bluuebunny/crossref_metadata_2025 huggingface dataset:
Activation of Muscarinic and of α1‐Adrenergic Receptors on Astrocytes Results in the Accumulation of Inositol Phosphates
Authors: Pearce Brian , Cambray‐Deakin Martin , Morrow Christine , Grimble Jill , Murphy Sean
Published: October 2006

Abstract: Astrocyte‐enriched cultures prepared from the newborn rat cortex incorporated [3H]myo‐inositol into intracellular free inositol and inositol lipid pools. Noradrenaline and carbachol stimulated the turnover of these pools resulting in an increased accumulation of intracellular [3H]inositol phosphates. The effects of noradrenaline and carbachol were dose‐dependent and blocked by specific α1‐adrenergic and muscarinic cholinergic receptor antagonists, respectively. The increase in [3H]inositol phosphate accumulation caused by these receptor antagonists was virtually unchanged when cultures were incubated in Ca2+ ‐free medium, but was abolished when EGTA was also present in the Ca2+ ‐free medium. Cultures of meningeal fibroblasts, the major cell type contaminating the astrocyte cultures, also accumulated [3H]myo‐inositol, but no increased accumulation of [3H]inositol phosphates was found in response to either noradrenaline or carbachol.
Laminar Distributions of Choline Acetyltransferase and Acetylcholinesterase Activities in the Inner Plexiform Layer of Rat Retina
Authors: Ross C. David , Dunning Daniel D. , Juengel Luann I. , Godfrey Donald A.
Published: October 2006

Abstract: Choline acetyltransferase and acetylcholinesterase activities were measured in samples taken at 7‐μm increments through the inner plexiform layer of rat retina. These enzyme activities were not uniformly distributed through the depth of the inner plexiform layer. Peaks of choline acetyltransferase activity occurred at about onethird and peaks of acetylcholinesterase activity at about one‐fifth of the depth into the inner plexiform layer from either side. The positions of the two peaks of choline acetyltransferase activity most likely correspond to the locations of processes from cholinergic amacrine somata in the inner nuclear layer, which spread in sublamina a, and processes from cholinergic amacrine somata “displaced’ in the ganglion cell layer which spread in sublamina b of the inner plexiform layer. The peaks of acetylcholinesterase activity may in addition correspond to the processes of cholinoceptive amacrine and ganglion cells. The magnitudes of choline acetyltransferase and acetylcholinesterase activities are as high as found anywhere in rat brain, emphasizing the important role of cholinergic mechanisms in visual processing through the rat inner plexiform layer.
Regulation of Serotonin Release from theIn VitroRat Hippocampus: Effects of Alterations in Levels of Depolarization and in Rates of Serotonin Metabolism
Authors: Auerbach Sidney , Lipton Peter
Published: October 2006

Abstract:We analyze the time course of 5‐hydroxytryptamine (5‐HT, serotonin) release from K+‐depolarized hippocampal slices using a two‐compartment kinetic model. The model is based on the assumptions that the rate of release is dependent on the amount of 5‐HT in a releasable pool and that this pool may be resupplied during depolarization by newly synthesized 5‐HT. Comparisons were made between predictions of the model and observed changes in 5‐HT metabolism and in 5‐HT release studied under a variety of experimental conditions. In accordance with predictions of the model, experimental manipulation of 5‐HT synthesis and breakdown rates did not affect release immediately after depolarization but did affect the release rate during prolonged depolarization. Increasing bath tryptophan from 0 to 10 μMapproximately doubled both 5‐HT synthesis and the release rate after 40 min of K+‐induced depolarization while having a smaller effect on release during the first 2 min. Inhibition of 5‐HT breakdown did not significantly affect release during the first 2 min of depolarization but increased it over threefold after 40 min. In contrast, altering the concentrations of K+or Ca2+in the incubation medium affected mainly the early phase of 5‐HT release and not the late phase. Reducing Ca2+from 2.4 to 0.4 mMreduced 5‐HT release by about 30% during the first 9 min of depolarization but did not affect release during the subsequent 30 min. Increasing the concentration of K+from 18 to 60 mMstimulated release by sixfold during the first 2 min but only twofold after a subsequent 30 min. These results support our kinetic model and suggest that regulation of 5‐HT metabolism at the site of the nerve terminal could be a mechanism for modulation of 5‐HT release during prolonged discharge of serotonergic neurons.
Alterations in mRNA Translation Products Associated with Regenerative Responses in the Retina
Authors: Stein‐Izsak C. , Harel A. , Solomon A. , Belkin M. , Schwartz M.
Published: October 2006

Abstract:Translation products of mRNA from retinas of goldfish optic nerve (representing a regenerative CNS) and adult rabbit optic nerve (representing a nonregenerative CNS which can be induced to express regenerative characteristics) were examined by one‐ and two‐dimensional gel electrophoresis. Translation products from retinas of the regenerating goldfish optic nerve included polypeptides barely detectable in the translation products of mRNA derived from retinas of uninjured controls. Some of these polypeptides, of apparent molecular weights 24–28, 43–49, 60, and 65 kilodaltons can be considered as growth‐associated polypeptides described in other regenerative and developing systems. The induction of regeneration‐associated characteristics in the injured adult rabbit optic nerve, “implanted” with diffusible substances from nonneuronal cells of regenerative or growing nerve, is reflected by changes in the mRNA translation products of the retina. Among such translation products are those of the following molecular weights: 16–18, 28, 32–35, 43–47, and 56–60 kilodaltons, and some higher‐molecular‐weight species.
Neurotransmitter Systems in Morphologically Undifferentiated Human Y‐79 Retinoblastoma Cells: Studies of GABAergic, Glycinergic, and β‐Adrenergic Systems
Authors: Madtes Paul , Kyritsis Athanassios , Chader Gerald J.
Published: October 2006

