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PubMed1981

Comparative ultrastructure of eyespot membranes in gametes and zoospores of the green alga Ulva lactuca (Ulvales).

Eyespot membranes in zoospores, and male and female gametes of the green alga Ulva lactuca, were studied comparatively by the freeze-fracture technique. The plasmalemma and the outer chloroplast envelope membrane overlying the eyespot lipid globules are specialized in all 3 types of reproductive cells. In the eyespot region the protoplasmic face (PF) of the outer chloroplast envelope membrane contains significantly more intramembraneous particles (IMP) compared to membrane areas outside the eyespot: in female gametes there are 2.5 times more IMP/micrometers 2, in zoospores 3 and in male gametes about 4. Small size-class IMP (4--6 nm diameter) are particularly abundant on both fracture faces of the outer chloroplast membrane, but size-class distribution is not significantly different between membrane areas inside and outside the eyespot region. The total number of IMP/eyespot on the PF of the outer chloroplast membrane was calculated to be 4900 in male gametes, 5500 in female gametes and 11 200 in zoospores. The results are discussed in accordance with the view that these membrane specializations participate in photoreception relating to green algal phototaxis. Evidence is presented that there is a correlation between IMP numbers per eyespot in the outer chloroplast envelope membrane and the different phototactic behaviour of gametes compared to zoospores in Ulva.

Chlorophyta↗
PubMed1981

Freeze-fracture architecture and polypeptide composition of thylakoid membranes from euploid Ricinus cells.

The freeze-fracture architecture and polypeptide composition of thylakoid membranes of euploid cells of Ricinus communis L. were examined. Electron microscopic examination of the chloroplasts of 1N 2N and 4N cells revealed little variation in the size of chloroplasts, lamellar structure and internal organization of plastids, despite increases in plastid numbers per cell observed to accompany the increase in nuclear ploidy. Thylakoid membranes from euploid cells were also similar in their freeze-fracture morphology. Two basic types of intramembranous particles were observed on the fracture faces of thylakoid membranes of euploid cells. The endoplasmic fracture (EF) face of experimentally unstacked thylakoid membranes of 1N, 2N and 4N cells contain 2 size categories of particles (115-121 A and 164-166 A), whereas the protoplasmic fracture (PF) face of these membranes contain a single size category of particles (85-88 A). The distribution and size of the EF- and PF-face particles were found to be similar among membranes from cells of the 3 ploidy levels. Analysis of the polypeptide composition of thylakoid membranes from 1N, 2N and 4N cells revealed no difference in the relative proportion of the constituent polypeptides of these membranes. The possible factors involved in the regulation of the development of thylakoid structure and composition in the presence of altered nuclear genome size are discussed.

Ricinus communis↗
PubMed1982

The motive force of the migrating pseudoplasmodium of Dictyostelium discoideum under dark and light conditions.

As a contribution to the understanding of the mechanism of the collective motion of the cell mass in the migrating pseudoplasmodium its motive force has been determined. The measurements in the present paper were made by an original method using centrifugal force. The maximum value of the motive force of a migrating pseudoplasmodium in the dark was estimated as 0.6-1.6 dyn. This value is comparable to the motive force of protoplasmic streaming in Physarum polycephalum. In addition, the motive force in phototactic migration, which has not previously been estimated, was measured. Illumination from the front increased the motive force, whereas illumination from the back produced the opposite results. That is to say, illumination could directly influence the motive force. Furthermore, some interesting results were obtained by measuring the changes in the velocity of the pseudoplasmodia placed under four conditions (combinations of illuminated or non-illuminated, and parallel or antiparallel to the direction of the external force). These results suggest the necessity of taking into consideration the direct effect of light upon the motility of the cells in the migrating pseudoplasmodium.

Cell Movement↗
PubMed1983

Immunocytochemistry of the acellular slime mould Physarum polycephalum. I. Preparation, morphology, and reliability of results concerning cytoplasmic actomyosin patterns in sandwiched plasmodia.

