bluuebunny/crossref_metadata_2025 huggingface dataset:
Binding Characteristics of t‐[35S]Butylbicyclophosphorothionate in Discrete Brain Regions of Rats Made Tolerant to and Dependent on Pentobarbital
Authors: Ito Yoshihisa , Saunders Paul A. , Lim Dong Koo , Ho Ing K.
Published: October 2006

Abstract: The effects of acute and continuous pentobarbital administration by pellet implantation on binding characteristics of t‐[35S]butylbicyclophosphorothionate ([35S]TBPS) in discrete regions of rat brains were examined. Acute administration of pentobarbital (60 mg/kg, s.c.) affected neither the KD nor the Bmax values of [35S]TBPS binding in any of the regions studied. The cerebella of pentobarbital‐tolerant rats had an increased density of [35S]TBPS binding sites with no change in their apparent affinity. There were no significant changes in the binding characteristics in the frontal cortex (FC), the striatum (ST), and the substantia nigra (SN) of these animals. Twenty‐four hours after removal of the pentobarbital pellets, a significant decrease in the latency of onset of first twitch response induced by pentylenetetrazol (PTZ) (50 mg/ kg, i.p.) was observed. In addition, the density of [35S]TBPS binding sites was significantly increased in the FC, the SN, and the cerebellum but not in the ST. In all brain regions studied, placebo pellet implantation and pentobarbital tolerance and dependence caused no changes in the apparent affinity of [35S]TBPS binding or the IC50 of pentobarbital for the inhibition of [35S]TBPS binding. These results suggest that [35S]TBPS binding was significantly increased following the withdrawal of the pentobarbital pellets without altering intrinsic coupling activity of barbiturate recognition sites and convulsant binding sites and that these increases in [35S]TBPS binding are related to the increased susceptibility to seizures induced by PTZ in rats made dependent on pentobarbital.
Effects of Cyclic Nucleotides and Calcium/Calmodulin on Protein Phosphorylation in the CNS of Hirudo medicinalis
Authors: Garcia‐Gil Mercedes , Berton Fulvia , Tongiorgi Enrico , Brunelli Marcello
Published: October 2006

Abstract: Protein phosphorylation plays an important role in the regulation of neural functions. We have studied the phosphorylation of proteins in homogenates of segmental ganglia of the leech Hirudo medicinalis. We describe a number of proteins whose phosphorylation is dependent on cal‐cium/calmodulin or cyclic nucleotides. Most of the proteins whose phosphorylation is increased in the presence of calcium seem to be substrates for cyclic nucleotide‐dependent protein kinases. Only two of the phosphoproteins described appear to be specific substrates for calcium/calmodulin protein kinase(s), and at least six phosphoproteins appear to be specific substrates for cyclic nucleotide‐dependent kinase(s). The leech nervous system, with large and identifiable neurons, provides a good tool for studies of neural functions, such as learning. The results are discussed in the context of the role of protein phosphorylation on learning processes.
Effect of Latent Iron Deficiency on 5‐Hydroxytryptamine Metabolism in Rat Brain
Authors: Shukla Arti , Agarwal K. N. , Chansuria J. P. N. , Taneja V.
Published: October 2006

Abstract:Eight weeks of latent iron deficiency in weaned rats maintained on an experimental low iron content diet (18–20 mg/kg) did not significantly alter the packed cell volume and hemoglobin concentration; however, the hepatic and brain nonheme iron contents decreased by 66% and 21% (p < 0.001), respectively. The tryptophan concentration decreased by 31% and 34% in liver and brain, respectively, in rats on experimental diet (p < 0.01). The brain 5‐hydroxytryptamine and 5‐hydroxyindoleacetic acid contents were reduced by 21% and 23% (p < 0.01 and p < 0.02), respectively. However, in the brain, weight, protein, DNA, and the activities of monoamine oxidase, aldehyde dehydrogenase, and liver tryptophan oxygenase were found to remain unaltered.

When rehabilitated with a diet containing 390 mg/kg iron, rats previously maintained on the experimental diet for 2 weeks showed partial recovery in tryptophan levels both in liver and brain. However, brain 5‐hydroxytryptamine and 5‐hydroxyindoleacetic acid levels remained unaltered. The hepatic iron content improved without any change in brain iron content. The latent iron deficiency produced significant alterations in the metabolism of 5‐hydroxytryptamine and brain iron content that could not be recovered 2 weeks after the iron rehabilitation.
Developmental Study of Myelin Basic Protein Variants in Various Regions of Pig Nervous System
Authors: Sheng H. Z. , de Rosbo N. Kerlero , Carnegie P. R. , Bernard C. C. A.
Published: October 2006

Abstract: A developmental study of myelin basic protein (MBP) variants in eight regions of pig nervous system (NS) was performed using a quantitative electroimmunoblotting procedure. Four major MBP forms with apparent molecular weights of 21.5K, 20.2K, 18.5K, and 17.3K were identified in both the CNS and the PNS and were detected as early as 22 days before birth. Quantification of the most abundant forms, the 21.5K and 18.5K MBPs, revealed characteristic profiles of accumulation of these two variants in different regions of the NS. The ratio of 21.5K:18.5K MBP varied with developmental time as well as with the various NS regions, peaking 20 days postnatally. The 17.3K MBP was observed from embryonic stages to adulthood, as were the 21.5K and 18.5K forms. In contrast, the 20.2K variant appeared most abundant from 10 days before to 22 days after birth and thereafter decreased in intensity so as to be no longer detectable in the brain of a 5‐year‐old pig. A similar pattern was also observed with an anti‐MBP‐reacting protein with an apparent molecular weight of 23K. Taken together, these results suggest that in the pig NS, the expression of MBP variants may be regulated both regionally and developmentally.
Postnatal Development of Thiamine Metabolism in Rat Brain
Authors: Matsuda Toshio , Doi Tadashi , Tonomura Hideo , Baba Akemichi , Iwata Heitaroh
Published: October 2006