Abstract: Elements of three neurotransmitter systems were investigated in morphologically undifferentiated human Y‐79 retinoblastoma cells in suspension culture. Specific γ‐aminobutyric acid (GABA) uptake, GABA binding, and glycine binding were absent from these cells, although the cells had been shown to exhibit an active uptake and release of [3H]glycine. Binding and competition studies using both α‐and β‐adrenergic ligands indicated the presence of a β‐adrenergic receptor. This finding was confirmed by treatment of the cells with β‐agonists in competition with a β‐antagonist and with an α‐antagonist; the level of cyclic AMP was competitively stimulated. Therefore, human Y‐79 cells in suspension culture contain β‐adrenergic receptors, and not glycinergic or GABAergic ones. Thus, the Y‐79 cells may be of use in studying the factors involved in developmental regulation of neurotransmitter function.
Interaction of [3H](–)‐SKF‐10,047 with Brain σ Receptors: Characterization and Autoradiographic Visualization
Authors: Zukin Stephen R. , Tempel Ann , Gardner Eliot L. , Zukin R. Suzanne
Published: October 2006

Abstract: The σ opiates differ from other opiates in their stimulatory and psychotomimetic actions. The σ opiate [3H](–)‐SKF‐10,047 has been used to characterize σ receptors in rat nervous tissue. Binding of [3H](–)‐SKF10,047 to rat brain membranes was of high affinity, saturable, and reversible. Scatchard analysis revealed the apparent interaction of this drug with two distinct binding sites characterized by affinities of 0.03 and 75 nM (5 mM Tris‐HCI buffer, pH 7.4, at 4°C). Competition analyses involving rank order determinations for a series of opiates and other drugs indicate that the high‐affinity binding site is the μ opiate receptor. The lower‐affinity site (revealed after suppression of μ and δ receptor binding) has been identified as the σ opiate/phencyclidine receptor. In vitro autoradiography has been used to visualize neuronanatomical patterns of receptors labeled using [3H](–)‐SKF‐10,047 in the presence of normorphine and [D‐Ala2,D‐Leu5]enkephalin to block μ and δ interactions, respectively. Labeling patterns differ markedly from those for μ, δ, or K receptors. The highest densities (determined by quantitative autoradiography) are found in the medial portion of the nucleus accumbens, amygdaloid nucleus, hippocampal formation, central gray, locus coeruleus, and the parabrachial nuclei, Receptors in these structures could account for the stimulatory, mood‐altering, and analgesic properties of the σ opiates. Although not the most selective σ opiate ligand, [3H](–)‐SKF‐10,047 binds to σ opiate receptors in brain, and this interaction can be readily distinguished from its interactions with other classes of brain opiate receptors.
Lithium Distribution in the Brain of Normal Mice and of “Quaking” Dysmyelinating Mutants
Authors: Heurteaux Catherine , Baumann Nicole , Lachapelle François , Wissocq Jean‐Claude , Thellier Michel
Published: October 2006

Abstract: Using nuclear reaction 6Li(n,α)3H and dielectric detectors, we have studied the distribution of Li in the brain of adult mice, following Li treatment of the animals. Two strains of animals were used in parallel: “quaking” dysmyelinating mutants and normally myelinated controls. The distribution appeared to be sharply regionalized in the brain of the normal mice (higher Li concentration in the gray rather than in the white matter, with the area postrema being particularly Li rich). In contrast, the Li distribution was practically homogeneous in the brain of the quaking dysmyelinating mutants, with a mean Li concentration comparable to that in the gray matter of the controls. The present method of Li detection has made it possible to estimate the Li equilibrium potentials (nerve cells with regard to plasma) in the different brain substructures. The results are consistent with (a) Li being actively extruded from nerve cells in all the cases and (b) myelination decreasing the relative importance of the passive component of Li transport in the nerve cells, as compared with the active component.
Taurine Levels in Discrete Brain Nuclei of Rats
Authors: Palkovits Miklós , Elekes István , Láng Tibor , Patthy András
Published: October 2006

Abstract Concentrations of taurine have been measured in 44 microdissected rat brain nuclei or areas. Taurine is ubiquitously present and distributed unevenly in the rat brain: the ratio of the highest (pyriform cortex) to lowest (midbrain reticular formation) concentrations is 4.7:1. High taurine levels were found in cerebral cortical areas, caudate‐puta‐men, cerebellum, median eminence, and supraoptic nucleus. Acute pain stress reduced taurine levels in the hypothalamus and the lower brainstem nuclei but not in cortical areas. Increased locomotor and behavioral activities following a high dose of amphetamine elevated taurine concentrations significantly in the substantia nigra and locus ceruleus.
Polyamine Localization and Biosynthesis in Chemically Fractionated Rat Retina
Authors: Ientile Riccardo , Russo Pietro , Macaione Salvatore
Published: October 2006

Abstract For elucidation of polyamine localization and biosynthesis in various cell types of rat retina, the putres‐cine, spermidine, and spermine contents as well as the or‐nithine decarboxylase and S‐adenosylmethionine decar‐boxylase activities have been measured in retinal cell layers obtained by the selective cytotoxic action of iodoacetate on photoreceptor cells and of monosodium glutamate on higher‐order retinal neurons. A notable depletion only in spermine content was associated with loss of the visual cell layer. Total ornithine decarboxylase and S‐adenosylmethionine decarboxylase activities per retina were significantly lower in all chemically fractionated tissue, but loss of the photoreceptor layer produced the greatest decrease. The specific activities of these enzymes did not show marked changes in rat retinas deprived of inner neurons. The data support the suggestions that polyamine synthesis, storage, and catabo‐lism have different distributions in the retinal layers and that the spermine levels and the high value of the spermine/ spermidine molar ratio might depend essentially on the proportion of rods to cones.
The Dopamine Metabolite 3‐Methoxytyramine Is Not a Suitable Indicator of Dopamine Release in the Rat Brain
Authors: Vulto Arnold G. , Westenberg Herman G. M. , Meijer L. Bert A. , Versteeg Dirk H. G.
Published: October 2006

