bluuebunny/crossref_metadata_2025 huggingface dataset:
The Relationship Between the Stimulation of Dopamine Synthesis and Release Produced by Amphetamine and High Potassium in Striatal Slices
Authors: Schwarz Roy D. , Uretsky Norman J. , Bianchine Joseph R.
Published: October 2006

The effects of amphetamine (amph) and high K+ on the synthesis and release of dopamine (DA) were compared in striatal slices. Both agents stimulated DA synthesis as well as release. For both agents, Ca2+ was required for the initiation of synthesis stimulation as well as for the maintenance of this stimulation. The addition of EGTA to medium containing slices that were already stimulated by 1.0μm‐amph or 55 mm‐K+ markedly reduced the stimulation of DA synthesis. Although it has been reported that high K+ activates soluble tyrosine hydroxylase (TH), neither high K1 nor amph appeared to increase the affinity of the synthetic cofactor, 6‐MPH4, or decrease the affinity of the catechol, DA, for TH. This finding was supported by the observation that the inhibitory effect of L‐DOPA on DA synthesis in slices, in which synthesis was stimulated by either agent, was not decreased. Although both 1.0μM‐amph and 55 mm‐K® stimulated the release of [3H]DA from striatal slices, the release produced by K+ was Ca2+‐dependent, whereas release produced by amph did not occur at any concentration tested. Studies on pH requirements for both synthesis and release also confirmed a similarity between amph and K1 in stimulating synthesis but not in stimulating release. These results suggest that depolarizing agents, such as high K1, couple synthesis and release of DA by a Ca2+‐dependent mechanism. In contrast, the simultaneous stimulation of synthesis and release by amph is not regulated by Ca2+.
Enzymes Related to Monoamine Transmitter Metabolism in Brain Microvessels
Authors: Hardebo J. E. , Emson P. C. , Falck B. , Owman Ch. , Rosengren E.
Published: October 2006

Abstract: The activities of tyrosine hydroxylase, aromatic l‐aminoacid decarboxylase, monoamine oxidase, and catechol‐O‐methyltransferase were measured in microvessel fractions (capillaries and venules), parenchymal arterioles, and pial vessels from rat brains, and the decarboxylase activity was compared in brain microvessels from rabbit, cat, dog, pig, cow, baboon, and man. Cranial sympathectomy was performed to estimate the neuronal contribution to the enzyme activities. All vascular regions had substantial activities of the various enzymes studied. The activity of aromatic l‐aminoacid decarboxylase in cerebral microvessels was high in rat, dog, pig, cow, and man; intermediate in rabbit and cat; and low in baboon. In addition to this enzyme, cerebral microvessels also contained tyrosine hydroxylase and monoamine oxidase. Aromatic aminoacid decarboxylase and monoamine oxidase serve an enzymatic barrier function at the microvascular level, whereas the main function of tyrosine hydroxylase is probably to synthesize monoamines within nerve terminals that remain in close association with microvessels under the conditions used for preparation of the microvascular fraction. In larger intracerebral and pial vessels monoamine oxidase was present both in the wall itself and in perivascular sympathetic nerves; the remaining two enzymes had a primarily neuronal localization. The latter types of vessels also contained catechol‐O‐methyltransferase in their walls.
Neurites and Somata of Sensory and Sympathetic Neurons in Culture Contain Multiple Species of Ganglio sides
Authors: Cornbrooks Carson J. , Bunge Richard P. , Gottlieb David I.
Published: October 2006

Abstract: We have used tissue culture methods to study the capacity of sensory and sympathetic neurons to synthesize gangliosides. Under appropriate culture conditions, explants of dorsal root or superior cervical ganglia generate an extensive halo of ncurites, which is substantially free of contaminating cells. The cultures incorporate enough [3H]glucosamine into glycolipids to allow biochemical characterization. Gangliosides synthesized by the cells are extracted and freed of radioactive precursors and other lipids by column chromatography. Synthesized material comigrates on (thin‐layer chromatograms with the gangliosides, GQ, GT, GD1a, GD1b, and GM1, In addition, a substantial amount of unidentified labeled material migrates in a region between the mono‐ and disialo bands. The ganglioside profiles show that a similar spectrum of gangliosides is found on the neurites and somata of a particular class of neuron. Furthermore, the ganglioside compositions of the two types of neuron studied appear to be similar. We conclude that both sensory and adrenergic autonomic neurons synthesize gangliosides of each of the major classes and that representatives of each class are found in both somata and neurites. Dispersed cell cultures of the superior cervical ganglion synthesize a repertoire of gangliosides similar or identical to that of explant cultures.
Substrate Specificity of [3H] Muscimol Binding to a Particulate Fraction of a Neuron‐enriched Culture of Embryonic Rat Brain
Authors: DeFeudis Francis V. , Ossola Lucienne , Schmitt Gaby , Mandel Paul
Published: October 2006

Abstract: The effects of some GABA analogues and some drugs on the binding of [3H]muscimol (3.08 nM) to thoroughly washed subcellular particles prepared from a neuron‐enriched culture of embryonic rat brain were examined using Na+‐free Tris‐citrate medium and a centrifugation method. Competition for [3H]muscimol binding sites by excess(10−5 M) unlabelled GABA provided estimates of “specific” binding. In accord with in vivo neuropharmacological studies on GABA receptors and with in vitro studies on cerebral membrane preparations, [3H]muscimol binding was potently inhibited by muscimol itself (IC50, 2.5 nM), GABA (1C50, 43 nM), isoguvacine (IC50, 61 nM), and 3‐aminopropanesulphonic acid (IC50, 160 nM), and less potently inhibited by the GABA antagonist bicuculline methobromide (IC50, 800 nM). δ‐ Aminovaleric acid (IC50, 2.6 μM), the glycinelp‐alanine antagonist strychnine (IC50, 6.6 μM), and the predominantly glial GABA uptake inhibitors β‐alanine (IC50, 23 μM) and p‐proline (IC50, 66 μM) also inhibited [3H]muscimol binding. Other inhibitors of Na+‐dependent GABA uptake, (±)‐nipecotic acid, L‐ 2,4‐diaminobutyric acid, and guvacine, as well as picrotoxinin, were relatively inactive as inhibitors of [3H]muscimol binding (IC50≥ 1 mM). In addition to revealing that GABA receptors are present on neuronal membranes before the formation of most synapses, this binding of [3H]muscimol that occurs to neuronal, but not to glial, membranes might be useful as a “neuronal marker” and for the further characterization and isolation of GABA receptors.
Taurine Interactions with Chick Retinal Membranes
Authors: López‐Colomé A. M. , Pasantes‐Morales H.
Published: October 2006

