bluuebunny/crossref_metadata_2025_split huggingface dataset:
Radioactive N,N‐Dimethylphenylethylamine: A Selective Radiotracer for In Vivo Measurement of Monoamine Oxidase‐B Activity in the Brain
Authors: Inoue Osamu , Tominaga Toshiyoshi , Yamasaki Toshio , Kinemuchi Hiroyasu 
Published: October 2006

Abstract: N‐[methyl‐14C]N,N‐dimethylphenylethylamine (DMPEA) was synthesized and its availability as a selective radiotracer for in vivo measurement of mouse brain monoamine oxidase (MAO) activity was examined. Relatively high incorporation of labelled DMPEA into brain (about 10% of the injected dose/per gram of brain) was observed just after its injection; however, radioactive dimethylamine, a metabolite produced from labelled DMPEA in the brain 1 h after DMPEA injection, was reduced in a dose‐dependent manner by pretreatment with various doses of a specific MAO‐B inhibitor, 1‐deprenyl, but was not reduced appreciably by pretreatment with a specific MAO‐A inhibitor, clorgyline. Pretreatment with 1‐deprenyl did not affect significantly the rate of incorporation of the radiotracer DMPEA into the brain, suggesting that reduction of the radioactivity in brain by this compound might be due to a decrease in the rate of production of the radioactive metabolite dimethylamine by brain MAO‐B. The amount of the radioactive metabolite trapped in the brain was found to be proportional to the brain MAO‐B activity remaining after pretreatment with 1‐deprenyl. In vitro deamination of DMPEA by mouse brain MAO showed a higher sensitivity to inhibition by 1‐deprenyl than that by clorgyline. These results indicate that DMPEA is a selective substrate for mouse brain MAO‐B both in vivo and in vitro and that the positron emitter [11C]DMPEA might be used instead of [14C]DMPEA as a radiotracer for in vivo measurement of MAO‐B activity in human brain.
Effects of Anti‐Glutamate‐Binding Protein Antibodies on Synaptic Membrane Ion Flux, Glutamate Transport and Release, and l‐Glutamate Binding Activities
Authors: Roy S. , Galton N. , Michaelis E. 
Published: October 2006

Abstract:Antibodies (Abs) raised against the l‐glutamate‐binding protein (GBP) purified from bovine brain were used to define the possible physiologic activity of GBP in synaptic membranes. Three processes were examined for their sensitivity to the Abs: the excitatory amino acid stimulation of thiocyanate (SCN−) flux, the transport of l‐glutamic acid across the synaptic membrane, and the depolarization‐induced release of l‐glutamate. Only the amino acid‐induced changes in ion flux were inhibited by the anti‐GBP Abs. The change in membrane potential produced by exposure of synaptic membranes to excitatory amino acids was measured as the increase in the uptake of the lipophilic anion SCN−. The l‐glutamate‐induced SCN− influx was 40 times more sensitive to inhibition by the anti‐GBP Abs than the stimulation of ion flux by kainate, and 60 times more sensitive than that produced by quisqualate. The anti‐GBP Abs did not inhibit the activation of ion flux produced by N‐methyl‐d‐aspartate. The inhibition of glutamate‐stimulated ion fluxes by the Abs was complete, whereas the inhibition of l‐glutamate binding to either the rat or bovine brain GBP was not. The results obtained indicated that although the majority of the anti‐GBP Abs were not directed against the glutamate recognition site of the GBP and of presumed synaptic membrane receptors, they were effective in blocking the activation of receptor‐associated ion channels. Thus, the GBP may be considered a component of some excitatory amino acid receptor complexes.
Isolated Brain Microvessels as In Vitro Equivalents of the Blood‐Brain Barrier: Selective Removal by Collagenase of the A‐System of Neutral Amino Acid Transport
Authors: Cardelli‐Cangiano P. , Fiori A. , Cangiano C. , Barberini F. , Allegra P. , Peresempio V. , Strom R. 
Published: October 2006

Abstract:On treatment with collagenase, brain microvessels, together with several protein components, lose some enzymatic activities such as alkaline phosphatase and γ‐glutamyltranspeptidase, whereas no change occurs in the activities of 5′‐nucleotidase and glutamine synthetase. The energy‐requiring „A‐system” of polar neutral amino acid transport is also severely inactivated, whereas the L‐system for the facilitated exchange of branched chain and aromatic amino acids is preserved. In the collagenase‐digested microvessels, this leads to loss of the transtimulation effect of glutamine on the transport of large neutral amino acids, because such transtimulation is due to a cooperation between the A‐ and L‐systems. By contrast, NH4+ maintains (and even enhances) its ability to stimulate the L‐system of amino acid transport, presumably through glutamine synthesis within the endothelial cells.
Axonal Transport of Slow Component a in Sciatic Nerves of Hypo‐and Hyperthyroid Rats
Authors: Sidenius Per , Nagel Poul , Larsen JØrn Rolighed , Boye Niels , Laurberg Peter 
Published: October 2006

