bluuebunny/crossref_metadata_2025_split huggingface dataset:
Antibodies Against Synthetic Peptides Predicted from the Nucleotide Sequence of D2 Receptor Recognize Native Dopamine Receptor Protein in Rat Striatum
Authors: Farooqui Shakeel M. , Brock Jeffery W. , Hamdi Anwar , Prasad Chandan 
Published: October 2006

AbstractTwo peptides corresponding to amino acid sequences predicted from the nucleotide sequence of the dopamine D2 receptor were synthesized. Peptide I (CGSEG‐KADRPHYC) and peptide II (NNTDQNECIIY), corresponding to 24–34 and 176–185 from the NH2 terminus, respectively, were conjugated to keyhole limpet hemocyanin and injected into rabbits. Peptide I showed a greater immunogenic response than did peptide II. Both peptide antibodies exhibited high titer for the homologous antigens, but showed little or no cross‐reactivity with heterogeneous peptides. Peptide I antibodies reacted with striatal membrane proteins of apparent molecular masses of 120, 90, 85, and 30 kDa on a western blot. Furthermore, the 90‐kDa band was identified as denatured D2 receptor by its high affinity for the D2 selective photoaffinity probe 125I‐AP‐azidospiperone (125I‐NAPS). Photoaffinity labeling of the 90‐kDa protein by 125I‐NAPS was reduced by 40% in the presence of the peptide I antibody. In addition, evidence is also presented to show the low level of 90‐kDa protein in cerebellum which contains little or no D2 ligand binding sites. The antibody to peptide I inhibited the binding of [3H] YM‐09151‐2, a dopamine D2 receptor selective antagonist, to striatal membranes in a concentration‐dependent manner, a 50% inhibition was obtained at a 1:500 dilution of the antisera with 20 pM ligand concentration. The data on the equilibrium inhibition kinetics of [3H] YM‐09151‐2 binding to striatal membranes were examined in the presence of antibody and showed a 25–30% decrease in Bmax (203.5 ± 11.0 and 164.6 ±3.3 fmol/mg of protein in presence of preimmune and immune sera, respectively) with no change in KD. These results suggest that polyclonal antisera raised against peptide I exhibited specific antibodies for the dopamine D2 receptor protein. The primary epitope for this antibody is at or near the ligand binding site which can be recognized in both denatured and native receptor protein in striatal membranes.
Quantitative and Qualitative Alterations of Neuronal and Glial Intermediate Filaments in Rat Nervous System After Exposure to 2, 5‐Hexanedione
Authors: Karlsson J.‐E. , Rosengren L. E. , Haglid K. G. 
Published: October 2006

AbstractThe precise mechanism for the neurotoxicity of 2, 5‐hexanedione is not known, but cross‐linking of neurofilament proteins has been suggested as one possibility. In this study the effects of long‐term exposure to 2, 5‐hexanedione were studied in the rat nervous system with special reference to regional changes in the quantities of neuronal and glial intermediate filaments. Using enzyme‐linked immunosorbent assays the concentrations of 68‐ and 200‐kDa neurofilament polypeptides were shown to be reduced in all brain regions studied. Similar results were obtained in the sciatic nerve. The concentration of glial fibrillary acidic protein was decreased in the cerebellar vermis and the dorsal cerebral cortex, whereas it was increased in the spinal cord, a result suggesting a regional variation in glial sensitivity. The intermediate filaments of the exposed animals were also immunoblotted using polyclonal antisera against the various neurofilament polypeptides and glial fibrillary acidic protein. In all tissues studied, several aggregates with molecular weights higher than those of the monomeric polypeptides were demonstrated. Contrary to clinical observations, these data indicate pronounced effects in both CNS and PNS and call for further studies on CNS effects in humans.
D1 Dopamine Receptors Can Interact with Both Stimulatory and Inhibitory Guanine Nucleotide Binding Proteins
Authors: Sidhu Anita , Sullivan Molly , Kohout Trudy , Balen Paul , Fishman Peter H. 
Published: October 2006