Abstract: The activities of thiamine diphosphatase (TDPase), thiamine triphosphatase (TTPase), and thiamine pyrophosphokinase and the contents of thiamine and its phosphate esters were determined in rat brain cortex, cerebellum, and liver from birth to adulthood. Microsomal TTPase activity in the cerebral cortex and cerebellum increased from birth to 3 weeks, whereas that in the liver did not change during postnatal development. Microsomal TDPase activity in the cerebral cortex showed a transient increase at 1–2 weeks, but that in the cerebellum did not change during development. In contrast to the activity of the brain enzyme, that of liver microsomal TDPase increased stepwise after birth. Thiamine pyrophosphokinase activity in the cerebellum increased from birth to 3 weeks and then decreased, whereas that in the cerebral cortex and liver showed less change during development. TDP and thiamine monophosphate (TMP) levels increased after birth and plateaued at 3 weeks whereas TTP and thiamine levels showed little change during development in the cerebral cortex and cerebellum. The contents of thiamine and its phosphate esters in the liver showed more complicated changes during development. It is concluded that thiamine metabolism in the brain changes during postnatal development in a different way from that in the liver and that the development of thiamine metabolism differs among brain regions.
Glycosaminoglycan Composition of PC 12 Pheochromocytoma Cells: A Comparison with PC12D Cells, a New Subline of PC12 Cells
Authors: Katoh‐Semba Ritsuko , Oohira Atsuhiko , Sano Mamoru , Watanabe Kimi , Kitajima Satoko , Kashiwamata Shigeo
Published: October 2006

Abstract: PC12D cells, a new subline of conventional PC12 cells, respond not only to nerve growth factor but also to cyclic AMP by extending their neurites. These cells are flat in shape and are similar in appearance to PC12 cells that have been treated with nerve growth factor for a few days. In both cell lines, we have characterized the glycosaminoglycans, the polysaccharide moieties of proteoglycans, which are believed to play an important role in cell adhesion and in cell morphology. Under the present culture conditions, only chondroitin sulfate was detected in the media from PC12 and PC12D cells, whereas both chondroitin sulfate and heparan sulfate were found in the cell layers. The levels of cell‐associated heparan sulfate and chondroitin sulfate were about twofold and fourfold higher in PCI 2D cells than in PC12 cells, respectively. Compared to PC12 cells, the amounts of [35S]sulfate incorporated for 48 h into chondroitin sulfate were twofold lower but those into heparan sulfate were 35% higher in PC12D cells. The amount of chondroitin sulfate released by PC12D cells into the medium was about a half of that released by PC12 cells. The ratio of [35S]sulfate‐labeled heparan sulfate to chondroitin sulfate was 6.2 in PC12D cells and 2.2 in PC12 cells. These results suggest that there may be some correlation between the increase in content of glycosaminoglycans and the change in cell morphology, which is followed by neurite outgrowth.
Reduction of Brain Glutathione by l‐Buthionine Sulfoximine Potentiates the Dopamine‐Depleting Action of 6‐Hydroxydopamine in Rat Striatum
Authors: Pileblad Erik , Magnusson Tor , Fornstedt Bodil
Published: October 2006

Abstract: Sprague‐Dawley rats were anesthetized with chloral hydrate, and plastic cannulae were permanently implanted into the lateral ventricles. The animals then were allowed to recover for 1–2 days. L‐Buthionine sulfoximine (l‐BSO), a selective inhibitor of glutathione (GSH) synthesis, and 6‐hydroxydopamine (6‐OH‐DA), a selective catecholaminergic neurotoxin, were administered intracerebroventricularly. The striatal concentrations of GSH and monoamines were determined by HPLC with electrochemical detection. Two injections of l‐BSO (3.2 mg, at a 48‐h interval) resulted in a 70% reduction of striatal GSH. 6‐OH‐DA (150 or 300 μg) reduced the concentrations of striatal dopamine and noradrenaline 7 days after the administration, but left the concentrations of 5‐hydroxytryptamine unaltered. L‐BSO treatment did not produce any changes in the levels of monoamines per se but it potentiated the catecholamine‐depleting effect of 6‐OH‐DA in the striatum. Thus, GSH appears to suppress the toxicity of 6‐OH‐DA, probably by scavenging the toxic species formed during 6‐OH‐DA oxidation. In view of these results one may suggest an important role for GSH in catecholaminergic neurons: protecting against the oxidation of endogenous catechols.
Properties of a Leu‐Phe‐Cleaving Endopeptidase Activity Putatively Involved in β‐Endorphin Metabolism in Rat Brain
Authors: Lebouille Jos L. M. , Hendriks Rudi W. , Soeter Nell M. , Burbach J. Peter H.
Published: October 2006

Abstract: Incubation of β‐endorphin with cytosolic and particulate fractions of rat brain resulted in the formation of several peptides, including γ‐endorphin [β‐endorphin‐(1–17)] and β‐endorphin‐(18–31), indicating the presence of enzyme activity cleaving the Leu17‐Phe18 bond of β‐endorphin. An assay for this Leu‐Phe cleaving activity, based on the cleavage of the 14C‐labeled substrate acetyl‐Val‐Thr‐Leu‐Phe‐[ε ([14C]CH3)2]Lys‐NHCH3, was used to examine the properties of this enzyme activity. β‐Endorphin‐(1–31) competitively inhibited the Leu‐Phe‐cleaving enzyme activity on the pentapeptide substrate. Over 90% of activity was recovered in the cytosolic fraction. Leu‐Phe‐cleaving activity behaved like a thiol endopeptidase because it was inhibited by low concentrations of N‐ethylmaleimide, p‐chloromercuribenzoate, p‐chloromercuribenzoyl sulfate, and low concentrations of Hg2+. Low concentrations of sulfhydryl compounds stimulated Leu‐Phe‐cleaving activity. The activity was optimal between pH 8.5 and 9.0. The Km of Leu‐Phe‐cleaving activity in the cytosolic fraction was 35 μM and in the particulate fraction 88 μM with Vmax values of 193 and 15 nmol mg protein−1 h−1, respectively. The apparent molecular mass of the Leu‐Phe‐cleaving enzyme was estimated by gel filtration to be ∼200 kilodaltons. These properties of Leu‐Phe‐cleaving activity indicate that the Leu‐Phe‐cleaving enzyme is distinct from any known brain endopeptidase.
Rapid Preparation of a Distinct Lysosomal Population from Myelinating Mouse Brain Using Percoll Gradients
Authors: Caimi L. , Marchesini S. , Aleo M. F. , Bresciani R. , Monti E. , Casella A. , Giudici M. L. , Preti A.
Published: October 2006