Abstract: Axonal transport of slow component a was studied in dorsal root afferents of the sciatic nerves of hypo‐and hyperthyroid rats. Three experimental groups of rats were made hypothyroid at the age of 12 weeks by the administration of 131I. From the age of 22 weeks to the end of the study, the groups were treated with daily subcutaneous injections of thyroxine in various doses to make them hypo‐(0 ug/100 g), normo‐(1 ug/100 g), and hyperthyroid (6 ug/100 g), respectively. The hypothyroid group had a moderate thyroid hormone deficiency (a serum triiodothyro‐nine level of 0.19 ± 0.10 nmol/L and a heart/body weight ratio of 1.87 ± 0.09 g/kg at time of killing compared with 0.60 ± 0.09 nmol/L and 2.18 ± 0.06 g/kg, respectively, for the control group). The hyperthyroid group was severely deranged, with serum triiodothyronine being 3.30 ± 0.37 nmol/L and a heart/body weight ratio of 3.11 ± 0.16 g/kg.The hypothyroid rats shov/ed a reduction in mean velocity for the transport of slow component a (0.80 ± 0.07 mm/day compared with 0.91 ± 0.05 mm/day in the controls). The width of the wave of activty was smaller for the hyperthyroid group than for the control group (6.6 ± 0.7 mm compared with 8.1 ± 1.2 mm), suggesting an increased clearance of the axonally transported activity in the proximal axon. A decrease in transport of slow component a in hypo‐thyroidism may be the explanation of peripheral neuropathy with axonal degeneration occasionally seen in patients with severe myxoedema.
Identification of the 5‐HT1A Receptor Binding Subunit in Rat Brain Membranes Using the Photoaffinity Probe [3H]8‐Methoxy‐2‐[N‐n‐Propyl, N‐3‐(2‐Nitro‐4‐Azidophenyl)Aminopropyl]Aminotetralin
Authors: Emerit M. B. , Mestikawy S. El , Gozlan H. , Cossery J. M. , Besselievre R. , Marquet A. , Hamon M. 
Published: October 2006

AbstractThe synthesis of a tritiated derivative of the 5‐HT1A photoaffinity probe 8‐methoxy‐2‐[N‐n‐propyl, N‐3‐(2‐nitro‐4‐azidophenyl)aminopropyl]aminotetralin ([3H]8‐methoxy‐3′‐NAP‐amino‐PAT) allowed the use of this probe for attempting the irreversible labeling of specific binding sites in rat brain membranes. Sodium dodecyl‐sulfate‐polyacrylamide gel electrophoresis of proteins solubilized from hippocampal microsomal membranes that had been incubated with 20 nM [3H]8‐methoxy‐3′‐NAP‐amino‐PAT under UV light revealed a marked incorporation of 3H label into a 63‐kilodalton protein termed PI . As expected of a possible correspondence between P1 and 5‐HT1A receptor binding sites, 3H labeling by the photoaffinity probe could be prevented by selective 5‐HT1A ligands such as 8‐hydroxy‐2‐(di‐n‐propylamino)tetralin, ipsapirone, buspirone, and gepirone and by N‐ethylmaleimide, but not by the 5‐HT2 antagonist ketanserin, noradrenaline‐and dopamine‐related drugs, monoamine oxidase inhibitors, and chlorimipramine. Furthermore, the regional and subcellular distributions of P1 were identical to those of specific 5‐HT1A binding sites. These results indicated that the binding subunit of the 5‐HT1A receptor is a 63‐kilodalton protein with a functionally important sulfhydryl group(s).
Calmodulin‐Dependent Phosphatases of PC12, GH3, and C6 Cells: Physical, Kinetic, and Immunochemical Properties
Authors: Farber Len H. , Wilson Frank J. , Wolff Donald J. 
Published: October 2006

