9) and 22:3(n‐9) and decreased levels of 20:4(n‐6) and 22:4(n‐6). However, acyl group changes varied among individual phosphoglycerides and were most obvious in the two species of ethanolamine phosphoglycerides. A decrease in 22:6(n‐3) level was also observed in some phosphoglycerides of the synaptosomal plasma membranes especially the diacyl‐sn‐glycerophosphorylserine. In this experiment, a new solvent system for chromatographic separation of the diacyl‐sn‐glycerophosphorylserine and diacyl‐sn‐glycerophosphorylinositol was reported. The separation technique was suitable for analysis of acyl group composition of individual phosphoglycerides by gas‐liquid chromatography. The results were consislent with a positive correlation of the non‐polar acyl groups of brain membranes with the active ion transport activity. The increase in enzymic activity during deficient state may be the result of a biological adaptation due to structural alteration of the brain membranes.
THE EFFECT OF SLEEP DEPRIVATION UPON THE IN VIVO AND IN VITRO INCORPORATION OF TRITIATED AMINO ACIDS INTO BRAIN PROTEINS IN THE RAT AT THREE DIFFERENT AGE LEVELS
Authors: Bobillier P. , Sakai F. , Seguin S. , Jouvet M.
Published: December 2006

Abstract— The effect of sleep deprivation on the in vivo and in vitro tritiated amino acid incorporation into brain proteins was studied in the rat at three age levels. Sleep deprivation was induced either by water tank or handling methods. Three experimental groups of animals were used: control, sleep deprived and post deprivation sleeping rats.

A significant decrease of protein synthesis was found in the cerebellum, telencephalon and in crude subcellular fractions of brainstem of adult rats selectively deprived of paradoxical sleep. However, no alteration of protein synthesis was observed either in vivo or in vitro, in the same brain regions or in the liver after the rebound of paradoxical sleep following deprivation.

In four crude subcellular protein fractions a specific increase of the in vitro labelled amino acid incorporation was observed in the brain stem of 24‐day‐old rats allowed to recuperate after sleep deprivation as compared with the deprived rats. No significant changes were seen in the telencephalon.

No alteration of incorporation was found in 7‐day‐old rats deprived of sleep.

The possible functional significance of these results is discussed in relation to stress and to variations in the size of the precursor pool for protein synthesis.
FACTORS AFFECTING THE SPONTANEOUS RELEASE OF [3H] γ‐AMINOBUTYRIC ACID FROM THE FROG RETINA IN VITRO
Authors: Kennedy A. J. , Voaden Mary J.
Published: December 2006

Abstract— The spontaneous efflux of [3H]GABA and its metabolites from the frog retina has been studied. The efflux of radioactivity was multiphasic in the presence or absence of amino‐oxyacetic acid (AOAA), an inhibitor of GABA metabolism, and was not affected by light or dark adaptation.

Strong retention of radioactivity was evident in the presence of AOAA, about 90% of the label remaining in the tissue after 4 h superfusion. Under these conditions, increases in the rate of release of radioactivity were evoked by electrical stimulation, 40 mm‐potassium. unlabelled GABA (5 mm), ouabain (5 × 10−5m) and the absence of calcium. The amount of [3H]GABA released by electrical stimulation was not markedly calcium dependent, whereas the response to 40 mm‐potassium was reduced by 96% in the absence of calcium.
THE DISTRIBUTION OF MUSCARINIC RECEPTOR SITES IN THE NERVOUS SYSTEM OF THE DOG
Authors: Hiley C. R. , Burgen A. S. V.
Published: December 2006

Abstract— The concentration of muscarinic receptors has been measured in 22 areas of the dog nervous system by measuring the atropine‐sensitive uptake of tritium‐labelled propylbenzilylcholine mustard. The highest concentration of receptor was found in the caudate nucleus, intermediate concentrations were found in five areas of cerebral cortex, the other basal ganglia and the superior colliculus. Significant concentrations were found in the corpus callosum and subcortical white matter, and are believed to be on axons derived from cholinoceptive neurons. The results are discussed in relation to other evidence concerning cholinergic transmission in the nervous system.
PROTEOLIPIDS IN THE DEVELOPING BRAINS OF NORMAL MICE AND MYELIN DEFICIENT MUTANTS1
Authors: Lerner Pauline , Campagnoni A. T. , Sampugna J.
Published: December 2006

Abstract— A developmental study of proteolipids from brains of normal mice and two myelin deficient mutants, jimpy and quaking, was performed. The proteolipids were obtained by diethyl ether precipitation of washed total lipid extracts from whole brains and were analysed on polyacrylamide gels containing sodium dodecyl sulphate. The amount of ether precipitable material extractable from normal brains increased almost six‐fold between 12 and 21 days posr partum. This increase was not observed with the mutant mice. Polyacrylamide gel electrophoretic analysis of the proteolipid fraction showed it to be heterogeneous, with eight major protein bands. Two of these proteins increased rapidly in quantity in normal mice between 13 and 21 days. These two proteins were present, in severely reduced quantities in the brains of jimpy and quaking mice at all ages examined. One of these proteolipids was the major species present in proteolipid extracts from the brains of normal mature mice. This protein coelectrophoresed with proteolipid isolated from purified myelin and has been tentatively identified as the myelin proteolipid. The other proteolipid which was deficient in jimpy and quaking brains was not characterized, but it appeared to be of extra‐myelin origin, and suggests that parts of the brain other than the myelin sheath may be involved in the jimpy and quaking disorders.
THE EFFECTS OF SOLUBILIZATION PROCEDURES ON THE RELEASE AND MOLECULAR STATE OF ACETYLCHOLINESTERASE FROM ELECTRIC ORGAN TISSUE
Authors: Dudai Y. , Silman I.
Published: December 2006

Abstract— A study was made of the effect of various solubilization procedures on the release of AChE from electric organ tissue of the electric eel and on the molecular state of the enzyme. The procedures employed included homogenization in different ionic media or in the presence of detergents, etuymic treatment and chemical modification. Studies were performed on intact electroplax, tissue homogenates and membrane fractions. The apparent AChE activity of intact cells, homogenates and membrane fractions was shown to be governed by diffusion‐controlled substrate and hydrogen ion gradients, generated by AChE‐catalyscd hydrolysis, leading to a lower substrate concentration and a lower pH in the vicinity of the particulate enzyme.