Small phaneroplasmodia of Physarum polycephalum migrate, under sandwich conditions between two agar sheets and a membrane of cellophane, as thin protoplasmic sheets. This method suitably simulates the situation in the natural habitat of acellular slime moulds; i.e. the narrow clefts of the forest soil. The highly differentiated system of cytoplasmic fibrils displayed under these conditions survives both long-term extraction with glycerol and fixation with methanol, procedures that remove the strong inherent autofluorescence, thus allowing the use of immunocytochemical studies. The complicated fibrillar system of sandwiched plasmodia consists of: (1) a membrane-associated cortical filament layer in the anterior region; (2) a more or less regular polygonal fibrillar network in the intermediate region; and (3) a helically twisted fibrillar system encircling endoplasmic pathways as well as isolated strands in the posterior region. So far, three different cytoskeletal proteins have been identified immunocytochemically as constituents of the fibrillar structures: actin, myosin and AM-protein (fragmin). No positive identification of alpha-actinin, filamin and tropomyosin was obtained using antibodies against vertebrate proteins. Electron microscopy of glycerol-extracted specimens treated with antibodies against actin and myosin revealed that the 6 nm filaments consist of actin, whereas the electron-dense material between single actin filaments appears to be myosin. The AM-protein modulating the polymer status of actin is located in all fibrillar structures.

Actins↗
PubMed1985

Nifedipine-sensitive calcium channels are involved in polar growth of lily pollen tubes.

Pollen germination and tube growth of Lilium longiflorum in vitro are affected by 10(-5) M-nifedipine. Germinating 'tubes' form broad protuberances along the whole colpus. Short tubes show a high tendency to grow 'amoeboid-like' and to branch; or a second tube emerges in another region of the colpus. Longer tubes (greater than or equal to 100 micron) broaden irregularly or swell at their tips. The diameter of the tube can vary drastically within the same tube. With increasing time of treatment many tubes burst. Normal tip growth stops within 10 min, but protoplasmic streaming continues even after 15 h. More or less regularly, wall thickenings are formed along the whole tube or on the flanks of the germinating region after some hours. The internal calcium gradient, visualized by chlorotetracycline (CTC) fluorescence, is also disturbed. Nifedipine treatment results in uniform or irregular CTC fluorescence. Branching tubes temporarily show a new subapical CTC gradient. After 6-8 h of nifedipine treatment many cells are no longer stainable with CTC. The results indicate the presence of nifedipine-sensitive calcium channels in pollen tubes.

Calcium↗
PubMed1986

Dynamic rearrangements of erythrocyte membrane internal architecture induced by infection with Plasmodium falciparum.

Cultured human erythrocytes infected with Plasmodium falciparum were studied by freeze-fracture electron microscopy. Special emphasis was placed upon the formation of the membrane surface excrescences ('knobs') found on red cells containing mature parasites. Knobs were visualized as conoid projections of the protoplasmic fracture face (PF) and depressions of the exoplasmic fracture face (EF). Knob formation was correlated with parasite growth and, on the basis of the organization of intramembranous particles (IMP) in the PF leaflet, a series of changes associated with parasite maturation was discerned: (1) a focal IMP cluster with minimal erythrocyte membrane elevation; (2) an elevated central IMP cluster surrounded by an IMP-free zone and concentric IMP ring; (3) maximal erythrocyte membrane deformation, concomitant with a loss of obvious IMP organization. Subtle changes in PF IMP organization were seen with knob formation and parasite maturation, including an apparent lateral partitioning of endogenous red cell membrane proteins between knobby or knob-free membrane areas in trophozoite-infected cells. IMP size distributions of the PF were shifted toward smaller particles in schizont-infected cells. Parasite development did not affect IMP densities in the PF; however, a decrease from 464 +/- 106 micron-2 to 374 +/- 94 micron-2 was seen in the EF of schizont-infected cells. IMP densities were similar over knobs and knob-free areas of either membrane leaflet, and there was no apparent EF IMP reorganization associated with the presence of knobs. These findings indicate that dynamic membrane changes are associated with knob formation and parasite maturation.

Cells, Cultured↗
PubMed1995

Analysis of the membrane structures involved in autophagy in yeast by freeze-replica method.

