bluuebunny/crossref_metadata_2025_split huggingface dataset:
Development of phenylethanolamine N‐methyltransferase (PNMT) in cultures of dissociated embryonic rat medulla oblongata
Authors: Bohn Martha Churchill , Engele Jürgen 
Published: April 2003

AbstractThe adrenergic phenotypic marker, phenylethanolamine N‐methyltransferase (PNMT) is expressed in a subgroup of catecholaminergic neurons in the brain, as well as in the chromaffin cells of the adrenal medulla. Although PNMT in the rat adrenal is regulated by glucocorticoids, PNMT in the rat brainstem appears not to be regulated by glucocorticoids. Furthermore, little is known about factors required for the differentiation of this specific class of central neuron. The identification of such factors has been hampered not only by the heterogeneity of cell types in the brainstem, of which only a smaller number express PNMT, but also by the lack of a well characterized in vitro system in which the development of these neurons can be studied under defined conditions. The present study addresses this issue by establishing and characterizing a culture system for the study of adrenergic neurons.Dissociated cultures were prepared from embryonic rat medulla oblongata and the expression and development of PNMT was studied using immunocytochemistry and radioisotopic assay of PNMT activity. The survival of PNMT‐immunoreactive (IR) neurons in vitro was found to be critically dependent on embryonic age. Numerous PNMT‐IR neurons were observed in cultures prepared only from embryos of 46–51 somites (embryonic day E13–13.5). In contrast, cultures containing numerous neurons immunoreactive for tyrosine hydroxylase (TH), the rate‐limiting enzyme in catecholamine biosynthesis, could be successfully established from medulla oblongata of any age between E13 and E16. In cultures of the E13 rat, PNMT was found to be catalytically active at 4 days in vitro and the levels of PNMT activity per neuron, as estimated from the number of PNMT‐IR neurons in sister cultures, increased 2‐fold in cultures grown for 8 days in the presence of 10% fetal bovine serum and 1.5‐fold in defined, serum‐free conditions. Treatment of cultures with either corticosterone or dexamethasone had no effect on PNMT activity, supporting previous studies suggesting that PNMT in central neurons is not regulated by glucocorticoids.These studies provide a convenient and defined in vitro system for studying factors that influence the development of central adrenergic neurons. The differential development of PNMT‐IR and TH‐IR neurons in these cultures suggests that the factors involved in adrenergic neuron survival and/or differentiation are not common to catecholaminergic neurons in general.
Neuronal sprouting and synapse formation in response to injury in the mouse organ of corti in culture
Authors: Sobkowicz Hanna M. , Slapnick Susan M. 
Published: April 2003

AbstractThe effect of mechanical injury on induction of regenerative phenomena within the neurosensory epithelium was investigated in cultures of neonatal mouse cochlea. The oldest examined culture in which new neuronal growth followed insult, was injured at 13 days in vitro and fixed 24 h later. By far, the most vigorous regenerative reaction was observed in a 3‐day culture 4 h post‐injury. The reaction included sprouting of nerve fibers injured directly, synapse formation between the surviving hair cells and sprouting neuronal growth cones, wrapping of growing nerve fibers by extending processes of hair cell cytoplasm, and collateral sprouting of synaptically‐engaged nerve endings and of nerve fibers in passage.
Effects of acetyl‐l‐carnitine on the survival of adult rat sensory neurons in primary cultures
Authors: Formenti A. , Arrigoni E. , Sansone V. , Martelli E. Arrigoni , Mancia M. 
Published: April 2003

AbstractAcetyl‐l‐carnitine produces a significant increase in the survival time‐course of adult rat sensory neurons maintained in primary cultures up to 40 days. The analysis of our data suggests that 200 μm acetyl‐l‐carnitine added to the medium, slows down neuronal decay especially in the first 10 days in vitro, sparing a fraction of cells which would otherwise be lost.Patch‐clamp recordings from these neurons show that superfusion with acetyl‐l‐carnitine (100–1000 μM) does not induce any membrane current. In addition an agonist muscarinic effect particularly concerning high‐voltage activated calcium channel modulation appears to be ruled out.In conclusion our data favour the role of acetyl‐l‐carnitine in the trophism of sensory neurons in adult rats. In agreement with other in vivo experiments our data reinforce the hypothesis that this substance might be involved in reducing neuronal loss observed in nervous system aging.
Hippocampal cholinergic alterations and related behavioral deficits after early exposure to ethanol
Authors: Pick Chaim G. , Cooperman Michael , Trombka David , Rogel‐Fuchs Yael , Yanai Joseph 
Published: April 2003