Treatment of homogenates with NaCl solutions or with NaCl solutions containing the nonionic detergent Triton X‐100 causes release of the native’molecular forms of the enzyme (primarily the 18 S species) which aggregate at low ionic strength. For optimal extraction both high ionic strength (e.g. 1 M‐NaCl) and the detergent are needed AChE is also solubilized by treatment of tissue homogenates with trypsin, bacterial protease or collagenase. The first two enzymes caused its release as an 11 S non‐aggregating form, while collagenase also produces a minor non‐aggregating ‐ 16 S component. Treatment of tissue homogenates with maleic anhydride causes release of AChE as a non‐aggregating 18 S species. On the basis of the solubilization experiments it is concluded that the interaction of AChE with the excitable membrane is primarily electrostatic. The possible orientation of the enzyme within the synaptic gap is discussed.
CHOLINE ACETYLTRANSFERASE ACTIVITY IN GUINEA‐PIG HEART IN VITRO
Authors: Jr. R. Roskoski. , Mayhr H. K. , Schmid P. G.
Published: December 2006

Abstract— Choline acetyltransferase (EC 2.3.1.6) catalyses the following reversible reaction: acetyl coenzyme A + choline acetylcholine + coenzyme A. Enzyme activity in the atria and ventricles of guinea‐pig heart varied independently of the biochemically related carnitine acetyltransferase (EC 2.3.1.7). Choline acelyltransferase activity was greatest in right atrium, intermediate in right ventricle and left atrium and lowest in left ventricle (405. 2‐33. 177 and I 33 nmol min‐1 g‐1, respectively). Carnitine acetyltransferasc activity was greatest in the right and left ventricle and least in the right and left atria (8‐86. 8‐27, 3‐18 and 2‐38 mmol min‐1g‐1. respectively). Carnitine acelyltransferase activity was 800‐ to 6000‐fold greater than that of the choline acetyltransferase. depending on the chamber. Bromoacctylcholine inhibited acetylcholine. but not acetylcarnitine biosynthesis in vitro. Contrariwise, acetylcarnitine inhibited carnitine, but not choline acetyltransferase. These results demonstrate the feasibility and specificity of measuring the differences in choline acetyltransferase activity in dialysed homogenates prepared from the four chambers of the heart.
α‐AMINOCYCLIC AND BICYCLIC ALKANE CARBOXYLIC ACIDS: DIFFERENTIAL EFFECTS ON SELECTED AMINO ACIDS OF RAT BRAIN CORTEX1
Authors: Zand R. , Sellingr O. Z. , Water R. , Harris R.
Published: December 2006

Abstract— The intraperitoneal administration of 1‐aminocyclopentane carboxylic acid, 1‐aminocyclohex‐ane carboxylic acid, l‐aminocycloheptane carboxylic acid, 1‐aminocyclooctane carboxylic acid, exo‐2‐aminobicyclo(2,2. l)heptane‐2‐carboxylic acid. endo‐2‐aminobicyclo(2,2.1)heptane‐2‐carboxylic acid. 2‐aminobicyclo(2.2.2)octane‐2‐carboxylic acid and 2‐aminobicyclo(3,2.l)octane‐2‐carboxylic acid to 18‐day‐old male rats selectively perturbed the levels of neutral amino acids in the cerebral cortex. While the effect of the above compounds was rather diversified and usually resulted in a reduction of amino acid levels. marked elevations of the levels of valine and isoleucine were also noted. 1‐Aminocycloheptane and cyclooctane carboxylic acids were particularly noteworthy, in that they elicited a marked reduction of the levels of cortical phenylalanine.
THE RELATIONSHIP BETWEEN SODIUM INFLUX AND VOLUME FLOW INTO THE CEREBRAL VENTRICLES OF CATS
Authors: Wald A. , Hochwald G. M. , Malhan Chaman
Published: December 2006

Abstract— The relationship between sodium ion (Na+) influx and vol. flow of fluid into the cerebral ventricles was measured during ventriculocisternal perfusion with sucrose solutions of various concentrations. The vol. flow of fluid into the ventricles of cats varied linearly from 0 to 90 μl/min with sucrose solutions of 6 to 780 mOsm/l. In the vol. flow range of 0 to 35 μl/min, Na+ influx was essentially constant independent of vol. flow rate with a mean value of 6.95 μEq/min. In the vol. flow range of 25 to 90 μl/min, Na+ influx increased linearly with flow rate. Under all conditions, Na+ influx was greater than that corresponding to newly formed fluid with a normal spinal fluid Na+ concentration. The virtual Na+ concentration of nascent fluid was effectively infinite when vol. flow was zero and had an asymptotic minimum value of 109 mEq/l as vol. flow increased above normal. These results demonstrate that Na+ influx into the ventricles may occur by diffusion from the surrounding brain and also with vol. flow of nascent fluid.
METABOLISM OF BRAIN SPHINGOMYELINS: HALF‐LIVES OF SPHINGOSINE, FATTY ACIDS AND PHOSPHATE FROM TWO TYPES OF RAT BRAIN SPHINGOMYELIN
Authors: Freysz L. , Lastennet A. , Mandel P.
Published: December 2006

Abstract— The metabolism of rat brain sphingomyelins containing short‐chain (C16‐C18) and long‐chain (C20‐ C24) fatty acids has been studied by determination of the content of radioactivity in the sphingo‐sinc. fatty acids and phosphate of the sphingomyelins over a period of 60 days following the intracisternal injection of [14C]acetate and [32P]phosphate. From the rate of decrease of the specific radioactivities of the different constituents of short‐chain fatty acid sphingomyelins, we have calculated a half‐life of 65 days for sphingosine. 41 days for fatty acids and 62 days for phosphate. For the long‐chain fatty acid sphingomyelins the half‐life of sphingosine was approximately 465 days. The fatty acids and phosphate from these sphingomyelins had fast and slow turnover pools. The half‐life for the fast pool was 7 days for the two constituents and the estimated half‐lives for the slow pool were 220 days for fatty acids and 480 days for phosphate.

