bluuebunny/crossref_metadata_2025_split huggingface dataset:
Phorbol ester‐induced change in astrocyte morphology: Correlation with protein kinase C activation and protein phosphorylation
Authors: Harrison B. C. , Mobley P. L. 
Published: January 2005

AbstractTreatment with 300 nM phorbol 12‐myristate 13‐acetate (PMA) transforms polygonal‐shaped cultured astrocytes into process‐bearing cells and produces a shift in protein kinase C (PK‐C) from the cytosol to the membrane. Exposure to PMA also produces increases in the phosphorylation of several proteins including vimentin, glial fibrillary acidic protein (GFAP), an acidic 80,000 molecular weight protein, and two 30,000 molecular weight proteins (pI5.5 and 5.7). The effects of PMA on the translocation of PK‐C and on protein phosphorylation precede the PMA‐induced changes in astrocyte morphology, and a close correlation exists between the concentration of PMA necessary to elicit half‐maximal and maximal effects on the shift of PK‐C to the membrane and on protein phosphorylation. In addition, the PMA‐induced alterations in cell morphology are not permanent, and within 24 hr after PMA treatment the cells have reverted almost to their original morphology. A second exposure to PMA at this time fails to elicit further change in cell shape and is also incapable of producing increases in the phosphorylation of proteins. It was determined that there is little, if any, PK‐C present in these PMA‐pretreated cells. The morphological responsiveness to PMA gradually returns in 5 to 8 days after the initial treatment with PMA, and this is accompanied by the recovery of PK‐C activity and the phosphorylation response. Therefore, these studies suggest that the effect of PMA on astrocyte morphology is mediated by the activation of PK‐C and subsequent protein phosphorylation.
Interspecies differences in the substance P‐ and vasoactive intestinal polypeptide‐like immunoreactivities in the olfactory bulb ofSalmo gairdneri and Barbus meridionalis
Authors: Alonso J. R. , Covenas R. , Lara J. , De Leon M. , Arevalo R. , Aijon J. 
Published: January 2005

AbstractThe distribution of substance P‐like (SP) and vasoactive intestinal polypeptide‐like (VIP) structures was studied using an indirect immunoperoxidase technique in the mediterranean barbel Barbus meridionalis and in the rainbow trout Salmo gairdneri. SP‐like positive fibers were observed in the inner strata of the mediterranean barbel olfactory bulb, mainly the granule cell and the plexiform layers. Ganglion cells and fibers of the terminal nerve were also labeled. No SP‐positive structure was found in the olfactory bulb of the rainbow trout. On the contrary, the VIP antiserum used displayed very strong immunostaining in the olfactory nerve fiber layer of Salmo gairdneri, whereas those fibers in Barbus meridionalis showed no immunoreactivity. After a complete transection of the olfactory tract and a survival time of 20 days, SP‐immunostained fibers were not observed. Thus, they can presumably be identified as centrifugal fibers, coursing from the telencephalic hemispheres through the olfactory tracts, into the olfactory bulb. The VIP immunoreactivity was confined to the olfactory fibers, both in the olfactory nerve and the olfactory bulb. The positive immunostaining disappeared after chemical lesion of the olfactory mucosa. These observations demonstrate that the olfactory bulb of freshwater teleosts exhibits a high degree of heterogeneity in its immunocytochemical distribution pattern, this pattern also differing from previous reports on higher vertebrates.
Asymmetry of interhemispheric responses evoked in the prefrontal cortex of the rat
Authors: Pérez H. , Ruiz S. , Ernández A. H. , Soto‐Moyano R. 
Published: January 2005

AbstractInterhemispheric responses of anesthetized rats were evoked in the prefrontal cortex of both cerebral hemispheres by electric stimulation of the homologous contralateral cortical region. Fatigability of interhemispheric responses recorded in the right hemisphere exhibited greater amplitude increments to double‐shock stimulation. No significant differences in rheobase, chronaxie, and threshold current were observed when comparing responsiveness of both prefrontal cortices to single stimuli. Results suggest that brain lateralization in the rat could depend, in part, on synaptic functional asymmetries of the prefrontal cortex.
Characterization of the distinctive neurofilament subunits of the soma and axon initial segments in the squid stellate ganglion
Authors: Tytell M. , Pant H. C. , Gainer H. , Hill W. D. 
Published: January 2005