Abstract: To study the vesicular lysosome‐associated transport and the metabolism of some brain macromolecules (in particular, sialoglycoconjugates), we developed a rapid procedure to obtain a distinct lysosomal population starting from myelinating mouse brain. This procedure is based on an initial differential centrifugation step producing a 1,000–17,500‐g fraction (P2), followed by isopycnic centrifugation of fraction P2 on a self‐generated colloidal silica gel (Percoll) gradient. The heaviest subfraction thus obtained is very rich in acid hydrolase activities like β‐galactosidase, arylsulfatase A, and acid phosphatase. The enrichment of these enzymes is ∼100‐fold as compared with the starting homogenate, whereas the markers of other subcellular organelles, such as mitochondria, plasma membranes, or the Golgi apparatus, are virtually absent. The lysosomal preparation contains ∼12–14% of the total acid hydrolase activities, with a protein yield of ∼0.12%. Electron microscopy shows that the lysosomal fraction is composed of an ∼90% pure population of lysosomes. Therefore, the procedure described here is suitable for obtaining a highly purified lysosome preparation from myelinating mouse brain.
Frontal Cortical and Left Temporal Glutamatergic Dysfunction in Schizophrenia
Authors: Deakin J. F. W. , Slater P. , Simpson M. D. C. , Gilchrist A. C. , Skan W. J. , Royston M. C. , Reynolds G. P. , Cross A. J.
Published: October 2006

Abstract: Glutamatergic mechanisms have been investigated in postmortem brain samples from schizophrenics and controls. D‐[3H]Aspartate binding to glutamate uptake sites was used as a marker for glutamatergic neurones, and [3H]kainate binding for a subclass of postsynaptic glutamate receptors. There were highly significant increases in the binding of both ligands to membranes from orbital frontal cortex on both the left and right sides of schizophrenic brains. The changes are unlikely to be due to antemortem neuroleptic drug treatment, because no similar changes were recorded in other areas. A predicted left‐sided reduction in D‐[3H]aspartate binding was refuted at 5% probability, but not at 10%. Previously reported high concentrations of dopamine in left amygdala were strongly associated with low concentrations of D‐[3H]aspartate binding in left polar temporal cortex in the schizophrenics. The findings are compatible with an overabundant glutamatergic innervation of orbital frontal cortex in schizophrenia. The results also suggest that schizophrenia may involve left‐sided abnormalities in the relationship between temporal glutamatergic and dopaminergic projections to amygdala.
Characterization and Regional Distribution of Individual Gangliosides in Goldfish Central Nervous System
Authors: Tanaka Takashi , Arai Yoko , Kishimoto Yasuo
Published: October 2006

Abstract: Gangliosides were partially purified from goldfish brain and fractionated by DEAE Fractogel column chromatography. Each fraction was then analyzed by HPTLC and also by HPLC after conversion of the gangliosides to 2,4‐dinitrophenylhydrazides. The tetrasialoganglioside GQ1c was found to constitute more than 50% of the total gangliosides. Gangliosides in smaller quantities were also tentatively identified. These included GT1b, GT1c, GT2, GT3, GD1a, and several others. By using this information, the amounts of individual gangliosides in various regions of goldfish central nervous system were compared. Although all areas of brain examined contained similar concentrations of gangliosides, with GQ1c as the predominant component, retina and optic nerve contained significantly lower concentrations of GQ1c, and GM3 was the major component.
Recovery of Proteolipid Protein in Mice Heterozygous for the Jimpy Gene
Authors: Benjamins Joyce A. , Studzinski Diane M. , Skoff Robert P. , Nedelkoska Liljana , Carrey Elizabeth A. , Dyer Charissa A.
Published: October 2006

Abstract: We have measured levels and synthesis of proteolipid protein (PLP) and its transport into myelin in female mice heterozygous for the jimpy gene and in their normal female littermates. In both cord and cerebrum, jimpy carriers show deficits in PLP during development followed by compensation in adulthood. Recovery of PLP occurs earlier in cord than in brain. At 13 days levels of PLP in carriers compared to controls are reduced to 0.60 and 0.44, respectively, in cord and cerebrum. By 100 days, normal levels of PLP are attained in cord (1.13) whereas levels of PLP in cerebrum are only 0.78 of control. By 200 days full recovery occurs in cerebrum, with a ratio of 1.21, suggesting a possible overcompensation. The yield of myelin from cerebrum was reduced to 0.78 in carriers compared to controls at 17 days. In brain slices, incorporation of [3H]leucine into homogenate PLP from carriers is the same as in controls, whereas [3H]Ieucine incorporation into myelin PLP is reduced to 0.68 of control. These results indicate that synthesis of PLP in the carriers is normal at 17 days, but transport of PLP into myelin is reduced. Similarly, acylation of homogenate PLP is normal, whereas acylation of myelin PLP is reduced, as measured by incorporation of [3H]palmitic acid. Transport of PLP into myelin was compared to transport of MBP; transport of both proteins was equally decreased as indicated by the similar ratio of labeled PLP to MBP in myelin from carriers compared to noncarriers. Thus, total synthesis of PLP is not reduced in the carriers, whereas transport of both PLP and MBP into myelin is decreased, as are levels of myelin. Our data from the metabolic studies and measurement of PLP show that myelination is retarded in the carriers. Recovery from the deficit in myelin appears complete by later ages. We find, however, no evidence for a selective defect in synthesis or acylation of PLP, or transport of PLP relative to MBP. Mechanisms that could be employed by the oligodendrocyte to compensate for the early deficits in the carriers are discussed relative to glial differentiation and the possible formation of defective PLP.
Isolation and Characterization of a Rat Brain Triakontatetraneuropeptide, a Posttranslational Product of Diazepam Binding Inhibitor: Specific Action at the Ro 5‐4864 Recognition Site
Authors: Slobodyansky E. , Guidotti A. , Wambebe C. , Berkovich A. , Costa E.
Published: October 2006