These results suggest that one can distinguish at least three metabolic pools of brain sphingomyelins: (a) sphingomyelins with long‐chain fatty acids situated in myelin whose half‐lives are 465 days for sphingosine, 220 days for fatty acids and 480 days for phosphate; (b) sphingomyelins with long‐chain fatty acids located mainly in non‐myelin structures having half‐lives of 465 days for sphingosine. 7 days for fatty acids and 7 days for phosphate; (c) sphingomyeiins with short‐chain fatty acids with half‐lives of 65 days for sphingosine. 41 days for fatty acids and 62 days for phosphate. The differences between the half‐lives of the three metabolic pools of sphingomyelin, together with the subcellular localizations of the two molecular species of these compounds, suggest that the metabolism of the different molecular species of sphingomyelin are independent and that in various subcellular fractions the long‐chain fatty acid and short‐chain fatty acid sphingomyelins have different turnover rates.
IMMUNOCHEMICAL STUDIES OF THE INTERACTION BETWEEN MYELIN BASIC PROTEIN AND S‐100 PROTEIN 1
Authors: Mahadik S. P. , Graf L. , Rapport M. M.
Published: December 2006

Abstract— The formation of a complex between myelin basic protein and S‐100 protein was detected from the change in migration of S‐100 protein on immunoelectrophoresis. A degree of specificity for the interaction was shown by two observations: (1) two other pure acidic proteins. III‐III‐2 and bovine serum albumin, did not show it and (2) complex formation was dependent on specific ions, either Ca2+ (10 mM) or Mn2+ (1 mM). Mg2+, Ba2+, and Li+ had no effect. Non‐specific interactions between S‐100 protein and other basic molecules (histones. polylysine) are not dependent on specific ions such as Ca2+ and Mn2+. The complex was stable at physiological salt concentrations and contained 3 mol of basic protein per mol of S‐100 protein. Complex formation was also detected from the alteration of migration rate of S‐100 protein in polyacrylamide gels. Serological activity (complement‐fixation) of S‐100 protein with anti‐S‐100 serum was reduced in the complex by 30%.
INFLUENCE OF COLD EXPOSURE ON THE ACTIVITIES OF SOLUBLE AND MEMBRANE‐BOUND DOPAMINE β‐HYDROXYLASE IN RAT HEART VESICLES
Authors: Brodde O.‐E. , Huvermann K. , Schümann H. J.
Published: December 2006

Abstract— A fraction containing noradrenaline storage vesicles of the sympathetic nerve terminals in the rat heart was obtained by differential centrifugation. In this preparation, 17% of the dopamine β‐hydroxylase was present in a soluble form. Cold exposure (3°C) for periods from 5 to 30 min led to an increase in the activity of soluble dopamine β‐hydroxylase by about 50%, while the activity of membrane‐bound dopamine β‐hydroxylase was simultaneously decreased by approx 30%. The nor‐adrenaline content of the vesicles rose concomitantly with the increase in the activity of soluble dopamine β‐hydroxylase. This rise in noradrenaline content was caused by an enhanced synthesis and not by an alteration in the subcellular distribution. The results are discussed with respect to the fate of dopamine β‐hydroxylase during enhanced sympathetic nerve activity.
CATABOLISM OF PROSTAGLANDIN ENDOPEROXIDES INTO PROSTAGLANDIN E2 AND F2x BY THE RAT BRAIN
Authors: Pace‐Asciak C. , Nashat M.
Published: December 2006

Abstract— Tritium labeled prostaglandin (PG) endoperoxides PGG2 and PGH2 were rapidly transformed (2 min, 37°C) in good yield (> 50%) by homogenates of whole rat brain into a mixture of products including prostaglandin E2 and F2x: under similar conditions (10min. 37°C) tritium labeled arachidonic acid remained essentially unoxidised. The ratio of PGE‐like products: PGF2x formed was approx 0.5 as determined by radio thin layer chromatography. This ratio changed to unity when homogenates of cerebral cortex or cerebral hemispheres were employed. On the other hand cerebellar homogenates formed PGF2x in much greater amounts. The structures of the products were confirmed by mass spectrometry and were further supported by experiments using octadeuterio‐endoperoxides. In the latter experiments the resulting PGE2 and PGF2x contained the expected seven and eight deuterium atoms respectively. Evidence for the formation of heptadeuterio PGD2. heptadeuterio‐6‐keto‐PGF1, and hexadeuterio 12‐hydroxyheptadecatrienoic acid was also obtained by mass spectrometry. These experiments demonstrate for the first time in brain tissue the biosynthesis of labeled prostaglandins from exogenous tritiated and deuterated precursors.
A MODULATING ROLE OF PITUITARY‐ADRENAL AXIS IN CEREBRAL METABOLISM OF ADENOSINE 3′,5′‐MONOPHOSPHATE
Authors: Nakagawa K. , Kuriyama K.
Published: December 2006

Abstract— The effect of adrenalectomy or hypophysectomy on the metabolism of adenosine 3′,5′‐monophosphate (cyclic AMP) in the cerebral cortex of male Wistar rats was investigated.