Abstract: Binding of [3H]taurine to whole retinal membranes and to membranes obtained from retinal subcellular fractions was studied. [3H]Taurine bound to chick retinal membranes with high affinity and specificity. Two types of [3H]taurine binding associated to retinal membranes were observed, one with a KD= 0.68 μM and the other one with a KD,= 9.32 μM. Both types of binding were highly Na−‐dependent. The Na+‐dependent taurine binding was antagonized by strychnine. Bound [3H]taurine was effectively displaced by β‐alanine but not by GABA or glycine. Taurine binding was preferentially localized in membranes obtained from the crude synaptosomal fraction, although it is also present in substantial amounts in all retinal membranes. A Na+‐independent [3H]taurine binding exhibiting properties which might represent interaction with postsynaptic receptor sites could not be demonstrated in the chick retina.
The Synthesis and Activity of cis‐and trans‐2‐(Aminomethyl) cyclopropanecarboxylic Acid as Conformationally Restricted Analogues of GABA
Authors: Allan R. D. , Curtis D. R. , Headley P. M. , Johnston G. A. R. , Lodge D. , Twitchin B.
Published: October 2006

Abstract: The synthesis of cis‐2‐(aminomethyl) cyclopropanecarboxylic acid, a new analogue of GABA in a folded conformation, is described, as is also an improved preparation of trans‐2‐(aminomethyl) cyclopropanecarboxylic acid. When adminstered microelectrophoretically the trans isomer was more potent than GABA as a bicuculline‐sensitive depressant of the firing of cat spinal neurons in vivo, whereas the cis‐isomer was less potent than GABA and its effects appeared not to be sensitive to bicuculline methochloride. Trans‐2‐(aminomethyl) cyclopropanecarboxylic acid was a weak inhibitor of the sodium‐dependent uptake of GABA by mini slices of rat cerebral cortex and a substrate for the GABA: 2‐oxoglutarate aminotransferase activity in extracts of rat brain mitochondria. The cis isomer did not influence GABA uptake or aminotransferase activity and neither isomer reduced glutamate decar‐boxylase activity in rat brain homogenates. Both cyclopropane isomers inhibited the sodium‐independent binding of GABA to synaptic membranes from rat brain and their relative potencies together with those found for the stereochemically related unsaturated derivatives, cis‐and trans‐4‐aminocrotonic acid, were broadly consistent with the activity observed for these compounds in vivo on cat spinal neurons. These studies reinforce the evidence that extended rather than folded conformations of GABA are active at most GABA recognition sites within the mammalian central nervous system.
Sulfation of Dopamine and Other Biogenic Amines by Human Brain Phenol Sulfotransferase
Authors: Renskers Kevin J. , Feor Kevin D. , Roth Jerome A.
Published: October 2006

Abstract: Phenol sulfotransferase was isolated in 100,000g supernatant fractions prepared from postmortem samples of human brain. Since phenol sulfotransferase (PST) has been shown to conjugate the amine neurotransmit‐ters in vivo, the abilities of eight different biogenic amines and structurally related compounds to act as substrates for PST were studied. These experiments demonstrate that at a concentration of 20 μM, dopamine (DA) was the best substrate examined and was followed in decreasing order of activity by 3‐methoxytyramine (3‐MT), tyramine, norepinephrine, 3‐methoxy‐4‐hydroxyphenylethyleneglycol, octopamine, 5‐hydroxytryptamine and dihydroxyphenylethyleneglycol. At a substrate concentration of 100 /UM the relative order of activity was altered, so that tyramine became the most rapidly conjugated substrate while the activity of DA and 3‐MT relative to the other substrates tested was diminished. This change in substrate affinity with differing substrate concentrations can be explained, at least for DA, by the occurrence of apparent substrate inhibition at concentrations above 25 to 30 μM. Using PST isolated in 100,000g supernatant fractions from human brain, the Km value for DA was found to be 5.0 μM, while the Km value for the sulfate‐donor 3′‐phosphoadenosine‐5′‐phosphosulfate was 0.25 μM. The ratio of 3‐O‐ to 4‐O‐DA‐sulfate formed in vitro by human brain PST was found to be about 4: 1. In addition, both the 3‐O‐ and 4‐O‐esters were found not to be deaminated by human brain mitochondrial MAO. The relative role of PST with respect to MAO and catechol‐O‐methyltransferase in the degradation of the biogenic amine neurotransmitters in human brain is discussed.
Ontogeny of Neurochemical Markers for Noradrenergic, GABAergic and Cholinergic Neurons in Neocortex Lesioned with Methylazoxymethanol Acetate
Authors: Johnston Michael V. , Coyle Joseph T.
Published: October 2006

