bluuebunny/crossref_metadata_2025 huggingface dataset:
[3H]8‐Hydroxy‐2‐(Di‐n‐Propylamino)Tetralin Binding to Pre‐ and Postsynaptic 5‐Hydroxytryptamine Sites in Various Regions of the Rat Brain
Authors: Hall M. D. , Mestikawy S. El , Emerit M. B. , Pichat L. , Hamon M. , Gozlan H.
Published: October 2006

The specific binding of [3H]8‐hydroxy‐2‐(di‐n‐propylamino)tetralin ([3H]8‐OH‐DPAT) to 5‐hydroxytryptamine (5‐HT)‐related sites was investigated in several regions of the rat brain. Marked differences were observed in the characteristics of binding to membranes from hippocampus, striatum, and cerebral cortex. Hippocampal sites exhibited the highest affinity (KD > 2 nM) followed by the cerebral cortex (KD > 6 nM) and the striatum (KD > 10 nM). Ascorbic acid inhibited specific [3H]8‐OH‐DPAT binding in all three regions but millimolar concentrations of Ca2+, Mg2+, and Mn2+ enhanced specific binding to hippocampal membranes, whereas only Mn2+ increased it in the cerebral cortex and all three cations inhibited specific binding to striatal membranes. Guanine nucleotides (0.1 mM GDP, GTP) inhibited binding to hippocampal and cortical membranes only. As intracerebral 5,7‐dihydroxytryptamine markedly decreased [3H]8‐OH‐DPAT binding sites in the striatum, but not in the hippocampus, the striatal sites appear to be on serotoninergic afferent fibers. In contrast, in the hippocampus the sites appear to be on postsynaptic 5‐HT target cells, as local injection of kainic acid decreased their density. Both types of sites appear to be present in the cerebral cortex. The postsynaptic hippocampal [3H]8‐OH‐DPAT binding sites are probably identical to the 5‐HT1A, subsites, but the relationship between the presynaptic binding sites and the presynaptic autoreceptors controlling 5‐HT release deserves further investigation.
Membrane Potential and Catecholamine Secretion by Bovine Adrenal Chromaffin Cells: Use of Tetraphenylphosphonium Distribution and Carbocyanine Dye Fluorescence
Authors: Friedman J. E. , Lelkes P. I. , Lavie E. , Rosenheek K. , Schneeweiss F. , Schneider RA. S.
Published: October 2006

Abstract: Changes in plasma membrane potential of isolated bovine adrenal chromaffin cells were measured independently by two chemical probe methods and related to corresponding effects on catecholamine secretion. The lipophilic cation tetraphenylphosphonium (TPP+) and the carbocyanine dye 3,3′‐dipropylthiadicarbocyanine [DiS‐C3‐(5)] were used. The necessity of evaluating the subcellular distribution of TPP+ among cytoplasmic, mitochondrial, secretory granule, and bound compartments was demonstrated and the resting plasma membrane potential determined to be – 55 mV. The relationship between membrane potential and catecholamine secretion was determined in response to variations in extracellular K+ and to the presence of several secretagogues including cholinergic receptor ligands, veratridine, and ionophores for Na+ and K+. The dependence of potential on K+ concentration fit the Goldman constant field equation with a Na/K permeability ratio of 0.1. The dependence of both K+ ‐ and veratridine‐evoked catecholamine secretion on membrane potential exhibited a potential threshold of about – 40 mV before a significant rise in secretion occurred. This is likely related to the threshold for opening of voltage‐sensitive Ca2+ channels. Acetylcholine and nicotine evoked a large secretory response without a sufficiently sustained depolarization to be detectable by the relatively slow potential sensitive chemical probes. Decamethonium induced a detectable depolarization of the chromaffin cells. Veratridine and gramicidin evoked both membrane depolarization and catecholamine release. By contrast the K ionophore valinomycin evoked significant levels of secretion without any depolarization. This is consistent with its utilization of an intracellular source of Ca2+ and the independence of its measured secretory response on extracellular Ca2+
Acetylcholine, ATP, and Proteoglycan Are Common to Synaptic Vesicles Isolated from the Electric Organs of Electric Eel and Electric Catfish as Well as from Rat Diaphragm
Authors: Volknandt W. , Zimmermann H.
Published: October 2006

AbstractCholinergic synaptic vesicles were isolated from the electric organs of the electric eel (Electrophorus electricus) and the electric catfish (Malapterurus electricus) as well as from the diaphragm of the rat by density gradient centrifugation followed by column chromatography on Sephacryl‐1000. This was verified by both biochemical and electron microscopic criteria. Differences in size between synaptic vesicles from the various tissue sources were reflected by their elution pattern from the Sephacryl column. Specific activities of acetylcholine (ACh; in nmol/mg of protein) of chromatography‐purified vesicle fractions were 36 (electric eel), 2 (electric catfish), and 1 (rat diaphragm). Synaptic vesicles from all three sources contained ATP in addition to ACh (molar ratios of ACh/ATP, 9–12) as well as binding activity for an antibody raised againstTorpedocholinergic synaptic vesicle proteoglycan. Synaptic vesicles from rat diaphragm contained binding activity for the monoclonal antibody asv 48 raised against a rat brain 65‐kilodalton synaptic vesicle protein. Antibody asv 48 binding was absent from electric eel and electric catfish synaptic vesicles. These antibody binding results, which were obtained by a dot blot assay on isolated vesicles, directly correspond to the immuno‐cytochemical results demonstrating fluorescein isothiocyanate staining in the respective nerve terminals. Our results imply that ACh, ATP, and proteoglycan are common molecular constituents of motor nerve terminal‐derived synaptic vesicles fromTorpedoto rat. In addition to ACh, both ATP and proteoglycan may play a specific role in the process of cholinergic signal transmission.
Perinatal Hypoxia‐Ischemia Disrupts Striatal High‐Affinity [3H]Glutamate Uptake into Synaptosomes
Authors: Silverstein Faye S. , Buchanan Karen , Johnston Michael V.
Published: October 2006