The bilateral removal of adrenal glands reduced significantly the activity of cerebral adenylate cyclase [EC 4.6.1.1]. whereas that of cyclic 3′.5′‐nucleotide phosphodiesterase [EC 3.1.4.17] remained unchanged. The formation of cyclic AMP measured in cerebral cortical slices from adrenalectomized or hypophysectomized rats was also diminished. Decreases in the activity of adenylate cyclase and formation of cyclic AMP following adrenalectomy were antagonized by in vivo administration of dexamethasone or aldosterone, while those observed in hypophysectomized rats were restored by ACTH or dexamethasone. It is suggested that the pituitary adrenal axis has a modulating role in the metabolism of cerebral cyclic AMP, possibly by changing adenylate cyclase activity.
PHOSPHOLIPASE A IN HUMAN BRAIN: A1‐TYPE AT ALKALINE pH
Authors: Rooke J. A. , Webster G. R.
Published: December 2006

Abstract— A novel phospholipase A1, present in human cerebral cortex and active at pH 9.25 upon ultrasonicated phosphatidylethanolamine is described. It has been purified 39‐fold from acetone‐dried powders of cortex by ammonium sulphate fractionation at 25% saturation, followed by Sephadex G150 gel filtration. A molecular weight of approx 500,000 has been found by Sepharose 6B gel filtration. The enzyme was slightly stimulated by 2 mM‐Ca2+ and was inhibited by 1 mM‐Hg2+ and by all detergents tested. The enzyme hydrolysed phosphatidylcholine at 18% of the rate for phosphatidylethanolamine, lysophospholipids to a lesser degree and neutral lipids not at all. There appeared to be a preference for fluid phosphatidylethanolamine substrates. The similarities of the enzyme to other phospholipases A1 are discussed.
REGIONAL, CELLULAR AND SUBCELLULAR DISTRIBUTION OF CALCIUM‐ACTIVATED CYCLIC NUCLEOTIDE PHOSPHODIESTERASE AND CALCIUM‐DEPENDENT REGULATOR IN PORCINE BRAIN1
Authors: Egrie J. C. , Campbell J. A. , Flangas A. L. , Siegel F. L.
Published: December 2006

Abstract— Cyclic nucleotide phosphodiesterase activity (calcium‐dependent and calcium‐independent) and CDR (calcium‐dependent regulator protein of phosphodiesterase) are present in all ten brain regions examined, with the specific activity of both being highest in areas predominating in grey matter and lowest in areas consisting largely of white matter. Fractionation of cerebral cortex into neuronal and glial perikarya shows that cyclic nucleotide phosphodiesterase and CDR are present at approximately equal levels in both cell types. Subcellular fractionation reveals that calcium‐sensitive enzyme specific activity, as well as CDR are highly localized in the 100,000 g supernatant fraction. Regional, cellular and subcellular distribution studies indicate that the distribution of CDR closely parallels the distribution of enzyme specific activity, suggesting that the levels of cyclic nucleotide phosphodiesterase and CDR may be synchronized.
EFFECT OF ZUCLOMIPHENE ON CHOLESTEROL FORMATION IN THE DEVELOPING CENTRAL NERVOUS SYSTEM OF THE RAT
Authors: Ramsey R. B. , Fredericks M. , Fischer V. W.
Published: December 2006

Abstract— The effect of zuclomiphene, a hypocholesterolemic agent, on developing rat CNS cholesterol biosynthesis was examined. Sterol content and composition was studied in relation to age in four regions of the CNS, cerebrum, brain stem, spinal cord and cerebellum. Sterol content of all four regions was slightly lower in drug‐treated animals than in controls. Brain stem and spinal cord were more susceptible to the effects of zuclomiphene than were cerebrum and cerebellum. Drug treatment resulted in the accumulation of desmosterol and zymosterol (5x‐cholesta‐8,24‐dien‐3β‐ol) in all CNS regions. After 15 days of drug treatment, desmosterol constituted more than 50% of the total sterol in the four examined regions. Six to 9% of the total sterol was zymosterol.

Examination by electron microscopy indicated only minimal morphological changes. Occasionally, neuronal membranous cytoplasmic inclusion bodies were evident.
IN VIVO METABOLISM OF [G‐3H]LIGNOCERIC ACID IN DEVELOPING RAT BRAIN1
Authors: Akanuma Hiroshi , Kishimoto Yasuo
Published: December 2006

Abstract— [G‐3H]Lignoceric acid (tetracosanoic acid) was injected into the brains of 20‐day‐old rats, and the animals were killed after 8, 24, or 72 h. Various lipids were isolated from these brains, and the distribution of radioactivity was determined. The injected free acid rapidly disappeared, and the radioactivity was incorporated into varying chain‐length nonhydroxy‐ and hydroxy saturated fatty acids of sphingolipids and phospholipids. Little radioactivity was found in unsaturated acids, sphingo‐sine, and cholesterol. A time‐dependent shift of the label among various fatty acids was relatively small 8 h after injection, probably because of the metabolic stability of the brain sphingolipids. In cerebrosides, the radioactivity was equally distributed between nonhydroxy and x‐hydroxy fatty acids of all chain lengths. C23 and C22 fatty acids contained equal total radioactivities; C23 and C24 fatty acids contained similar specific activities. These results confirm the significant role of a‐hydroxylation and 2‐oxidation in the synthesis of very long‐chain fatty acids in brain. In total lipid fatty acids, docosanoic acid (22:0) contained more radioactivity than its α‐oxidation precursor, α‐hydroxytricosanoic acid (23h:0) at all times. In sphingolipid fatty acids, the specific activity of 21:0 was always higher than that of its ct‐oxidation precursor 22:0. These observations indicate that part of the 22:0 and 21:0 was derived by β‐oxidation from the injected lignoceric acid or its α‐oxidation product, respectively.
DIFFERENTIATION OF DOPAMINERGIC AND NORADRENERGIC NEURONS IN RAT SPINAL CORD
Authors: Commissiong John W. , Galli Corrado L. , Neff Norton H.
Published: December 2006