Abstract: Methylazoxymethanol acetate (MAM), a potent, rapidly eliminated nucleic acid alkylating agent, produces microencephaly in rat pups when injected into their dams on day 15 of gestation. In the adult microencephalic rats, neuronal loss is largely confined to telencephalic structures, such as the superficial neocortical laminae, whose neuroepithelial progenitor cells were undergoing vigorous replication during the chemical exposure. Histological examination of the forebrain 2 days after injection revealed early selective damage to the ventricular geminal zone with relative sparing of cortical plate neurons generated on earlier days. The degree of specificity of MAM's action on neurochemically defined neuronal populations was examined by measuring presynaptic markers for GABAergic, noradrenergic and cholinergic neurons in atrophic lateral cortex from 20 days gestation to adulthood. Although treatment reduced GABAergic markers (GABA, its synthetic enzyme and synaptosomal uptake process) in proportion to loss of cortex mass (‐67%), the maturational pattern for remaining GABAergic neurons was virtually normal. Although the maturational sequence of noradrenergic markers was similar to control, the concentration of endogenous norepinephrine, [3H]norepinephrine uptake and tyrosine hydroxylase specific activity were two‐ to fourfold higher than control at each time. However, total noradrenergic markers per cortex section were nearly identical to control throughout development, indicating that development of the noradrenergic axonal arbor in neocortex was insensitive to loss of neurons in the terminal field. Maturation of cholinergic markers (endogenous acetylcholine, its synthetic enzyme and [3H]choline uptake) in the atrophic cortex was biphasic: concentrations were similar to control values for the first 12 postnatal days, but gradually rose to levels twofold higher than control. These results indicate that neurochemical alterations observed in cortex from prenatally MAM‐treated rats are primarily the result of early selective elimination of neuronal subpopulations. Fetal MAM exposure appeared to have minimal effects on biochemical differentiation of neurons remaining intact in the atrophic cortex. MAM appears to be a useful toxin for producing selective loss of neuronal groups based on their time of generation in the fetus.
pH‐Induced Alterations in Dopamine Synthesis Regulation in Rat Brain Striatal Synaptosomes
Authors: Patrick Robert L. , Rendel Michael T.
Published: October 2006

Abstract: Dopamine synthesis regulation as a function of pH has been examined in rat brain striatal synaptosomes. Synthesis stimulation produced by lowering the incubation pH from 7.2 to 6.2 is accompanied by a significant increase in apparent A'm for tyrosine and in apparent Vmax. While these kinetic alterations are similar to those produced by the depolarizing agent veratridine, it does not appear that synthesis is stimulated at pH 6.2 via synaptosomal depolarization since (1) synthesis stimulation still occurs at pH 6.2 in a calcium‐free medium in contrast to the calcium‐dependency of veratridine‐ induced stimulation and (2) tyrosine uptake is not inhibited by incubation at pH 6.2, but is markedly inhibited by veratridine. In order to study how the regulatory properties of synaptosomal preparations vary according to pH, the ability of synaptosomal dopamine synthesis to respond to various agents was tested between pH 7.2 and 6.2. The stimulatory effects of veratridine, amphetamine, phenylethylamine and dibutyryl cyclic AMP at pH 7.2 were significantly diminished at pH 6.2. In addition, incubation at pH 6.2 antagonized the veratridine‐induced inhibition of tyrosine uptake, suggesting an interference with the depolarization process. The inhibitory effects of dopamine and tyramine at pH 7.2 were also antagonized at pH 6.2. In contrast to the effects of pH 6.2 buffer, incubation at pH 6.6 does not markedly alter responses to the various drugs. The results suggest that, although basal dopamine synthesis rates can be increased by lowering the pH, synaptosomal regulatory properties are significantly altered as the pH is lowered below 6.6.
Regulation of the First Step of the Initiation of Brain Protein Synthesis by Guanosine Diphosphate
Authors: Dwyer Barney , Wasterlain Claude G.
Published: October 2006

Abstract: Initiation factor elF‐2‐like activity has been measured in the 0.5 M‐KCl wash of rat brain microsomes. Ternary complex formation (elF‐2 GTP Met‐tRNAr), one of the early steps in protein synthesis initiation, is optimal in a high‐[K+], low‐[Na+] environment. Mg2+, Ca2+, Li+, spermine and spermidine reduce and the antibiotic aurin tricarboxylic acid can effectively eliminate ternary complex formation. The formation of ternary complex requires GTP or its nonhydrolyzable analog, GMP‐P(NH)P. Ternary complex formation is particularly sensitive to the ratio of GDP to GTP. When the ratio of GDP to GTP added is 1: 10, ternary complex formation is inhibited between 40 and 50% over a 30‐fold concentration range of GTP. Other nucleotides exert little inhibition. These results suggest that the regulation of brain protein synthesis initiation may be tightly linked to the ratios of guanosine nucleotide concentrations in brain tissue.
Properties of the Sodium Extrusion Mechanism Controlled by Nerve Growth Factor in Chick Embryo Dorsal Root Ganglionic Cells
Authors: Skaper S. D. , Varon S.
Published: October 2006

Abstract: Cell dissociates from embryonic chick dorsal root ganglia, incubated for 6 h with 22Na+, accumulated four to six times more radioactivity in the absence than in the presence of Nerve Growth Factor (NGF). The accumulation of radioactivity paralleled the external Na+ concentration, indicating that the cells may have been reaching equilibrium with the medium. Delayed presentation of NGF to 22Na+‐loaded cells caused a rapid loss of radioactivity, even with extracellular 22Na+ still present, demonstrating that NGF caused an overall efflux of Na+ rather than an accelerated equilibration. The Na+ exclusion from 22Na+‐loaded cells was dependent upon NGF concentration. Use of nutrient‐rich medium, serum, and certain hormones and other proteins did not prevent the Na+ accumulation in the absence of NGF or its reversion by delayed NGF administration. Incubation of the ganglionic cells with ouabain or dinitrophenol during the 22Na+ loading period (no NGF) increased the rate, but not the magnitude, of loading. The same incubation carried out in a Na+‐free medium and followed by 22Na+ presentation resulted in fast radioactive loading that was identical to that occurring in drug‐free, NGF‐deprived cells and was not prevented by presentation of NGF together with the 22Na+. These data are consistent with a model in which NGF acts through a Na+ pump rather than by restricting Na+ influxes.
Effect of l‐Tryptophan on Mouse Brain 5‐Hydroxytryptamine: Comparison of Values Obtained Using a Fluorimetric Assay and a Liquid Chromatographic Assay with Electrochemical Detection
Authors: Marsden C. A.
Published: October 2006