Abstract We examined the impact of hypoxia‐ischemia on high‐affinity [3H]glutamate uptake into a synaptosomal fraction prepared from immature rat corpus striatum. In 7‐day‐old pups the right carotid artery was ligated, and pups were exposed to 8% oxygen for 0, 0.5, 1, or 2.5 h, and allowed to recover for up to 24 h before they were killed. High‐affinity glutamate uptakes in striatal synaptosomes derived from tissue ipsilateral and contralateral to ligation were compared. After 1 h of hypoxia plus ischemia, high‐affinity glutamate uptake in the striatum was reduced by 54 ± 13% compared with values from the opposite (nonischemic) side of the brain (p < 0.01, t test versus ligates not exposed to hypoxia). There were similar declines after 2.5 h of hypoxiaischemia. Activity remained low after a 1 h recovery period in room air, but after 24 h of recovery, high‐affinity glutamate uptake was equal bilaterally. Kinetic analysis revealed that loss of activity could be attributed primarily to a 40% reduction in the number of uptake sites. Hypoxia alone had no effect on high‐affinity glutamate uptake although it reduced synaptosomal uptake of [3H]3,4‐dihydroxyphenyl‐ethylamine. Addition of 1 mg/ml of bovine serum albumin to the incubation medium preferentia'ly stimulated high‐affinity glutamate uptake in hypoxic‐ischemic brain compared with its effects in normal tissue. These studies demonstrate that hypoxia‐ischemia reversibly inhibits high‐affinity glutamate uptake and this occurs earlier than the time required to produce neuronal damage in the model.
Effects of Angiotensin II on [3H]Noradrenaline Release and Phosphatidylinositol Hydrolysis in the Parietal Cortex and Locus Coeruleus of the Rat
Authors: Huang Yu , Rogers Jane , Henderson Graeme
Published: October 2006

Abstract: Angiotensin II (ANGII) (3–100 nM) facilitated the potassium‐evoked (22.5 mM) release of [3H]‐noradrenaline ([3H]NA) from slices of parietal cortex in a concentration‐dependent manner, but did not significantly alter the release of [3H]NA evoked in a similar manner from locus coeruleus slices. The facilitatory action of ANGII was blocked by saralasin (0.1–3 μM). Neither nimodipine (10–30 μM) nor phenylmethylsulphonyl fluoride (1 mM) altered either [3H]NA release or the facilitatory action of ANGII in the parietal cortex. Carbachol (0.01 ‐3 mM) and raised potassium (22.5 mM), but not ANGII (3–100 nM), stimulated the production of inositol phosphates in parietal cortex slices. The potassium‐evoked increase in inositol phosphate production was unaffected by ANGII (3–100 nM). In the locus coeruleus, ANGII (3–100 nM) did not stimulate inositol phosphate production. The mechanism underlying the ANGII facilitation of (3H]NA release from the parietal cortex does not appear to involve either nimodipine‐sensitive calcium channels or, as far as we have been able to determine, the release of calcium from intracellular stores following the breakdown of phospho‐inositides.
Chromogranin A Synthesis and Secretion in Chromaffin Cells
Authors: Eiden Lee E. , Iacangelo Anna , Hsu Chang‐Mei , Hotchkiss Adair J. , Bader Marie‐France , Aunis Dominique
Published: October 2006

Abstract: A sensitive and selective radioimmunoassay for chromogranin A (Chrg A) has been developed to quantitate content, release, and biosynthesis of this secretory protein in neuroendocrine tissues. An antiserum raised against Chrg A from bovine adrenal medulla was found to detect predominantly only the Mr 70–75 kilodalton Chrg A in its native form, allowing the use of this antiserum as a quantitatively specific probe for Chrg A in cell‐free extracts of the adrenal medulla and chromaffin cells. Chrg A comprises about 10% of the total protein of the chromaffin cell. It is released in parallel with Met‐enkephalin and catechol‐amines from the bovine chromaffin cell in primary culture in response to nicotine and nicotinic cholinergic agonists. From 14 to 22% of total Chrg A is released from the cell during a 15‐min exposure to a maximally stimulatory dose of nicotine (10–100 μM). Chrg A release on nicotinic stimulation is blocked by D‐600 and hexamethonium to the same extent as Met‐enkephalin and catecholamine release. The parallel time course and percent release of Chrg A and Met‐enkephalin indicate that these secretory polypeptides are contained in, and released from, functionally identical cellular compartments. Chrg A and Met‐enkephalin pentapeptide sequences are present in the chromaffin cell at a ratio of about 2:1, although Chrg A is far more abundant on a mass basis. Chrg A and Met‐enkephalin biosynthesis appear to be differentially regulated within the chromaffin cell, since chronic treatment of cells with nicotine and forskolin causes an elevation of Met‐enkephalin pentapeptide without a concomitant elevation of intracellular levels of Chrg A.
Polyamine Regulation of Protein Phosphorylation in the Brain of the Tobacco Hornworm, Manduca sexta
Authors: Birnbaum Mark J. , Combest Wendell L. , Bloom Timothy J. , Gilbert Lawrence I.
Published: October 2006

Abstract: An analysis of the effects of polyamines on protein phosphorylation in cytosolic fractions of the pupal brain of Manduca sexta showed that spermine elicited an increase in casein phosphorylation in a dose‐dependent manner (maximum three‐ to fourfold at 2.0 mM), whereas spermidine was less effective and putrescine was without effect. In contrast, with phosvitin as the exogenous substrate, higher doses of polyamines, especially spermine, inhibited phosphorylation. High salt conditions abolished the polyamine response. Cytosol protein kinase activity eluted from DEAE‐cellulose at 0.2–0.3 M NaCl. This activity was enhanced in the presence of spermine, and inhibited in the presence of heparin (IC50∼ 30 ng/ml). The enzyme was characterized by a sedimentation coefficient of 6.5S, and a Stokes radius of 49 Å, consistent with a Mr of 130,000. Both GTP (Km, 55 μM) and ATP (Km, 34 μM) were utilized as phosphoryl donors (Vmax for ATP being four‐fold higher than that observed for GTP). These results indicate the presence in the insect brain of an enzyme very similar to vertebrate casein kinase II. Sodium dodecyl sulfate‐polyacrylamide gel electrophoresis and autoradiography demonstrated that low concentrations of spermine (100 μM) strongly enhanced the phosphorylation of three high‐molecular‐weight cytosolic proteins (305,000, 340,000, and 360,000) localized in the insect nervous system.
Biochemistry of an Olfactory Purinergic System: Dephosphorylation of Excitatory Nucleotides and Uptake of Adenosine
Authors: Trapido‐Rosenthal Henry G. , Carr William E. S. , Gleeson Richard A.
Published: October 2006