AbstractPretreatment of striatal membranes with N‐ethylmaleimide in the presence of a D1specific agonist inactivated endogenous guanine nucleotide binding proteins (G proteins), but not D, dopamine receptors, resulting in a loss of high‐affinity agonist binding sites. Such D1 receptors were solubilized, mixed with exogenous G proteins from cells not containing D1receptors, and reconstituted into phospholipid vesicles. These reconstituted receptors were able to couple to the exogenous G proteins, and the proportion of agonist high‐affinity sites of the receptor (40–57%) was similar to levels obtained with naive receptors coupling to endogenous G proteins (40%) upon solubilization and reconstitution. These hybrid high‐affinity sites were fully modulated by guanine nucleotides. Pretreatment of cells with pertussis toxin prior to extraction of G proteins resulted in a 50% decrease in the proportion of high‐affinity sites; these sites remained sensitive to guanine nucleotides. When D, receptors were reconstituted with extracts of cyc− cells, which lack stimulatory G proteins, the proportion of high‐affinity sites was reduced to 31% of the total. Pertussis toxin treatment of the cyc− cells completely abolished the formation of high‐affinity sites. These results demonstrate that D1‐dopaminergic receptors are able to couple to not only stimulatory G proteins (Gs), but also to inhibitory G proteins (Gi).
Prohormone Thiol Protease and Enkephalin Precursor Processing: Cleavage at Dibasic and Monobasic Sites
Authors: Krieger Timothy J. , Mende‐Mueller Liane , Hook Vivian Y. H. 
Published: October 2006

AbstractProduction of active enkephalin peptides requires proteolytic processing of proenkephalin at dibasic Lys‐Arg, Arg‐Arg, and Lys‐Lys sites, as well as cleavage at a monobasic arginine site. A novel “prohormone thiol protease” (PTP) has been demonstrated to be involved in enkephalin precursor processing. To find if PTP is capable of cleaving all the putative cleavage sites needed for proenkephalin processing, its ability to cleave the dibasic and the monobasic sites within the enkephalin‐containing peptides, peptide E and BAM‐22P (bovine adrenal medulla docosapeptide), was examined in this study. Cleavage products were separated by HPLC and subjected to microsequencing to determine their identity. PTP cleaved BAM‐22P at the Lys‐Arg site between the two basic residues. The Arg‐Arg site of both peptide E and BAM‐22P was cleaved at the NH2‐terminal side of the paired basic residues to generate [Met]‐enkephalin. Furthermore, the monobasic arginine site was cleaved at its NH2‐terminal side by PTP. These findings, together with previous results showing PTP cleavage at the Lys‐Lys site of peptide F, demonstrate that PTP possesses the necessary specificity for all the dibasic and monobasic cleavage sites required for proenkephalin processing. In addition, the unique specificity of PTP for cleavage at the NH2‐terminal side of arginine at dibasic or monobasic sites distinguishes it from many other putative prohormone processing enzymes, providing further evidence that PTP appears to be a novel prohormone processing enzyme.
Uptake and Distribution of Iron and Transferrin in the Adult Rat Brain
Authors: Morris C. M. , Keith A. B. , Edwardson J. A. , Pullen R. G. L. 
Published: October 2006

AbstractBrain uptake of iron‐59 and iodine‐125‐labelled transferrin from blood in the adult rat has been investigated using graphical analysis to determine the blood‐brain barrier permeability to these tracers in experiments that lasted between 5 min and 8 days. The blood‐brain barrier permeability (Kin) to 59Fe was 89 ± 10−5 ml/min/g compared to the value of 7 ± 10−5 ml/min/g for 125I‐transferrin, which is similar to that of albumin, a plasma marker. The autoradiographic distribution of these tracers in brain was also studied to determine any regional variation in brain uptake after the tracers had been administered either system‐ically or applied in vitro. No regional uptake was seen for 125I‐transferrin even after 24 h of circulation. In contrast, 59Fe showed selective regional uptake by the choroid plexus and extra‐blood‐brain barrier structures 4 h after administration. After 24 h of circulation, 59Fe distribution in brain was similar to the transferrin receptor distribution, as determined in vitro, but was unlike the distribution of non‐haem iron determined histochemically. The data suggest that brain iron uptake does not involve any significant transcytotic pathway of transferrin‐bound iron into brain. It is proposed that the uptake of iron into brain involves the entry of iron‐loaded transferrin to the cerebral capillaries, deposition of iron within the endothelial cells, followed by recycling of apotransferrin to the circulation. The deposited iron is then delivered to brain‐derived transferrin for extracellular transport within the brain, and subsequently taken up via transferrin receptors on neurones and glia for usage or storage.
A Novel Glial Growth Inhibitory Factor, Gliostatin, Derived from Neurofibroma
Authors: Asai Kiyofumi , Hirano Takayoshi , Kaneko Shuji , Moriyama Akihiko , Nakanishi Keiko , Isobe Ichiro , Eksioglu Yaman Z. , Kato Taiji 
Published: October 2006