Abstract: 5‐Hydroxytryptamine (5‐HT) in mouse brain and spinal cord was assayed in the same samples using a fluorimetric assay and a high pressure liquid chromatographic (HPLC) assay with electrochemical detection. The HPLC assay was able to detect levels of 5‐hydroxytryptamine as low as 0.2‐0.5 pmol. With the column (Vydac cation exchange), solvent system (acetate/citrate buffer, 0.1 or 0.2 M, pH 4.8‐5.2) extraction procedure and electrode potential (+0.55 V) used, the HPLC assay was specific for 5‐HT. When the electrode potential was increased to +0.9 V tryptamine could also be detected, with a longer retention time than 5‐hydroxytryptamine. The percentage increase in mouse brain 5‐hydroxytryptamine after pargyline (75 mg/kg) and pargyline +l‐tryptophan (100 mg/kg) was very similar whether measured by fluorimetry or HPLC, although the fluorimetric assay gave consistently higher absolute values (24–32%) in both control and drug‐treated animals. l‐Tryptophan (25, 50 and 100 mg/kg) also increased brain 5‐hydroxytryptamine with similar percentage increases with either assay method. There was a significant correlation (P < 0.001) between the values obtained with the two assay methods. The results confirm the use of HPLC with electrochemical detection as a sensitive and specific assay method for 5‐hydroxytryptamine and indicate its potential use for the assay of tryptamine, and the importance of determining the electroactivity and retention characteristics of any drugs used.
Presence of Protein I, a Phosphoprotein Associated with Synaptic Vesicles, in Cerebellar Granule Cells
Authors: Dolphin Annette C. , Greengard Paul
Published: October 2006

Abstract: The cerebellar levels of Protein I, a synapse‐specific neuronal phosphoprotein, have been investigated in the cerebellar mouse mutants staggerer (sg), weaver (wv), nervous (nr), and Purkinje cell degeneration (pcd). The Protein I concentration was reduced by about 66% in sg and wv mutants, representing a 90% loss of Protein I per cerebellum. A heterozygote effect was observed in the wv mutant. These results indicate that a great majority of Protein I in the normal cerebellum may be present in the granule cells. in nr mutants the cerebellar Protein I concentration was reduced by only 12% in 62‐day‐old mice, suggesting that Purkinje cells contribute little to cerebellar Protein I. However, a greater reduction was observed in pcd mutants, which may reflect on the nature of the pcd mutation.
Enhancement of 45Ca2+ Binding to Acidic Lipids by Barbiturates, Diphenylhydantoin, and Ethanol
Authors: Chweh A. Y. , Leslie S. W.
Published: October 2006

Abstract: Effects of barbiturates, diphenylhydantoin, and ethanol on 45Ca2+ binding to acidic lipids have been examined in an organic solvent‐aqueous partition system. Hexobarbital, pentobarbital, and phenobarbital, at concentrations of 0.3 and/or 0.6 mM, enhanced the binding of 45Ca2+ to phosphatidic acid, phosphatidylserine, and sulfatide but not to phosphatidylinositol or cardiolipin. Diphenylhydantoin, 0.3 mM, enhanced 45Ca2+ binding to phosphatidic acid and phosphatidylserine but not to sulfatide. Ethanol at 80 mM did not enhance 45Ca2+ binding to phosphatidic acid, but ethanol decreased the binding to cardiolipin and increased it to sulfatide.
Characterization of the Binding of the GABA Agonist [3H]Piperidine‐4‐Sulphonic Acid to Bovine Brain Synaptic Membranes
Authors: Krogsgaard‐Larsen P. , Snowman A. , Lummis S. C. , Olsen R. W.
Published: October 2006

Abstract: The binding of radioactive piperidine‐4‐sulphonic acid ([3H]P4S) to thoroughly washed, frozen, and thawed membranes isolated from cow and rat brains has been studied. Quantitative computer analysis of the binding curves for four regions of bovine brain revealed the general presence of two binding sites. In these brain regions less satisfactory computer fits were obtained for receptor models showing one or three binding sites or negative cooperativity. With the use of Tris‐citrate buffer at 0°C the two affinity classes for P4S in bovine cortex membranes revealed the following binding parameters: KD= 17 ± 7 nM (Bmax= 0.15 ± 0.07 pmol/mg protein) and KD= 237 ± 100 nM (Bmax= 0.80 ± 0.20 pmol/mg protein). Heterogeneity was also observed for association and dissociation rates of [3H]P4S. The slow binding component (kon= 5.6 × 107 or 8.8 × 107 M‐1 min‐1, kOff= 0.83 min‐1, and KD= 14.7 or 9.4 nM, determined by two different methods in phosphate buffer containing potassium chloride) corresponds to the high‐affinity component of the equilibrium binding curve (KD= 11 nM, Bmax= 0.12 pmol/mg protein in the same buffer system). The association and dissociation rates for the subpopulation of rapidly dissociating sites, apparently corresponding to the low‐affinity sites, were too rapid to be measured accurately. The binding of [3H]P4S appears to involve the same two populations of sites with Bmax values similar to those for [3H]GABA binding to the same tissue, although the kinetic parameters for the two ligands are somewhat different. Furthermore, comparative studies on the inhibition of [3H]P4S and [3H]GABA binding by various GABA analogues, strongly suggest that P4S binds to the GABA receptors. The different effects of P4S and GABA on benzodiazepine binding are discussed.
Identification and Determination of Tele‐Methylhistamine in Cerebrospinal Fluid by Gas Chromatography‐Mass Spectrometry
Authors: Swahn Carl‐Gunnar , Sedvall Göran
Published: October 2006