Abstract: The olfactory organ of the spiny lobster, Panu‐lirus argus, is composed of chemosensory sensilla containing the dendrites of primary chemosensory neurons. Receptors on these dendrites are activated by the nucleotides AMP, ADP, and ATP but not by the nucleoside adenosine. It is shown here that the lobster chemosensory sensilla contain enzymes that dephosphorylate excitatory nucleotides and an uptake system that internalizes the nonexcitatory dephosphorylated product adenosine. The uptake of [3H]‐adenosine is saturable with increasing concentration, linear with time for up to 3h, sodium dependent, insensitive to moderate pH changes and has a Km of 7.1 μM and a Vmax of 5.2 fmol/sensillum/min (573 fmol/μg of protein/min). Double‐label experiments show that sensilla dephosphorylate nucleotides extracellularly; 3H from adenine‐labeled AMP or ATP is internalized, whereas 32P from phosphate‐labeled nucleotides is not. The dephosphorylation of AMP is very rapid; 3H from AMP is internalized at the same rate as 3H from adenosine. Sensillar 5′‐ectonucleotidase activity is inhibited by ADP and the ADP analog α,β‐methylene ADP. Collectively, these results indicate that the enizymes and the uptake system whereby chemosensory sensilla of the lobster inactivate excitatory nucleotides and clear adenosine from extracellular spaces are very similar to those present in the internal tissues of vertebrates, where nucleotides have many neuroactive effects.
Selective Depletion of Dopamine, Octopamine, and 5‐Hydroxytryptamine in the Nervous Tissue of the Cockroach (Periplaneta americana)
Authors: Sloley B. Duff , Orikasa Shuzo
Published: October 2006

Abstract: High‐performance liquid chromatography with electrochemical detection was used to measure the concentrations of 3,4‐dihydroxyphenylethylamine (dopamine), 5‐hydroxytryptamine (5‐HT), p‐hydroxyphenylethanolamine (octopamine), α‐methyl‐p‐tyrosine, and tryptophan in the cerebral ganglia of cockroaches (Periplaneta americana) after peripheral administration of α‐methyl‐p‐tyrosine and α‐methyltryptophan. In addition, the levels of dopamine, 5‐HT, octopamine, α‐methyl‐p‐tyrosine, and tryptophan were determined after injection of α‐methyl‐p‐tyrosine, 6‐hydroxydopamine, or 5,7‐dihydroxytryptamine directly into the cerebral ganglia by means of microinjection needles. Peripheral administration of α‐methyl‐p‐tyrosine (400–1,600 μg/insect) caused a reduction in dopamine and 5‐HT concentrations in cockroach cerebral ganglia, although the reduction in dopamine concentrations was more pronounced. Peripheral injections of α‐methyl‐p‐tyrosine also reduced octopamine levels in the cerebral ganglia. Peripheral injection of α‐methyltryptophan (400–1,600 μg/insect) caused a marked reduction in 5‐HT and tryptophan concentrations in cockroach cerebral ganglia without altering dopamine or octopamine concentrations. Central injections of α‐methyl‐p‐tyrosine (80 μg/insect) reduced dopamine concentrations in the cerebral ganglia. However, neither 6‐hydroxydopamine (20 μg/insect) nor 5,7‐dihydroxytryptamine (20 μg/insect) caused reductions in amine levels when applied near or directly into the cerebral ganglia. The results suggest that specific lesions of aminergic neurons in insects by either 6‐hydroxydopamine or 5,7‐dihydroxytryptamine are impractical. The specific, long‐lasting depletion of 5‐HT by α‐methyltryptophan suggests that this chemical may be useful in elucidating the functions of 5‐HT in insects.
Modulation of the Benzodiazepine/γ‐Aminobutyric Acid Receptor Chloride Channel Complex by Inhalation Anesthetics
Authors: Moody Eric J. , Suzdak P. D. , Paul S. M. , Skolnick P.
Published: October 2006

Abstract: Inhalation anesthetics, such as diethyl ether, halo‐thane, and enflurane, increase 36C1− uptake into rat cerebral cortical synaptoneurosomes in a concentration‐dependent, picrotoxin‐sensitive fashion. At concentrations consistent with those that stimulate 36C1− uptake, inhalation anesthetics also inhibit the binding of t‐[35S]butylbicyclophosphoro‐thionate ([35S]TBPS) to well‐washed cortical membranes. Scatchard analysis of [35S]TBPS binding indicates that these agents reduce the apparent affinity of this radioligand and have little effect on the Bmax. The ability of inhalation anesthetics to directly stimulate 36C1− uptake and inhibit [35S]TBPS binding is a property shared by nonvolatile anesthetics. Nonetheless, there are differences between nonvolatile agents (such as barbiturates and alcohols) and in halation anesthetics, because the former compounds augment muscimol (a GABAmimetic) stimulated 36C1− uptake, whereas the latter group (such as ether and enflurane) inhibit this effect. These findings demonstrate that therapeutically relevant concentrations of inhalation anesthetics perturb the benzodiazepine/γ‐aminobutyric acid receptor chloride channel complex, and suggest this oligomeric protein may be a common mediator of some aspects of anesthetic action.
Neutral Amino Acid Transport in Astrocytes: Characterization of Na+‐Dependent and Na+‐Independent Components of α‐Aminoisobutyric Acid Uptake
Authors: Brookes N.
Published: October 2006