AbstractThe stellate ganglion, which gives rise to the giant axons of the squid, was dissected into two parts, one containing primarily cell bodies and the other axon initial segments. A neurofilament protein‐enriched extract of each was prepared and compared biochemically and immunochemically with an axoplasmic neurofilament preparation and with the glial sheath that surrounds the axons. Both parts of the ganglion lacked the 220 kDa subunit of axoplasmic neurofilaments (NFs). However, they did contain a protein of about 190 kDa that reacted with the Pruss anti‐inter‐mediate filament antibody (aIFA; Pruss et al.: Cell 27:419–428, 1981), but not with a phosphorylationdependent NF antibody (Cohen et al.: I Neurosci 7: 2056–2074, 1987). Dephosphorylation of the axoplasmic NF 220 Vielded a product that comigrated on twodimensional (2D) gel electrophoresis with the 190 kDa ganglion protein, suggesting that the latter represented the incompletely phosphorylated precursor of NF220. The major low molecular weight aIFA‐reactive species in the ganglion preparation was a polypeptide of about 65kDa. A relatively small quantity of that polypeptide was also found in axoplasm and it comigrated in 2D gels with an aIFA‐reactive polypeptide from the glial sheath. These results indicate that the site of modification of the 190 kDa NF precursor to the 220 kDa axonal form is probably at the point where the axon initial segments leave the ganglion, which is several mm distal to its site of synthesis in the cell body. Furthermore, the filament network of the axoplasm and possibly the cell bodies includes a gliallike intermediate filament protein in addition to the NF protein subunits.
Stimulation of adrenergic development in neural crest cultures by a reconstituted basement membrane‐like matrix is inhibited by agents that elevate cAMP
Authors: Maxwell G. D. , Forbes M. E. 
Published: January 2005

AbstractPrevious work (Maxwell and Forbes: Development 101:767–776, 1987) has shown that an overlay of reconstituted basement membrane‐like (RBM) gel dramatically increased the number of catecholaminepositive (CA+) cells which differentiated in neural crest cultures. We report here that this increase was inhibited when cultures were grown for 7 days in the presence of agents that elevate cAMP, such as 8‐bromo‐cAMP and 3‐isobutyl‐1‐methylxanthine. The action of 8‐bromo‐cAMP was dose dependent with a half‐maximal effect at about 50 μM. The development of CA+ cells was dramatically reduced when 8‐bromo‐cAMP was present from days 0–4 in vitro, but was relatively unaffected if 8‐bromo‐cAMP was present from days 4–7 in vitro. The development of tyrosine hydroxylase immunoreactive cells was also inhibited by 8‐bromo‐cAMP. The addition of 8‐bromo‐cAMP increased the number of melanocytes and resulted in either no change or only modest reductions in the number of E/C8 and neurofilament immunoreactive cells, indiciting that the effect on CA+ cell ontogeny was selective. In contrast to the effectof 8‐bromo‐cAMP, addition of 8‐bromo‐cGMP did not inhibit CA+ cell development in the presence of the RBM gel nor did it stimulate CA+ cell development in the absence of the RBM gel overlay. Our results suggest that cAMP may be an important regulator of phenotypic expression in at least some neural crest cell lineages. In particular, these observations provide evidence that agents that elevate cAMP levels can act to prevent an essential part of the normal cellular response leading to the normal cellular response leading to the appearance of CA+ phenotype in neural crest cultures.
Neurite outgrowth promoting activity of G4 and its inhibition by monoclonal antibodies
Authors: Chang S. , Rathjen F. G. , Raper J. A. 
Published: January 2005

AbstractThe chick G4 molecule is a cell surface glycoprotein which is a member of the L1/NILE/NgCAM/8D9 group of neural cell adhesion molecules. Polyclonal antisera against G4 have been shown to decrease sympathetic neurite outgrowth on sympathetic axons and to de‐bundle retinal axons growing on a tectal membrane substrate. We have extended the specificity of these results by showing that a panel of monoclonal antibodies against G4 is also effective in reducing sympathetic neurite outgrowth on sympathetic axons. Furthermore, purified G4 adsorbed onto an inert substrate promotes extensive neurite outgrowth. Monoclonal antibodies to G4 completely inhibit the activity of purified G4. These data show that G4 is a cell surface neurite outgrowth promoting molecule, a function which was first suggested by antibody perturbation experiments and now is confirmed directly.
Isolation, culture, and characterization of adult rat oligodendrocytes
Authors: Vick R. S. , Chen S.‐J. , DeVries G. H. 
Published: January 2005