Abstract: This report describes the purification and characterization from rat brain of triakontatetraneuropeptide (TTN, DBI 17‐50), a major biologically active processing product of diazepam binding inhibitor (DBI). Brain TTN was purified by immunoaffinity chromatography with polyclonal octa‐decaneuropeptide, DBI 33‐50) antibodies coupled to CNBr‐Sepharose 4B followed by two reverse‐phase HPLC steps. The amino acid sequence of the purified peptide is: Thr‐Gln‐Pro‐Thr‐Asp‐Glu‐Glu‐Met‐Leu‐Phe‐Ile‐Tyr‐Ser‐His‐Phe‐Lys‐Gln‐Ala‐Thr‐Val‐Gly‐Asp‐Val‐Asn‐Thr‐Asp‐Arg‐Pro‐Gly‐Leu‐Leu‐Asp‐Leu‐Lys. Synthetic TTN injected intra‐cerebroventricularly into rats induces a proconflict activity (IC50 0.8 nmol/rat) that is prevented by the specific “peripheral” benzodiazepine (BZ) receptor antagonist isoquinoline carboxamide, PK 11195, but not by the “central” BZ receptor antagonist imidazobenzodiazepine, flumazenil. TTN displaces [3H]Ro 5‐4864 from synaptic membranes of olfactory bulb with a Ki of approximately 5 μM. TTN also enhances picrotoxinin inhibition of γ‐aminobutyric acid (GABA)‐stimulated [3H]flunitrazepam binding. These data suggest that TTN, a natural DBI processing product acting at “Ro 5‐4864 preferring” BZ binding site subtypes, might function as a putative neuromodulator of specific GABAA receptor‐mediated effects.
Excitatory Amino Acid‐Induced Toxicity in Chick Retina: Amino Acid Release, Histology, and Effects of Chloride Channel Blockers
Authors: Zeevalk Gail D. , Hyndman Arnold G. , Nicklas William J.
Published: October 2006

Abstract: Acute excitotoxicity in embryonic chick retina and the ability of C1− channel blockers to prevent toxicity were evaluated by measurement of endogenous amino acid release and histology. Treatment of retina with kainate, quisqualate, or N‐methyl‐D‐aspartate resulted in a large dose‐dependent release of γ‐aminobutyric acid and taurine, moderate release of glutamine and alanine, and no measurable release of glu‐tamate or aspartate. Concentrations inducing maximal γ‐aminobutyric acid release were 50 μM quisqualate, 100 μM kainate, and 100 μM N‐methyl‐D‐aspartate. Treatment with 1 mM glutamate resulted in significant γ‐aminobutyric acid release, as well as an elevation in medium aspartate levels. Typical excitotoxic retinal lesions were produced by the agonists and, at the lower concentrations tested, revealed a regional sensitivity. There was a positive correlation between the amount of γ‐aminobutyric acid release and the extent of tissue swelling, suggesting that release may be secondary to toxic cellular events. Omission of C1− completely blocked cytotoxic effects due to kainate or glutamate. Likewise, addition of the C1−/bicarbonate anion channel blocker 4,4′‐di‐isothiocyanatostilbene‐2,2′‐disulfonate at 600 μM protected retina from cytotoxic damage from all excitotoxic analogs and restored amino acid levels to baseline values. Furosemide. which blocks Na+/K+/2C1− cotransport, was only minimally effective in reducing amino acid release induced by the agonists. Consistent with the latter, histological examination showed the continued presence of the lesion but with general reduction of cellular edema. These results indicate that although influx of C1− is a central component of the acute excitotoxic phenomenon, mechanisms other than passive Cl−flux may be involved.
Postnatal Evolution of the γ‐Aminobutyric Acid/Benzodiazepine Receptor Complex in a Model of Inherited Epilepsy: The Quaking Mouse
Authors: Caboche Jocelyne , Mitrovic Nadia , Le Saux Françoise , Besson Marie‐Jo , Sauter André , Maurin Yves
Published: October 2006

Abstract: Binding assays of [3H]muscimol and [3H]‐flunitrazepam have been performed on brain homogenates of brainstem, cerebellum, and forebrain of genetically epileptic quaking (qk) mutant mice 20, 40, 70, and 90 days old and their corresponding controls of the same strain (C57BL/ 6J). The endogenous γ‐aminobutyric acid (GABA) content has been determined in various brain regions of 70‐day‐old qk and control mice. Finally, the behavioral effects of diazepam, of the mixed GABAA/GABAB receptor agonist progabide, and of the selective GABAB receptor agonist baclofen have been assessed in adult qk mutants. Our results strongly suggest a lack of involvement of GABAergic neurotransmission in the inherited epilepsy of the qk mutant mouse.
Opioid Receptors of Neuroblastoma Cells Are in Two Domains of the Plasma Membrane that Differ in Content of G Proteins
Authors: Ott Susanne , Costa Tommaso , Herz Albert
Published: October 2006