Abstract It has been postulated that changes in the concentration of 3‐methoxytyramine (3‐MT) in the brain might reflect changes in the release of 3,4‐dihydroxyphenylethylamine (DA, dopamine) and, therefore, might be used as an index of dopaminergic activity in the brain. 3‐MT is known to accumulate rapidly after death. Killing by microwave irradiation (MWR) is considered to be the method of choice to obtain “undisturbed” 3‐MT concentrations. We measured striatal 3‐MT concentrations even lower than those following MWR when the brains were excised and frozen in dry ice very rapidly (typical time between decapitation and freezing of the brain 22 s). There was a linear increase in striatal 3‐MT concentration when the time between decapitation and freezing was varied between 13 and 300 s. Extrapolation to time zero indicated negligible amounts of 3‐MT at the time of decapitation. In addition, it was observed that DA, 3,4‐dihydroxyphenylacetic acid, and homovanillic acid decompose during the cooling phase after heating the brain by microwave. It is concluded that MWR induces arti‐factual changes in the postmortem levels of DA and metabolites. Consequently 3‐MT cannot be considered to be a reliable indicator of DA release in the rat brain.
Evidence for a Selective Localization of Voltage‐Sensitive Ca2+ Channels in Nerve Cell Bodies of Corpus Striatum
Authors: Sanna E. , Head G. A. , Hanbauer I.
Published: October 2006

Abstract Specific binding sites for [3H]nitrendipine, an organic Ca2+ channel antagonist, were abolished in crude syn‐aptosomal membranes of kainic acid‐lesioned caudate nuclei. In contrast, specific lesions of dopaminergic or serotonergic axon terminals in caudate nuclei failed to alter the density or the affinity of [3H]nitrendipine binding sites. In addition, the basal and veratridine‐stimulated 45Ca2+ accumulations were greatly impaired in slices prepared from kainic acid‐lesioned caudate nuclei. The veratridine‐elicited accumulation of 45Ca2+ in control slices was attenuated by addition of tetrodotoxin in the incubation medium. The present data provide evidence that most of the [3H]nitrendipine binding sites and the voltage‐dependent Ca2+ channels are located in intrinsic neurons or interneurons in caudate nucleus. In contrast, destruction of dopaminergic or serotonergic nerve terminals emanating from other brain areas and innervating the caudate nucleus failed to change the apparent Bmax value for [3H]nitrendipine binding.
A Kinetic Study of Stimulus‐Induced Vesicle Recycling in Electromotor Nerve Terminals Using Labile and Stable Vesicle Markers
Authors: Ágoston D. V. , Dowe G. H. C. , Fiedler W. , Giompres P. E. , Roed I. S. , Walker J. H. , Whittaker V. P. , Yamaguchi T.
Published: October 2006

Abstract The kinetics of recovery, by recycling electromotor synaptic vesicles, of the biophysical parameters of the reserve population has been studied in perfused blocks of electric organ of Torpedo marmorata prestimulated in vivo, followed by density gradient separation of the extracted vesicles in a zonal rotor using labile (acetylcholine and ATP) and stable (proteoglycan) vesicle markers. Stimulation in vivo at 0.15 Hz for 3.3 h depleted tissue acetylcholine much less than stimulation at 1 Hz for 1 h but nevertheless generated a much larger pool of recycled vesicles that recovered more slowly. At the lower rate of stimulation, recovery of the biophysical characteristics of the reserve population by the recycled vesicles, identified by their content of newly synthesized transmitter, was essentially complete by 8 h. The stable proteoglycan marker was immunochemically assayed and was bimodally distributed in the vesicle‐containing portion of the density gradient even in experiments with unstimulated or recovered tissue. The second peak corresponded with that of newly synthesized transmitter and was thus identified as containing the recycled vesicles. Its normalized acetylcholine/proteoglycan ratio was lower than that of the first peak, which is consistent with earlier findings that recycled vesicles, before recovery, are only partially loaded with transmitter. However, as expected, the proportion of total vesicular proteoglycan and acetylcholine associated with the recycled vesicle fraction was very much lower in preparations derived from unstimulated or recovered tissue than in those from recently stimulated tissue.
Release of Ethanolamine, but Not of Serine or Choline, in Rat Pontine Nuclei on Stimulation of Afferents from the Cortex, In Vivo
Authors: Perschak Henry , Wolfensberger Max , Do Kim Quang , Dunant Yves , Cuénod Michel
Published: October 2006

Abstract: Release of ethanolamine, serine, and choline in rat pontine nuclei on electrical stimulation of afferents from the cortex was investigated using in vivo push‐pull cannula techniques. Ethanolamine was determined by using gas chromatographic techniques; serine was measured with a HPLC system; and choline was assayed with a luminescence method. Resting elution rates of ethanolamine, serine, and choline were 50.8 ± 8.4, 34.8 ± 12.6, and 1.16 ± 0.20 pmol/5 min, respectively. Stimulation of the cortico‐pontine tract evoked a highly significant 3,4‐fold increase in release of ethanolamine, whereas serine and choline release was unaffected. Reactions in membrane phospholipids are most likely involved in the stimulation‐dependent release of ethanolamine and special consideration was given to base‐exchange reactions. Alternatively, a release from intracellular, possibly synaptic stores cannot be excluded.
Differential Changes in the Content of Amino Acid Neurotransmitters in Discrete Regions of the Rat Brain Prior to the Onset and During the Course of Homocysteine‐Induced Seizures
Authors: Allen Ian C. , Grieve Angus , Griffiths Roger
Published: October 2006