AbstractInvestigation into CNS demyelinating diseases, which usually occur in adults, can be facilitated by the use of a good in vitro model. We have established a methodology whereby oligodendrocytes from adult rat CNS can be cultured in vitro, and we have characterized these cultures morphologically and immunologically. Approximately 1 g of spinal cord and brainstem per adult rat was removed and dissociated mechanically and enzymatically. After filtration of the white matter homogenate, myelin was removed by 0.9 M sucrose density centrifugation. The cells were further purified by centrifugation through a 0.3%/4% discontinuous gradient of bovine serum albumin (BSAZ). The pellet was resuspended and placed in an untreated 6‐well culture dish overnight to allowthe astrocytes to attach. The non‐adherent cells were replated on poly‐l‐lysine‐treated coverslips. Approximately 8.25 × 105 cells were recovered per animal. The adult oligodendrocytes initially appeared as rounded cell bodies, but after 2–5 days in vitro (DIV), the oligodendrocytes extended 6–10 thick processes. A membrance sheath between these processes was immunostainable with either anti‐galactocerebroside (GC), anti‐04, anti‐myelin basic protein (MBP), or anti‐2′3′ cyclic nucleotide 3′ phosphohydrolase (CNPase) and was also evident in scanning EM. Older cultures (up nto 60 DIV) maintained whorls of myelin and transmission EM revealed a major dense line distance of approximately 103 AS with up to 11 concentric layers of membrance. Immunologically, the adult oligodendrocytes are GC+, 04+, MBP+, CNPase+, and GFAP−. The method described will allow adult rat oligodendrocytes to be isolated and maintained in culture; these cultures retain the characteristics of differentiated adult oligodendrocytes.
Method for isolation of kappa‐opioid binding sites by dynorphin affinity chromatography
Authors: Simon J. , Benyhe S. , Hepp J. , Varga E. , Medzihradszky K. , Borsodi A. , Wollemann M. 
Published: January 2005

AbstractA kappa‐opioid receptor subtype was purified from a digitonin extract of frog brain membranes, using affinity chromatography. The affinity resin was prepared by coupling dynorphin (1–10) to AH Sepharose 4B. The purified receptor binds 4,750 pmol [3H]ethylketocyclazocine (EKC) per mg protein (5,600‐fold purification over the membrane‐bound receptor) with a Kd of 9.1 nM. The addition of cholesterol‐phosphatidylethanolamine (2:1) enhanced 3.6‐fold the binding activity of the purified material, which gives a purification very close to the theoretical. The purified receptor protein exhibits high affinity for kappa‐selective ligands. The purified fraction shows one major band (65,000 Mr) in sodium dodecyl sulfate (SDS) gel electrophoresis.
Conditioned medium from activated splenocytes increases substance P in sympathetic ganglia
Authors: Jonakait G. Miller , Schotland S. 
Published: January 2005

AbstractFollowing removal of the presynaptic input to the superior cervical ganglion (SCG) of the neonatal rat, there is an increase in substance P (Kessler et al.:Science 214:335–336, 1981; Kessler and Black: Brain Res 234:182–187, 1982) and the mRNA coding for its prohormone precursor (Roach et al.: Proc Natl Acad Sci USA 84:5078–5081, 1987). However, the functional significance of this increase has been unclear. We report here that SP increases dramatically in cultures of SCG grown in the presence of conditioned medium from con‐A‐stimulated splenocytes. The effect is mimicked by growing SCG explants in the presence of human recombinant interleukin‐1 (hrIL‐1) but not hrIL‐2. Nerve growth factor (NGF) is not involved in mediating this effect since antibodies to NGF included in the culture fail to alter the lympho‐kine‐induced increase in SP. Moreover, the effect is somewhat specific for SP since the activities of tyrosine hydroxylase, tryptophan hydroxylase, and choline acetyltransferase (enzymes in the biosynthetic pathways for norepinephrine, serotonin, and acetylcholine) are not similarly elevated. Dorsal root ganglia respond with only modest increases in SP. The action of lymphokines in stimulating SP may, therefore, be a ganglion‐specific action in promoting recovery following injury.
Method for the identification of brain macrophages/phagocytic cells in vitro
Authors: Jordan F. L. , Wynder H. J. , Booth P. L. , Thomas W. E. 
Published: January 2005