Abstract: Opioid receptors of NG 108–15 cell membranes are distributed in two membrane fractions sedimenting at 20,000 g (P2) and 200,000 g (P3). The number of receptors is identical in P2 and P3, but in P2 all sites are present in one high‐affinity state (2 nM), whereas in P3 60% of these receptors display lower affinity (150 nM). Upon addition of GTP or pretreatment with pertussis toxin, 80% of the sites exist in low affinity in both P2 and P3. Therefore, the effect of GTP and pertussis toxin on agonist binding appears to be smaller in P2 than in P3. In contrast, sodium inhibits agonist binding in P2 and P3 to the same extent and with identical potency. Opioid‐mediated stimulation of GTPase is much greater in P2 than in P3, whereas inhibition of adenylate cyclase does not differ in the two fractions. Using site‐specific antibodies and pertussis toxin‐catalyzed ADP‐ribosylation, we found that the amount of G proteins in P3 is only 30–50% of that in P2. Treatment of intact cells with the hydrophilic protein‐modifying agent sulfosuccinimido‐biotin results in biotinylation of proteins from both fractions and in a similar reduction of opioid binding in P2 and P3. Likewise, exposure of intact cells to the alkylating opioid antagonist, chlornaltrexamine, produces identical degrees of receptor inactivation in P2 and P3. The rate of in vivo pertussis toxin‐mediated modification of G proteins is not different in the two fractions. We conclude from these data that (1) a larger proportion of receptors is uncoupled from G proteins in P3 than in P2; (2) the effects of GTP and Na+ on opioid receptor binding can be attributed to different membrane components; and (3) receptors in P3 appear to be present in the plasma membranes and, thus, do not represent internalized receptors.
Na+ Dependence of Tyrosine Transport Across the Synaptosomal Membrane Reflects Changes in the Morphology of Synaptosomes
Authors: Diez‐Guerra F. Javier , Gimenez Cecilio
Published: October 2006

Abstract: The Na+ dependence of tyrosine uptake into rat brain synaptosomes and synaptosomal plasma membrane vesicles (SPMV) was examined in the present study. At low tyrosine concentrations, the isoosmotic substitution of Na+ by sucrose in the incubation medium led to an increase of tyrosine uptake in synaptosomes and to a decrease in SPMV. The removal of extracellular Ca2+ and Mg2+ and addition of isoosmotic sucrose completely prevented the augmented tyrosine uptake in Na+‐free incubated synaptosomes. Morphological differences were found at the electron‐microscopic level when synaptosomes were incubated in Na+‐free and Na+‐containing media. The internal volume measured for synaptosomes incubated in a Na+‐free medium was almost half of that obtained in a Na+‐containing medium, in good agreement with the observations made with the electron microscope. Also, the omission of Ca2+ and Mg2+ resulted in a specific swelling of only the synaptosomes incubated in Na+‐free medium. When synaptosomes and SPMV were preloaded with several neutral amino acids, the tyrosine uptake rate was greatly increased, indicating fully operational exchange mechanisms for these amino acids. We propose that the enhancement of high‐affinity synaptosomal tyrosine uptake observed in Na+‐free medium is a consequence of a specific shrinkage of the synaptosomes and a parallel increase of the exchange rate with endogenous neutral amino acids.
Effects of Chronic Chlorpromazine Treatment on Peripheral Benzodiazepine Binding Sites in Heart, Kidney, and Cerebral Cortex of Rats
Authors: Gavish Moshe , Weizman Ronit
Published: October 2006

Abstract: Daily intraperitoneal administration to rats of 5 mg/kg of chlorpromazine (CPZ) for 21 days induced a significant up‐regulation (51%) of peripheral benzodiazepine binding sites (PBSs) in cerebral cortex and a down‐regulation of PBSs in the heart (25%) and kidney (14%), whereas no alteration in [3H]flunitrazepam binding in cerebral cortex was observed. [3H]PK 11195 binding to cerebral cortex returned to normal following 5 days of CPZ withdrawal, whereas the density of PBSs in the heart and kidney remained reduced. The affinity of PBSs for the ligand [3H]PK 11195 in the cerebral cortex and heart was not affected by the drug treatment or withdrawal. The CPZ‐induced alterations in PBSs may be relevant to the effects of the drug on CNS and/or peripheral organs.
Blockage of Nerve Growth Factor Action in PC12h Cells by Staurosporine, a Potent Protein Kinase Inhibitor
Authors: Hashimoto Seiichi , Hagino Akihiko
Published: October 2006

Abstract: Staurosporine, which has a structure similar to that of K‐252a, a potent protein kinase inhibitor that blocks nerve growth factor (NGF) action in PC12 and PC12h cells, is also known as a potent inhibitor of several protein kinases. This study shows that in PC12h cells staurosporine has a dual action: at lower concentrations than that required by K‐252a, it is an inhibitor of NGF induction of neurite formation and of changes in the phosphorylation of specific proteins, whereasat concentrations higher than that required to inhibit NGF‐induced neurite outgrowth, it rapidly enhances outgrowth by itself.
Kinetics of Carboxylmethylation of the Charge Isoforms of Myelin Basic Protein by Protein Methyltransferase II
Authors: Caamaño Claudio A. , Azcurra Julio M. , Sellinger Otto Z. , Zand Robert
Published: October 2006

Abstract: The charge isoforms (C1‐C5) of bovine myelin basic protein (MBP) were used as substrates for the rat brain enzyme protein carboxylmethyltransferase (PM II). The objective of these experiments was to ascertain whether the kinetic behavior of the MBP isoforms reflected differences in the structures of this molecular family. Initial velocity plots as a function of the MBP‐isoform concentration showed significnt differences (p > 0.05) among the assayed isoforms except for isoforms C2 and C4. Under the conditions of our experiment all the curves exhibited a consistent sigmoidicity. The kinetic data were best fitted by a model, previously described for the enzyme D‐β‐hydroxybutyrate dehydrogenase, in which two independent sites must be randomly occupied before any catalytic activity can occur. This mechanism is substantially different from that proposed by other investigators for similar PM II enzymes and other substrates. The differences in the rates of isoform carboxylmethylation are largely accounted for by the different apparent dissociation constants Ks and is explained on the basis of inherent structural differences among the charge isoforms.
Purified Rat Brain Myelin Contains Measurable Acyl‐CoA:Lysophospholipid Acyltransferase(s) but Little, if Any, Glycerol‐3‐Phosphate Acyltransferase
Authors: Vaswani Kuldeep K. , Ledeen Robert W.
Published: October 2006