Abstract: Changes in amino acid concentrations were investigated in selected regions of rat brain prior to the onset and during the course of epileptiform seizures induced by l‐homocysteine. The concentration of γ‐aminobutyric acid (GABA) decreased preictally in substantia nigra (‐ 18%), caudate putamen (‐ 26%), and inferior colliculus (‐ 46%). After seizure onset, the GABA content was further reduced in substantia nigra (‐ 31%) and additionally in hippocampus (‐ 18%). Preictal taurine levels were elevated in globus pallidus (+ 26%) and caudate putamen (+ 13%) but returned to normal after seizure onset. However, in hippocampus, taurine decreased both preictally (‐ 22%) and after seizure onset (‐ 56%). Glycine was reduced preictally only in globus pallidus (‐ 13%). After seizure onset the direction of its concentration change varied in the brain regions studied. Glutamate levels decreased preictally in hippocampus (‐ 10%) and hypothalamus (‐ 46%) but increased in globus pallidus (+ 14%). Normal levels were detectable after seizure onset in hypothalamus and globus pallidus but a further reduction in hippocampus (‐ 59%) and significant reductions in substantia nigra (‐ 15%) and caudate putamen (‐ 17%) were detected. Aspartate was elevated in hippocampus, both preictally (+ 49%) and after seizure onset (+ 21%) while at the same phases in globus pallidus a consistent reduction (‐ 30%) was observed. The glutamine content increased preictally in globus pallidus (+ 41%) and hypothalamus (+ 36%), and in all brain areas during the ictal phase of seizure, the hippocampus exhibiting a dramatic increase (∼300%). The contents of serine and alanine were altered in most regions studied only after seizure onset, with the exception of the hippocampus, where a decrease (‐ 41%) of serine was observed preictally.
Effect of Adenosine, Adenosine Derivatives, and Caffeine on Acetylcholine Release from Brain Synaptosomes: Interaction with Muscarinic Autoregulatory Mechanisms
Authors: Pedata Felicita , Giovannelli Lisa , De Sarno Patrizia , Pepeu Giancarlo
Published: October 2006

Abstract: Synaptosomes, prepared from rat cerebral cortex and hippocampus, were preincubated with [methyl‐3H]choline. The effect of adenosine, cyclohexyl‐adenosine, N‐ethylcarboxamide adenosine, 2′‐deoxyadenosine, and oxotremorine on K+‐evoked 3H efflux was investigated. High‐voltage electrophoretic separation showed that in the presence of physostigmine, the K+‐evoked 3H efflux from hippocampal synaptosomes was 90% [3H]acetylcholine and 10% [3H]choline. Adenosine (30 μM) and oxotremorine (100 μM) both decreased [3H]acetylcholine release from hippocampal synaptosomes. The effect was inversely proportional to the KCI concentration and disappeared at a KCI concentration of 50 mM. Cyclohexyladenosine was ∼3,000 times more active then adenosine, whereas N‐ethylcarboxamide adenosine and 2′‐deoxyadenosine were inactive. This indicates that A1 adenosine receptors were involved in the inhibitory effect. Caffeine antagonized the adenosine effect, and at a concentration of 100 μM, it stimulated [3H]acetylcholine efflux. The inhibitory effect of oxotremorine was as great in cortical as in hippocampal synaptosomes. In contrast, adenosine was much less active in cortical than in hippocampal synaptosomes. When inhibitory concentrations of adenosine and oxotremorine were added together into the incubation medium, the effect of adenosine on [3H]acetylcholine release was consistently reduced. An interaction between muscarinic and A1 adenosine presynaptic receptors at a common site modulating acetylcholine release can be assumed.
Regulation of Nerve Growth Factor Action on Nsp100 Phosphorylation in PC12h Cells by Calcium
Authors: Hashimoto Seiichi , Iwasaki Chieko , Kuzuya Hiroshi , Guroff Gordon
Published: October 2006

Abstract: Previous work from these laboratories has shown that in PC12 cells the phosphorylation of a specific soluble protein is decreased by treatment with nerve growth factor. This protein, designated Nsp100, and its kinase have been separated and partially purified from PC12 cells. The present studies have been designed to investigate the role of calcium in this action of nerve growth factor. It is shown here, using PC12h cells, that A23187, a calcium ionophore, and high levels of K+, a depolarizing stimulus, also decrease phosphorylation of Nsp100. Furthermore, the actions of nerve growth factor as well as those of A23187 and high levels of K+ are prevented by treatment of the cells with the calcium chelator EGTA. It is also shown that agents that raise levels of cyclic AMP in the cells, specifically dibutyryl cyclic AMP and cholera toxin, also decrease phosphorylation of Nsp100 but, in addition, increase phosphorylation of tyrosine hydroxylase. The action of these latter agents on Nsp100 is blocked by EGTA, but their action on tyrosine hydroxylase is not, indicating that even agents such as cholera toxin act on Nsp100 through a Ca2+‐dependent mechanism.
Activation of Myelin Protein Kinase by Diacylglycerol and 4β‐Phorbol 12‐Myristate 13‐Acetate
Authors: Murray Norman , Steck Andreas J.
Published: October 2006

Abstract: In an effort to gain a more complete understanding of the regulation of myelin basic protein phosphorylation, we have been interested in defining further the mode of regulation of the myelin protein kinase involved in this posttranslational modification. Here we report the partial purification of a protein kinase from rat brain myelin. By gel filtration, it was determined that the molecular weight of this enzyme was in the range of 70–80 × 103 daltons. Furthermore, it was established that at low calcium concentrations, this enzyme was markedly activated by phosphatidylserine in combination with either 4β‐phorbol 12‐myristate 13‐acetate or diolein. The enzyme was not affected by cyclic AMP or by calcium, alone or in combination with calmodulin. On the basis of these findings this enzyme can be identified as a protein kinase C‐like enzyme.
Amino Acid Changes in Autopsied Brain Tissue from Cirrhotic Patients with Hepatic Encephalopathy
Authors: Lavoie Joël , Giguère Jean‐Francois , Layrargues Gilles Pomier , Butterworth Roger F.
Published: October 2006