AbstractThe present study was designed to ascertain septohippocampal cholinergic alterations and their related behavioral deficits after early exposure to ethanol. Mouse pups were exposed to ethanol, 3 g/kg by daily subcutaneous injection on postnatal days 2–14. At age 50 days, the ethanol‐exposed mice had significant reductions from control levels in eight‐arm maze performance. For example, on the fourth testing day, the number of correct entries in the ethanol group was 21% below control levels (P < 0.05) and the number of trials needed to enter all arms was 48% above control (P < 0.001). It took the ethanol‐exposed mice twice the time to reach criterion than it did control (P < 0.01). A 33% increase from control level in muscarinic receptor number (βmax) was found in the treated mice of age 22 days and a 64% increase at age 50 days (P < 0.001). However, no differences between control and treated groups could be detected in the presynaptic component of the cholinergic innervation, choline acetyltransferase activity. The results suggest that early ethanol exposure acts on hippocampal function similarly to phenobarbital, probably via alterations in postsynaptic processes in the septohippocampal cholinergic pathways.
Culture of rat brain preoptic area neurons: Effects of sex steroids
Authors: Takagi Keiichi , Kawashima Seiichiro 
Published: April 2003

AbstractThe aim of the present study was to investigate the direct effects of testosterone (T), estradiol‐17α (E2) and 5α‐dihydrotestosterone (5α‐DHT) on the survival of neurons and the morphology of neuronal processes (the total process length, the number of processes and the number of process branchings) of the cells derived from neonatal rat preoptic area (POA) to analyze the bases of morphological sexual dimorphism of the POA. The POA cells were exposed to sex steroids during 2–14 days of culture in serum‐free medium. Neuronal survival was significantly greater in cultures treated with 1.0 nM T than controls. Although T at any concentration tested (0.1,1.0 or 10.0 nM) failed to affect the mean value of the process length, a significant difference was observed in the frequency distribution of total process length between 1.0 nM T‐treated cultures and control cultures. By 1.0 nM T, the number of process branchings was also significantly increased. In contrast, the number of neurons and the total process length of neurons were not affected by 5α‐DHT. However, the number of process branchings was significantly increased by 0.1 nM 5α‐DHT. E2 had no significant effect on any of these parameters. These results suggest that T directly enhances the survival of neurons and increases the dendritic and/or axonic field of the POA, which serves, at least partly, for the development of morphological sexual dimorphism in the rat.
Functional PAF receptors in glia cells: Binding parameters and regulation of expression
Authors: Brodie Chaya 
Published: April 2003

AbstractPlatelet activating factor is a unique phosphoglycerine which possesses a variety of biological functions exerting its biological effects via specific surface receptors. In the central nervous system, platelet activating factor has been suggested to play a role during injury especially in conditions of ischemia and trauma‐induced neuronal damage. The specific cell populations expressing platelet activating factor receptor, however, have not been identified. In this study, the binding properties of platelet activating factor receptors in C6 glioma cells and primary cultures of astrocytes and oligodendrocytes were characterized by using the ligand [3H]WEB 2086. Early‐passage glial cells which exhibit oligodendrocytic phenotype, expressed lower levels of [3H]WEB 2086 binding than either late‐passage cells which exhibit astrocytic phenotypes or primary astroglia cells. No specific binding was observed in primary cultures of oligodendrocytes. The Bmax (136 ± 15.3 fmol/mg protein) and Kd (29 ± 3.2 nM) levels obtained for primary astroglia cells were similar to those described for other cell types. The expression of platelet activating factor receptor in early‐passage glia cells was up‐regulated by treatment with insulin which induces astrocytic differentiation. In contrast, db‐cyclic AMP exerted an inhibitory effect on the level of platelet activating factor receptor in both early‐ and late‐passage cells. The level of functional platelet activating factor receptor in C6 cells as measured by the ability of platelet activating factor to induce 45Ca2+ influx was increased in cells expressing astrocytic phenotypes and was decreased in db‐cyclic AMP‐treated cells. In accordance with lack of specific [3H]WEB 2086 binding, platelet activating factor did not induce a detectable response of Ca2+ influx in cultures of oligodendrocytes. This report provides the first direct demonstration of selective expression of functional platelet activating factor receptors and their properties in astroglia cells. The findings support the suggestion that platelet activating factor may play an important role as a mediator of injury and immune responses in the nervous system.
The critical period for ethanol effects on cholinergic neuronal expression in neuroblast‐enriched cultures derived from 3‐day‐old chick embryo: NGF ameliorates the cholinotoxic effects of ethanol
Authors: Rahman Hafeez , Kentroti Susan , Vernadakis Antonia 
Published: April 2003

