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61.
Developmental regulation of 16S acetylcholinesterase and acetylcholine receptors in a mouse muscle cell line 总被引:18,自引:0,他引:18
Nibaldo C. Inestrosa Jeffrey B. Miller Laura Silberstein Lea Ziskind-Conhaim Zach W. Hall 《Experimental cell research》1983,147(2):393-405
We have studied the appearance, distribution and regulation of acetylcholinesterase (AChE) and acetylcholine receptors (AChRs) in a mouse skeletal muscle cell line (C2), that was originally isolated and described by Yaffe & Saxel [54]. In culture, cells from this line form spontaneously contracting myotubes, with overshooting action potentials that are TTX-sensitive. After fusion of myoblasts into myotubes, there was a dramatic increase in the amount of both AChE and AChR. Three forms of AChE, distinguished by their sedimentation on sucrose gradients, were synthesized: 4-6S, 10S, and 16S. The 4-6S and 10S forms appeared 1 day after the cells began to fuse, whereas the 16S form appeared only 2 days after fusion began. Maximal levels of the 16S AChE form (25-30% of the total) were obtained by reducing the concentration of horse serum in the fusion medium. Prevention of myoblast fusion by reducing the calcium levels in the medium decreased the total AChE by 70%, and only the 4-6S form was synthesized. Blocking spontaneous contractile activity of the myotubes by tetrodotoxin (TTX) led to a 50% reduction in all three esterase forms. Thus, the 16S, or endplate form of AChE is not specifically regulated by electrical or contractile activity in the C2 cell line. After fusion the number of AChRs increased rapidly for 3-4 days and then stabilized. Receptor clusters, ranging from 10-30 micron in length, appeared 1 day after myoblast fusion began. When cells were grown in medium containing reduced Ca2+, the total number of AChRs was decreased by 20-50%. Reduction of Ca2+ after myotubes and AChR clusters had formed resulted in dispersal of AChR clusters. Inhibition of muscle contractions with TTX did not affect the number of AChRs or their distribution. 相似文献
62.
Jorge H. Medina María Laura Novas† Eduardo De Robertis† 《Journal of neurochemistry》1983,41(3):703-709
Abstract: This study was designed to analyze possible differences in the binding of [3H]flunitrazepam ([3H]FNZP) and [3H]ethyl - β - carboline - 3 - carboxylate ([3H]β-CCE), to rat brain membranes, in various experimental conditions. In cerebral cortex, hippocampus, cerebellum, and orain stem the number of binding sites for [3H]β-CCE was higher than for [3H]FNZP; both were displaced by clonazepam. Until the 7th day of postnatal brain development the numbers of [3H]FNZP and [3H]β-CCE sites were equivalent; but later on, the β-carboline sites increased to a higher level. Noradrenergic denervation by 6-hydroxydopamine was followed in the hippocampal formation. Already after 2 days, there was a decrease in [3H]FNZP sites, which reached 70% of control after 14 days. Similar results were obtained with DSP-4 denervation. This change was only in Bmax and not in KD, In contrast, the [3H]β-CCE sites did not change with denervation. Neonatal injection of l - 2,4,5 - trihydroxyphenylalamine or DSP-4 produced in the adult a decrease in [3H]FNZP sites in the cerebral cortex, in parallel with the noradrenergic denervation. On the other hand, there was an increase in the cerebellum and brain stem, in correspondence with the hyperinnervation by sprouting. In these rats, the number of sites for [3H]β-CCE did not change in the different brain regions. With 0.1% Triton X-100, applied to synaptosomal membranes, [3H]FNZP binding was reduced by 35%, while that of [3H]β-CCE was not significantly changed. These results suggest that there is heterogeneity of binding sites for benzodiazepine receptors in rat brain. A tentative interpretation of the experiments involving noradrenergic denervation and hyperinnervation, as well as those with Triton X-100, is that [3H]FNZP binds to pre- and postsynaptic receptors, while [3H]β-CCE binds mainly to postsynaptic benzodiazepine receptors. 相似文献
63.
Anna Maria Tagliasacchi Laura Maria Costantina Forino Manuela Frediani Silvana Avanzi 《Protoplasma》1983,115(2-3):95-103
Summary The pattern of DNA and RNA puffs in pair VII of polytene chromosomes has been investigated in the suspensor ofPhaseolus coccineus during early embryo development. The pattern of3H-TdR and3H-U incorporation has been also detected. Collected data indicate that: 1. both heterochromatic regions, p11 and q(111+112), of chromosome pair VII, organize large DNA puffs; 2. DNA puffs of both regions are specific of different embryo differentiation steps; 3. a seasonal influence on the DNA puffing seems also to be present, as demonstrated by the comparison of the results collected in two different crops; 4. the incorporation experiment by3H-TdR evidences that not all DNA puffs show clustered labeling; 5. the RNA puffing of the two regions seems also to be specific of determined embryo stages. 相似文献
64.