Abstract: Brain tissue was obtained at autopsy from nine cirrhotic patients dying in hepatic coma and from an equal number of controls, free from neurological, psychiatric, or hepatic diseases, matched for age and time interval from death to freezing of dissected brain samples. Glutamine, glutamate, aspartame, and γ‐aminobutyric acid (GABA) levels were measured in homogenates of cerebral cortex (prefrontal and frontal), caudate nuclei, hypothalamus, cerebellum (cortex and vermis), and medulla oblongata as their O‐phthalaldehyde derivatives by HPLC using fluorescence detection. Glutamine concentrations were found to be elevated two‐ to fourfold in all brain structures, the largest increases being observed in prefrontal cortex and medulla oblongata. Glutamate levels were selectively decreased in prefrontal cortex (by 20%), caudate nuclei (by 27%), and cerebellar vermis (by 17%) from cirrhotic patients. On the other hand, GABA content of autopsied brain tissue from these patients was found to be within normal limits in all brain structures. It is suggested that such region‐selective reductions of glutamate may reflect loss of the amino acid from the releasable (neurotransmitter) pool. These findings may be of significance in the pathogenesis of hepatic encephalopathy resulting from chronic liver disease. Key Words: Hepatic encephalopathy—Hyperammonemia— Cerebral amino acids—Glutamine—Glutamate—γ‐Aminobutyric acid. Lavoie J. et al. Amino acid changes in autopsied brain tissue from cirrhotic patients with hepatic encephalopathy.
Purification of a Ciliary Neurotrophic Factor from Bovine Heart
Authors: Walters Diane J. , Hendry Ian A.
Published: October 2006

Abstract: A neurotrophic factor that promotes the survival of cholinergic parasympathetic ciliary neurons has been purified ∼20,000‐fold from bovine cardiac tissue under nondenaturing conditions using heparin‐affinity chromatography. Up to 22 μg of purified factor having a specific activity of 4 × 105 trophic units/mg can be obtained from 250 g of heart muscle. Sodium dodecyl sulfate (SDS)‐polyacrylamide gels of the purified material show a broad band that is sometimes resolvable into a closely spaced pair of bands of 22 and 23 kilodaltons. Partially purified factor can be resolved into two peaks of activity (pI 5.6 and 5.0) by highresolution anion‐exchange chromatography and chromato focusing, although these procedures have not proved useful as purification methods because of the large losses of activity incurred. It is likely that these two peaks represent the two bands seen on SDS‐polyacrylamide gels. The bovine cardiac factor(s) differs from similar factors purified from chick optic tissues and pig brain in that it is irreversibly denatured by SDS.
Increase of Catecholamines in Mouse Brain by Systemic Administration of γ‐Glutamyl L‐3,4‐Dihydroxyphenylalanine
Authors: Ichinose Hiroshi , Togari Akifumi , Suzuki Hidenori , Kumagai Hideaki , Nagatsu Toshiharu
Published: October 2006

Abstract: We investigated the effect of systemic administration of γ‐glutamyl L‐3,4‐dihydroxyphenylalanine (γ‐Glu‐DOPA) on catecholamine contents in the brain. γ‐Glu‐DOPA was transformed to dopamine (DA) in vitro with brain homogenate by the sequential action of γ‐glutamyl transpeptidase and aromatic L‐amino acid decarboxylase. Intraperitoneal injection of γ‐Glu‐DOPA to mice increased DA markedly and noradrenaline (NA) moderately in the brain. The increase of endogenous DA was followed by elevation of the main DA metabolites (3,4‐dihydroxyphenyl‐acetic acid and homovanillic acid). These increases were in a dose‐dependent manner. The maximal elevation of DA was observed within 30 min after administration of γ‐Glu‐DOPA, but a substantial increase of NA was observed 2 h after the administration. These results suggest that γ‐Glu‐DOPA may be applicable to the treatment of Parkinson's disease.
Composition of Lipids in Elasmobranch Electric Organ and Acetylcholine Receptor Membranes
Authors: Rotstein Nora P. , Arias Hugo R. , Barrantes Francisco J. , Aveldaño M. I.
Published: October 2006

Abstract: The composition of phospholipids from electric organ and from membranes enriched in acetylcholine receptors (AChRs) is analyzed in three elasmobranch fish (Torpedo marmorata, Torpedo californica, and Discopyge tschudii). Irrespective of their purity, AChR‐containing membranes are similar to electric organ in lipid and fatty acid composition. The following characteristics are common to the three species: (a) Choline, ethanolamine, and serine glycerophospholipids account for 80–90% of the phospholipids. (b) Their major fatty acid constituents are monoenes, saturates, and long‐chain (n‐3) polyenes (especially docosahexaenoate). (c) A large proportion of (he ethanolamine glycerophospholipids (30–50%) is made up by plasmenylethanolamine, which contains fewer polyenes than phosphatidylethanolamine per mole of lipid. (d) Poly‐phosphoinositides represent 20–30% of the inositides of electric organ, (e) Phosphatidylinositol and phosphatidate have large proportions of 20‐and 22‐carbon polyenes. (f) Diphosphatidylglycerol and triacylglycerols are rich in oleate but also contain long‐chain polyenes. (g) Sphingo‐myelin has monoenes and saturates ranging from 14 to 26 carbons. Species‐related variations are observed (a) in the ratios between some phospholipid classes and subclasses and (b) in the relative abundance of the major polyunsatu‐rated acyl chains of phospholipids. Despite these differences, the average unsaturation and length of fatty acids in major phospholipid classes are similar for the three species.
S‐100 Protein in Clonal Astroglioma Cells Is Released by Adrenocorticotropic Hormone and Corticotropin‐Like Intermediate‐Lobe Peptide
Authors: Suzuki Fujiko , Kato Kanefusa , Kato Taiji , Ogasawara Nobuaki
Published: October 2006