Abstract: Neutral amino acid transport is largely unexplored in astrocytes, although a role for these cells in blood‐brain barrier function is suggested by their close apposition to ce‐rebrovascular endothelium. This study examined the uptake into mouse astrocyte cultures of α‐aminoisobutyric acid (AIB), a synthetic model substrate for Na+‐dependent system A transport. Na+‐dependent uptake of AIB was characteristic of system A in its pH sensitivity, kinetic properties, regulatory control, and pattern of analog inhibition. The rate of system A transport declined markedly with increasing age of the astrocyte cultures. There was an unexpectedly active Na+‐independent component of AIB uptake that declined less markedly than system A transport as culture age increased. Although the saturability of the Na+‐independent component and its pattern of analog inhibition were consistent with system L transport, the following properties deviated: (1) virtually complete inhibition of Na+‐independent AIB uptake by characteristic L system substrates, suggesting unusually high affinity of the transporter; (2) apparent absence of transstimulation of AIB influx; (3) unusually concentrative uptake at steady state (the estimated distribution ratio for 0.2 mM AIB was 55); and (4) susceptibility to inhibition by N‐ethylmaleimide. Direct study of the uptake of system L substrates in astrocytes is needed to confirm the present indications of high affinity and concentrative Na+‐independent transport.
Selective Destruction of Cultured Dopaminergic Neurons from Fetal Rat Mesencephalon by 1‐Methyl‐4‐Phenylpyridinium: Cytochemical and Morphological Evidence
Authors: Sanchez‐Ramos Juan R. , Michel Patrick , Weiner William J. , Hefti Franz
Published: October 2006

Abstract: Dopaminergic neurons in cultures of dissociated cells from fetal rat mesencephalon were exposed to the principal metabolite of the neurotoxin 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP), 1‐methyl‐4‐phenyl‐pyridinium ion (MPP+), and several of its structural analogues. At concentrations between 0.01 and 0.1 μM, MPP+ inhibited catecholamine accumulation as visualized by cytofluorescence. Between 0.1 and 10.0 μM, MPP+ resulted in disappearance of tyrosine hydroxylase immunoreactivity without affecting other cells in the cultures. At concentrations higher than 10 μM, MPP+ was toxic to all cells present in the cultures. The effect of low concentrations of MPP+ on catecholamine cytofluorescence of the dopaminergic neurons was partially reversible. The intermediate concentrations produced irreversible structural changes of tyrosine hydroxylase‐positive cells, resulting in complete disappearance of these neurons. The morphological changes were specific to the dopaminergic neurons and were not evident in other cells viewed with phase contrast microscopy. Of the structural analogues tested, the 1‐ethyl analogue of MPP+ was effective in selectively destroying dopaminergic neurons in our culture system. The antioxidants l‐acetylcarnitine, β‐carotene, and α‐tocopherol failed to protect against MPP+ neurotoxicity when co‐incubated with the toxin.
Pyruvate Dehydrogenase Activity in Osmotically Shocked Rat Brain Mitochondria: Stimulation by Oxaloacetate
Authors: Haas Richard H. , Thompson Geoffrey , Morris Bernard , Conright Kelly , Andrews Torre
Published: October 2006

Pyruvate dehydrogenase complex activity (PDHC) measured by CO2 release isotopic assay has generally been much lower than activity measured by the spec‐trophotometric arylamine acetyltransferase assay (ArAT). Decarboxylation of [l‐14C]pyruvate was measured in osmotically shocked rat brain cortical mitochondria. Activity is dependent on the concentration of the substrate pyruvate. Activity of 74.6 units ± 12.3 SD (n = 22) was observed at 4 mM pyruvate (1 unit = 1 nmol pyruvate decarboxy‐lated/min/mg protein). Activity was dependent on added NAD, CoA, and thiamine pyrophosphate, implying increased mitochondrial permeability after osmotic shock. Freeze/thaw with sonication of the mitochondrial preparation reduced PDHC activity to 11.5 units ± 3.0 SD (n = 4). Oxaloacetate produced a marked stimulation of activity. The optimal assay contained 3 mM oxaloacetate, and without oxaloacetate activity fell to 15.4 units ± 9.9 SD (n = 8). These studies highlight the importance of optimal substrate concentrations in the CO2 release isotopic PDHC method. Higher PDHC activity is found with intact mitochondria and thus activity values should be interpreted in the light of the presence or absence of intact mitochondria in individual preparations.
Kinetic Characterization of Inhibition of γ‐Aminobutyric Acid Uptake into Cultured Neurons and Astrocytes by 4,4‐Diphenyl‐3‐Butenyl Derivatives of Nipecotic Acid and Guvacine
Authors: Larsson O. M. , Falch E. , Krogsgaard‐Larsen P. , Schousboe A.
Published: October 2006

The effects of N‐(4,4‐diphenyl‐3‐butenyl) derivatives of nipecotic acid (SKF‐89976‐A and SKF‐100844‐A) and guvacine (SKF‐100330‐A) on neuronal and astroglial γ‐aminobutyric acid (GABA) uptake were investigated. In addition, the uptake of SKF‐89976‐A was studied using the tritiated compound. All of the compounds were found to be competitive inhibitors of GABA uptake irrespective of the cell type, with Ki values similar to or lower than those of the parent amino acids. Moreover, none of the compounds exhibited selectivity with regard to inhibition of neuronal and glial GABA uptake. In spite of the competitive nature of SKF‐89976‐A, the compound was not transported by the GABA carriers in the two cell types, because no saturable uptake could be demonstrated.
Adenosine Receptors Activate Adenylate Cyclase and Enhance Secretion from Bovine Adrenal Chromaffin Cells in the Presence of Forskolin
Authors: Chern Yi‐Juang , Kim Kyong‐Tai , Slakey Linda L. , Westhead Edward W.
Published: October 2006

