bluuebunny/crossref_metadata_2025_split huggingface dataset:
Protein synthesis in the neuropil of the rat dentate gyrus during synapse development
Authors: Phillips L. L. , Pollack A. E. , Steward O. 
Published: January 2005

AbstractPrevious studies have shown that there are dramatic accumulations of polyribosomes under developing synapses on dendrites of CNS neurons. The present study was designed to evaluate what types of proteins might be synthesized by the synapse‐associated polyribosomes. Hippocampal slics from rat pups sacrificed at 4,7,10,12,14, and 21 days after birth as well as slices from adult animals were incubated in a modified Eagle's medium containing 3H‐leucine. After a 30 min exposure to radiolabeled amino acids, the slices were microdissected, separating the dendritic enriched molecular layer from the cell bodies of the dentate gyrus and the hippocampus proper. The level of protein synthetic activity was assessed by comparing the incorporation in cell body and dendritic laminae. Polypeptides present in each dissected zone were separated electrophoretically on 1D SDS‐polyacrylamide gels according to their molecular weight and the newly synthesized proteins were analyzed through gel fluorography.The overall level of 3H‐amino acid incorporation into protein (measured as cpm/μg protin) was higher than that of the adult at all postnatal ages. When the entire slice was analyzed, the maximum incorporation was at 12 days after birth. In the dissected subregions of the slice the peak protein synthetic activity in cell‐body‐enriched regions of dentate gyrus and hippocampus proper was at 4–7 days postnatal, declining between 7 and 21 days to values comparable to the adult. By contrast, protein synthesis in the molecular layer of the dentate gyrus did not peak until 12 days after birth, decreasing toward adult rates after 14 days. The overall pattern of Coomassie stained polypeptides present in the dentate molecular layer was comparable at all ages examined. Moreover, one‐dimensional gel analysis showed no qualitative differences in the proteins that were synthesized in the three dissected zones across ages. Gel fluorography identified four mid‐molecular weight ranges of polypeptides (43, 50, 52, and 56 kD) and a group of high molecular weight proteins (190‐220 kD) that were the most heavily labeled species in the dissected molecular layer. Quantitative gel analysis revealed that three of the midrange bands (43, 52, and 56 kD) accounted for a different percentage of the over all labeled protein across developmental age; prominent changes were found in the 43 kD band, where increased radiobeling was directly correlated with postnatal periods of rapid synaptogenesis.We conclude that there is a high level of protein synthesis in dendritic laminae during developmental synaptogenesis. This burst of synthesis appears to result in the increased generation of a set of proteins which are normally synthesized within both granule cell body and dendrite enriched regions. The 43 kD member (s) of this set of polypeptides shows increased labeling during the most rapid phase of synaptogenesis.
Rapid, sensitive, and simple method for quantification of both neurotoxic and neurotrophic effects of NMDA on cultured cerebellar granule cells
Authors: Didier M. , Heaulme M. , Soubrié P. , Bockaert J. , Pin J.‐P. 
Published: January 2005

AbstractA simple and sensitive method adapted from the staining of living cells with fluorescein diacetate was developed to rapidly estimate the number of living cells remaining in a culture dish 24 hr after a few min of NMDA treatment of cerebellar neurons. This method consists of the measurement, after cell lysis, of the total amount of fluorescein produced from fluorescein diacetate by the living granule cells present in each culture dish. We show that this method can also be used to quantify the survival effect of chronic exposure of granule cells to either K+ or NMDA In both cases, the fluorescence measured was found to be proportional to the number of fluorescein‐labelled cells counted under a fluorescence microscope, indicating that the present method can be used to quantify both toxic and trophic effects of NMDA on cerebellar granule cells. This study confirms that these two NMDA effects occur at the same NMDA concentration, and both are inhibited by MK 801 in the same concentration range. We showed, moreover, that granule neurons developed in the presence of NMDA are much less sensitive to NMDA toxicity than neurons developed in K+‐enriched medium.
Induction of oligodendrocyte differentiation by activators of adenylate cyclase
Authors: Raible D. W. , McMorris F. A. 
Published: January 2005