AbstractStudies from our laboratory have established that ethanol exerts morphological and biochemical neurotoxic effects during early neuroembryogenesis in the chick brain both in ovo and in culture. In the present study, we further localized the critical period for ethanol effects on cholinergic neuronal expression using neuroblast‐enriched cultures derived from 3‐day‐old chick embryos. Moreover, we report that NGF attenuated the cholinotoxic effects of ethanol. We used the following experimental paradigms: cultures treated with ethanol alone either C0‐C3 or C4‐C10; NGF alone CO‐C4 or C4‐C10; ethanol and NGF given concomitantly; ethanol given first then replaced with NGF in the medium; or NGF given first then replaced with ethanol in the medium. The results revealed: (1) the cholinotoxic effect of ethanol occurs between culture days CO and C4 with day 3 appearing to be most critical. (2) similarly, the critical period for the cholinotoxic effects of NGF is during early neuroblast differentiation, culture days CO‐C4. (3) NGF can prevent the cholinotoxic effects of ethanol only if both ethanol and NGF are given concomitantly or if ethanol is given first, then culture is replaced with NGF‐containing medium.
Pattern of polyamines and related monoacetyl derivatives in chick embryo retina during development
Authors: Taibi G. , Schiavo M.R. , Calvaruso G. , Tesoriere G. 
Published: April 2003

AbstractPolyamines and related monoacetyl derivatives were studied in chick embryo retina during development (6th–19th day). Putrescine, which is high in the first phase of retinogenesis, is necessary to sustain both tissue proliferation and via N‐acetylputrescine, γ‐aminobutyric acid synthesis. A later increase in spermidine and particularly spermine may play a role in the last phase of development when the retina reaches maturation. The presence of N‐acetylspermidine already at the 8th day indicates that in chick embryo retina, putrescine synthesis can depend on two separate pathways. The first involves ornithine decarboxylase activity; the second, spermidine/spermine N1‐acetlytransferase and probably polyamine oxidase that converts spermidine to putrescine via N1‐acetylspermidine.
Development of glutamate‐stimulated phosphatidylinositol metabolism in primary neuronal and astrocyte cultures
Authors: DeCoster Mark A. , Yourick Debra L. 
Published: April 2003

AbstractIt was the purpose of the present study to evaluate glutamate‐stimulated phosphatidylinositol metabolism in primary mixed astrocyte/neuron and neuron‐enriched cortical cultures through different stages of development in vitro. Glutamate (0–200 μM) stimulated inositol phosphate accumulation in a concentration‐dependent fashion at 6, 13 and 20 days in vitro. Pure astrocyte cultures exhibited glutamate‐stimulated phosphatidylinositol hydrolysis only at high concentrations (100–400 μM), indicating that these cells contribute little to the overall inositol phosphate accumulation measured in mixed neuronal cultures treated with low glutamate concentrations. Comparison of mixed neuronal cultures with and without antimitotic treatment revealed that increasing astrocyte number suppressed glutamate‐stimulated responses, presumably via glutamate uptake. In contrast to previous reports, glutamate‐stimulated inositol phosphate accumulation, when expressed as a function of cell number, increased with increasing days in vitro.
Plasticity of autonomic nerves: Differential effects of long‐term guanethidine sympathectomy on the sensory innervation of the rat uterus during maturation
Authors: Brauer M.M. , Lincoln J. , Milner P. , Sarner S. , Blundell D. , Passaro M. , Corbacho A. , Burnstock G. 
Published: April 2003