Abstract— Norepinephrine (NE), dopamine (DM) and 3‐methoxy‐4‐hydroxyphenylacetic acid (HVA) content have been measured in different parts of rat spinal cord and cerebellum by a gas chromatographic mass spectrometric method. In cerebellum, which does not contain dopaminergic neurons, the ratio of NE to DA content was 47, whereas in parts of the spinal cord this ratio varied between 11 and 19. In the cord after desipramine (25 mg/kg, i.p.) plus 6‐hydroxydopamine (6‐HDA, 100/jg intracisternally), there was a significant depletion of DM but not of NE. Conversely, after benztropine (25 mg/kg, i.p.) plus 6‐HDA there was a significant depletion of NE but not of DM. Chlorpromazine (10 mg/kg, i.p.) or clozapine (25 mg/kg, i.p.) caused a significant increase in spinal cord HVA concentration 1 h after treatment. Evidence is presented which suggests that the increased HVA measured in the cord did not originate in the brain. After electrolytic lesion of the locus coeruleus there was a significant reduction of NE but not of DM. Spinal cord DM and NE were depleted by reserpine in a dose‐dependent manner, the threshold dose for DM depletion being less than that for NE depletion. Seven days after cord transection at T10 spinal cord DM was significantly reduced in the lumbar region. These results suggest that dopaminergic neurons exist in rat spinal cord independently of noradrenergic neurons and that the DM is likely to be present in the terminals of descending axons.
CHLOROTETRACYCLINE AS AN INDICATOR OF THE INTERACTION OF CALCIUM WITH BRAIN MEMBRANE FRACTIONS
Authors: Carvalho C. A. M.
Published: December 2006

Abstract— The fluorescence of chlorotetracycline (CTC) in the presence of synaptosomes isolated from sheep brain is selectively increased by Ca2+ under conditions in which Mg2+, Na+, K+, Li+ or choline have only a small effect. The monovalent cations release bound Ca2+ from synaptosomes, and this effect is reflected by a decrease in the CTC fluorescence. Under optimal conditions there is a near parallelism between Ca2+ and CTC binding to the synaptosomes membranes, and Li+ is the monovalent cation tested which interferes the most with the binding of both substances. These results obtained in a predominantly sucrose medium become less distinct when media simulating physiological composition are utilized, which limits the usefulness of the method. Brain mitochondria and myelin also bind Ca2+ and CTC. The ratio of the fluorescence signal (or CTC bound) to Ca2+ bound is highest of all for mitochondrial membranes, and the apparent fluorescence quantum yield of CTC is also the highest in these membranes, which suggests that the Ca2+ in these membranes is localized in a more apolar region than is the case for synaptosomes and myelin.
EFFECT OF FATTY ACIDS ON RAT BRAIN PARTICULATE HEXOKINASE
Authors: Domaska‐Janik Krystyna , Broniszewska‐Ardelt Barbara , Wroablewski Jarda T.
Published: December 2006

Abstract— The effect of free fatty acids on rat brain particulate hexokinase was studied in vitro. Hexokinase bound with brain mitochondrial fraction was found to be sensitive to the action of free fatty acids, resulting in the solubilization of at least part of bound enzyme activity into the supernatant. The decrease of total enzyme activity observed at the highest free fatty acid concentration was probably due to the inhibition of hexokinase. The physiological consequence of hexokinase solubilization by low concentrations of free fatty acids, similar to that observed in vivo, is discussed in relation to activity changes of soluble and particulate enzyme forms demonstrated previously under hypoxic conditions.
GABA‐TRANSAMINASES OF HUMAN BRAIN AND PERIPHERAL TISSUES—KINETIC AND MOLECULAR PROPERTIES
Authors: White Helen L. , Sato Takao L.
Published: December 2006

Abstract— Kinetic experiments with 4‐aminobutyrate‐2‐ketoglutarate transaminase (GABA‐T), partially purified from human brain tissue, supported a Bi Bi Ping‐Pong type of enzyme mechanism in which the enzyme oscillates between forms bound to pyridoxal phosphate and pyridoxamine phosphate. Extrapolated Km values were 0.31 mm for γ‐aminobutyrate, 0.16 mm for α‐ketoglutarate, and 3.8 μm for pyridoxal phosphate. Very similar kinetic parameters were observed with rat brain enzyme. Apparent molecular weight of human GABA‐T by gel filtration was 70,000 ± 3000. Electrofucusing experiments indicated a single ionic form with isoelectric pH = 5.7. Enzyme activity was inhibited by Tris, halides, cadmium and cupric ions, and known GABA‐T inhibitors.

GABA‐transaminating enzymes isolated from human kidney and liver were found to be similar to the brain enzyme with respect to substrate affinities, cofactor requirements, isoelectric pH values, molecular weights, and response to inhibitors.
DIHYDROPTERIDINE REDUCTASE MAY FUNCTION IN TETRAHYDROFOLATE METABOLISM
Authors: Pollock Robert J. , Kaufman Seymour
Published: December 2006

Abstract— The tetrahydrofolate‐dependent serine hydroxymethyl transferase (l‐serine: tetrahydrofolate 10‐hydroxymethyl transferase, EC 2.1.2.1) reaction in rat or human brain homogenates incubated aerobically is dependent on added reducing agents for full activity in order to protect the readily oxidized substrate, tetrahydrofolate. In this role, 0.1 mm‐NADH is as affective as 10mm‐2‐mercaptoethanol and it can be shown that the NADH prevents destruction of tetrahydrofolate incubated with brain homogenates. If the dihydropteridine reductase (NADPH:6,7‐dihydropteridine oxidoreductase, EC 1.6.99.7) activity of the brain homogenate is inhibited by a specific antiserum, NADH, but not 2‐mercaptoeth‐anol, is no longer effective. Furthermore, an homogenate of a brain biopsy from a human lacking dihydropteridine reductase requires added dihydropteridine reductase for maximal stimulation by NADH of the serine hydroxymethyl transferase reaction. We conclude that dihydropteridine reductase mediates the NADH stimulation and can play a role in preserving tetrahydrofolate from oxidation. The rinding of greatly reduced folate levels in the brain biopsy from the human lacking dihydropteridine reductase supports this postulated role of dihydropteridine reductase in folate metabolism.
TIME‐DEPENDENT PARTITIONING OF GLUCOSE CARBONS METABOLIZED BY THE SUPERIOR CERVICAL GANGLION FROM CALVES
Authors: Quadflieg K. H. , Brand K.
Published: December 2006