Abstract: S‐100 protein in clonal GA‐1 and C6 rat glioma cell lines was released in serum‐free medium supplemented with adrenocorticotropic hormone (ACTH). The induction of S‐100 protein release by ACTH was dose‐dependent, showing a half‐maximal release at about 5 μM, and the S‐100 protein concentration in the medium increased sharply within 3 min, but slightly during further incubation. The S‐100 protein release was apparently accompanied by a decrease in the membrane‐bound form of S‐100 protein in the cell. The S‐100 protein release was induced not by the ACTH1–24 fragment, which exhibits the known effects of ACTH, but by the ACTH18–39 fragment, which is designated as corticotropin‐like intermediate‐lobe peptide (CLIP). These results indicate that the C‐terminal half of ACTH is responsible for the S‐100 protein release. The enhancement of S‐100 protein release by ACTH was also observed in normal rat glioblasts. The release induced by ACTH was apparently specific to S‐100 protein, because little release of the cytoplasmic enzymes, creatine kinase, and enolase was observed under the same conditions. High concentrations (5 mM) of dibutyryl cyclic AMP or dibutyryl cyclic GMP were also found to induce S‐100 protein release; however, catecholamines (epinephrine, norepi‐nephrine, isoproterenol, and dopamine), acetylcholine, and glutamic acid did not enhance the release. The S‐100 protein release was also stimulated by 5 mM ATP, ADP, and GTP, probably by lowering the concentration of Ca2+ in the medium, because addition of Ca2+ inhibited the release by the nucleotides (but not by the cyclic nucleotides or ACTH), and 1 mM EGTA could induce the S‐100 protein release from GA‐1 glioma cells.
Characterization of Two [3H]Ketanserin Recognition Sites in Rat Striatum
Authors: Roth B. L. , McLean S. , Zhu X.‐Z. , Chuang D.‐M.
Published: October 2006

Two [3H]ketanserin recognition sites are present in the rat striatum. The high‐affinity site (Kd, 0.39 nM) is similar to the 5‐hydroxytryptamine2 (5‐HT2) site previously characterized by various investigators. The low‐affinity site (Kd, 21.8 nM) has a unique pharmacologic specificity and is preferentially localized to rat striatum and septum. Conventional 5‐HT2 antagonists as well as 5‐HT and 5‐HT uptake inhibitors are ineffective at inhibiting [3H]ketanserin binding to this low‐affinity site. Also, chronic treatment with p‐chlorophenylalanine, which depletes brain 5‐HT, upregulates only the high‐affinity site. Thus, in the striatum and septum, [3H]ketanserin labels a unique recognition site. This site has recently been shown to be associated with dopaminergic nerve endings and may regulate biogenic amine release.
Expression of Glial Fibrillary Acidic Protein in Rat C6 Glioma Relates to Vimentin and Is Independent of Cell‐Cell Contact
Authors: Backhovens H. , Gheuens J. , Siegers H.
Published: October 2006

Glial fibrillary acidic protein (GFAP) was induced in rat C6 glioma cells grown in M199 and HAM F10 media by addition of 1 mM dibutyryl cyclic AMP. The amount of GFAP per cell increased 7‐and 33‐fold in M199 and HAM F10 media, respectively. GFAP could be induced in each phase of the cell culture except for the lag phase, where GFAP synthesis was delayed until the onset of the logarithmic growth. The induction took place under conditions where the total protein content of the cell decreased. Mear surement of the amount of vimentin indicated that GFAP was induced under conditions of low vimentin concentration. Our results do not support the hypothesis that GFAP induction depends on cell‐cell contact or cell proliferation. They indicate a shift from vimentin to GFAP synthesis by an as yet unknown mechanism.
Thiamine Triphosphate and Membrane‐Associated Thiamine Phosphatases in the Electric Organ of Electrophorus electricus
Authors: Bettendorff Lucien , Michel‐Cahay Colette , Grandfils Christian , Rycker Cécile De , Schoffeniels Ernest
Published: October 2006

The main electric organ of Electrophorus electricus is particularly rich in thiamine triphosphate, which represents 87% of the total thiamine content in this tissue. The thiamine pyrophosphate concentration, however, is very low in the eel electric organ and skeletal muscle as compared with other eel or rat tissues. Furthermore, electroplax membranes contain a whole set of enzymes responsible for the dephosphorylation of thiamine tri‐, pyro‐, and monophos‐phate. Thiamine triphosphatase has a pH optimum of 6.8 and is dependent on Mg2+. The real substrate of the enzyme is probably a 1:1 complex of Mg2+ and thiamine triphosphate. Thiamine pyrophosphatase is activated by Ca2+. The apparent Km for thiamine triphosphate and Vmax are found to be, respectively, 1.76 mM and 5.95 nmol/mg of protein/min. Thiamine triphosphatase activity is inhibited at physiological K+ concentrations (up to 90 mM) and increasing Na+ concentrations (50% inhibition at 300 mM). ZnCl2 (10 mM) inhibits 90% of the enzyme activity. ATP and ITP are also strongly inhibitory. No significant effect of neurotoxins is seen. Membrane‐associated thiamine triphosphatase is affected differently by proteolytic enzymes and is partially inactivated by pretreatment with phospholipase C and neuraminidase. The physiological significance of thiamine triphosphatase is discussed in relation to a specific role of thiamine in the nervous system.
Nature and Immunochemical Characteristics of a Ca2+/Calmodulin Kinase Activity Endowed in a Highly Insoluble Protein Purified from Adult Rat Brain
Authors: Bezamahouta C. , Zanetta J. P. , Revel M. O. , Zwiller J. , Meyer A. , Malviya A. N. , Vincendon G.
Published: October 2006