Abstract: The presence of tele‐methylhistamine in human cerebrospinal fluid has been established. The concentration was determined with the use of deuterated tele‐methylhistamine. The preparation of the deuterated standard is described. The concentration range in samples from neuropsychiatric patients was 0.1‐2.5 ng/ml. The structure of the pentafluoropropionyl derivative used for gas chromatography was studied with the aid of proton nuclear magnetic resonance spectroscopy.
Turnover of Free Sialic Acid, CMP‐Sialic Acid, and Bound Sialic Acid In Rat Brain
Authors: Ferwerda Wijnholt , Blok Corrie M , Heijlman Jan
Published: October 2006

Abstract: Adult male rats were injected intraventricularly with N‐[3H]acetylmannosamine. After different time intervals the rats were killed and free sialic acid, CMP‐sialic acid, lipid‐ and protein‐bound sialic acid were isolated from brain and the specific radioactivities determined. Maximal specific radioactivity was reached after approximately 4 h for CMP‐sialic acid, after 10–12 h for free sialic acid and after approximately 42 h for lipid‐and protein‐bound sialic acid. After some days the specific radioactivities of all four pools were the same and decreased equally, with a calculated turnover rate of approximately 3.5 weeks. The conclusion was that this phenomenon was the result of reutilisation of sialic acid and/or precursors. Therefore, the calculated turnover is not the turnover of bound sialic acid, but merely the rate of leakage of sialic acid and/or precursors out of the brain, so that no real turnover can be measured by this method. The first few hours after injection the specific radioactivity of CMP‐sialic acid rose above that of free sialic acid. It is supposed that a compartmentalization exists of free sialic acid. The newly synthesized sialic acid molecules are not secreted into the cytoplasmic pool but are preferentially used for the synthesis of CMP‐sialic acid. The results and conclusions are discussed in view of the general problems concerning turnover measurements of glycoconjugates.
Degradation of the P0, P1, and Pr, Proteins in Peripheral Nervous System Myelin by Plasmin: Implications Regarding the Role of Macrophages in Demyelinating Diseases
Authors: Cammer Wendy , Brosnan Celia F , Bloom Barry R , Norton William T
Published: October 2006

Abstract: Activated macrophages secrete a variety of neutral proteinases, including plasminogen activator. Since macrophages are implicated in primary demyelination in the peripheral nervous system (PNS) in Guillain‐Barré syndrome and experimental allergic neuritis, we have investigated the ability of plasmin and of conditioned media from cultured macrophages, in the presence of plasminogen, to degrade the proteins in bovine and rat PNS myelin. The results indicate that (a) the major glycoprotein P0 and the basic P1, and Pr, proteins in PNS myelin are extremely sensitive to plasmin, perhaps more so than is the basic protein in CNS myelin; (b) the initial product of degradation of P0 by plasmin has a molecular weight higher than that of the “X” protein; (c) large degradation products of P0 are relatively insensitive to further degradation; and (d) the neuritogenic P2 protein in PNS myelin is quite resistant to the action of plasmin. Results similar to those with plasmin were obtained with conditioned media from macrophages and macrophage‐like cell lines together with plasminogen activator, and the degradation of the PNS myelin proteins, P0 and P1, under these conditions was inhibited by p‐nitrophenylguanidinobenzoate, an inhibitor of plasmin and plasminogen activator. The results suggest that the macrophage plasminogen activator could participate in inflammatory demyelination in the PNS.
Characteristics of the Inhibition of Rat Brain Monoamine Oxidase In Vitro by MD780515
Authors: Kan J. P , Benedetti M. Strolin
Published: October 2006

Abstract: The inhibition of type A and B MAO in rat forebrain crude membrane preparation by MD780515. (3‐{4‐[(3‐cyanophenyl)methoxy]phenyl)‐5‐(methoxymethyl)‐2‐oxazolidinone Centre de Recherche Delalande, France) has been investigated in vitro with 5‐hydroxytryptamine and β‐phenylethyl‐amine as substrates. The inhibition of the high‐affinity binding of [3H]harmaline, a specific marker of type A MAO, was also studied. In the experimental conditions used, MD780515 appeared to be a pure mixed MAO inhibitor (MAOI) of 5‐HT deamination, both Km, and Vmax being altered [K1 (Dixon) =Ki, (slope) = 2 nM; Ki (intercept) = 12 nM]. Phenylethylamine oxidation could be considered to be noncompetitively inhibited by MD780515 (Ki (slope) = 78 nM; Ki, (intercept) = 103 nM). Dixon and intercept replots were hyperbolic, suggesting that, at high concentrations, PEA could be deaminated by both forms of MAO. This hypothesis was confirmed by biphasic inhibition curves of 80 μM‐PEA obtained when MD780515, clorgyline, harmaline and deprenyl were used as MAOIs. MD780515 was a potent inhibitor (IC50= 1–2 nM) of [3H]harmaline binding. Comparatively, clorgyline, ‘cold’ harmaline and Lilly 51641 inhibited 3H ligand binding, with IC50 of 5, 7 and 40 nM respectively. In conclusion, MD780515 is a reversible, specific and potent type A MAOI.
The Effect of Acetylcholine Release on Choline Fluxes in Isolated Synaptic Terminals
Authors: Marchbanks R. M. , Wonnacott S. , Rubio M. A.
Published: October 2006