AbstractOligodendrocyte differentiation is accelerated by analogs of cyclic AMP (cAMP), such as N62‐′O‐dibutyryladenosine 3′,5′‐cyclic monophosphate (dbcAMP) or 8‐bromo cAMP, in developing rat glial cell cultures (Raible and McMorris, 1989). However, it is not known whether the immature developing oligodendrocytes have G protein‐coupled adenylate cyclase capable of generating cAMP, as would be required if development of the cells is normally regulated by endogenous cAMP. In the present study, we demonstrate that the diterpene forskolin, a potent activator of adenylate cyclase, or cholera toxin, which activates adenylate cyclase through G protein modification, can induce a precocious increase in oligodendrocyte number. When both forskolin and cholera toxin are added together, oligodendrocyte differentiation is induced to a level similar to that observed in response to cAMP analogs. These results establish for the first time that modulation of the endogenous cAMP regulatory system alters the schedule of oligodendrocyte differentiation.
Potassium‐stimulated release of taurine from cultured cerebellar granule neurons is associated with cell swelling
Authors: Schousboe A. , Morán J. , Pasantes‐Morales H. 
Published: January 2005

AbstractEffects of increased concentrations of potassium and of hyposmolar conditions on release of taurine were investigated in cerebellar granule neurons cultured from mice. It was found that increases in the external potassium concentration as well as decreases in osmolarity dose‐dependently increased release of exogenously supplied [3H]‐taurine and endogenous taurine from the neurons. The release of endogenous taurine elicited by a reduction of the osmolarity of the incubation media to 70% or 50% was much more pronounced than that of other amino acids, particularly glutamine, the release of which was not affected at all. The potassium‐stimulated release of [3H]‐taurine was strictly chloride dependent and it was inhibited by an increase of the osmolarity of the media as well as by 4,4′‐diisothyocyanostilbene‐2,2′‐disulfonate (DIDS) (100 μM). Moreover, a similar increase in the potassium concentration led to an increase in intracellular volume (swelling), a process which was also chloride dependent. It is concluded that potassium‐stimulated taurine release from cerebellar granule neurons is associated with cell volume changes and that taurine is likely to play a role as an osmotically active substance in these neurons.
4‐Aminopyridine interrupts the modulation of acetylcholine release mediated by muscarinic and opiate receptors
Authors: Törőcsik A. , Vizi E. S. 
Published: January 2005

AbstractThe effect of 4‐aminopyridine, a potassium channel blocker on the muscarinic and opiate modulation of acetylcholine release, was investigated. Rat frontal cortical slices were loaded with [3H]choline, super‐fused continuously, and stimulated electrically. 4‐Aminopyridine enhanced the stimulation‐evoked release of tritium without affecting basal release. The electrically evoked release of radioactivity was reduced by the muscarinic agonist oxotremorine and the delta selective opiate receptor agonist Metenkephalin, and was enhanced—in the presence of the cholinesterase inhibitor physostigmine—by the muscarinic antagonist atropine. These effects were completely abolished by 4‐aminopyridine. Since 4‐aminopyridine blocks potassium permeability of the neuron, it is suggested that the changes in potassium permeability and the consequent alteration of membrane polarization are involved in the presynaptic modulation of acetylcholine release.
Specific antibodies for tyrosinated and detyrosinated tubulin recognize retina tubulin subpopulations that do not participate in the posttranslational tyrosination/detyrosination cycle
Authors: Arregui C. , Barra H. S. 
Published: January 2005

AbstractWe have used the monoclonal YL 1/2 (Tyr antibody) and polyclonal (Glu antibody) antibodies, specific for tyrosinated and detyrosinated tubulin, respectively, to determine the levels and cellular distribution of these tubulin species in chick retina during development. At embryonic day 4, detyrosinated tubulin was restricted to the ganglion cells of the fundic region. As development progresses, immunofluorescence also appears, first, in the outermost zone of the retina and then in the plexiform layers. The Tyr antibody staining was found in the different layers and it was fairly homogeneous in distribution. Analysis by dot immunobinding showed that the ratios of tyrosinated to detyrosinated tubulin obtained at different ages do not agree with those obtained previously by an enzymatic method based on the incorporation of [14C]tyrosine found that the lack of coincidence is due to the fact that a fraction of the tubulin species determined by the Tyr and Glu antibodies does not participate in the posttranslational tyrosination/detyrosination cycle. This is a novel concept that should be considered in the interpretations of immunofluorescence studies concerning the cellular distribution of tyrosinated and detyrosinated tubulin.
Carbohydrate epitopes involved in neural cell recognition are conserved between vertebrates and leech
Authors: Bajt M. L. , Schmitz B. , Schachner M. , Zipser B. 
Published: January 2005