Abstract: Previous reports from several laboratories have demonstrated the presence of many lipid‐metabolizing enzymes in myelin, including all the enzymes needed to convert diacylglycerol to phosphatidylcholine and phosphatidylethanolamine. Axonal transport studies had suggested the presence of additional enzymes which incorporate acyl chains into specific phospholipids of myelin. We report here evidence for one such group of enzymes, the acyl‐CoA:lysophospholipid acyltransferases. At the same time, activity of acyl‐CoA: sn‐glycerol‐3‐phosphate acyltransferase was negligible in myelin. Oleoyl‐CoA and arachidonoyl‐CoA were both active substrates for transfer of acyl chains to lysophosphatidylcholine and lysophosphatidylinositol. Activity in myelin varied from 7 to 19% of microsomal activity, values well above the likely level of microsomal contamination as judged by microsomal markers. Additional evidence for a myelin locus came from assays at sequential stages of purification and from mixing experiments. Arachidonoyl‐CoA was somewhat more reactive than oleoyl‐CoA toward lysophosphatidylcholine; the myelin Km for these two CoA derivatives was 98 μM and 6.6 μM, respectively. Activity with lysophosphatidylinositol as substrate was approximately 40% of that with lysophosphatidylcholine in myelin, whereas activities with lysophosphatidylethanolamine and lysophosphatidylserine were considerably less.
Sodium‐Dependent Uptake of Nucleosides by Dissociated Brain Cells from the Rat
Authors: Johnston Mark E. , Geiger Jonathan D.
Published: October 2006

Abstract: Sodium‐dependent 3H‐labeled nucleoside transport was studied using a mixed population of dissociated brain cells from adult rats. The accumulation of [3H]adenosine during brief (15‐s) incubation periods was significantly greater in the presence of 110 μM Na+ than in its absence. This occurred at substrate concentrations that ranged from 0.25 to 100 μM. Similar findings were observed for the rapid accumulation of [3H]uridine. Kinetically, the rapid accumulation of [3H]adenosine in both the absence and the presence of Na+ was best described by a two‐component system. In the presence of Na+, the KT and Vmax values for the high‐affinity component were 0.9 μM and 8.9 pmol/mg of protein/15 s, and those for the low‐affinity component were 313 μM and 3,428 pmol/mg of protein/15 s, respectively. In the absence of Na+, the KT value for the high‐affinity component was significantly higher (1.8 μM). [3H]Uridine accumulation was best described kinetically by a one‐component system that in the presence of Na+ had KT and Vmax values of 1.0 mM and 2.6 nmol/mg of protein/15 s, respectively. As was found for [3H]adenosine, in the absence of Na+, the KT value was significantly higher (1.8 mM). The sodium‐dependent transport of [3H]adenosine was inhibitable by ouabain and 2,4‐dinitrophenol. Of the three nucleoside transport inhibitors tested, only nitrobenzylthioinosine demonstrated high affinity and selectivity in blocking the sodium component. Thus, high‐affinity sodium‐dependent nucleoside transport systems, in addition to facilitated diffusion systems, exist on brain cells from adult rats.
Regulation of σ‐Receptors: High‐ and Low‐Affinity Agonist States, GTP Shifts, and Up‐Regulation by Rimcazole and 1,3‐Di(2‐Tolyl)guanidine
Authors: Beart Philip M. , O'Shea Rbss D. , Manallack David T.
Published: October 2006

The regulation of the central σr‐binding site was investigated using both in vitro and in vivo manipulations in conjunction with radioligand binding. The displacement of the binding of R(+)‐[3H]3‐[3‐hydroxyphenyl][N‐(l‐pro‐pyl)piperidine (R(+)‐{3H]3‐PPP} to cortical homogenates by a range of drugs was consistent with the site labelled being a σ‐receptor. (+)‐SKF 10,047, (‐)‐SKF 10,047, (±)‐cyclazo‐cine, phencyclidine, and dexoxadrol displaced R(+)‐[3H]3‐PPP with pseudo‐Hill coefficients of less than . Further analysis employing nonlinear curve fitting techniques demonstrated that displacement data for these compounds were described better by a model whereby R(+)‐[3H]3(‐PPP was displaced from two discrete sites; approximately 6[5% of the total sites were in the high‐affinity state. In the presence of 10 mM Mg2+ and 0.3 mM GTP, displacement curves for (+)‐SKF 10,047 and (±)‐cyclazocine were shifted to the right. These findings were due to the shift of some 15% of the high‐affinity binding sites to a low‐affinity state. Saturation experiments revealed that 0.3 mM GTP acted competitively to decrease the affinity of R(+)‐[3H]3‐PPP for the σ suites. The σ‐binding site was thus likely to be linked to a guanine nucleotide regulatory (G) protein. Thus σ drugs could be subdivided on the basis of their GTP sensitivity and psdudo‐Hill coefficients, and by analogy with other receptors R(+)‐3‐PPP. (+)‐SKF 10,047, and (±)‐cyclazocine, may be putative σ‐agonists. l,3‐Di(2‐tolyl)guanidine (DTG), rimcazole, and haloperidol displaced R(+)‐[3H]3‐PPP with pseudo‐Hill coefficients of approximately unity and thus may be σ‐antagonists. Subchronic treatment with rimcazole was characterized by slight sedation and a concomitant up‐regulation, with a decrease in the affinity, of σr‐binding sites. The schedule of rimcazole also increased dopamine turnover in the nucleus ac‐cumbens; both the concentration of 3,4‐dihydroxyphenyl‐acetic acid (DOPAC) and the DOPAC/dopamine ratio were elevated. DTG produced similar alterations to the binding parameters of the σ‐binding site; however, changes were not observed in general behavior or accumbal dopamine turnover. (T‐Receptors are likely to be linked to a G protein and are functionally involved in the CNS.
Tyrosine Hydroxylase Content of Residual Striatal Dopamine Nerve Terminals Following 6‐Hydroxydopamine Administration: A Flow Cytometric Study
Authors: Wolf Marina E. , Zigmond Michael J. , Kapatos Gregory
Published: October 2006