Glucosylation of Teichoic Acid: Solubilization and Partial Characterization of the Uridine Diphosphoglucose:Polyglycerolteichoic Acid Glucosyl Transferase from Membranes of Bacillus subtilis 总被引:4,自引:2,他引:2
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Polyglycerolteichoic acid:glucosyl transferase (TAG transferase), one of the three enzymes involved in the pathway leading to the glucosylation of teichoic acid in Bacillus subtilis 168, was investigated. During the early stages of the growth of B. subtilis, TAG transferase is predominantly a soluble enzyme found in the cytoplasm. As growth proceeds, the amount of soluble enzyme decreases and the proportion of insoluble, membrane-bound TAG transferase increases, reaching a maximal value at the close of the logarithmic phase. Data are presented which suggest that these are two forms of the same enzyme, or have some common component. The effects of chaotropic agents, such as sodium trichloroacetate and sodium perchlorate, on the cytoplasmic membrane were also studied. These data show that such compounds can effectively remove the TAG transferase from the membrane in a water-soluble form. A study of some of the physical properties of this solubilized enzyme suggests that there is little difference between the two forms of the enzyme. Experiments are described which indicate that the glucosyl transfer by both the membrane-bound and soluble enzymes is not mediated by lipids. 相似文献
65.
Nature of Escherichia coli B(P1) Yielder Cells at the Time of Infection with Restricted T1 总被引:1,自引:1,他引:0
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In the infection of Escherichia coli B(P1) with restricted T1, it was shown that yielder cells consist of both special and nonspecial cells. Special or predetermined yielders occurred only among the earliest yielders. In most instances, yielder-cell formation was most easily explained by assuming that the first step was a chance escape of the restricted phage DNA from the degrading enzyme of the restricting cell. 相似文献
66.
Locus-specific Changes in Cell Wall Composition Characteristic of Osmotic Mutants of Neurospora crassa 总被引:4,自引:4,他引:0
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Laura R. Livingston 《Journal of bacteriology》1969,99(1):85-90
The osmotic phenotype of Neurospora crassa is characterized by inhibition of growth at high osmolalities of growth medium. Mutations at six osmotic loci of linkage group I were examined to assess the biochemical and physiological effects of these mutants. Isolated cell walls from 23 osmotic strains were compared with the wild type with regard to quantitative levels of the following components: percentage of total dry weight, total glucose, alkali-soluble glucose, nonglucose carbohydrates, amino acids, glucosamine, galactosamine, and a compound tentatively identified as quinovosamine. The last component has not previously been observed in N. crassa cell walls. Although the cell wall dry weight content of osmotic mutants was not altered, walls isolated from all of the osmotic strains had less alkali-insoluble glucose than those from the wild type. In addition, all of the loci except cut exhibited other consistent differences from the wild type. The os-1, os-3, and os-5 mutants had low levels of alkali-soluble glucose. The os-3 and os-5 mutants had high levels of nonglucose carbohydrates, and flm-2 had a low level of nonglucose carbohydrates. The os-4 mutants had low levels of galactosamine and amino acids and high levels alkali-soluble glucose. An os-1 mutant, B135, produced less of the whole alkali-soluble fraction of the cell wall. 相似文献
67.
Egg Membrane Lytic Activity of Sperm Extract and its Significance in Relation to Sperm Entry in Hydroides hexagonus (Annelida) 总被引:4,自引:4,他引:0
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Previous electron microscope studies indicated that the individual spermatozoön of Hydroides hexagonus forms a hole in the vitelline membrane by means of lysis. Other observations established that the hole is real, being visible in living material during sperm entry. During the present investigation sea water extracts from frozen-thawed sperm were tested for lytic effect on the membrane. In normal living eggs the membrane appears as a single thick envelope, but in electron micrographs of sections it is seen to consist of a narrow outer border layer, a wide principal or middle layer, and a narrow inner border layer. After immersion in sperm extract the outer border layer elevates but does not dissolve, the middle layer liquefies and disappears, and the inner border layer seems not to change. This is interpreted as lysis of the middle layer. The extract exerted the same effect on fertilized and unfertilized eggs. In electron micrographs the sections treated with extract greatly resemble that part of the membrane which has been penetrated by the individual spermatozoön. It is concluded that the individual spermatozoön, too, exerts a lytic effect. Together, the present and two earlier studies are considered clearly to demonstrate that in Hydroides the individual spermatozoön does indeed make an entry hole in the egg membrane by applying lytic material to that part of the membrane in its own vicinity. 相似文献
68.