AbstractWe are reporting on the evolutionary conservation of carbohydrate epitope families from vertebrate to leech. 1) The sulfated L2/HNK‐1 carbohydrate epitope Abo T, Balch CM (1981): J Immunol 127:1024–1029; Kruse J,Mailhammer R, Wernecke H, Faissner A, Timpl R, Schachner M (1984): Nature 311:153–155 is detected on glycoproteins of leech neurons using monoclonal antibodies (mAbs) L2 (336) and HNK‐1. 2) Three rat mAbs, L3, L4, and L5, bind to leech nerve and muscle. The L3, L4, and L5 epitopes are localized to a group of mannosidic leech glycoproteins originally identified through mAbs Lan3‐2 (Hogg N, Flaster M, Zipser B (1983): J Neurosci Res 9:445–457 and Laz6‐189 (McRorie JW III, Zipser B (1988): “Cell Culture Approaches to Invertebrate Neuroscience.” London: Academie Press, pp 33–52. MAb Lan3‐2, which binds to a mannosidic epitope of the 130 kD sensory protein, has recently been shown to perturb the penetration of sensory afferents into the synaptic area of the central neuropile (Zipser B, Morell R, Bajt ML (1989): Neuron 3:621–630). The L3, L4, and L5 mAbs have been described to recognize different mannosidic epitopes on glycoproteins, some of which have been identified as neural cell adhesion molecules, and on astrocyte‐specific proteoglycan from mouse brain (Kucherer A, Faissner A, Schachner M (1987): J Cell Biol 104:1597–1602; Fahrig T, Schmitz B, Weber D, (Kücherer‐Ehret A, Faissner A, Schachner M (1990): Eur J Neurosci 2:153–161. Streit A, Faissner A, Gehrig B, Schachner M (1990): J Neurochem In Press). The superposition of five different mannosidic epitopes on the axons of sensory afferents suggests complex, concerted participation of mannosidic epitopes in neuronal pathfinding and target recognition.
Glass micro‐fibers: A model system for study of early events in myelination
Authors: Bullock P. N. , Rome L. H. 
Published: January 2005

AbstractA system was developed to analyze early events in the process of myelination. Primary cultures of rat oligodendrocytes were maintained in the presence of glass micro‐fibers which served as artificial axons. A culture chamber was constructed which allowed the close apposition of fibers and cells in a three‐dimensional arrangement designed to resemble an in vivo environment. Cells cultured in the presence of glass micro‐fibers coated with a glial cell matrix extract were induced to organize into clusters around the fibers. Examination of oligodendrocyte‐fiber sandwiches by SEM revealed the presence of a number of cell contacts with the fibers. TEM images showed that, in most cases, fibers were surrounded by the cells and not multiply wrapped. Only occasionally was a loose wrapping of cell membrane observed around the fibers. Cells cultured in the presence of matrix‐coated glass micro‐fibers showed an increased production of sulfolipids that was at least partially dependent on the presence of the matrix coating. Coating of these “artificial axons” may aid in the identification of signal molecules produced by neurons which enable them to be myelinated.
Type 1 astrocytes and oligodendrocyte‐type 2 astrocyte glial progenitors migrate toward distinct molecules
Authors: Armstrong R. C. , Harvath L. , Dubois‐Dalcq M. E. 
Published: January 2005

AbstractDuring central nervous system (CNS) development, glial precursors proliferate in subventricular zones and then migrate throughout the CNS to adopt their final destinations and differentiate into various types of mature glial cells. Although several growth factors promoting the proliferation and/or differentiation of glial precursors have been identified, very little is known about the nature of signals that guide glial cell migration in the CNS. Therefore, we have investigated whether polypeptide growth factors and/or extracellular matrix molecules may mediate the migration of two major glial cell types, type 1 astrocytes and oligodendrocyte‐type 2 astrocyte (O‐2A) progenitor cells. We show that, in a microchemotaxis chamber assay, type 1 astrocytes move toward laminin and complement‐derived C5a. Astrocyte migration toward laminin is inhibited by a laminin‐specific pentapeptide, YIGSR‐NH2. In contrast, O‐2A progenitors migrate toward platelet‐derived growth factor (PDGF), which also functions as a mitogen for these cells. Using a new method to simultaneously assay migration and DNA synthesis, we also demonstrate that O‐2A progenitors can migrate toward PDGF even when DNA replication is inhibited with an antimitotic agent. Thus, migration of different types of glial cells can be induced in vitro by specific signaling molecules, which are present in the developing brain and may stimulate migration of glial cells prior to CNS myelination.
Isolation and localization of a slow troponin (TnT) gene on chromosome 19 by subtraction hybridization of a cDNA muscle library using myotonic dystrophy muscle cDNA
Authors: Samson F. , Lee J. E. , Hung W. ‐Y. , Potter T. G. , Herbstreith M. , Roses A. D. , Gilbert J. R. 
Published: January 2005