AbstractCalmodulin‐dependent phosphoprotein phosphatase (CaMDP) activity has been found in each of three cultured cell lines: rat pheochromocytoma (PC 12), glioma (C6), and pituitary adenoma (GH3) cells. These CaMDP activities bind to immobilized calmodulin in the presence of Ca2+ and are eluted by EGTA. Sucrose density centrifugation revealed that the phosphatase activities exhibited sedimentation coefficients of 4.37, 4.23, and 4.59 for proteins derived from C6, GH3, and PC 12 cells, respectively. The Stokes radii measured for the PC 12 and C6 activities were 41.8 and 40.0 A, respectively. The estimated molecular weights calculated for the enzymes from these data are 79, 100 and 72, 200. The phosphatase activities required the presence of divalent cations such as Ca2+ or Mn2+ for expression of activity, which was optimal only in the presence of calmodulin. The apparent Km for phosphorylated myelin basic protein substrate was 8 μM. Affinity‐purified antibodies to the B subunit of bovine brain CaMDP were found by immunoblot (Western blot) to cross‐react with a single protein among proteins extracted from PC 12, C6, and GH3 cells that had been resolved by two‐dimensional electrophoresis. In each case, the cross‐reacting protein exhibited an Mr of 16,000 and an isoelectric point of 4.7, values virtually identical to those reported previously for the B subunit of bovine brain CaMDP (sometimes called calcineurin). This cross‐reacting protein was found among cellular proteins eluted from immobilized calmodulin by EGTA. Immuno‐cytochemical localization of the cross‐reacting protein in undifferentiated PC 12 cells or in cells differentiated in response to nerve growth factor revealed its presence diffusely throughout the cytoplasm. These experiments support the contention that each of these cell lines contains a calmodulin‐regulated phosphatase homologous physically and kinetically, and immunologically related to bovine brain CaMDP.
Alkylation of Free Sulfhydryls Fortifies Electroplax Subsynaptic Structures
Authors: Gysin Reinhard , Ranagan Steven D. 
Published: October 2006

AbstractThe cysteine‐rich 43,000‐dalton peripheral membrane protein, v1, is localized at the cytoplasmic face of electroplax and muscle cholinergic synapses, where it is thought to play an important role in the endplate supramolecular structure. The peripheral membrane protein properties of v1 are inferred by its removal from nicotinic cholinergic membranes by the action of mild alkali or lithium diiodosa‐licylate. An interesting property of v1 is its high concentration of free sulfhydryl groups, whose exact role in synaptic structure is still largely unknown. Alkylation of free sulfhydryls with N‐ethylmaleimide (3 mM) has a profound effect on the association of v1 with synaptic membranes, rendering v1 unextractable by pH 11 treatment or by lithium diiodosalicylate and, concomitantly, decreasing v1 's electrophoretic mobility on sodium dodecyl sulfate‐polyacrylamide gels. lodoacetamide and iodoacetate have similar effects, but at concentrations 10‐to 100‐fold higher than required for N‐ethylmaleimide. Furthermore, sulfhydryl modification also stabilizes the association of nicotinic receptor subunits with the detergent‐insoluble cytoskeleton. N‐Ethylma‐leimide treatment increases the fraction of insoluble receptor molecules on extraction with Triton X‐100, sodium cholate, or octylglucoside. These results suggest an important role of sulfhydryl groups in the structural stability of the postsynaptic cholinergic membrane.
Nerve Growth Factor Receptors in Human Neuroblastoma Cells
Authors: Marchetti Dario , Perez‐Polo J. Regino 
Published: October 2006

AbstractReceptors for the nerve growth factor protein (NGFR) present in the human neuroblastoma cell line LAN‐1 were characterized. LAN‐1 cells display high‐affinity (type I, with KD value of 5.9 × 10‐11M) and low‐affinity (type II, with KD value of 9.2 × 10‐9M) binding to NGF. NGFR were fractionated by preparative isoelectric focusing in a granulated gel (PEGG). High‐affinity binding was found in the 5.9–6.2 pH region of the PEGG, and low‐affinity binding in the 4.6–4.8 and 8.8–9.3 pH ranges. After further analysis by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) we observed both 92.5‐and 200‐kDa molecular species associated with NGF binding activity. The 200‐kDa protein was found in fractions displaying high‐affinity NGF binding and the 92.5‐kDa protein in fractions displaying low‐affinity NGF binding. Equilibrium binding analysis of NGF in PEGG fractions confirmed the presence of two specific saturable binding sites with KD values similar to those observed for whole dissociated cells. When NGFR II activity from the acidic region of the PEGG chromatogram was incubated with NGFR II from the basic region of the PEGG chromatogram, there was no change in NGF binding or in the number of apparent NGF receptors. However, incubation of these same fractions with a fraction having only NGFR I showed an apparent increase in high‐affinity NGF binding and a decrease in low‐affinity NGF binding. Immunoprecipitation of this “mixed” fraction and analysis on SDS‐PAGE under reduced and nonreduced conditions showed 200‐kDa and 92.5‐kDa proteins under nonreduced conditions and a 92.5‐kDa protein under reduced conditions. Our findings are consistent with the hypothesis that there are two distinct NGF receptors in NGF‐responsive cells. The interconvertibility of low‐and high‐affinity receptors and the possible existence of a modulator type protein or of “silent” type receptors are also in agreement with our findings.
Na+‐Dependent Transport of Taurine by Membrane Vesicles of Neuroblastoma ± Glioma Hybrid Cells
Authors: Yuasa Shigeki , Hamprecht Bernd 
Published: October 2006