AbstractNeurofibroma tissue was investigated for the presence of glial growth modulators that would suppress the proliferation of glial cells. A novel endogenous polypeptide inhibitor of proliferation and DNA synthesis in glial cells, gliostatin, was purified from the extracts of neurofibroma by a procedure comprising dye and anion‐exchange column chromatography, and HPLC. A monoclonal antibody raised against partially purified gliostatin showed no cross‐reactivity with known cytokines, but adsorbed the growth inhibitory activity of gliostatin and immunochemi‐cally visualized the putative gliostatin bands on western blot analyses. Although the product showed an apparent Mr of 100,000 accompanied by an inhibitory activity on gel filtration column chromatography, it migrated at a lower apparent Mr of 50,000 under the reducing conditions on western blotting, indicating that a homodimeric structure of native gliostatin consisted of 50‐kDa subcomponents. Gliostatin was a potent growth inhibitor acting at nanomolar concentrations against all glial tumor cells and glia maturation factor‐stimulated astroblasts, but not neuronal cells.
Changing Fatty Acid Content of Growth Cone Lipids Prior to Synaptogenesis
Authors: Martin Rex E. , Bazan Nicolas G. 
Published: October 2006

AbstractThe developing mouse was used to assess biochemical changes in membrane lipids during the period when nerve growth cones become synapses. Growth cone particles and synaptosomes were simultaneously obtained from common brain homogenates. Incorporation of the essential fatty acid, docosahexaenoic acid (22:6ω‐3), was correlated with the developmental changes in endogenous fatty acid content of growth cones and synaptosomes. Analysis of endogenous lipid content indicated that, at all ages studied, the growth cones contained more arachidonoyl acyl chains (20:4ω‐6) than did synaptosomes. Before the onset of synaptogenesis, levels of arachidonoyl chains increased and levels of 22:6, oleoyl and linoleoyl chains decreased in synaptosomes. Although stearoyl and palmitoyl (16:0) remained stable in synaptosomes, 16:0 decreased in growth cones. With the exception of 16:0 and 20:4, endogenous fatty acyl content of growth cones and synaptosomes became similar by postnatal day 10, which coincides with the onset of synaptogenesis. When 5‐day‐old mouse pups were injected intraperitoneally with [3H]22:6, the incorporation into growth cone and synaptosome phospholipids was greatest in phosphatidylethanolamine, followed by phosphatidylser‐ine and phosphatidylcholine. Nominal labeling was present in phosphatidic acid and phosphatidylinositol. Labeling in neutral lipids was less than that of phospholipids, with tria‐cylglycerol incorporating most of the neutral lipid label, followed by diacylglycerol and free 22:6. Only the growth cone fraction contained detectable amounts of 22:6‐labeled cholesterol esters. The distribution of 22:6 label in plasma 72 h after injection indicated that ∼60% of the label was in phospholipids with ∼40% in neutral lipids and <5% in free fatty acids. Collectively, the data indicate that changes in fatty acid composition occur during the conversion of growth cones to synapses.
Enhancement by Cytidine of Membrane Phospholipid Synthesis
Authors: G.‐Coviella Ignacio López , Wurtman Richard J. 
Published: October 2006

AbstractCytidine, as cytidine 5′‐diphosphate choline, is a major precursor in the synthesis of phosphatidylcholine in cell membranes. In the present study, we examined the relationships between extracellular levels of cytidine, the conversion of [14C]choline to [14C]phosphatidylcholine, and the net syntheses of phosphatidylcholine and phosphatidyleth‐anolamine by PC12 cells. The rate at which cytidine (as [3H]cytidine) was incorporated into the PC12 cells followed normal Michaelis‐Menten kinetics (Km= 5 μM; Vmax= 12 × 10−3 mmol/mg of protein/min) when the cytidine concentrations in the medium were below 50 μM; at higher concentrations, intracellular [3H]cytidine nucleotide levels increased linearly. Once inside the cell, cytidine was converted mainly into cytidine triphosphate. In pulse‐chase experiments, addition of cytidine to the medium caused a time‐ and dose‐dependent increase (by up to 30%) in the incorporation of [14C]choline into membrane [14C]‐phosphatidylcholine. When the PC12 cells were supplemented with both cytidine and choline for 14 h, small but significant elevations (p > 0.05) were observed in their absolute contents of membrane phosphatidylcholine, phospha‐tidylethanolamine, and phosphatidylserine, all increasing by 10–15% relative to their levels in cells incubated with choline alone. Exogenous cytidine, acting via cytidine triphosphate, can thus affect the synthesis and levels of cell membrane phospholipids.
Dopamine Depletion Preferentially Impairs D1 over D2‐Receptor Regulation of Striatal In Vivo Acetylcholine Release
Authors: Bertorelli Rosalia , Zambelli Matilde , Chiara Gaetano Di , Consolo Silvana 
Published: October 2006