AbstractSubtraction hybridization techniques were used to isolate 91 cDNA clones which are overexpressed in normal control skeletal muscle relative to muscle from patients with myotonic muscular dystrophy. The gene responsible for myotonic dystrophy (DM) has been localized to the 19q13.2–13.3 region of chromosome 19. To test as a candidate gene for DM, clones which represent differences in transcription are analyzed for localization to chromosome 19. One clone, designated MSL 366, was found to be on the long arm of chromosome 19 distal to the CKMM gene at 19q13.2. Sequence analysis confirmed that MSL 366 is the cDNA for human slow skeletal muscle troponin T. A genomic clone has been isolated and linkage studies with DM are in progress.
Glial fibrillary acidic protein and βA4 protein deposits in temporal lobe of aging brain and senile dementia of the Alzheimer type: Relation with the cognitive state and with quantitative studies of senile plaques and neurofibrillary tangles
Authors: Harpin M. L. , Delaère P. , Javoy‐Agid F. , Bock E. , Jacque C. , Delpech B. , Villarroya H. , Duyckaerts C. , Hauw J. J. , Baumann N. 
Published: January 2005

AbstractThe aim of this study was to compare brain glial fibrillary acidic protein (GFAP) levels to the modifications of cognitive functions (Blessed test score [BTS]), the density of the main neuropathological lesions (senile plaques [SP] and neurofibrillary tangles [NFT]), and the density of the two main subtypes of βA4 deposits (classic plaques and diffuse deposits) in a series of patients with normal aging and senile dementia of the Alzheimer type of various degrees of severity. GFAP levels (enzymelinked immunosorbent assay [ELISA] technique) and the densities of changes were measured in the temporal lobe of 12 women over 75 years of age. Under these conditions, the ELISA assay could determine GFAP in brain homogenates (aqueous‐Triton buffer soluble extract) in a range from 2.5 ng to 600 ng per assay. Least affected patients (with a BTS of 19 and over) all ranged below 60 μg/mg protein. Most affected patients (with a BTS under 6) ranged above 150 μg/mg protein. However, interindividual variations were wide. A significant correlation between the BTS and the amount of GFAP could be found only when using the non parametric test of Spearman. There was a significant positive correlation between the amount of GFAP and the density of (1) SP, (2) NFT both revealed by Bodian's silver stain, and (3) classic βA4 plaques shown by immunocytochemistry. On the contrary, no correlation was observed with diffuse βA4 deposits. One case with very large amounts of diffuse βA4 deposits without SP or NFT showed no associated GFAP reactivity. This suggests that GFAP production is a critical event in the formation of classic SP.
Calcium‐dependent regulation of the enkephalin phenotype by neuronal activity during early ontogeny
Authors: Agoston D. V. , Eiden L. E. , Brenneman D. E. 
Published: January 2005

AbstractGenetic components of the neuronal phenotype are regulated by epigenetic factors—trophic molecules and neuronal activity—during neurodifferentiation. Developing neurons in dissociated cultures of embryonic mouse spinal cord show spontaneous electrical activity after one week in culture. We now report that the blockade of this spontaneous electrical activity for two days with tetrodotoxin (TTX) causes virtually complete down‐regulation of preproenkephalin A gene transcripts in embryonic spinal cord cultures. This TTX‐induced down‐regulation is fully reversed upon reinitiation of neuronal activity (removal of TTX from cultures). This reversible, tetrodotoxin‐induced down‐regulation kephalin mRNA is confined to a restricted period of early neurodevelopment (days 7 to 21 in culture). Since depolarization triggers calcium entry through voltage‐activated calcium channels, we have investigated the involvement of calcium in the mechanism of this activity‐ and age‐dependent regulation of preproenkephalin A expression. The selective activation of the L‐type of voltage‐sensitive calcium channels by a dihydropyridine derivative [( + ) 202‐791] prevented this TTX‐induced down‐regulation without reducing methionine enkephalin secretion. This effect was observed only when the drug was applied to electrically active cultures, prior to the addition of TTX. Simultaneous application of ( + ) 202‐791 and TTX, or pretreatmont with TTX, failed to prevent TTX‐induced down‐regulation. Thus, activity‐dependent phenotypic plasticity of met‐enkephalinergic neurons in spinal cord is:(1) maximum at an early age of neuronal development (<10 days in culture) and becomes less apparent in old cultures (>30 days); (2) reversible throughout; and (3) mediated by calcium entry through L‐type channels.
Transplantation of a temperature‐sensitive, nerve growth factor‐secreting, neuroblastoma cell line into adult rats with fimbria–fornix lesions rescues cholinergic septal neurons
Authors: Whittemore Scott R. , Holets V. R. , Keane R. W. , Levy D. J. , McKay R. D. G. 
Published: January 2005