Abstract— Glucose metabolism in the superior cervical ganglion for calves has been studied by incubating slices with [1‐14C]‐, [6‐14C]‐ and [U‐14C]‐labelled glucose at 37°C and pH 7.4. Glucose utilization and the metabolic partitioning of glucose carbon in products during different incubation periods ranging from 5 to 60 min were determined by isotopic methods.

Separation and identification of labelled compounds have been achieved by anion and cation exchange chromatography as well as by TLC and enzymatic analyses.

From the data obtained a carbon balance could be constructed showing lactate to be the major product of glucose metabolism followed by CO2 and amino acids. Measuring the release of 14CO2 from differently 4C‐labelled glucose, the existence of an active pentose phosphate pathway in the ganglion could be demonstrated although this pathway seems to contribute only to a small extent to glucose metabolism. The marked decrease of the C‐U: C‐6 and the C‐U:C‐1 ratios in 14CO2 observed in the course of incubation is discussed in terms of a time‐dependent change in the rate of synthesis of amino acids which are directly connected with intermediates of the citric acid cycle.
MULTIPLE FORMS OF PROTEIN KINASES IN MYELIN AND MICROSOMAL FRACTIONS OF BOVINE BRAIN
Authors: Miyamoto E. , Miyazaki K. , Hirose R. , Kashiba A.
Published: December 2006

Abstract— The activity profiles of the solubilized protein kinases from the microsomal and myelin fractions of bovine brain were examined by column chromatography and sucrose density gradient centrifugation. The main peak of adenosine 3′,5′‐monophosphate (cyclic AMP)‐dependent activity with histone as substrate for each membrane enzyme was eluted with about 0.2 m‐NaCl on a DEAE‐cellulose column. A peak of activity stimulated with cyclic AMP was also eluted with about 0.1 m‐NaCl for the microsomal enzyme. A peak with protamine and casein as substrate for the microsomal or myelin enzyme, respectively, was larger than that with histone as substrate for each enzyme. The first peak with histone as substrate on a DEAE–cellulose column appeared as two peaks on the Sepharose 6B column. The second peak with histone as substrate on DEAE–cellulose column was shown to be a holoenzyme consisting of regulatory and catalytic subunits. The holoenzyme and subunits were eluted at similar positions to each other between both membrane enzymes on Sepharose 6B column. The holoenzyme sedimented as two peaks of activity on sucrose density gradient centrifugation, both of which were stimulated with cyclic AMP. The preincubation of the holoenzyme with cyclic AMP resulted in shifting to a position of a smaller molecular size.

The results indicate the occurrence of multiple forms of protein kinases in membrane fractions of brain with respect to substrate specificity and physical property.
Kinetics of Cerebral Uptake Processes In Vitro of L‐Glutamine, Branched‐Chain L‐Amino Acids, and L‐Phenylalanine: Effects of Ouabain
Authors: Benjamin A. M. , Verjee Z. H. , Quastel J. H.
Published: December 2006

Abstract: Estimates have been made of the amounts and rates of uptake of radioactive branched‐chain i‐amino acids, L‐phenylalanine, and L‐glutamine into incubated rat brain cortex slices. Estimates have also been made of the binding of these amino acids to brain cell fragments. It is shown that such binding, as well as the process of passive diffusion, is not affected by the presence of ouabain (0.2 mM), which suppresses the energy‐dependent concentrative uptakes of the amino acids investigated. The maximum specific binding of L‐glutamine is about three times that of the other amino acids and amounts to about 11% of the total uptake of the amino acid by rat brain cortex slices in 12 min from a medium containing 0.25 mM‐glutamine. The sodium‐ion concentration of the medium appears not to play a significant role in determining the rate of L‐glutamine uptake in brain slices except at relatively low concentrations (<20 mequiv./l). The presence of Na+, however, is essential for the attainment of a tissue‐to‐medium concentration ratio greater than 2.0 for L‐glutamine. At relatively low concentrations (0.25 mM) the rapidity of uptake of L‐glutamine into a suspension of nerve terminals exceeds that into brain cortex slices. The uptakes of L‐glutamine (Km's = 0.66 mM and 2.25 mM) and of the branched chain L‐amino acids (Km's approx. 0.3 mM and 2 mM) by rat brain cortex slices are characterized by a double affinity system, but that of L‐phenylalanine has only one affinity system (Km= 0.23 mM). The Km's have been calculated after subtracting the ouabain‐insensitive passive uptakes of the amino acids from the total observed uptakes.
ATP‐Stimulated Ca2+ Transport into Cholinergic Torpedo Synaptic Vesicles
Authors: Michaelson D. M. , Ophir I. , Angel I.
Published: December 2006