AbstractThe roles of D2 and D1 dopaminergic receptors on the regulation of striatal acetylcholine (ACh) release in vivo were examined for a period of 120 min after acute (2 h) or prolonged (16 h) depletion of brain dopamine (DA) by a‐methyl‐p‐tyrosine. The reduction of DA transmission did not affect basal ACh output after 2 h but markedly lowered ACh release by 16 h (50%). Acute α‐methyl‐p‐tyrosine pre‐treatment prevented the reduction of ACh release by the D, antagonist SCH 23390 and its increase by the D2 antagonist, remoxipride, consistent with a drastic reduction of DA transmission at both DA receptors. However, 16 h after α‐methyl‐p‐tyrosine, the effect of remoxipride on ACh release was restored, but SCH 23390 still had no effect, suggesting that the D2 inhibitory tone on ACh release had recovered, whereas the reduction of the D1 facilitatory influence persisted. The D1 facilitatory control of ACh neurotransmis‐sion thus appears to be more sensitive than the D2 inhibitory control to a reduction in DA transmission. The new model of DA‐ACh interaction resulting from these data casts fresh light on the relationship between changes in DA transmission and extrapyramidal motor function.
Glutamate Release in Experimental Ischaemia of the Retina: An Approach Using Microdialysis
Authors: Louzada‐Junior P. , Dias J. J. , Santos W. F. , Lachat J. J. , Bradford H. F. , Coutinho‐Netto J. 
Published: October 2006

AbstractA rabbit eye model of neural ischaemia is described that uses an increased pressure in the anterior eye chamber to block the capillary supply to the retina. A microdialysis probe placed very close to the retinal surface was used to monitor release of amino acids during ischaemia. A large (two‐ to threefold) increase in the release of glutamate and O‐phosphoserine (twofold), but not of six other amino acids monitored, occurred during initial ischaemia. During reperfusion after release of intraocular pressure, much larger (five‐ to 10‐fold) increases in the release of these amino acids were observed. Parallel ischaemic retinal tissue damage was observed. This damage was prevented by keta‐mine applied locally via a superfusion needle, suggesting that glutamate released during ischaemia, and particularly during reperfusion, was responsible for cell death.
Protein Phosphatases 1 and 2A Dephosphorylate B‐50 in Presynaptic Plasma Membranes from Rat Brain
Authors: Han Yi‐fan , Wang Wei , Schlender Keith K. , Ganjeizadeh Mehdi , Dokas Linda A. 
Published: October 2006

AbstractThe protein B‐50 is dephosphorylated in rat cortical synaptic plasma membranes (SPM) by protein phospha‐tase type 1 and 2A (PP‐1 and PP‐2A)‐like activities. The present studies further demonstrate that B‐50 is dephosphorylated not only by a spontaneously active PP‐1‐like enzyme, but also by a latent form after pretreatment of SPM with 0.2 mM cobalt/20 μg of trypsin/ml. The activity revealed by cobalt/trypsin was inhibited by inhibitor‐2 and by high concentrations (μM) of okadaic acid, identifying it as a latent form of PP‐1. In the presence of inhibitor‐2 to block PP‐1, histone H1 (16–64 μg/ml) and spermine (2 mM) increased B‐50 dephosphorylation. This sensitivity to poly‐cations and the reversal of their effects on B‐50 dephosphorylation by 2 nM okadaic acid are indicative of PP‐2A‐like activity. PP‐1‐ and PP‐2A‐like activities from SPM were further displayed by using exogenous phosphorylase a and histone H1 as substrates. Both PP‐1 and PP‐2A in rat SPM were immunologically identified with monospecific antibodies against the C‐termini of catalytic subunits of rabbit skeletal muscle PP‐1 and PP‐2A. Okadaic acid‐induced alteration of B‐50 phosphorylation, consistent with inhibition of protein phosphatase activity, was demonstrated in rat cortical synaptosomes after immunoprecipitation with affinity‐purified anti‐B‐50 immunoglobulin G. These results provide further evidence that SPM‐bound PP‐1 and PP‐2A‐like enzymes that share considerable similarities with their cytosolic counterparts may act as physiologically important phosphatases for B‐50.
Participation of Prostaglandin E2 in Dopamine D2 Receptor‐Dependent Potentiation of Arachidonic Acid Release
Authors: Di Marzo Vincenzo , Piomelli Daniele 
Published: October 2006