Abstract: As in intact tissues, choline influx into synaptosomes is enhanced after a period of depolarization induced release of acetylcholine. The activation of uptake is dependent on the presence of Ca2+ and inhibited by high Mg2+ concentrations in the medium during depolarization. Choline transport in erythrocytes was not activated by prior treatment with potassium. The permeability constant of the synaptosome membrane to choline was found to be 2.7 × 10−8 cm·s−1 and to acetylcholine 1.8 ′ 10−8 cm·s−1. Choline influx has been studied after pre‐loading synaptosomes with choline. Different radiolabels were used to measure efflux of preloaded choline and influx simultaneously. Isotopic dilution in flux studies was estimated and corrected for. Influx was stimulated by high internal concentrations of choline, and efflux similarly stimulated by high outside concentrations of choline. The maximal influx and efflux at saturating opposite concentrations of choline were equal with a value of about 500 pmol·min−1 per mg synaptosomal protein. A reciprocating carrier would explain the equality of the maximal influx and efflux. Acetylcholine competes with choline for binding to the carrier but is itself hardly transported. Increased acetylcholine concentrations were shown to inhibit both choline influx and efflux from the trans position. Raising intrasynaptosomal acetylcholine concentrations by pre‐loading abolished the stimulation of influx by prior depolarization. It is proposed that high concentrations of acetylcholine immobilize the carrier on the inside of the synaptic membrane. The stimulation of choline influx consequent upon depolarization is caused by release of ACh which results in relief of this immobilisation. The enhanced supply of choline achieved by this mechanism is likely to be important in maintaining stores of the acetylcholine in vivo.
Comparison of DABA and GABA Transport into Plasma Membrane Vesicles Derived from Synaptosomes
Authors: Roskoski Robert
Published: October 2006

Abstract: Transport of GABA by a high‐affinity transport system (Km≃ 10−5 M) is thought to terminate the action of this postulated neurotransmitter. 2,4‐Diaminobutyric acid (DABA), a structural analogue, is taken up by neuronal elements and inhibits GABA uptake. Localization of [3H]DABA by auto‐radiography has been used to identify neurons with the GABA high‐affinity transport system. After reconstitution of lysed synaptosomal fractions in potassium salts, transfer of these membrane vesicles to sodium salts produces sodium and potassium ion gradients which drive [3H]GABA and [3H]DABA transport. For each, transport requires external sodium, is abolished by ionophores that dissipate the Na+ gradient, and is enhanced by conditions which make the intravesicular electromotive force more negative. Some characteristics of the transport of these substances, however, differ. For example, external chloride is required for GABA, but not DABA, transport. Internal potassium is required for DABA, but not GABA, transport. DABA is a competitive inhibitor (Ki≃ 0.6 MM) of GABA transport into membrane vesicle and synaptosomes. GABA, however, is a feeble inhibitor of DABA uptake into the membrane vesicles. These differences suggest that the two substances are transported by different mechanisms and possibly by different carriers. In addition to these experiments, using enzymatic‐fluorometric techniques, it was shown that the artificially imposed ion gradients drive net chemical transport of GABA into the vesicles.
Effect of Bilirubin on the Arrhenius Plots for Na, K‐ATPase Activities of Young and Adult Rat Cerebra
Authors: Kashiwamata Shigeo , Asai Mitsuoki , Semba Ritsuko K.
Published: October 2006

Abstract: The effect of bilirubin on the temperature dependence of Na, K‐ATPase activity was investigated with NaI‐treated microsomes prepared from young (13‐ to 23‐day‐old) and adult (about 1‐year‐old) rat cerebra. The Arrhenius plots for the enzymic activities in the young and adult rats showed break temperatures at 26 and 18°C, respectively. In the young rat enzyme, bilirubin caused a shift of the break temperature to 23 and 22°C at concentrations of 60 and 120 μM, respectively, with a significant increase of the activation energy below the break temperature. However, no significant changes in the break temperature and the activation energy were observed in the adult rat enzyme exposed to the same concentrations of the pigment. The results suggest that the lipid environment surrounding the enzyme and modulating its activity in the plasma membranes may differ between the young and adult rat cerebra, and that bilirubin may change the physical state of the lipids related to the activity of the young rat Na, K‐ATPase.
Studies on γ‐Aminobutyric Acid Transport in Cobalt Experimental Epilepsy in the Rat
Authors: Ross Stephen M. , Craig Charles R.
Published: October 2006

Abstract: Crude brain homogenates and cerebral tissue slices from rats with cobalt metal implanted in right and left cerebral cortices were used to examine high‐ and low‐affinity GABA transport. High‐affinity GABA transport was maximally reduced to 34% of controls 7 days after cobalt implantation, a time that coincides with peak seizure activity in this model. Kinetic analysis of high‐affinity GABA transport, using brain homogenates, revealed a significant change in Vmax 7 days after cobalt implantation. (Vmax= 446.4 ± 26.2 pmol/mg prot./min, cobalt, versus 787.8 ± 67.3, control). An analysis of the low‐affinity system revealed no depression of Km, or Vmax parameters. Administration of valproic acid at a concentration as high as 1 mM in vitro or a dose of 300 mg/kg in vivo had no effect on high‐ or low‐affinity GABA transport. The results obtained from cobalt‐treated rats provide additional evidence for an involvement of GABA in experimental epilepsy.
Relationship Between Plasma and Brain Tryptophan in Pigs During Experimental Hepatic Coma Before and After Hernodialysis with Selective Membranes
Authors: Delorme M. L. , Denis J. , Nordlinger B. , Boschat M. , Opolon P.
Published: October 2006