Abstract: Cells of the adrenal medulla release not only catecholamines but also high concentrations of neuropeptides and nucleotides. Chromaffin cells, like many neuronal cells, have a diversity of receptors: adrenergic receptors, peptide receptors, histamine receptors, and dopamine receptors. We recently reported that these cells have nucleotide receptors that can mediate inhibition of the secretory response. The present studies show that adenosine, in the presence of enabling concentrations of forskolin, can potently enhance response to nicotinic stimulation. Neither adenosine nor forskolin alone produces a significant effect. A marked rise in intracellular cyclic AMP (cAMP) concentration is associated with the enhancement of secretion caused by forskolin plus adenosine. A phosphodiesterase inhibitor, Ro 20–1724, used together with forskolin produces significant increases in both cellular cAMP content and catecholamine secretion. However, the adenosine agonist 5′‐N‐ethylcarboxyadenosine elevates cellular cAMP content in the presence of forskolin without having any positive effect on secretion. This finding suggests that the rise in cAMP level may not be the sole cause of the increase in secretion by adenosine.
Butyrylcholinesterase in Human Brain and Acetylcholinesterase in Human Plasma: Trace Enzymes Measured by Two‐Site Immunoassay
Authors: Brimijoin Stephen , Hammond Pamela
Published: October 2006

Abstract: Enzyme‐linked immunosorbent assays for acetylcholinesterase (AChE) and for butyrylcholinesterase (BuChE) were markedly more specific than conventional assays using selective enzyme inhibitors. The new assays were used with blood and brain samples containing traces of one enzyme dominated by large amounts of the other. The results showed that human plasma does contain AChE (8 ng/ml), even though its major cholinesterase is BuChE (3,300 ng/ml). BuChE immunoreactivity was not detected in human red blood cells but occurred in all brain regions. The cerebellum was the richest region tested (540 ng of BuChE/g of tissue), whereas the cerebral cortex was the poorest (240 ng of BuChE/g). However, because of the small local AChE content (99 ng/g), BuChE was the major cortical cholinesterase. The picture was reversed in the putamen, where BuChE immunoreactivity (340 ng/g) was far outweighed by that of AChE (6,100 ng/g).
Relationship Between the Nerve Growth Factor‐Regulated Clone 73 Gene Product and the 58‐Kilodalton Neuronal Intermediate Filament Protein (Peripherin)
Authors: Aletta John M. , Angeletti Ruth , Liem Ronald K. H. , Purcell Clement , Shelanski Michael L. , Greene Lloyd A.
Published: October 2006

Abstract: Exposure of PC12 cells to nerve growth factor (NGF) has been shown to induce an rnRNA that encodes a novel neuronal intermediate filament protein. The findings presented here concern the identity of this filament protein. The major protein in NGF‐treated PC12 cell cytoskeletons derived by extraction with 1% Triton X‐100 is of apparent Mr= 58,000, focuses by isoelectric focusing as several closely spaced spots of pl 5.6–5.8, and is elevated relative to non‐NGF‐treated cells. Partial microsequencing of this material reveals 2 internal sequences that are identical to a 14‐residue sequence encoded by the NGF‐regulated clone 73 mRNA, but not to sequences of other known proteins. An antiserum raised against a 19‐residue synthetic peptide corresponding to the deduced C‐terminus of the protein encoded by the NGF‐regulated clone 73 mRNA specifically recognizes the 58,000‐Mr protein. Properties of the 58‐kilodalton protein strongly suggest that it corresponds to an intermediate filament protein (peripherin) previously identified in PC12 cells and in peripheral and certain CNS neurons. Identification of the intermediate filament protein encoded by an NGF‐induced message should facilitate studies of its regulation and function.
Phorbol Ester‐Induced Upregulation of Angiotensin II Receptors in Neuronal Cultures Is Potentiated by a Calcium Ionophore
Authors: Sumners Colin , Rueth Susan M. , Myers Linda M. , Kalberg Christopher J. , Crews Fulton T. , Raizada Mohan K.
Published: October 2006

Previous studies have suggested that protein ki‐nase C is important in the regulation of angiotensin II receptors in neuronal cultures, because the C‐kinase agonists, phorbol esters, are able to increase the number of these receptors. In the present study, we have further investigated the role of protein kinase C in angiotensin II receptor regulation. This enzyme is calcium dependent, and so we investigated the effects of A23187, a calcium ionophore, on phorbol ester‐stimulated and basal angiotensin II receptor regulation. A23187, at concentrations that increased 45Ca2+ influx, caused a dose‐dependent potentiation of phorbol‐12‐myristate‐13‐acetate (TPA)‐stimulated upregulation of angiotensin II receptors. This potentiation by A23187 was a further increase in angiotensin II receptor number and was abolished in calcium‐free medium. In the absence of TPA, A23187 caused a decrease in angiotensin II receptor number, an effect not observed in calcium‐free medium. The results suggest at least two pathways for angiotensin II receptor regulation in neuronal cells: (a) by calcium‐dependent protein kinase C and (b) via an influx of calcium into the cell.
Characterization and Regional Distribution of α2‐Adrenoceptors in Postmortem Human Brain Using the Full Agonist [3H]UK 14304
Authors: Meana J. Javier , Barturen Fernando , García‐Sevilla Jesús A.
Published: October 2006