AbstractThe use of labelled latex beads, with acute incubation conditions, for the identification of active macrophages in mixed cell cultures based on their phagocytic activity is described. Fluorescent beads provided the best results and selectively labelled active macrophage cell in cultures of blood monocytes. When this technique was applied to primary cultures of rat cerebral cortex, a specific cell type was significantly labelled. This was a small round cell, previously uncharacterized, which in addition to phagocytic activity indicated by the ingestion of beads also possessed macrophage biochemical markers. Thus, the procedure appears useful for phagocytic macrophage identification in vitro, and should be generally applicable to any tissue culture system. Additionally, this procedure can be rendered compatible with cell viability and, therefore, be utilized for long‐term monitoring of macrophages.
Axonal modulation of myelin gene expression in the peripheral nerve
Authors: LeBlanc A. C. , Poduslo J. F. 
Published: January 2005

AbstractMyelin gene expre(P0, MBP, P2, and MAG) was investigated during Wallerian degeneration and in the presence or absence of subsequent axonal regeneration and remyelination. The steady state levels of mRNA and protein were assessed in the crushed or permanently transected rat sciatic nerve at 0, 1, 4, 7, 10, 12, 14, 21, and 35 days after injury. The mRNA and protein steady state levels of the myelin specific genes, P0 and the MBPs, decreased to low yet detectable levels during Wallerian degeneration and returned to normal levels with subsequent axonal regeneration. The steady state level of P2 protein also followed a similar pattern of expression. The steady state level of MAG mRNA decreased to unddectable levels by 4 days of injury in the permanently transected nerve. After crush injury, re‐expression of MAG to levels comparable to those of normal nerves preceded that of P2 by 2 days and that of P0 and the MBPs by 3 weeks during axonal regeneration and remyelination. These results support the proposed roles for MAG in the formation of initial Schwann cell‐axonal contact required for myelin assembly, for P2 in fatty acid transport during myelination, and for P0 and the MBPs in the maintenance of the integrity and compactness of the myelin sheath. In addition, these results indicate that the expression of the myelin specific genes, P0 and MBP, is constitutive and that the level of myelin specific mRNAs is modulated by axonal contact and myelin assembly.
Gene transfer into brain tumor cell lines: Reporter gene expression using various cellular and viral promoters
Authors: Oellig C. , Seliger B. 
Published: January 2005

AbstractIn the present study we determined the optimal conditions for transferring DNA into rat and human brain tumor cell lines of glial and neuronal origin using electroporation as the transfection method. Gene transfer efficiency was measured in terms of transient chloramphenicol acetyltransferase (CAT) activity and stable neomycin expression. Moreover, the activity of a variety of cellular and viral promoters in brain tumor cell lines of distinct origin was characterized. The results revealed various expression patterns, including glial as well as neuronal specific promoter activity.
Axons modulate myelin protein messenger RNA levels during central nervous system myelination in vivo
Authors: Kidd G. J. , Hauer P. E. , Trapp B. D. 
Published: January 2005

AbstractExpression of myelin protein genes by myelinating Schwann cells in vivo is dependent on axonal influences. This report investigated the effect of axons on myelin protein mRNA levels in the central nervous system (CNS). In situ hybridization studies of rat spinal cord sections localized mRNAs encoding proteolipid protein (PLP) and myelin basic protein (MBP) 20 and 40 days after unilateral rhizotomy. Compared with control tissue, hybridization intensity was reduced in transected tissue, but there was little change in the number of oligodendrocytes labeled. Cellular RNA was extracted from transected and age‐matched control optic nerves 5, 10, 20, and 40 days after surgery, and levels of the following mRNAs were determined by slot blot procedures: PLP, MBP, myelin‐associated glycoprotein (MAG), and 2′,3′ cyclic nucleotide 3′‐phosphodiesterase (CNP). In transected nerves, PLP and MBP mRNA levels were approximately 85%, 45%, and 25% of control values at 5, 20 and 40 days posttransection, respectively. Axonal transection had a lesser effect on CNP and MAG mRNA levels, which declined to approximately 60% of control levels at 40 days. Immunocytochemical studies indicated that the number of oligodendrocytes was not decreased 40 days after optic nerve transection. These data demonstrate that axons modulate myelin protein mRNA levels in oligodendrocytes. In contrast to Schwann cells, however, oligodendrocytes continue to express significant levels of myelin protein mRNA in vivo following loss of axonal contact.
Effects of short‐ and long‐term ganglioside treatment on the recovery of neurochemical markers in the ibotenic acid‐lesioned rat striatum
Authors: Contestabile A. , Virgili M. , Migani P. , Barnabei O. 
Published: January 2005