Under starvation conditions, the yeast S. cerevisiae sequesters its own cytoplasmic components by forming autophagosomes with double membrane in the cytoplasm. The autophagosome then fuses with the vacuolar membrane and delivers its own cytoplasmic components into the vacuole in the form of an autophagic body with a single membrane (Baba, M., Takeshige, K., Baba, N., and Ohsumi, Y. 1994. J. Cell Biol., 124: 903-913). We examined membrane structures involved in the autophagy induced by nitrogen-starvation by using freeze-replica method. The most conspicuous characteristic of the autophagic body is that the intramembrane particles were rarely detected on either the protoplasmic or exoplasmic face of its fracture membrane. This morphological feature of the fractured face was clearly different from other intracellular organelles. Next we examined the autophagosomal membrane. The inner membrane of autophagosome was also intramembrane particle-free, and its morphological feature was identical to the membrane of autophagic body. At the fusion site between autophagosome and vacuole we obtained direct evidence that two different membranes, the outer membrane of autophagosome and vacuolar membrane, became continuous by using freeze-etching technique. From these results we concluded that the autophagic body originated from the inner membrane of the autophagosome, and its membrane reflects an intrinsic feature of autophagosomal membrane. The outer membrane of autophagosome had only a few intramembrane particles and may be differentiated from the inner membrane. In cells under nitrogen-starvation condition, the density of intramembrane particles of vacuolar membrane decreased beyond that in control cells.

Autophagy↗
PubMed1983

Evidence for a vertical displacement of intramembranous particles on the plasma membrane of Tetrahymena cells by a local anesthetic.

We examined the effect of a local anesthetic, dibucaine, on the plasma membrane of Tetrahymena pyriformis strain NT-1 using freeze-fracture electron microscopy. Intramembranous particles (IMPs) were distributed homogeneously on the plasma membrane of untreated cells. But, when Tetrahymena cells had been treated with 1.3 mM dibucaine for 5 min at growth temperature, freeze-fracture micrographs of the plasma membrane showed marked alterations. Although IMPs showed an almost homogeneous distribution, their density was elevated markedly on the protoplasmic fracture (PF) face but greatly reduce on the exoplasmic fracture (EF) face. Areas around deciliated portions had a reverse IMP density distribution for the PF and EF faces. These results suggest that dibucaine induced vertical displacement of the IMPs in the plasma membrane.

Animals↗
PubMed1984

Effect of the local anesthetic dibucaine on the surface membrane in sterol-manipulated Tetrahymena pyriformis studied by freeze-fracture electron microscopy.

The effect of the local anesthetic dibucaine on the membrane ultrastructure of sterol-manipulated Tetrahymena pyriformis (NT-1 strain) was studied by freeze-fracture electron microscopy. Dibucaine-treated, ergosterol-replaced Tetrahymena cells had marked alterations in their plasma membranes. IMP-free small depressions (exoplasmic fracture face) and protrusions (protoplasmic fracture face) were formed on the plasma membranes which was in contact with the outer alveolar membrane. In addition, large IMP-free surface "blebs" covered with hexagonally-arranged depressions and protrusions appeared on both the plasma and outer alveolar membranes. These "blebs" were pinched off when the membranes were severely affected. Our previous study (28) demonstrated that the plasma membrane of dibucaine-treated native Tetrahymena cells that contain tetrahymanol showed vertical displacement of its intramembranous particles and that subsequently a smooth, flat surface appeared. Therefore, the structural changes in ergosterol-replaced membranes produced by dibucaine differ strikingly from changes in the native membranes. The remarkable difference in the ultrastructural deformation of the plasma membrane probably is due to a difference in the membrane lipid composition induced by sterol-manipulation.

Animals↗
PubMed1997

Neurotropic pyrimidine heterocyclic compounds. II. Effects of novel neurotropic pyrimidine derivatives on astrocytic morphological differentiation.

Astrocytes play an important role in supporting nerve regeneration after brain injuries. In this study, the effects of novel neurotropic synthesized pyrimidine compounds on astrocytic morphological differentiation were examined. Treatment of protoplasmic cultured astrocytes with 2-piperidino-5,6-dihydro-7-methyl-6-oxo(7 H)pyrrolo[2,3-d]pyrimidine maleate (MS-430) and the related compounds caused astrocytic process formation in 60 min. The morphology of MS-430-treated cells was similar to that of dibutyryl cAMP (DBcAMP)-treated cells. The astrocytic process formation by MS-430 was observed within 60 min and the maximum effect was obtained at the drug concentration of 0.5-1.0 mM. Rhodamine-phalloidin staining showed that astrocytic cytoskeletal actin was reorganized by MS-430 and DBcAMP. MS-430 did not increase cAMP accumulation in cultured astrocytes. These results suggest that the neurotropic pyrimidines induced astrocytic morphological differentiation through a cAMP-independent mechanism.