Fluorescence‐activated cell sorting based on im‐munolabeling with a monoclonal antibody to tyrosine hydroxylase and a fluorescein‐conjugated secondary antibody was used to identify striatal synaptosomes derived from ni‐grostriatal dopamine nerve terminals. The amount of tyrosine hydroxylase immunoreactivity in dopaminergic striatal synaptosomes prepared from control rats was compared to the amount in dopaminergic synaptosomes prepared from rats that had received intraventricular injections of 6‐hydroxy‐dopamine. Although the absolute number of dopaminergic synaptosomes was decreased in lesioned animals, those residual dopamine terminals present contained more tyrosine hydroxylase than did dopamine terminals from control rats. Both the decrease in the absolute number of dopamine terminals and the increase in tyrosine hydroxylase immunoreactivity in residual terminals were proportional to the extent of the lesion, as determined by measurement of striatal dopamine levels. These results suggest that an increase in the amount of tyrosine hydroxylase protein in residual terminals may represent one compensatory mechanism by which residual dopamine neurons maintain normal striatal function after partial destruction of the nigrostriatal dopamine projection.
Monoamine Oxidase (MAO)‐A but Not MAO‐B Inhibitors Potentiate Tyramine‐Induced Catecholamine Release from PC12 Cells
Authors: Youdim Moussa B. H.
Published: October 2006

The previous report that PC12 (pheochromocytoma) cells have a K+‐induced, as well as a tyramine‐induced, catecholamine release mechanism has been confirmed. Selective monoamine oxidase (MAO)‐A (clorgyline and moclobemide) and not MAO‐B inhibitors (l‐deprenyl, AGN 1135, and Ro 16–6491) potentiate the catecholamine‐releasing action of tyramine significantly more than that of K+. The potentiation of tyramine‐induced [3H]noradrenaline release from PC 12 cells by MAO‐A inhibitors has been linked to the presence of MAO‐A in these cells, for which tyramine and noradrenaline are substrates. In the above respects, it is the PC 12 cell that resembles more closely the peripheral adrenergic neuron, rather than the chromaffin cell, which is endowed with MAO‐B and lacks the tyramine‐releasable pool of catecholamines.
Effects of Choline Administration on In Vivo Release and Biosynthesis of Acetylcholine in the Rat Striatum as Studied by In Vivo Brain Microdialysis
Authors: Koshimura Kunio , Miwa Soichi , Lee Ken , Hayashi Yasunori , Hasegawa Hiroshi , Hamahata Keigo , Fujiwara Motohatsu , Kimura Mieko , Itokawa Yoshinori
Published: October 2006

The purpose of the present study is to clarify the effects of the administration of choline on the in vivo release and biosynthesis of acetylcholine (ACh) in the brain. For this purpose, the changes in the extracellular concentration of choline and ACh in the rat striatum following intracerebroventricular administration of choline were determined using brain microdialysis. We also determined changes in the tissue content of choline and ACh. When the striatum was dialyzed with Ringer solution containing 10 μM physostigmine, ACh levels in dialysates rapidly and dose dependently increased following administration of various doses of choline and reached a maximum within 20 min. In contrast, choline levels in dialysates increased after a lag period of 20 min following the administration. When the striatum was dialyzed with physostigmine‐free Ringer solution, ACh could not be detected in dialysates both before and even after choline administration. After addition of hemicholinium‐3 to the perfusion fluid, the choline‐induced increase in ACh levels in dialysates was abolished. Following administration of choline, the tissue content of choline and ACh increased within 20 min. These results suggest that administered choline is rapidly taken up into the intracellular compartment of the cholinergic neurons, where it enhances both the release and the biosynthesis of ACh.
Identification of Glucagon Receptors in Rat Retina
Authors: Fernandez‐Durango R. , Sanchez D. , Fernandez‐Cruz A.
Published: October 2006

In this study, we characterize the glucagon receptors on rat retinal particulate preparations. The specific binding of 125I‐glucagon was saturable and reversible. Apparent equilibrium conditions were established within 30–45 min. Analysis of binding data is compatible with the existence of two classes of binding sites: a high‐affinity class with a KD of 7 ± 0.8 nM and a Bmax of 2.3 ± 0.2 pmol/mg of protein and a low‐affinity class with a KD of 84.4 ± 2.5 nM and a Bmax of 16.5 ± 2.3 pmol/mg of protein. The 125I‐glucagon binding to retinal particulate preparation was not inhibited by 1 μM concentrations of insulin, atrial natriuretic factor, angiotensin II, somatostatin, and vasoactive intestinal peptide. However, synthetic human pancreatic growth hormone‐releasing factor, hGRF‐44, inhibited binding, although the concentration required for half‐maximal displacement was 10‐fold higher than that for native glucagon. Glucagon binding was GTP sensitive. Inclusion of 0.1 mM GTP in the binding assay produced an increase in the concentration of unlabeled glucagon required for half‐maximal displacement of 125I‐glucagon, from 23 to 220 nM. Glucagon stimulated adenylate cyclase formation in retinal particulate preparations. The concentration of glucagon required for half‐maximal activation of retinal adenylate cyclase was 16.2 nM. These results suggest that glucagon may play a role as a neurosignal transmitter in rat retina.
Neurotensin Binding to Dopamine
Authors: Adachi Derek K. , Kalivas Peter W. , Schenk James O.
Published: October 2006