Abstract: Experimental acute liver ischemia in pigs induces an increment in plasma free tryptophan with decreased total tryptophan. Brain tryptophan is elevated in all brain areas. A slight, but significant increase of brain serotonin is demonstrated in the striatum only, while 5‐HIAA (5‐hydroxyindoleacetic acid) is significantly lower in the hypothalamus. Other brain areas do not show significant changes in serotonin and 5‐HIAA levels. Neither the high plasma free tryptophan levels, nor the decreased sum of neutral competitive amino acids are consistent with such an elevation of brain tryptophan. Hemodialysis was carried out with two different kinds of membranes: cuprophan (with an efficient removal of molecules up to molecular weight 1300) and AN 69 polyacrylonitrile (efficient removal up to 15,000). Ammonia and aminoacid clearance are similar for both membranes. After AN 69, plasmatic free tryptophan and brain tryptophan are lower than after liver devascularization, but still higher than normal. Serotonin significantly increases in the cortex, midbrain and hypothalamus without concomitant rise of 5‐HIAA levels. After cuprophan hemodialysis, plasma total tryptophan is lower than in normal and even comatose animals, whereas free tryptophan is normal. Intracerebral tryptophan is similar to AN 69 dialysed animals, but in the hypothalamus it is similar to nondialysed animals. Brain serotonin levels are not modified. 5‐HIAA decreases in the hypothalamus. This finding suggests that middle molecules (which are not cleared out with cuprophan hemodialysis) are involved in the intracerebral transfer of tryptophan and the metabolism of serotonin, mainly in the hypothalamus.
A More Sensitive Radioimmunoassay for Neuron‐Specific Enolase Suitable for Cerebrospinal Fluid Determinations
Authors: Parma Alexandra M. , Marangos Paul J. , Goodwin Frederick K.
Published: October 2006

Abstract: Neuron‐specific enolase (NSE) and non‐neuronal enolase (NNE) have been shown to be highly specific neuronal and glial products respectively and are therefore useful as biochemical markers of the two major cell types in the vertebrate central nervous system. An iodinated radioimmunoassay (RIA) procedure for human NSE (NSE‐H) with approximately 50‐fold greater sensitivity than the previously available tritiated assay is described. This assay is capable of detecting 100 pg of NSE‐H per assay. NSE levels in human cerebrospinal fluid (CSF) which were previously undetectable with the tritiated RIA are now easily measured and have been shown to be approximately 2 ng/ml of CSF. Furthermore, results obtained with the newly described assay procedure on more concentrated brain tissue extracts are comparable to the tritiated RIA. The iodinated NSE RIA is also shown to be capable of accurately detecting added amounts of NSE in human CSF, indicating the potential clinical usefulness of this assay in determining elevated levels of NSE in CSF.
Chloroquine Intoxication Induces Ganglioside Storage in Nervous Tissue: A Chemical and Histopathological Study of Brain, Spinal Cord, Dorsal Root Ganglia, and Retina in the Miniature Pig
Authors: Klinghardt Georg W. , Fredman Pam , Svennerholm Lars
Published: October 2006

Abstract :

The effect of chronic chloroquine intoxication on lipid composition, particularly the gangliosides, was studied in the nervous system of miniature pigs, type Göttingen. The tissues examined were cerebrum, spinal cord, dorsal root ganglia and retina. Chloroquine was given in the diet in doses of 2.0‐3.5 g/kg food. The intoxication of the pigs was started at the age of 100–240 days and continued for 177‐219 days. The control pigs received the same diet without chloroquine. The ganglioside concentration was increased in all the tissues examined. Dorsal root ganglia and retina were the tissues affected most and showed a twofold increase. This corresponded to the light and electron microscopically demonstrated extensive storage process in the perikarya of dorsal root ganglion cells and inner ganglion cells of the retina. Under light microscopy the storage material was granular, intensely PAS‐positive and dissolved by paraffin embedding. The electron microscopical equivalent consisted of conglomerates of membranous lysosomal residual bodies. In cerebrum the ganglioside concentration was increased by 12%. Storage in the brain varied widely between different systems and types of cells. The allocortex was much more affected than the isocortex. Certain inhibitory ganglion cell types, such as the basket cells, exhibited the most massive storage of all. The spinal medulla was morphologically less involved but showed approximately the same ganglioside increase, though not statistically significant. With the exception of cerebrum the increase in the tissues examined involved all the individual gangliosides, most severely ganglioside GM2 and three fucogangliosides. In cerebrum only the ganglioside GM2 was increased more than the other gangliosides. Chloroquine intoxication did not affect the concentration of phospholipids or cholesterol in the cerebrum, spinal cord or dorsal root ganglia, but in retina the acidic phospholipids were significantly increased.
Glutamic Acid Decarboxylase and γ‐Aminobutyric Acid in Huntington's Disease Fibroblasts and Other Cultured Cells, Determined by a [3H]Muscimol Radioreceptor Assay
Authors: Hamel Edith , Goetz Ingeburg E. , Roberts Eugene
Published: October 2006

Abstract A sensitive and reproducible [3H]muscimol radioreceptor assay was developed for measuring low levels of both glutamic acid decarboxylase activity and γ‐aminobutyric acid. By using this technique, endogenous γ‐aminobutyric acid and glutamic acid decarboxylase activity were detected in two rat neuroblastomas, B35 and B50, a human medulloblastoma cell line, TE671, and cultured human skin fibroblasts. Glutamic acid decarboxylase activities and γ‐aminobutyric acid levels were compared for human skin fibroblasts obtained from patients with Huntington's disease and their controls in a well‐controlled, blind study. However, no significant difference was found for either measure between Huntington and control cells. Glutamic acid decarboxylase activity was relatively low in all cell types examined except for the TE671 cells, which had more than four times the activity found in the other cells. This human medulloblastoma cell line appears to be a good model for studying γ‐aminobutyric acid metabolism and the control of glutamic acid decarboxylase expression.
Cholesterol Esterifying Enzyme in Normal and Degenerating Peripheral Nerve
Authors: Yao Jeffrey K. , Dyck Peter James
Published: October 2006