Abstract: Activation of the Ca2+/Mg2+ ATPase associated with highly purified Torpedo synaptic vesicles results in 45Ca2+ uptake. The accumulated 45Ca2+ is released by hypoosmotic buffer and by the Ca2+ ionophore A23187. Density‐gradient centrifugation and permeation chromatography reveal that vesicular acetylcholine and the membrane‐bound 45Ca2+ co‐migrate, thus implying that 45Ca2+ is transported into cholinergic vesicles. ATP‐dependent 45Ca2+ uptake follows saturation kinetics, with KmCa2+= 50 μM, KmATP= 5 μM, and Vmax= 3 ± 0.3 nmol Ca2+/mg protein/min. Treatment of the vesicles with mersalyl, dicyclohexyl‐carbodiimide, and quercetin leads to inactivation of the Ca2+/Mg2+ ATPase and to comparable inhibition of 45Ca2+ transport. Ruthenium red and ouabain have no effect on either of these activities. Nigericin in the presence of external K+ is a potent inhibitor of 45Ca2+ translocation, whereas gramicidin activates transport. The proton translocator carbonylcyanide p‐trifluoromethoxy‐phenylhydrazone (FCCP) and FCCP + the ionophore valinomycin partially inhibit 45Ca2+ transport. By contrast, the above ionophores do not affect Ca2+/Mg2+ ATPase activity. Tentative mechanisms for ATP‐dependent Ca2+ transport into cholinergic synaptic vesicles and the physiological significance of this process are discussed.
The Use of Sep‐Pak™ C18 Cartridges During the Isolation of Gangliosides
Authors: Williams Marcia A. , McCluer Robert H.
Published: December 2006

Abstract: The use of Sep‐Pak™ C18 reverse‐phase cartridges to adsorb gangliosides from aqueous solutions was studied. When upper phases formed from chloroform‐methanol tissue extracts or aqueous salt solutions containing gangliosides are rapidly passed through the cartridges, the lipids are retained and the non‐lipid components can be washed through. Gangliosides and other retained lipids can subsequently be eluted with methanol or chloroform‐methanol.
Uptake of Nucleotides and Catecholamines by Chromaffin Granules from Pig and Horse Adrenal Medulla
Authors: Carmichael S. W. , Weber A. , Winkler H.
Published: December 2006

Abstract: The uptake of nucleotides and Catecholamines into chromaffin granules from adrenals of pigs and horses is similar to that previously seen in bovine chromaffin granules. The rate of [3H]ATP uptake at 2 mM‐ATP concentration was 0.42 ± 0.06 and 0.15 ± 0.02 nmol/mg protein/min for pig and horse granules, respectively. The apparent Km's were 1.37 mM for pig granules, 0.89 mM for horse granules, and 1.2 mM for ox granules. The sensitivity of the uptake for nucleotides and catecholamine to specific inhibitors was found to be similar in granules from pig and ox, indicating that the same mechanisms of uptake are involved in both species.
Stimulus‐Secretion Coupling in Isolated Adrenal Chromaffin Cells: Calcium Channel Activation and Possible Role of Cytoskeletal Elements
Authors: Schneider Allan S. , Cline Hollis T. , Rosenheck Kurt , Sonenberg Martin
Published: December 2006

Abstract: The catecholamine secretory function of a preparation of isolated bovine adrenal chromaffin cells has been further characterized under conditions designed to elucidate the mechanism of calcium channel activation and the possible role of cytoskeletal elements in stimulus‐secretion coupling. Three related sets of data were obtained: (1) Differences in kinetics, Ca dependence, strength, and additivity of the secretory response to acetylcholine (ACh) versus excess K; (2) the effects on secretion of the Ca channel‐blocking agents, Ni, Mg, and verapamil; and (3) the Ca dependence of vinblastine action on ACh‐ and K‐evoked secretion. The results suggest that a major portion of the Ca influx required for catecholamine release enters the cell via voltage‐dependent Ca channels with some additional Ca influx via the ACh receptor channel. Comparison of the present secretion data with corresponding known electrophysiological properties of isolated chromaffin cells provides added evidence for a role of chromaffin cell action potentials in regulation of Ca influx and the secretory response. Elevated Ca concentrations enhanced K‐evoked secretion to levels comparable to that of ACh but did not induce a vinblastine block of K‐evoked release. This provides further evidence against a role of microtubules in the common exocytosis event per se. However, a role of cytoskeletal elements in directing the movement of secretory granules, or an action of vinblastine at cholinergic receptors, remain distinct possibilities.
Effect of Guanine Nucleotides on Dopaminergic Agonist and Antagonist Affinity for [3H]Sulpiride Binding Sites in Rat Striatal Membrane Preparations
Authors: Freedman S. B. , Poat Judith A. , Woodruff G. N.
Published: December 2006

Abstract: [3H]Sulpiride bound to rat striatal membrane preparations with a saturable, high affinity component. This binding was displaced potently by dopamine antagonists (both classic neuroleptics and the benzamide, sulpiride) and less potently by dopamine agonists. GTP and its stable analogue Gpp(NH)p did not affect [3H]sulpiride binding to the membranes but altered the affinity for dopaminergic agonists. This effect was specific in that antagonist binding was not affected and only GTP, GDP, and Gpp(NH)p produced the effect. Similar alterations in ligand binding affinity caused by guanine nucleotides have been observed for binding sites linked to an adenylate cyclase. Such an interpretation for the case of [3H]sulpiride is contrary to suggestions that sulpiride labels only those dopamine receptors that are not cyclase linked.
Regional Distribution of Methionine Adenosyltransferase in Rat Brain as Measured by a Rapid Radiochemical Method
Authors: Hiemke Christoph , Ghraf Rüdiger
Published: December 2006

Abstract: The distribution of methionine adenosyltransferase (MAT) in the CNS of the rat was studied by use of a rapid, sensitive and specific radiochemical method. The S‐adenosyl‐[methyl‐14C]l‐methionine ([14C]SAM) generated by adenosyl transfer from ATP to [methyl‐14C]l‐methionine is quantitated by use of a SAM‐consuming transmethylation reaction. Catechol O‐methyltransferase (COMT), prepared from rat liver, transfers the methyl‐14C group of SAM to 3,4‐dihydroxybenzoic acid. The 14C‐labelled methylation products, vanillic acid and isovanillic acid, are separated from unreacted methionine by solvent extraction and quantitated by liquid scintillation counting. Compared to other methods of MAT determination, which include separation of generated SAM from methionine by ion‐exchange chromatography, the assay described exhibited the same high degree of specificity and sensitivity but proved to be less time consuming. MAT activity was found to be uniformly distributed between various brain regions and the pituitary gland of adult male rats. In the pineal gland the enzyme activity is about tenfold higher.
The Activity of 2′,3′‐Cyclic Nucleotide 3′‐Phosphodiesterase in Rat Tissues
Authors: Weissbarth Sulamith , Maker Howard S. , Raes Ingrid , Brannan Timothy S. , Lapin Evelyn P. , Lehrer Gerard M.
Published: December 2006