Using sequential extraction procedure of proteins from adult rat forebrain, a protein of Mr 52,000, insoluble in neutral detergents, capable of binding calmodulin in the presence of Ca2+, was isolated. Antibodies to this antigen had the capacity to inhibit the Ca2+/calmodulin‐dependent kinase activity associated with this protein. This protein (52K) (in many respects identical to the major protein of postsynaptic densities) shares by itself the Ca2+/calmodulin‐dependent kinase activity, thus differing from soluble Ca2+/calmodulin‐dependent kinases isolated by others. Despite its insolubility in most detergents, the 52K protein is not particularly rich in hydrophobic amino acids. Its richness in cysteine and proline residues suggests that the active conformation of the enzyme is sustained by numerous disulfide bridges.
Uptake, Release, and Subcellular Localization of l‐Methyl‐4‐Phenylpyridinium in Blood Platelets
Authors: Cesura A. M. , Ritter A. , Picotti G. B. , Da Prada M.
Published: October 2006

Abstract: : The neurotoxic compound 1‐[methyl‐3H]‐4‐phenylpyridinium ([3H]MPP+) was actively taken up by human, rabbit, and guinea pig platelets incubated in plasma. In human platelets, the apparent Km of this uptake (22.6 μM) was 50 times higher than that for serotonin [5‐hydroxytryptamine (5‐HT)]. The uptake of [3H]MPP+ by human platelets was inhibited by selective 5‐HT uptake blockers [cianopramine, (−)‐paroxetine, and clomipramine], by metabolic inhibitors (KCN and ouabain), and by drugs that interfere with amine storage in the 5‐HT organelles (reserpine, mepacrine, and Ro 4–1284). Impairment of the trans‐membrane proton gradient by ionophores (monensin and nigericin) induced a marked release of radioactivity from platelets preincubated with [3H]MPP+. Fractionation of homogenates of rabbit platelets preincubated with [3H]MPP+ showed that the drug was concentrated to a great extent in the 5‐HT organelle fraction. MPP+ competitively inhibited [14CJ5‐HT uptake by human platelets and reduced the endogenous 5‐HT content of human, rabbit, and guinea pig platelets. These investigations show that MPP+ is transported into the platelets via the 5‐HT carrier and is accumulated predominantly in the subcellular organelles that store 5‐HT and other monoamines. It is suggested that an accumulation of MPP+ in amine storage vesicles of neurons may be involved in the effects of the drug in the CNS, e.g., by protecting other subcellular compartments from exposure to high concentrations of MPP+, by sustaining a gradual release of the toxin, or both.
Increased Fucosylation of Chick Brain Proteins Following Training: Effects of Cycloheximide
Authors: McCabe Norah , Rose Steven P. R.
Published: October 2006

Abstract: When chicks are trained to avoid pecking a bead coated with methylanthranilate in a one‐trial passive avoidance task there is an increase in fucose incorporation in vivo and in vitro in the right forebrain base of methylanthranilate (M)‐trained compared to water (W)‐trained chicks. The relation of this increase to de novo protein synthesis in vivo and in vitro has been examined. Cycloheximide (Cx), 1 mM, inhibited in vitro fucosylation of chick brain slices by 60% after 3 h. However, the training‐related increase in in vitro fucosylation still persisted. When Cx was injected intraventricularly 10 min before training, the subsequent increase in in vitro fucosylation due to training was still apparent. When Cx was injected and [14C]leucine and [3H]fucose incorporation studied in vivo in M‐trained and W‐trained chicks, there was no increase in fucosylation due to training in the Cx‐treated M‐trained over the W‐trained chicks. These results are taken to indicate that in vitro fucosylation and its increase subsequent to training is not protein synthesis‐dependent, but that both in vivo and in vitro there are interactions between Cx and fucosylation steps that are independent of Cx's effects on protein synthesis.
Inhibition of Monoamine Oxidase by N‐Methylisoquinolinium Ion
Authors: Naoi Makoto , Hirata Yoko , Nagatsu Toshiharu
Published: October 2006

Abstract: N‐Methylisoquinolinium ion (N‐MIQ) has been found to inhibit the biosynthesis of catecholamines; it inhibited tyrosine hydroxylase activity in striatal tissue slices. In this article, the effects of N‐MIQ and an analogue, N‐methylquinolinium ion, on monoamine oxidase (MAO) activity were examined to see their effects on the catabolism of catecholamines. MAO‐A in human placental mitochondria was strongly inhibited by N‐MIQ in competition with the substrate. The apparent Ki value of N‐MIQ was found to be 20.4 ±1.1 μM, whereas that of N‐methylquinolinium ion was 54.6 ± 4.5 μM. MAO‐B in human brain synaptosomes and liver mitochondria was found to be inhibited by N‐MIQ, but the inhibition proved to be noncompetitive. The inhibition of MAO‐B by N‐MIQ was completely reversible by dialysis of the incubation mixture. MAO‐A in human brain and liver mitochondria was more sensitive to the inhibitor than MAO‐B. By quantitative analysis of N‐MIQ, using HPLC, it was found not to be catabolized by the incubation with mitochondria, suggesting that the inhibition was due to N‐MIQ itself and not due to any metabolic product. The inhibition of MAO by N‐MIQ is discussed in terms of its possible involvement of the etiology of parkinsonism.
Phosphorylation of Tyrosine Hydroxylase by Cyclic GMP‐Dependent Protein Kinase
Authors: Roskoski Robert , Vulliet P. Richard , Glass David B.
Published: October 2006