AbstractThe HT4 cell line was derived from infection of a mouse neuroblastoma cell line with a retrovirus that encoded the temperature‐sensitive (ts) mutant of SV40 large T antigen. At nonpermissive temperature, HT4 cells differentiated with neuronal morphology, expressed neuronal antigens, synthesized nerve growth factor (NGF) mRNA, and secreted biologically active NGF in vitro. We sought to establish whether transplanted HT4 ceils expressed class I major histocompatibility complex (MHC) antigens, a partial requirement for recognition by cytotoxic T lymphocytes (CTL), and thus be susceptible to xenograft rejection. Differentiated HT4 cells expressed marginally detectable levels of class I MHC antigens, but demonstrated higher levels of class I MHC expression after treatment with interferon‐γ. However, HT4 cells were resistant to direct lysis by perforin the pore‐forming protein of CTLs, and thus may have potential use in xenograft experiments. To address whether HT4 cells secrete NGF in vivo, HT4 cells were transplanted into adults rats with unilateral fimbria‐fornix transections. A ts cell line derived from P4 cerebellum, BTl, that does not differentjate with neuronal phenotype or synthesize NGF in vitro, was transplanted as a control. Six weeks posttransplant. HT4 cells had integrated into host CNS without forming tumors. In BT1 transplants, the number of medial septal acetylcholinesterase (AChE)‐positive cells was reduced to 26‐39% of the contralateral control side, depending on the rostrocaudal level. In HT4 transplants, the number of cholinergic septal neurons was 58‐78% of the contralateral side. This percentage was significantly (P < 0.005) greater than that seen with BT1 transplants, indicating that transplanted HT4 cells secrete NGF in vivo and rescue cholinergic septal neurons following fimbria‐fornix transection.
Tyrosinated and detyrosinated microtubules in axonal processes of cerebellar macroneurons grown in culture
Authors: Arregui C. , Busciglio J. , Caceres A. , Barra H. S. 
Published: January 2005

AbstractWe have used the monoclonal antibody YL 1/2 (Tyr) specific for tyrosinated tubulin, and a polyclonal antibody (Glu) specific for detyrosinated tubulin to visualize the distribution of microtubules and microtubule assembly sites during axonal outgrowth. Cerebellar macroneurons growing in culture initially extend several short and thin neurites which have the potential to differentiate either as axons or dendrites (Ferreira and Caceres: Developmental Brain Research 49:205‐213, 1989). At the onset of axonal outgrowth the Tyr antibody labels the minor neurites, the axon, and its growth cone, while the Glu antibody only shows immunoreactivity in the axonal shaft. After nocodazole treatment, the Tyr staining disappears, whereas that produced by the Glu antibody remains practically unchanged. When nocodazole was removed, tyrosinated microtubules reappeared first at the tip of the axon, in a more distal region than that occupied by detyrosinated microtubules; another focal site of tyrosinated tubulin incorporation was detected in the cell body. Incorporation of tyrosinated tubulin into growing axons was also studied after taxol treatment. After long incubation periods in the presence of taxol, the Tyr staining disappeared from the axon but remained in the cell body; however, immunoreactivity in this site was negative when the cells were preincubated in the presence of protein synthesis inhibitors. Release from taxol results in the reappearance of Tyr immunoreactivity at the distal end of the axon. Taken collectively, the present results indicate (1) that in cerebellar macroneurons axonal differentiation is accompanied by a temporal and spatial differentiation of microtubules fcnd (2) that there is an active site of tyrosinated tubulin assembly at the tip of axonal processes, and they suggest that the highly tyrosinated domain in this region is a consequence of rapid microtubule turnover and tubulin tyrosine ligase activity.