Abstract: The activity of the myelin‐associated enzyme 2′,3′‐cyclic nucleotide 3′‐phosphodiesterase (CNP) was measured in 14 rat tissues and in subcellular fractions of rat liver by a sensitive fluorometric method, using cyclic NADP as substrate. CNP activity in brain (339 μmol/h/mg protein) was fourfold that of the sciatic nerve. The activities in tissues outside the nervous system ranged from a low of 0.42 μmol/h/mg protein in the unwashed red blood cell to a high of 9.96 in the spleen. The activity was highest in tissues containing cells with membranes capable of undergoing transformation and elaboration (spleen and thymus) and low in those in which the cell membranes are morphologically stable (muscle and red cell). The enzyme was found in all major liver subtractions, with the highest activities in the microsomal and nuclear fractions. Despite the large difference in the maximal velocities of CNP in brain and liver, the affinity of the liver enzyme for the substrate (km) was similar to that of brain enzyme. Brain CNP was stable over a 48‐h postmortem period.
Inhibitors of a Rat Brain Enkephalin Aminopeptidase
Authors: Wagner Grant W. , Dixon Jack E.
Published: December 2006

Abstract: Eight protease inhibitors of microbiological origin were examined as potential inhibitors of a homogeneous rat brain enkephalin aminopeptidase. Bestatin [(2S,3R)‐3‐amino‐2‐hydroxy‐4‐phenylbutanoyl]‐l‐leucine and analogs of bestatin having basic, acidic, and other neutral amino acids substituted for the Leu residue exhibited inhibition constants ranging from 3.3 ± 10−5 to 8.3 ± 10−8m. The best inhibitor had a positively charged amino acid (Lys) substituted for Leu. A series of phenylalanyl dipeptides were examined as substrates with the aminopeptidase. The amino acid residue on the carboxyl side of the peptide bond undergoing cleavage was varied systematically in the dipeptides to include neutral, acidic, and basic residues. Again, a positively charged amino acid (Arg) adjacent to the bond undergoing scission was kinetically preferred. These results may be used to design highly specific inhibitors of the enkephalin aminopeptidase.
Biochemical Demonstration of the Myelin‐Associated Glycoprotein in the Peripheral Nervous System
Authors: Figlewicz Denise A. , Quarles Richard H. , Johnson David , Barbarash Gary R. , Sternberger Nancy H.
Published: December 2006

Abstract: Recent immunocytochemical studies indicated that the myelin‐associated glycoprotein (MAG) is localized in the periaxonal region of central nervous system (CNS) and peripheral nervous system (PNS) myelin sheaths but previous biochemical studies had not demonstrated the presence of MAG in peripheral nerve. The glycoproteins in rat sciatic nerves were heavily labeled by injection of [3H]fucose in order to re‐examine whether MAG could be detected chemically in peripheral nerve. Myelin and a myelin‐related fraction, WI, were isolated from the nerves. Labeled glycoproteins in the PNS fractions were extracted by the lithium diiodosalicylate (LIS)‐phenol procedure, and the extracts were treated with antiserum prepared to CNS MAG in a double antibody precipitation. This resulted in the immune precipitation of a single [3H]fucose‐labeled glycoprotein with electrophoretic mobility very similar to that of [14C]fucose‐labeled MAG from rat brain. A sensitive peptide mapping procedure involving iodination with Bolton‐Hunter reagent and autoradiography was used to compare the peptide maps generated by limited proteolysis from this PNS component and CNS MAG. The peptide maps produced by three distinct proteases were virtually identical for the two glycoproteins, showing that the PNS glycoprotein is MAG. The MAG in the PNS myelin and Wl fractions was also demonstrated by Coomassie blue and periodic acid‐Schiff staining of gels on which the whole US‐phenol extracts were electrophoresed, and densitometric scanning of the gels indicated that both fractions contained substantially less MAG than purified rat brain myelin. The presence of MAG in the periaxonal region of both peripheral and central myelin sheaths is consistent with a similar involvement of this glycoprotein in axon‐sheath cell interactions in the PNS and CNS.
Bulk Preparation of CNS Cytoskeleton and the Separation of Individual Neurofilament Proteins by Gel Filtration: Dye‐Binding Characteristics and Amino Acid Compositions
Authors: Chiu Fung‐Chow , Norton William T.
Published: December 2006

Abstract: The three major proteins of mammalian neurofilaments, of molecular weight 70,000, 160,000, and 210,000, have been resolved by sodium dodecyl sulfate (SDS)‐polyacrylamide gel eJectrophoresis, and more recently, by ion‐exchange chromatography in urea solution. We describe here a method to separate the neurofilament proteins by gel filtration without the use of SDS. A bulk preparation of cytoskeleton from rat spinal cord was first characterized. This preparation was then solubilized in a buffer containing 8 M urea and subjected to gel filtration. Individual neurofilament proteins, in milligram quantities, were harvested following the pooling of appropriate fractions. Gel electrophoresis showed a high degree of homogeneity in each of the three pooled fractions. Dye binding studies demonstrated that the protein of molecular weight 210,000 was relatively underrepresented when stained with Coomassie Blue, while all three neurofilament proteins showed similar dye binding properties with Fast Green. Amino acid analysis indicated that (1) all three neurofilament proteins contained a high content of acidic residues; (2) the molecular weight 210.000 protein contained >8 mol% proline; and (3) no simple oligomeric relationship existed among the neurofilament triplets.