Rotating disk electrode voltammetry at glassy carbon electrodes and ultraviolet/visible spectroscopy were used to demonstrate that dopamine binds to neurotensin with a dissociation constant of 7.5 × 10‐8. By measuring the binding constants of various neurotensin analogs, it was found that the ‐Arg8‐Arg9‐portion of the neurotensin sequence is critical for binding dopamine. Neurotensin also formed a complex with 4‐ethylcatechol, 4‐methylcatechol, 3‐methoxytyramine, and norepinephrine. Although changes in the side chain did not alter the binding constant, methoxylation of the catechol moiety significantly increased the dissociation constant. These data along with additional studies of dopamine interactions with arginine derivatives suggest that the guanidino groups of arginine and the catechol hydroxyls of dopamine are responsible for mediating the observed binding. It is hypothesized that the capacity of neurotensin to bind directly to dopamine may be partly responsible for its previously observed antagonism of dopamine‐induced locomotor activity.
Autoradiographic Visualization of A1 Adenosine Receptors in Rat Brain with [3H]8‐Cyclopentyl‐1,3‐Dipropylxanthine
Authors: Weber Ruthild G. , Jones C. Richard , Lohse Martin J. , Palacios José M.
Published: October 2006

A1 adenosine receptors were labeled in rat brain sections with the antagonist [3H]8‐cyclopentyl‐1,3‐dipropylxanthine ([3H]DPCPX) and visualized at the light microscopic level using autoradiography. The specific binding of [3H]DPCPX to the sections showed the pharmacological characteristics of A1 adenosine receptors and was accompanied by very low levels of nonspecific binding. Whereas GTP had no significant effect on [3H]DPCPX binding to rat brain membranes, the addition of 100 μM GTP increased the apparent affinity of [3H]DPCPX to tissue sections fivefold (from 1.83 to 0.35 nM), enhancing it to the affinity measured in membranes. However, GTP altered neither the binding capacity nor the distribution of binding sites in tissue sections. It is suggested that a competitive antagonism with endogenous adenosine explains the lower affinity of [3H]DPCPX in the absence of GTP. The autoradiographic pattern of [3H]DPCPX binding was characteristic for A1 adenosine receptors. Distinct labeling of the different layers of the cerebellar cortex was shown by photomicrographs generated with the coverslip technique. In addition, several fiber tracts were found to be labeled. The high selectivity for A1 adenosine receptors and low nonspecific binding of [3H]DPCPX, the ability to produce high‐resolution autoradiograms, together with the fact that the effects of endogenous adenosine can be eliminated by the addition of GTP make [3H]DPCPX a very useful tool in the autoradiographic study of A1 adenosine receptors.
Extremely compact massive galaxies at z ∼ 1.4
Authors: Trujillo I. , Feulner G. , Goranova Y. , Hopp U. , Longhetti M. , Saracco P. , Bender R. , Braito V. , Della Ceca R. , Drory N. , Mannucci F. , Severgnini P.
Published: October 2006

The optical rest-frame sizes of 10 of the most massive (∼5 × 1011h−270 M⊙) galaxies found in the near-infrared MUNICS survey at 1.2 < z < 1.7 are analysed. Sizes are estimated in both the J and K′ filters. These massive galaxies are at least a factor of 4+1.9−1.0 (±1σ) smaller in the rest-frame V-band than local counterparts of the same stellar mass. Consequently, the stellar mass density of these objects is (at least) 60 times larger than that of massive ellipticals today. Although the stellar populations of these objects are passively fading, their structural properties are rapidly changing since that redshift. This observational fact disagrees with a scenario where the more massive and passive galaxies are fully assembled at z∼ 1.4 (i.e. a monolithic scenario) and points towards a dry merger scenario as the responsible mechanism for the subsequent evolution of these galaxies.
Interaction Between Crystalline Penicillin and Human Plasma Proteins
Authors: Chow Bacon F. , McKee Clara M.
Published: October 2006

Penicillin combines with human serum albumin, as demonstrated by dialysis experiments. Unlike the sulfonamide-albumin complex, the penicillin-albumin possesses antibiotic activity. Such a complex can be freed of unbound penicillin by repeated precipitation at 50 per cent. alcohol and obtained as a dried powder. Mouse excretion studies indicated that the penicillin-albumin complex was excreted more slowly than the sodium salt of penicillin.
Translocation of Carbohydrates in Maize
Authors: Loomis W. E.
Published: October 2006

(1) The pith of maize shows a changing sugar content with changing sugar movement in the phloem which would not be predicted from our present picture of the morphology of the maize bundle.

(2) By all tests of changing concentration, sucrose is the important carbohydrate of translocation in maize. Interconversion of the several sugars is too rapid, however, to permit a final conclusion.

(3) An hypothesis of translocation in maize must not only account for movement against an osmotic gradient, but against gradients of each of the substances which might possibly be translocated. Such secretory translocation certainly occurs between the leaf mesophyll and the phloem and probably along the phloem itself.

(4) Translocation in maize is polarized, out of the leaf, out of the xylem and toward the developing fruit. Polarized translocation out of the leaf is established during the later stages of tissue differentiation. Polarized translocation toward the fruit is established in the early phases of embryo development and does not develop in the absence of pollination.
Thiamine Deficiency and High Estrogen Findings in Uterine Cancer and in Menorrhagia
Authors: Ayre J. Ernest , Bauld W. A. G.
Published: October 2006

The finding of abnormal estrogenic activity coupled with thiamine deficiency in cases of menorrhagia and uterine cancer suggests a possible etiological correlation between the dietary deficiency, the abnormal estrogen level, and the pathological lesion.

The specific element deficient in these cases was thiamine, while the other B factors were normal.

Preliminary report of the evidence is made in this small series while more extensive studies on a large series of cases are being pursued.

Cornification in cytology smears was used to study estrogenic activity, since the present study was prompted by cytological findings; the method is simple, practical, and reasonably accurate. The urinary estimation measures only the amount excreted, and if liver impairment actually is present, the quantity excreted would not give a true index of the amount retained in the body.

Further studies are being undertaken in which estrogenic, urinary, and cornification levels are being compared before and after thiamine administration in cases proven to be deficient.