Abstract The full agonist [3H]UK 14304 [5‐bromo‐6‐(2‐im‐idazolin‐2‐yl‐amino)‐quinoxaline] was used to characterize α2‐adrenoceptors in postmortem human brain. The binding at 25°C was rapid (t1/2, 4.6 min) and reversible (t1/2, 14.1 min), and the Kd determined from the kinetic studies was 0.48 nM In frontal cortex, the rank order of potency of adrenergic drugs competing with [3H]UK 14304 or [3H]clonidine showed the specificity for an α2A‐adrenoceptor: UK 14304 ≅ yohimbine ≅ oxymetazoline ≅ clonidine > phentolamine ≅ (–)‐adrenaline > idazoxan ≅ (–) ‐noradrenaline > phenylephrine > (±)‐adrenaline ≫ corynanthine > prazosin ≫ (±)‐propranolol. GTP induced a threefold decrease in the affinity of [3H]UK 14304, with no alteration in the maximum number of binding sites, suggesting that the radioligand labelled the high‐affinity state of the α2‐adreno‐ceptor. In the frontal cortex, analyses of saturation curves indicated the existence of a single population of noninter‐acting sites for [3H]UK 14304 (Kd= 0.35 ± 0.13 nM; Bmax= 74 ± 9 fmol/mg of protein). In other brain regions (hypothalamus, hippocampus, cerebellum, brainstem, caudate nucleus, and amygdala) the Bmax ranged from 68 ± 7 to 28 ± 4 fmol/mg of protein. No significant changes in the Kd values were found in the different regions examined. The binding of [3H]UK 14304 was not affected by age, sex or postmortem delay. [3H]UK 14304 should be a useful tool to assess brain α2‐adrenoceptor density in a variety of neuropsychiatric disorders.
Enhancement by Potassium of Carbachol‐Stimulated Inositol Phospholipid Breakdown in Rat Cerebral Cortical Miniprisms: Comparison with Other Depolarising Agents
Authors: Tiger G. , Björklund P.‐E. , Cowburn R. F. , Fowler C. J.
Published: October 2006

Abstract: Increasing the [K+] in the assay medium from 5.7 to 17.8 mM produces a large enhancement of the inositol phospholipid breakdown response to the muscarinic agonist carbachol in rat cerebral cortical miniprisms, with minor effects on basal inositol phospholipid breakdown. This effect is also found with Rb+. The enhancement by a raised [K+] is not accompanied by a change in the composition of the labelled polyphosphoinositides. The carbachol‐stimulated inositol phospholipid breakdown at 17.8 and 42.7 mM K+ was antagonised by veratrine (5–80 μM), 4‐aminopyridine (5 mM), and tetraethylammonium (20 mM). These compounds, however, also inhibited the binding of [3H]quinuclidinyl benzilate to cortical membranes. BRL 34915 (0.2–20 μM) was without significant effect on carbachol‐stimulated inositol phospholipid breakdown at either 5.7 or 17.8 mM K+. Mg2+ (10 mM) considerably reduced the carbachol‐stimulated inositol phospholipid breakdown at 17.8, but not 42.7, mM K+. Inositol phospholipid breakdown was also stimulated, albeit to a small extent, by L‐glutamate (100–3,000 μM) and quisqualate (1–100 μM), with the stimulation being additive to that produced by carbachol at both 5.7 and 17.8 mM K+. N‐Methyl‐D‐aspartate (10–1,000 μM in Mg2+‐free medium) had no significant effect on basal inositol phospholipid breakdown and had little or no effect on carbachol‐stimulated inositol phospholipid breakdown at either 5.7 or 17.8 mM K+. It is concluded that it may not be correct to ascribe wholly the enhancement by K+ of carbachol‐stimulated inositol phospholipid breakdown to the tissue‐depolarising actions of this ion and that other actions of K+ may be involved.
Protein Kinase C Activation Enhances K+‐Stimulated Endogenous Dopamine Release from Rat Striatal Synaptosomes in the Absence of an Increase in Cytosolic Ca2+
Authors: Chandler L. Judson , Leslie Steven W.
Published: October 2006

Abstract: The possibility that protein kinase C modulates neurotransmitter release in brain was investigated by examining the effects of 12‐O‐tetradecanoylphorbol 13‐acetate (TPA) on Ca2+ transport and endogenous dopamine release from rat striatal synaptosomes. TPA (0.16 and 1.6 μM) significantly increased dopamine release by 24 and 33%, respectively, after a 20‐min preincubation with TPA followed by 60 s of depolarization with 30 mM KCl. Depolarization‐induced 45Ca2+ uptake, measured simultaneously with dopamine release, was not significantly increased by TPA. Neither 45Ca2+ uptake nor dopamine release was altered under resting conditions. When the time course of K+‐stimulated 45Ca2+ uptake and dopamine release was examined, TPA (1.6 μM) enhanced dopamine release after 15, 30, and 60 s, but not 1, 3, or 5 s, of depolarization. A slight increase in 45Ca2+ uptake after 60 s of depolarization was also seen. The addition of 30 mM KCl to synaptosomes which had been preloaded with the Ca2+‐sensitive fluorophore fura‐2 increased the cytosolic free Ca2+ concentration ([Ca2+]i) from 445 nM to 506 nM after 10 s of depolarization and remained elevated after 60 s. TPA had no effect on [Ca2+]i under depolarizing or resting conditions. Replacing extracellular Ca2+ with 100 μM EGTA reduced K+‐stimulated (60 s) endogenous dopamine release by 53% and decreased [Ca2+]i to 120 nM. In Ca2+‐free medium, 30 mM KCl did not produce an increase in the [Ca2+]i. TPA (1.6 μM) did not alter the [Ca2+]i under resting or depolarizing conditions, but did increase K+‐stimulated dopamine release in Ca2+‐free medium. The inactive 4α‐phorbol had no effect under any of the above experimental conditions. The demonstration that TPA induced an increase in dopamine release without an increase in the [Ca2+]i may suggest that protein kinase C increases the sensitivity of the exocytotic release process to Ca2+.
Vascular Cholinesterases and Choline Uptake in Isolated Rat Forebrain Microvessels: A Possible Link
Authors: Shimon Mazal , Egozi Yakov , Kloog Yoel , Sokolovsky Mordechai , Cohen Sasson
Published: October 2006