AbstractThe sensory nerves, containing substance P and calcitonin gene‐related peptide, and noradrenaline‐containing sympathetic nerves of the rat uterus were analyzed following long‐term sympathectomy with guanethidine in prepubertal (four weeks), young adult (eight weeks) and fully adult animals (18 weeks). Immunohistochemical and histochemical methods were used in association with nerve density measurements and biochemical assays. The main findings were as follows: (1) long‐term guanethidine treatment completely abolished the noradrenergic innervation of the uterine horn and parametrial tissue and markedly reduced the tissue levels of noradrenaline in both regions at the three ages analysed; (2) in the uterine hom guanethidine treatment had no effect on the tissue levels of either calcitonin gene‐related peptide or substance P or on the density of calcitonin gene‐related peptide‐containing nerves, at any of the three ages studied; (3) in the parametrial tissue increased levels of calcitonin gene‐related peptide were observed at 8 and 18 weeks of age, together with a significant increase in the density of calcitonin gene‐related peptide‐containing nerves. Substance P levels showed a transient increase in this tissue at eight weeks. In conclusion, long‐term sympathectomy with guanethidine resulted in an increase in calcitonin gene‐related peptide and substance P in sensory nerves in the parametrial tissue, but not in the uterine horn. The changes in the parametrial tissue only occurred after puberty. It is suggested that sensory nerves in the uterine horn may be less responsive to sympathetic denervation since loss of sympathetic nerves occurs as part of a normal physiological process during pregnancy in this region.
Major increase in brain natriuretic peptide indicates right ventricular systolic dysfunction in patients with heart failure
Authors: Mariano‐Goulart Denis , Eberlé Marie‐Claude , Boudousq Vincent , Hejazi‐Moughari Azadeh , Piot Christophe , de Kerleau Charles Caderas , Verdier Régis , Barge Marie‐Luce , Comte Frédéric , Bressot Nicole , Rossi Michel , Kotzki Pierre‐Olivier 
Published: April 2003

AbstractThis study sought to investigate whether the presence of right ventricular systolic dysfunction with pre‐existing left ventricular systolic dysfunction is associated with higher plasma brain natriuretic peptide (BNP) levels, compared with patients with isolated left ventricular dysfunction. Eighty‐five patients referred for evaluation of isotopic ventricular function were prospectively included in the study. Left (LVEF) and right (RVEF) ventricular ejection fractions were evaluated by gated blood pool scintigraphy and compared with plasma BNP levels. BNP correlated negatively with LVEF, except in patients with ischaemic heart disease (P=0.09) and in patients with LVEF<40% (P=0.11). In contrast, BNP levels correlated negatively with RVEF for all subgroups. Among patients with RVEF<40%, no significant BNP difference was found between patients with or without additional left ventricular systolic dysfunction (P=0.51). Among patients with LVEF<40%, plasma BNP levels were significantly higher in patients with RVEF<40% than in patients with RVEF≥40% (P=0.004) whereas age, renal function, clinical findings, ventricular volumes, LVEF or medication were not significantly different. In conclusion, an important increase in BNP levels in patients with left ventricular systolic dysfunction should be considered by cardiologists as an indication of high risk of right ventricular dysfunction and should justify further investigation.
Improved amino acid flexibility parameters
Authors: Smith David K. , Radivojac Predrag , Obradovic Zoran , Dunker A. Keith , Zhu Guang 
Published: April 2003

AbstractProtein molecules exhibit varying degrees of flexibility throughout their three‐dimensional structures, with some segments showing little mobility while others may be so disordered as to be unresolvable by techniques such as X‐ray crystallography. Atomic displacement parameters, or B‐factors, from X‐ray crystallographic studies give an experimentally determined indication of the degree of mobility in a protein structure. To provide better estimators of amino acid flexibility, we have examined B‐factors from a large set of high‐resolution crystal structures. Because of the differences among structures, it is necessary to normalize the B‐factors. However, many proteins have segments of unusually high mobility, which must be accounted for before normalization can be performed. Accordingly, a median‐based method from quality control studies was used to identify outliers. After removal of outliers from, and normalization of, each protein chain, the B‐factors were collected for each amino acid in the set. It was found that the distribution of normalized B‐factors followed a Gumbel, or extreme value distribution, and the location parameter, or mode, of this distribution was used as an estimator of flexibility for the amino acid. These new parameters have a higher correlation with experimentally determined B‐factors than parameters from earlier methods.
Peptidase family U34 belongs to the superfamily of N‐terminal nucleophile hydrolases
Authors: Pei Jimin , Grishin Nick V. 
Published: April 2003