Abstract: Tyrosine hydroxylase purified from rat pheochro‐mocytoma was phosphorylated and activated by purified cyclic GMP‐dependent protein kinase as well as by cyclic AMP‐dependent protein kinase catalytic subunit. The extent of activation was correlated with the degree of phosphate incorporated into the enzyme. Comparable stoichio‐metric ratios (0.6 mol phosphate/mol tyrosine hydroxylase subunit) were obtained at maximal concentrations of either cyclic AMP‐dependent or cyclic GMP‐dependent protein kinases. The enzymes appeared to mediate the phosphorylation of the same residue based on the observation that incorporation was not increased when both enzymes were present. The major tryptic phosphopeptide obtained from tyrosine hydroxylase phosphorylated by each protein kinase exhibited an identical retention time following HPLC. The purified phosphopeptides also exhibited identical isoelectric points. These data provide support for the notion that the protein kinases are phosphorylating the same residue of tyrosine hydroxylase.
Phospholipid Metabolism in Mouse Sciatic Nerve In Vivo
Authors: Gould Robert M. , Connell Fred , Spivack Warren
Published: October 2006

Abstract: To probe the activities of various pathways of lipid metabolism in peripheral nerve, six phospholipid‐directed precursors were individually injected into the exposed sciatic nerves of adult mice, and their incorporation into phos‐pholipids and proteins was studied over a 2‐week period. Tritiated choline, inositol, ethanolamine, serine, and glyc‐erol were mainly used in phospholipid synthesis; in contrast, methyl‐labeled methionine was primarily incorporated into protein. Phosphatidylcholine was the main lipid formed from tritiated choline, glycerol, and methionine precursors. Phosphatidylserine, phosphatidylethanolamine, and phosphatidylinositol were the main lipids formed from serine, ethanolamine, and inositol, respectively. With time there was a shift in label among phospholipids, with higher proportions of choline appearing in sphingomyelin, glycerol in phosphatidylserine, ethanolamine in phosphatidylethanolamine (plasmalogen), and inositol in polyphosphoinosi‐tides, especially phosphatidylinositol 4,5‐bisphosphate. We suggest that the delay in formation of these phospholipids, which are concentrated in peripheral nerve myelin, may, at least in part, be due to their formation at a site(s) distant from the sites where the bulk of Schwann cell lipids are made. We propose that separating the synthesis of these my‐elin‐destined lipids to near the Schwann cell's plasma membrane would facilitate their concentration in peripheral nerve myelin sheaths. At earlier labeling times, ethanolamine and glycerol were more actively incorporated into phosphatidylcholine and phosphatidylinositol, respectively, than later. The transient labeling of these phospholipids may reflect some unique role in peripheral nerve function.
A Ca2+‐Dependent Protein Kinase Activity Associated with Serotonin Binding Protein
Authors: Adlersberg Mella , Liu Kuo‐Peing , Hsiung Shu‐Chi , Ehrlich Yigal , Tamir Hadassah
Published: October 2006

Abstract: The endogenous phosphorylation of serotonin binding protein (SBP), a soluble protein found in central and peripheral serotonergic neurons, inhibits the binding of 5‐hydroxytryptamine (5‐HT, serotonin). A protein kinase activity that copurifies with SBP (SBP‐kinase) was partially characterized and compared with calcium/calmodulin‐dependent protein kinase II (CAM‐PK II). SBP itself is not the enzyme since heating destroyed the protein kinase activity without affecting the capacity of the protein to bind [3H]5‐HT. SBP‐kinase and CAM‐PK II kinase shared the following characteristics: (1) size of the subunits; (2) autophosphorylation in a Ca2+‐dependent manner; and (3) affinity for Ca2+. In addition, both forms of protein kinase phosphorylated microtubule‐associated proteins well and did not phosphorylate myosin, phosphorylase b., and casein. Phorbol esters or diacylglycerol had no effect on either of the protein kinases. However, substantial differences between SBP‐kinase and CAM‐PK II were observed: (1) CAM enhanced CAM‐PK II activity, but had no effect on SBP‐kinase; (2) synapsin I was an excellent substrate for CAM‐PK II, but not for SBP‐kinase; (3) 5‐HT inhibited both the autophosphorylation of SBP‐kinase and the phosphorylation of SBP, but had no effect on CAM‐PK II. These data indicate that SBP‐kinase is different from CAM‐PK II. Phosphopeptide maps of SBP and SBP‐kinase generated by digestion with S. aureus V8 protease are consistent with the conclusion that these proteins are distinct molecular entities. It is suggested that phosphorylation of SBP may regulate the transport of 5‐HT within neurons.
A Comparative Study and Partial Characterization of Multi‐Uptake Systems for γ‐Aminobutyric Acid
Authors: Wood J. D. , Sidhu H. S.
Published: October 2006

Abstract: Previous work by the authors had indicated that synaptosome‐enriched preparations from the cerebral cortex of the rat contained a high‐, a medium‐, and a low‐affinity uptake system for γ‐aminobutyric acid (GABA). The present study demonstrated that this phenomenon also prevailed in synaptosomes from rat diencephalon, mesencephalon, and cerebellum, although the Fmax values for the high‐and medium‐affinity systems in the cerebellum were very low relative to those of the other regions. When a different type of preparation containing nerve endings (glomeruli) was obtained from the cerebellum, it possessed a Vmax value for the high‐affinity system that was more similar to that for the corresponding system in synaptosomes from the other brain regions. In contrast to the above situation, synaptosomes from rat olfactory bulb lacked the low‐affinity uptake system, as did synaptosomes from dog olfactory bulb. The aspartate/glutamate uptake systems, as measured with D‐aspartate, provided a regional pattern quite different from those of GABA uptake. Only two uptake systems, a high‐and a low‐affinity system, were observed in all regions tested. All three GABA uptake systems were present in cortical synaptosomes from the mouse, hamster, and guinea pig, and all three systems were sodium dependent, energy dependent, temperature sensitive, and totally inhibited by nipecotic acid.