AbstractStimulation of dopamine D2 receptors potentiates Ca2+ ionophore‐ or ATP‐induced arachidonic acid (AA) release in D2 receptor cDNA‐transfected Chinese hamster ovary (CHO) cells [CHO(D2)]. By using a combination of chromatographic, biochemical, and radioimmunochemical techniques, we show here that prostaglandin (PG) E2 is a major product of AA metabolism in CHO(D2) cells stimulated with the Ca2+ ionophore A23187. Formation of this PG was markedly increased by the concomitant application of quinpirole, a D2 receptor agonist. In addition, PGE2 enhanced D2‐dependent amplification of AA release, either when it was added (EC50= 100 nM) or when it was produced endoge‐nously, as shown by experiments carried out with the cyclooxygen‐ase inhibitor indomethacin. The results suggest that PGE2 may participate in D2 receptor‐mediated potentiation of AA release in CHO(D2) cells. They also support a functional role for this PG in the modulation of dopaminergic transmission in areas of the CNS, such as amygdala and hypothalamus, where high levels of both PGE2 and dopamine D2 receptors are found.
Evidence for D2 Receptor Regulation of Dopamine Release in the Goldfish Retina
Authors: Rashid Kashif , Baldridge William H. , Ball Alexander K. 
Published: October 2006

AbstractThe possible existence of a dopamine D2 receptor‐mediated regulation of dopamine release was investigated in the goldfish retina. Isolated retinas were preloaded with [3H]dopamine and superfused with D2 dopamine receptor agonists or antagonists to determine if there was an effect on [3H]dopamine release. The D2 receptor antagonist sulpiride increased both baseline [3H]‐ dopamine release and [3H]dopamine release induced by an increase in extracellular potassium concentration. The D2 receptor agonists LY‐171555 and RU‐24213 did not reduce baseline [3H]dopamine release but completely inhibited [3H]dopamine release induced by an increase in [K±]o. This action of the D2 agonists was blocked by sulpiride. These studies demonstrate the existence of D2 receptor, possibly autoreceptor, regulation of dopamine release in the teleost retina.
Developmental Changes in τ Phosphorylation: Fetal τ Is Transiently Phosphorylated in a Manner Similar to Paired Helical Filament‐τ Characteristic of Alzheimer's Disease
Authors: Brion Jean‐Pierre , Smith Carthage , Couck Anne‐Marie , Gallo Jean‐Marc , Anderton Brian H. 
Published: October 2006

AbstractRat and human fetal brain τ were probed with a panel of monoclonal antibodies (tau‐1, AT8, 8D8, RT97, SMI31, SMI34) that distinguish between paired helical filament (PHF)‐τ of Alzheimer's disease and normal adult brain τ. These antibodies discriminate between normal and PHF‐τ because their epitopes are phosphorylated in PHF‐τ. Although only one molecular isoform of τ was shown to be expressed in fetal brain, two fetal τ species could be distinguished on sodium dodecyl sulfate‐polyacrylamide gel electrophoresis and the slower migrating species was recognized by all of the PHF‐τ‐specific antibodies. Moreover, this immunoreactivity was shown to be phosphorylation dependent. Our observations suggest that the abnormal phosphorylation of τ in Alzheimer's disease may be the result of reactivation of pathways governing the phosphorylation of τ in the developing brain.
Differential Processing of the Ferritin Heavy Chain mRNA in Human Liver and Adult Human Brain
Authors: Dhar M. S. , Joshi J. G. 
Published: October 2006

AbstractNorthern blot analyses of the poly(A)± RNAs from human brain and liver, using a human brain ferritin heavy chain (FTH) cDNA as the probe, shows the presence of two transcripts of 1.4 and 1.1 kb. The larger, 1.4‐kb RNA, is expressed predominantly in the brain, whereas the smaller, 1.1 kb, is expressed abundantly in the liver. Screening of two normal human brain cDNA libraries yielded two types of human brain FTH cDNAs. One type corresponds to the previously characterized 1.1‐kb RNA from liver and lymphocytes. The other is also identical to the previously characterized FTH cDNA except that it contains an additional 279‐bp sequence at the 3′ untranslated region. This additional sequence shows 94.1%, 62.5%, and 58.9% identity to the 3′ flanking sequence of the human liver and mouse and rat FTH genomic clones, respectively. A fragment of a genomic clone containing the 279‐bp sequence was also isolated and sequenced. These data suggest that differential processing of the primary transcript for the FTH mRNA in human brain and liver could generate two mature mRNAs of 1.4 and 1.1 kb. This could be due to the use of alternative polyadenylation sites in the pre‐mRNA.