AbstractPeptidase family U34 consists of enzymes with unclear catalytic mechanism, for instance, dipeptidase A from Lactobacillus helveticus. Using extensive sequence similarity searches, we infer that U34 family members are homologous to penicillin V acylases (PVA) and thus potentially adopt the N‐terminal nucleophile (Ntn) hydrolase fold. Comparative sequence and structural analysis reveals a cysteine as the catalytic nucleophile as well as other conserved residues important for catalysis. The PVA/U34 family is variable in sequence and exhibits great diversity in substrate specificity, to include enzymes such as choloyglycine hydrolases, acid ceramidases, isopenicillin N acyltransferases, and a subgroup of eukaryotic proteins with unclear function.
Crystal structures of a pantothenate synthetase from M. tuberculosis and its complexes with substrates and a reaction intermediate
Authors: Wang Shuishu , Eisenberg David 
Published: April 2003

AbstractPantothenate biosynthesis is essential for the virulence of Mycobacterium tuberculosis, and this pathway thus presents potential drug targets against tuberculosis. We determined the crystal structure of pantothenate synthetase (PS) from M. tuberculosis, and its complexes with AMPCPP, pantoate, and a reaction intermediate, pantoyl adenylate, with resolutions from 1.6 to 2 Å. PS catalyzes the ATP‐dependent condensation of pantoate and β‐alanine to form pantothenate. Its structure reveals a dimer, and each subunit has two domains with tight association between domains. The active‐site cavity is on the N‐terminal domain, partially covered by the C‐terminal domain. One wall of the active site cavity is flexible, which allows the bulky AMPCPP to diffuse into the active site to nearly full occupancy when crystals are soaked in solutions containing AMPCPP. Crystal structures of the complexes with AMPCPP and pantoate indicate that the enzyme binds ATP and pantoate tightly in the active site, and brings the carboxyl oxygen of pantoate near the α‐phosphorus atom of ATP for an in‐line nucleophilic attack. When crystals were soaked with, or grown in the presence of, both ATP and pantoate, a reaction intermediate, pantoyl adenylate, is found in the active site. The flexible wall of the active site cavity becomes ordered when the intermediate is in the active site, thus protecting it from being hydrolyzed. Binding of β‐alanine can occur only after pantoyl adenylate is formed inside the active site cavity. The tight binding of the intermediate pantoyl adenylate suggests that nonreactive analogs of pantoyl adenylate may be inhibitors of the PS enzyme with high affinity and specificity.
Neonatal Lymphocytes Dominate against Lymphocytes of Their Own Mother but Not against Allogenic Maternal or Adult Lymphocytes in Bidirectional Mixed Lymphocyte Cultures
Authors: Brune Thomas , Beier Kirsten , Exeler Rita , Harms Erik , Louwen Frank 
Published: April 2003

<i>Objective:</i> Recent studies have shown a regular prenatal transfer of maternal immunocompetent cells into the fetal circulation. However, these cells engraft and proliferate only in a few exceptional cases if the fetus reaches an immunocompetent state. Thus the fetus has to have an immunologic defense mechanism against the engraftment of maternal cells. In the current study we investigated whether the fetus has such an immune defense and whether this defense mechanism specifically attacks cells of the mother. <i>Patients, Material and Methods:</i> Blood samples were obtained from 15 mothers and 15 newborns directly after delivery. We compared individual vitality and spontaneous cytotoxicity between fetal and maternal lymphocytes in a cell ratio of 1:1 in nonstimulated bidirectional mixed lymphocyte cultures (MLC). The distribution of each cell population within the MLC was visualized by fluorescence in situ hybridization and X/Y-DNA probes. This was compared to MLCs between unrelated fetal and maternal as well as between unrelated adult lymphocytes. <i>Results:</i> After 72 h, a significant cell shift was observed only in the MLC with neonatal lymphocytes mixed with cells of their own mother; there was a significantly higher number of neonatal cells (0.71 vs. 0.29) present. All other groups continued to have a cell distribution of 1:1. <i>Conclusion:</i> Our results show that neonatal lymphocytes specifically dominate against maternal but not allogenous maternal or adult lymphocytes in nonstimulated bidirectional MLCs.