AbstractThe transport of taurine into membrane vesicles prepared from neuroblastoma × glioma hybrid cells 108CC5 was studied. A great part of the taurine uptake by the membrane preparation is due to the transport into an osmotically sensitive space of membrane vesicles. Taurine uptake by membrane vesicles is an active transport driven by the concentration gradient of Na+ across the membrane (outside concentration > inside). The Km value of 36 μM for Na+‐dependent taurine uptake indicates a high‐affinity transport system. The rate of taurine transport by the membrane vesicles is enhanced by the K+ gradient (inside concentration > outside) and the K+ ionophore valinomycin. Taurine transport is inhibited by several structural analogs of taurine: hypotaurine, β‐alanine, and taurocyamine. All these results indicate that the taurine transport system of the membrane vesicles displays properties almost identical to those of intact neuroblastoma × glioma hybrid cells.
Cholecystokinin Stimulates Dopamine Synthesis in Synaptosomes by a Cyclic AMP‐Dependent Mechanism
Authors: Chowdhury Muhammad , Steardo Luca , Fillenz Marianne 
Published: October 2006

AbstractThe effects of different fragments of cholecystokinin (CCK) on dopamine synthesis were studied in synaptosomal preparations from the striatum, substantia nigra, and frontal cortex. In striatal synaptosomes, dopamine synthesis rate measured by dopamine accumulation was 12.5% lower than that measured by 3,4‐dihydroxyphenylalanine (DOPA) accumulation; however, K+‐accelerated synthesis was the same for both methods. Synthesis rate was independent of exogenous tyrosine levels. In the three regions studied, the combined stimulatory effects of 8‐Br‐cyclic AMP and high K+ were additive. CCK‐5, CCK‐3, CCK‐27–33, and CCK‐8 (sulphated) enhanced synthesis, CCK‐5 being the most potent fragment. The nonsulphated octapeptide had no effect. In all three regions, CCK‐5 and high K+ had an additive effect on dopamine synthesis; CCK‐5 and 8‐Br‐cyclic AMP together produced the same enhancement of synthesis as CCK‐5 alone. CCK‐5 produced similar dose‐dependent increases in dopamine synthesis and cyclic AMP accumulation in striatal synaptosomes, and both effects were blocked by the CCK antagonist proglumide.
Expression of Myelin Components in Mouse Schwann Cells in Culture
Authors: Burroni D. , White F. V. , Ceccarini C. , Matthieu J.‐M. , Costantino‐Ceccarin E. 
Published: October 2006

AbstractMouse Schwann cells cultured in vitro are capable of expressing basal levels of the major myelin components P1, P2, P0, and galactocerebroside. Numerical counts of immunostained cultures indicated that between 22 and 40% of the cells are positive up to 21 days for all of the components indicated. Electrophoretic analysis of Schwann cells labeled with a 14C‐amino acid mixture revealed the presence of proteins with relative mobilities identical to those of P0 and P1. Positive identification of the two proteins was indicated by immunoprecipitation of P1 and immunoblotting of P0. These data show that in the absence of neurites, Schwann cells in culture can express low levels of myelin characteristic components even in the absence of myelin assembly.
Rapid Determination of Norepinephrine, Dopamine, Serotonin, Their Precursor Amino Acids, and Related Metabolites in Discrete Brain Areas of Mice Within Ten Minutes by HPLC with Electrochemical Detection
Authors: Murai Shigeo , Saito Hiroko , Masuda Yoshikatsu , Itoh Tadanobu 
Published: October 2006