Jeffery R. Cook Barbara E. Crute Laura M. Patrone Joseph Gabriels Maureen E. Lane Robert G. van Buskirk 《In vitro cellular & developmental biology. Plant》1989,25(10):914-922
Summary We have analyzed the ability of the physical substratum to modulate both the ultrastructural and protein synthetic characteristics
of the Madin-Darby canine kidney (MDCK) renal cell line. When MDCK cells were seeded on Millipore Millicell CM microporous
membrane cell culture inserts they demonstrated a more columnar organization with an increase in cell density sixfold greater
than the same cells seeded on conventional plastic substrata. After 1 wk postseeding on the microporous membrane a partial
basal lamina was noted, with a contiguous basement membrane being apparent after 2 wk. One-dimensional sodium dodecyl sulfate
gel electrophoresis was used to analyze detergent-solubilized proteins from MDCK cells maintained on plastic substrata vs.
microporous membranes. When proteins were pulse-labeled with [35S]methionine, a 55 kDa protein was evident in the cytosolic extract of cells grown on collagen, laminin, and nontreated plastic
substrata; but this labeled protein was not evident in similar extracts from cells grown on collagen and laminin-coated microporous
membranes. To test if the polarized, basement-membrane secreting phenotype of the MDCK cells could be generated on a microporous
membrane without pretreatment with any extracellular matrix (ECM) components, cells were seeded on the Millipore Millicell
HA (cellulosic) microporous membrane. This type of substrata does not need a coating of ECM components for cell attachment.
A partial basement membrane was formed below cells where the basal surface of the cell was planar, but not in areas where
the cell formed large cytoplasmic extensions into the filter. This led us to the conclusion that the microporous nature of
the substrata can dictate both ultrastructural and protein synthetic activities of MDCK cells. Furthermore, we suggest that
both the planar nature of the basal surface and the microporosity of the substrate are corequisites for the deposition of
the basement membrane. 相似文献
69.
The ultrastructure of the mature spermatozoa and spermatogenesis of the bivalve Scrobicularia plana are described. Support cells extend from the basal lamina to the lumen of the testis and are laterally connected to the germinal epithelium. Germ cells present intercellular bridges and flagella since the spermatogonial stage. While spermatogonia and spermatocytes appear connected to support cells by desmosome-like junctions, elongated spermatids are held at the acrosomal region by support cell finger-like processes. During spermiogenesis, the acrosomal vesicle differentiates from a golgian saccule and then migrates to the nuclear apex. A microtubular manchette arising from centrioles surrounds the acrosomal vesicle, the nucleus, and the mitochondria at the time these three organelles start their elongation, disappearing after that. The mature spermatozoon of S. plana lacks a distinct midpiece because the mitochondria extend from the region of the pericentriolar complex along the nucleus anteriorly for approximately 1.4 μm. The features of this bivalve type of modified spermatozoon are compared with those of other animal groups having similar modifications. 相似文献
70.
Bruna Tadolini Diana Fiorentini Laura Landi Luciana Cabrini 《Free radical research》1989,5(4):245-252
To find experimental conditions to selectively study the propagation phase of lipoperoxidation we studied the lipoperoxidation, catalyzed by FeCl2, of liposomes in a buffering condition where Fe2+ autoxidation and oxygen active species generation does not occur. Liposomes from egg yolk phosphatidylcholine. prepared by vortex mixing, do not oxidize Fe2+: on the contrary they oxidize Fe2+ when prepared by ultrasonic irradiation. Dimyristoyl phosphatidylcholine liposomes prepared by ultrasonic irradiation do not oxidize Fe2+. During sonication polyunsaturated fatty acid residues autoxidize and lipid hydroperoxides (LOOH) are generated. Only when LOOH are present in the liposimes Fe2+ oxidizes and its rate of oxidation depends on the amount of LOOH in the assay. The reaction results in the generation of both LOOH and thiobarbituric acid reactive material (TBAR): it is inhibited by butylated hydroxytoluene and has a acidic pH optimum; it is not inhibited by catalase and OH' scavengers. The reaction studied. thus, appears to be the chain branching and propagation phase of lipoperoxidation. When we studied the dependence of Fe2+ oxidation, LOOH and TBAR generation on FeCl2 concentration, we observed that at high FeCl2 concentrations the termination phase of lipoperoxidation was prevalent. Thus. by selecting the appropriate FeCl2 concentration the proposed experimental system allows study of either the propagation or the termination phase of lipoperoxidation. 相似文献