Abstract: The two parameters of the active [methyl‐3H]choline uptake into isolated rat forebrain microvessels, Km and Vmax, were determined for 1‐, 3‐, l0‐, and 24‐month‐old Charles River male rats and compared with the activities of the enzymes choline acetyltransferase (ChAT), acetylcholinesterase (AChE), and butyrylcholinesterase (BuChE) in these microvessels over the same time course. The value of Km remained constant over the entire period, but that of Vmaxincreased from 8.5 ± 1.0 to 80.6 ± 16.4 nmol g−1 (mean ± SEM) over the first 3 months of life. Over the same period, the increase in ChAT activity, from an initial value of 7.1 ± 1.6 to 10.2 ± 0.3 nmol g−1 min−1, was not proportional to that of choline uptake. Levels of BuChE activity (0.9‐1.3 μmolg−1 min−1) were almost unchanged throughout the entire 24‐month period, but those of AChE showed a steady and significant increase from 1 to 24 months, remaining relatively high at senescence (4.7 μmolg−1 min−1), when choline uptake had decreased to one‐third of its optimal value. Selective inhibition of AChE with l,5‐bis(4‐allyldimethylammonium‐phenyl)pentan‐3‐one dibromide (0.5 μM) in unruptured capillaries from 3‐month‐old rats resulted in a decrease in Vmax of choline uptake from ∼81 to 59 nmol g−1 min−1 or with 9‐amino‐l,2,3,4‐tetrahydroacridine (10 μM) in capillaries from 2‐month‐old rats from ∼ 30 to 15 nmol g−1 min−1. Selective inhibition of BuChE with tetraisopropyl pyrophos‐phoramide (100 μM) resulted in an increase in Vmax from ∼81 to 96 nmol g−1 min−1. It is possible that the two vascular enzyme systems are coupled to a hypothetical endothelial choline transporter, but with an action opposite to each other.
Effect of Different Photoperiod Exposure on [3H]Imipramine Binding and Serotonin Uptake in the Rat Brain
Authors: Rovescalli Alessandra C. , Brunello Nicoletta , Riva Marco , Galimberti Roberto , Racagni Giorgio
Published: October 2006

Abstract: Seasonal rhythmicity in the occurrence of acute depressive episodes and the therapeutic efficacy of light exposure suggest the possible involvement of the pineal gland or other biological oscillators in the pathophysiology of depressive illness. We have performed studies to clarify whether different light/dark (LD) cycle schedules may induce changes in the biochemical targets of antidepressants in the rat CNS. In particular, we have investigated the effect of short‐ (LD 8: 16) or long‐day (LD 14:10) photoperiods on different biochemical parameters of serotonergic neurons. A significant increase in the density of [3H]imipramine ([3H]IMI) binding and in the Vmax of 5‐[3H]hydroxytryptamine (5‐[3H]HT) uptake was found in the hypothalamus of LD 8:16‐ with respect to LD 14:10‐exposed rats, whereas no difference was found in the kinetic properties of postsynaptic 5‐HT receptors and in 5‐HT metabolism in the hypothalami and cerebral cortices of rats exposed to the two different photoperiods. A seasonal rhythm of [3H]IMI binding sites and 5‐HT uptake seems to exist only in certain brain areas, such as the hypothalamus, because no differences were found in the cerebral cortex of LD 14:10‐ and LD 8:16‐accustomed rats. [3H]IMI binding and 5‐HT uptake were significantly increased in the hypothalamus of rats accustomed to a light/dark‐inverted cycle (DL 10:14) and killed 6 h after the stopping of lighting in comparison to rats exposed to normal LD 14:10 cycles and killed 6 h after the beginning of lighting. Therefore, a circadian modification of the serotonergic presynaptic sites seems to be present and related to light/dark exposure. Because the existence of endogenous compounds able to modulate [3H]IMI binding and 5‐HT uptake, other than 5‐HT, has been postulated in the mammalian brain, the involvement of these substances in the periodic changes observed could be suggested.
Effects of Vitamin A and Its Analogs on Nonenzymatic Lipid Peroxidation in Rat Brain Mitochondria
Authors: Das Nagaratnam Patmanatha
Published: October 2006

Abstract: Vitamin A (retinol) and some of its analogs exhibited varying degrees of inhibition on induced iron and ascorbic acid lipid peroxidation of rat brain mitochondria. Malonyldialdehyde production was used as an index of the extent of in vitro lipid peroxidation. The fat‐soluble vitamins retinol, retinol acetate, retinoic acid, retinol palmitate, and retinal at concentrations between 0.1 and 10.0 mmol/L inhibited brain lipid peroxidation. Retinol and retinol acetate were the most effective inhibitors. It is concluded from this study that retinol and its analogs can be considered as potential antioxidant factors, more potent than some of the well‐known antioxidants such as α‐tocopherol and butylated hydroxytoluene.
Adenosine Formation and Release by Embryonic Chick Neurons and Glia in Cell Culture
Authors: Meghji Parviz , Tuttle Jeremy B. , Rubio Rafael
Published: October 2006

Abstract: Adenosine formation and release were studied in 48‐h‐old cultured ciliary‐ ganglia and confluent peripheral and CNS glial cultures from embryonic chicks. Metabolic poisoning induced by 30 mM 2‐deoxygIucose and 2 μg/ml oli‐gomycin reduced ATP concentration by 90%. An increase in adenosine accounted for 15–40% of the fall in ATP. Dilazep (3 × 10−6M), a nucleoside transport inhibitor, decreased both incorporation of adenosine (an index of nucleoside transport) and release of adenosine by 80–90%. Dilazep trapped the newly formed adenosine intracellularly. A concentration of α,β‐methylene ADP that inhibited ecto‐5′‐nucleotidase by 80–90% did not alter the concentration of adenosine or AMP in neurons, glia, or medium. The results demonstrate that adenosine is formed intracellularly and exported out of the cell via the nucleoside transporter. The participation of ecto5′‐nucleotidase was excluded.
Expression of Glial Fibrillary Acidic Protein and Neurofilament mRNA in Gliosis Induced by Experimental Autoimmune Encephalomyelitis
Authors: Aquino Dennis A. , Shafit‐Zagardo Bridget , Brosnan Celia F. , Norton William T.
Published: October 2006