AbstractA rapid and simple chromatographic procedure using HPLC with electrochemical detection is described for simultaneous determination of the substrates from precursor amino acids to metabolites related to synthesis and metabolism of three monoamine neurotransmitters—norepinephrine (NE), dopamine (DA), and 5‐hydroxytryptamine (5‐HT, serotonin)—in discrete brain areas of the mouse. Under the present instrumental and mobile phase conditions, the procedure permits simultaneous determination of three monoamines (NE, DA, and 5‐HT), two precursor amino acids (tyrosine and tryptophan), and four respective metabolites (3‐methoxy‐4‐hydroxyphenylglycol, 3,4‐dihydroxyphenylacetic acid, homovanillic acid, and 5‐hydroxyindoleacetic acid) within 10 min in one chromato graphic run. By varying column temperature, this procedure also permits simultaneous determination of 10–14 monoamine‐related substrates including the nine substrates described above within 15–21 min. The validity of the present procedure is demonstrated by analyzing the effect of an α2‐adrenergic agonist (clonidine) and an α2‐antagonist (yohimbine) in mouse hypothalamus.
Biochemical Defect of the hph‐1 Mouse Mutant Is a Deficiency in GTP‐Cyclohydrolase Activity
Authors: McDonald J. David , Cotton Richard G. H. , Jennings Ian , Ledley Fred D. , Woo Savio L. C. , Bode Vernon C. 
Published: October 2006

AbstractA hyperphenylalaninemic mouse mutant, hph‐1, has been identified in the progeny of mice treated with the mutagen ethylnitrosourea. Phenylalanine hydroxylase activity levels in mutant liver lysates are reduced relative to normal, but correction for the amount of enzyme protein present demonstrates that the specific activity of this enzyme is normal in mutant mice. Quinonoid‐dihydropteridine reductase activity is also normal. GTP‐cyclohydrolase activity levels are essentially absent early in life and greatly diminished later in life. This finding has significant implications for the study of catecholamine neurotransmitter synthesis because GTP‐cyclohydrolase catalyzes an important step in the de novo synthesis of tetrahydrobiopterin, an enzyme cofactor required for the synthesis of 3,4‐dihydroxyphenylalanine (DOPA) and serotonin.
5‐Hydroxytryptamine‐Stimulated Inositol Phospholipid Hydrolysis in the Mouse Cortex Has Pharmacological Characteristics Compatible with Mediation via 5‐HT2 Receptors but This Response Does Not Reflect Altered 5‐HT2 Function After 5,7‐Dihydroxytryptamine Lesioning or Repeated Antidepressant Treatments
Authors: Godfrey P. P. , McClue S. J. , Young M. M. , Heal D. J. 
Published: October 2006

Abstract5‐Hydroxytryptamine (5‐HT; 3 × 10‐8‐1 × 10‐5M) produced a dose‐dependent increase in phosphatidyl‐inositol/polyphosphoinositide (PI) turnover in mouse cortical slices, as measured by following production of 3H‐la‐belled inositol phosphates (IPs) in the presence of 10 mM LiCl. Analysis of individual IPs, in slices stimulated for 45 min, indicated substantial increases in inositol monophos‐phate (IP1; 140%) and inositol bisphosphate (IP2; 95%) contents with smaller increases in inositol trisphosphate (IP3; 51%) and inositol tetrakisphosphatc (IP4; 48%) contents. The increase in IP3 level was solely in the 1.3,4‐isomer. This response was inhibited by the nonselective 5‐HT antagonists methysergide, metergoline, and spiper‐one. It was also inhibited by the selective 5‐HT2 antagonists ketanserin and ritanserin but not by the 5‐HT1 antagonists isapirone, (‐)‐propranolol, or pindolol. 5‐HT‐stimulated IP formation was also unaltered by atropine, prazosin, and mepyramine. Lesioning brain 5‐HT neurones using 5,7‐di‐hydroxytryptamine (5.7‐DHT; 50 μg i.c.v.) produced a 210% (p < 0.01) increase in the number of 5‐HT2‐mediated head‐twitches induced by 5‐methoxy‐N,N‐dimethyltrypt‐amine (2 mg/kg). However, 5,7‐DHT lesioning had no effect on 5‐HT‐stimulated PI turnover in these mice. Similarly, an electroconvulsive shock (90 V, 1 s) given five times over a 10‐day period caused an 85% (p < 0.01) increase in head‐twitch responses but no change in 5‐HT‐stimulated PI turnover. Decreasing 5‐HT2 function by twice‐a‐day injection of 5 mg/kg of zimeldine or desipramine (DMI) produced 59% (p < 0.01) and 56% (p < 0.01), respectively, reductions in head‐twitch behaviour. Although zimeldine had no effect on 5‐HT‐stimulated IP formation, DMI treatment inhibited this by 33% at 3 μM 5‐HT (p < 0.02). Previous studies have shown an excellent correlation between head‐twitch responses and cortical 5‐HT2 receptor number in C57/B1/6 mice. Hence, it is concluded that although the 5‐HT‐stimulated PI turnover in cortical slices has the characteristics of a 5‐HT2 receptor‐mediated response, it does not, in general, reflect adaptive changes in 5‐HT2 receptor number and function.