Abstract: The cholesterol esterifying enzyme which incorporates exogenous free [1‐14C]oleate into cholesteryl ester is present in rat sciatic endoneurium. Cholesterol esterification is optimal at pH 4.8. Exogenous ATP, CoA, and oleyl‐CoA do not greatly affect its activity. Various detergents and bile salts are inhibitory. Enzyme activity does not change appreciably during storage at 4°C for up to 4 days or at ‐70°C for up to 1 month. Of the subcellular fractions, the microsomal fraction exhibits the highest specific activity. Over 75% of enzyme activity is recovered, with equal amounts in the microsomal and soluble fractions. During nerve fiber degeneration an increase (more than fivefold) in cholesterol esterifying activity, which peaks 6 days after crush, is observed. Elevated levels of enzyme activity persist for 90 days after crush, by which time nerve regeneration is well established. Thus, it is concluded that an increase in cholesterol esterifying activity in degenerating nerve is primarily responsible for cholesterol esterification during Wallerian degeneration. The maximum increase in cholesterol esterifying activity is associated temporally with axonal degeneration and, particularly, with the formation of myelin ovoids.
Protection of Neocortical Tissue Prisms from Freeze‐Thaw Injury by Dimethyl Sulphoxide
Authors: Haan E. A. , Bowen D. M.
Published: October 2006

Abstract: Neocortical tissue prisms prepared from rat and human brain were frozen to ‐196°C by a two‐step freezing procedure and 10% dimethyl sulphoxide as cryoprotectant. Frozen and thawed rat neocortical prisms incorporated glucose into acetylcholine and carbon dioxide at 89% and 86% of control values, respectively, and noradrenaline uptake into frozen and thawed rat prisms was 94% of the control value. Frozen and thawed prisms from three human neocortical specimens showed a similar degree of protection from freeze‐thaw injury.
Calcium‐Stimulated Adenosine Triphosphatases in Synaptic Membranes
Authors: Sorensen Roger G. , Mahler Henry R.
Published: October 2006

Abstract: We have investigated the properties of several ATPases present in synaptic membrane preparations from the cerebral cortex of rat. In addition to the intrinsic (Na++ K+)–ATPase and a low level of contaminating Mg2+–ATPase of mitochondrial origin, both of which could be controlled by the addition of ouabain and azide, respectively, four activities were studied: (1) a Mg2+–ATPase; (2) a Mg2+–independent activity requiring Ca2+ ions at high concentrations; (3) a (Ca2++ Mg2+)–ATPase with a high affinity for Ca2+, which was enhanced further (4) by the inclusion of calmodulin (33 DM for half–maximal activity). In the presence of 0.5 mM–EGTA in the buffer used, half saturation for these respective metal ions was observed at 0.9 mM for (1), 1.0 DIM for (2), and –0.3 mM for (3) and (4); the latter values correspond to concentrations of free Ca2+ of 0.38 and 0.18 /XM for (3) and (4), respectively. The level of activities observed, all in nmol × min−1× mg−′, under optimal conditions at 37°C, was in a number of preparations (n in parenthesis): for (1) 446 ± 19 (19); for (2) 362 ± 18 (3); for (3) 87 ± 13 (12); and for (4) 161 ± 29 (12). The (Ca2++ Mg2+)‐ATPase, both in the presence and absence of calmodulin, could be inhibited specifically by a number of agents (approximate I0.5 in parentheses) which, at these concentrations, showed little or no potency against the other activities; among them were vanadate (≥10 μm), La3+ (75 μm), trifluoperazine, and other phenothiazines (50μm). These properties suggest that the (Ca2++ Mg2+)‐ATPase described may be responsible for calcium transport across one (or more) of the several membranes present in nerve endings and contained in the preparation used.
Interactions of Di‐n‐Propylacetate, Gabaculine, and Aminooxyacetic Acid: Anticonvulsant Activity and the γ‐Aminobutyrate System
Authors: Wood J. D. , Kurylo E. , Tsui S.‐K.
Published: October 2006

Abstract: Di‐n‐propylacetate (DPA), aminooxyacetic acid (AOAA), and gabaculine were administered alone or in combination to Swiss mice. Six hours after administration of the drugs the anticonvulsant action (against isonicotinic acid hydrazide‐induced seizures) of AOAA and DPA combined was less than that of AOAA alone. The cause of this phenomenon appeared to be an interaction between DPA and AOAA with respect to inhibition of GABA‐T activity, resulting in a long‐term diminished inhibition by AOAA, which in turn led to a lessening of the AOAA‐induced elevation in the GABA content of nerve endings (synaptosomes). An excellent correlation was observed between the delay in onset of seizures and the elevation of synaptosomal GABA content.
Ontogenesis of Muscarinic Receptors and Acetylcholinesterase Activity in Various Areas of Chick Brain
Authors: Jerusalinsky Diana , Aguilar J. S. , Brusco Alicia , Robertis E. De
Published: October 2006

Abstract: Muscarinic receptors, labeled with [3H]quinuclidinyl benzylate (3H]QNB), and acetylcholinesterase activity were studied in five areas of the developing chick brain: (1) hyperstriatum and neostriatum, (2) paleostriatum, (3) optic lobes, (4) mesodiencephalon and (5) cerebellum. The protein content of these areas, expressed as mg/g tissue and total protein, was determined between day ‐10 and adulthood. Differences in both determinations were observed among the areas. The binding of [3H]QNB was expressed as density (fmol/mg protein) and total number of receptors (fmol/total protein) in the area. Considerable variations were observed among the areas. The cerebellum showed the lowest receptor density and a large decrease in density and total number of receptors in the adult, which may reflect a change in neuronal population. Acetylcholinesterase, in certain areas, accompanied the changes in receptor concentration, but the timing and rate of increase had special features in each case. The most striking one was the cerebellum, in which the enzyme increased steadily postnatally, while the muscarinic receptors dropped to very low values.