We have previously shown that the content of glial fibrillary acidic protein (GFAP) gradually increases in the spinal cord of Lewis rats with acute experimental autoimmune encephalomyelitis (EAE), reaching a level 1.5–2 times greater than that in controls by 35 days postimmunization (dpi). We report here that the increase in GFAP mRNA level followed a completely different time course and reached higher levels relative to controls than did that of the protein. Total RNA was isolated using a modified version of current methods using phenol/chloroform extractions to ensure optimal recovery from spinal cord. Control animals yielded 323 ± 35 μg (mean ± SD; n = 34) of total RNA/spinal cord throughout the experimental period. EAE animals contained up to three times as much total RNA during 11–14 dpi, a finding largely reflecting the infiltration of inflammatory cells. By 65 dpi, total RNA levels closely approached control values. As early as 10 dpi, increased amounts of GFAP mRNA were detected in EAE animals relative to controls. During 11–14 dpi, GFAP mRNA levels reached six‐ to eightfold greater than values in controls and then slowly declined throughout the remainder of the time course, with a fourfold increase still detected at 65 dpi. However, coinciding with the height of inflammation and clinical signs at 12 dpi, the GFAP mRNA content dropped to ∼50% of the level at 11 dpi but rose again at 13 dpi. This dip was mirrored by a similar decrease in neurofilament mRNA content, but otherwise the level of this message remained relatively constant and equal to that in controls. The changes in GFAP message content contrasted sharply with those in protein content, which increases slowly and does not reach maximal levels until after 4 weeks. Elevated levels of GFAP mRNA, protein, and immunostaining persisted through 65 dpi, >40 days after clinical signs had resolved.
Soil Microbiology as a Field Of Science
Authors: Waksman Selman A.
Published: October 2006

It can now be definitely recognized that the soil microbiologist is in a position to make important contributions not only to our knowledge of soil processes and plant growth, but also to microbiology, especially microbial physiology, and to the utilization of microorganisms for various industrial, public health and other processes. The soil microbiologist is able to contribute in many ways to man's capacity to survive, by learning to control the activities of injurious microorganisms and by favoring the processes brought about by the beneficial organisms.

The reason why the broader concept "microbiology" rather than the narrower term "bacteriology" has been used throughout this discussion is that the soil microbiologist has often to pay as much attention to the fungi as to the bacteria, and occasionally also to the protozoa, the algae and even the nematodes and other worms, inhabiting the soil in large numbers. Only a recognition of all these lower forms of life and their many interrelationships can help to elucidate this complex and important science.
Prolongation of Islet Allograft Survival Following in Vitro Culture (24°C) and a Single Injection of ALS
Authors: Lacy Paul E. , Davie Joseph M. , Finke Edward H.
Published: October 2006

Isolated rat islets remain morphologically and functionally intact during a 7-day period of in vitro culture at 24°C. In vitro culture of islets at 24°C for 7 days prior to transplantation, in conjunction with a single injection of antiserum to lymphocytes into the diabetic recipient, results in islet allograft survival of 100 days when the islets are transplanted across a major histocompatibility barrier.
Vesicular Binesis: Glucose Effect on Insulin Secretory Vesicles
Authors: Gabbay Kenneth H. , Korff J. , Schneeberger Eveline E.
Published: October 2006

Binesis is a process whereby the membrane of the insulin secretory vesicle in the beta cell forms a lingula that indents the vesicular membrane of an adjoining secretory vesicle or the plasma membrane. Vesicular binesis in beta cells increases when islets of Langerhans are incubated at a stimulatory glucose concentration (300 milligrams per 100 milliliters). These vesicular membrane alterations may be the morphological concomitants of activation of the insulin secretory vesicle, and indicate an active role for the vesicle and its membrane in the release mechanism.
Sleep During Transcendental Meditation
Authors: Pagano Robert R. , Rose Richard M. , Stivers Robert M. , Warrenburg Stephen
Published: October 2006

Five experienced practitioners of transcendental meditation spent appreciable parts of meditation sessions in sleep stages 2, 3, and 4. Time spent in each sleep stage varied both between sessions for a given subject and between subjects. In addition, we compare electroencephalogram records made during meditation with those made during naps taken at the same time of day. The range of states observed during meditation does not support the view that meditation produces a single, unique state of consciousness.
Chromatin Fragments Resembling ν Bodies
Authors: Senior Marilyn B. , Olins Ada L. , Olins Donald E.
Published: October 2006

Fragments of chromatin resembling the spheroid chromatin units (ν bodies) have been isolated from formaldehyde-fixed and sonicated chicken erythrocyte nuclei. Ultracentrifugal analyses demonstrated that monomer ν bodies have a molecular weight of about 300,000 per particle, exhibit a protein to DNA ratio (by weight) of 1.22:1, and contain a DNA fragment with a molecular weight of approximately 140,000 per ν body.
2,3-Diphosphoglycerate: A Physiological Inhibitor of Platelet Aggregation
Authors: Iatridis Sotirios G. , Iatridis Panayotis G. , Markidou Sofia G. , Ragatz Barth H.
Published: October 2006

2,3-Diphosphoglycerate (2,3-DPG) may inhibit the platelet release reaction and the irreversible aggregation of human blood platelets induced by adenosine diphosphate, epinephrine, or norepinephrine. The effects of 2,3-DPG on platelet aggregation were more pronounced in cases with low hematocrit (< 30 percent). Dipyridamole and vincaminor potentiated the antiaggregating effect of 2,3-DPG. Erythrocytes (10
3
to 10
4
per microliter) exhibited a similar antiaggregating effect, especially when secured from anemic patients.
Auditory Discrimination and Recall in Monkeys
Authors: Dewson James H. , Burlingame Allen C.
Published: October 2006

The first auditory recall functions have been derived for monkeys; the same animals also demonstrate perceptual abilities closely approximating those of man. An efficient, powerful psychophysical technique is used to specify and force predicted levels of difficulty of task performance across time, animals, and problems.
Contradiction of Auditory and Visual Information by Brain Stimulation
Authors: Kleinman D. , John E. R.
Published: October 2006

Cats with permanently implanted electrodes were trained to discriminate between trains of flashes or clicks at two different repetition frequencies. After substantial overtraining with these sensory stimuli, high levels of stimulus generalization were obtained to electrical stimulation of the reticular formation. Subsequent stimulation of the reticular formation at either frequency simultaneously with contradictory flicker or click stimulation at the opposite frequency resulted in control of the behavior by the reticular stimulus. Lateral geniculate stimulation failed to show this effect.