AbstractThe striatum of adult rats was bilaterally lesioned with stereotaxic injections of ibotenic acid in a dose (16 nmoles) that resulted in subtotal lesions. Some rats received systemic ganglioside treatment starting the day before operation and lasting for 6 or 24 days after operation; they were compared with lesioned rat receiving systemic saline injections as well as with corresponding groups of sham‐operated animals. Specific neurochemical markers for cholinergic neurons (choline acetyltransferase, ChAT), GABAergic neurons (glutamate decarboxylase; GAD), and astrocytes (glutamine synthetase; GS) were assayed to asses the neurochemical recovery promoted by ganglioside treatment. Twenty‐four, but not six, days after operation a significant increase of ChAT and GAD was measured in the striatum of lesioned rats treated with gangliosides in comparison with the saline group. Furthermore, a significant increase of both enzymes occurred in the striatum of lesioned rats receiving ganglioside treatment for 24 days in comparison with rats receiving gangliosde treatment for 6 days only. A small but significant increase of ChAT was measured in the striatum of sham‐operated rats after 24 days of ganglioside treatment in comparison with the corresponding saline group. Finally, the increase of GS caused by the glial reaction to the ibotenic acid lesion was not affected by ganglioside treatment. The results indicate that a relatively long‐lasting ganglioside treatment stimulates the recovery of specific neuronal transmitter markers and that some effect is, in addition, exerted on unlesioned cholinergic striatal neurons.
Developmental and regional expression of basic fibroblast growth factor mRNA in the rat central nervous system
Authors: Ernfors P. , Lönnerberg P. , Ayer‐LeLievre C. , Persson H. 
Published: January 2005

AbstractA cDNA clone encoding rat basic fibroblast growth factor (bFGF) was used as a hybridization probe to determine the level of bFGF mRNA during rat brain development as well as in different adult rat brain regions. In the rat brain, a 3.7kb bFGF RUNA was detected together with lower levels of two minor bFGF mRNA species of 1.8kb and 1.5kb, respectively. The 3.7kb bFGF mRNA was detected in the rat brain already at embryonic day 16, the earliest time point tested. The embryonic brain contained 1.5 to 2 times higher levels of the 3.7kb bFGF mRNA than the adult brain. The amount of the 3.7kb bFGF mRNA in the adult rat brain was approximately 50 times higher than the level of β‐nerve growth factor mRNA in the rat brain. bFGF mRNA was found in all 12 brain regions tested in the adult rat brain with the highest level in colliculi, cerebral cortex, thalamus, and olfactory bulb. The lowest levels were found in pons and medulla oblongata. All three bFGF mRNA species showed the same regional distribution in the brain. In contrast to nerve growth factor mRNA, the level of bFGF mRNA in the neonatal hippocampus was slightly decreased 10 days after a cholinergic denervation by transection of the fimbria‐fornix.
Basic fibroblast growth factor in neuronal cultures of human fetal brain
Authors: Torelli S. , Dell'Era P. , Ennas M. G. , Sogos V. , Gremo F. , Ragnotti G. , Presta M. 
Published: January 2005

AbstractThe presence of basic fibroblast growth factor (bFGF) was investigated in neuronal cells derived from 12 and 18 week‐old human fetal brain cultures. To this purpose, the ability of bFGF to stimulate plasminogen activator (PA) production in fetal bovine aortic endothelial GM 7373 cells was used as an assay for this molecule in neuronal cell extracts. The identity of the PA‐stimulating activity of neuronal cell extract with bFGF was confirmed by its high affinity for heparin and by its cross‐reactivity with polyclonal antibodies to human placental bFGF. These antibodies recognized a Mr 18,000 cell‐associated protein experiments. All the neurons showed bFGF immunoreactivity as demonstrated by immunocytochemical staining, while nonneuronal cells were unstained. The data demonstrate for the first time that cultured human fetal brain neurons contain and synthesize bFGF.