Animals↗
PubMed1997

Purification and properties of beta-fructofuranosidase from Clostridium perfringens.

beta-Fructofuranosidase [EC 3.2.1.26] in Clostridium perfringens was induced in the presence of sucrose and suppressed in the presence of glucose or maltose. The enzyme seems to be present in protoplasm in a soluble state. The beta-fructofuranosidase from C. perfringens cells grown on sucrose was purified by ammonium sulfate precipitation. DEAE-cellulose chromatography, Sephadex G-150 gel filtration, and hydroxylapatite chromatography to a homogeneous state. The molecular weight was 37,000 by gel filtration using Sephadex G-150 and by SDS-polyacrylamide gel electrophoresis. The amino acid composition is not much different from those of other microorganisms, but the Glx content was a little higher. The enzyme was inhibited by heavy metals, such as Hg2+, Cu2+, and Ag+, as well as pCMB; the activity was restored by incubating with mercaptoethanol. Fructose and amines including Tris and aniline had inhibitory effects.

Amino Acids↗
PubMed1990

Rules for distinguishing toxicants that cause type I and type II narcosis syndromes.

Narcosis is a nonspecific reversible state of arrested activity of protoplasmic structures caused by a wide variety of organic chemicals. The vast majority of industrial organic chemicals can be characterized by a baseline structure-toxicity relationship as developed for diverse aquatic organisms, using only the n-octanol/water partition coefficient as a descriptor. There are, however, many apparent narcotic chemicals that are more toxic than baseline narcosis predicts. Some of these chemicals have been distinguished as polar narcotics. Joint toxic theory and isobole diagrams were used to show that chemicals strictly additive with phenol were generally more toxic than predicted by narcosis I models and characterized by a different mode of action called narcosis II syndrome. This type of toxicity is exemplified by certain amides, amines, phenols, and nitrogen heterocycles. Evidence is provided that suggests that narcosis II syndrome may result from the presence of a strong hydrogen bonding group on the molecule, and narcosis I syndrome results from hydrophobic bonding of the chemical to enzymes and/or membranes. This shift in toxic action is apparently indistinguishable for narcotic chemicals with log P greater than about 2.7. General rules for selecting the appropriate models are proposed.

Animals↗
PubMed1991

Evaluation of enzyme-linked immunosorbent assay in comparison with complement fixation test for the diagnosis of subclinical paratuberculosis in cattle.

An enzyme-linked immunosorbent assay (ELISA) was evaluated and compared in parallel with the standard complement fixation test (CFT) for the diagnosis of bovine subclinical paratuberculosis. Bovine sera preabsorbed with the mixture of Mycobacterium phlei and kaolin suspension were assayed for antibody activities to the crude protoplasmic antigen of Mycobacterium paratuberculosis in the ELISA. ELISA antibody titer was expressed as ELISA antibody index (EAI) value: EAI = (At-An)/(Ap-An), where At, Ap and An are the absorbance values of a 1:200 dilution of unknown test sera, a 1:400 dilution of positive control serum, and a 1:200 dilution of negative control serum. An EAI of 0.6 or greater was established as a reasonable cutoff point for a positive antibody titer by ELISA. Of the 156 sera from cattle with subclinical M. paratuberculosis-infection, 106 (67.9%) were positive by ELISA and 41 (26.3%) by CFT. Of the 3,880 sera from cattle in the herds which had no history or evidence of paratuberculosis, 3,875 (99.9%) were negative by ELISA, and 3,787 (97.6%) by CFT. Positive ELISA titers were detectable 1 to 5 months earlier than positive CFT titers in experimentally infected cattle, and 7 to 10 months earlier in naturally infected cattle. These results indicate that the ELISA should replace the CFT as the routine test of choice for the diagnosis of bovine paratuberculosis.

Animals↗
PubMed2000

Malassezia pachydermatis isolated from a South American sea lion (Otaria byronia) with dermatitis.

A fungus was isolated from the skin of an Otaria byronia and from the water of the pool in which the animal was kept. It formed creamy colonies with soft texture on Dixon agar and grew well without supplements of long-chain fatty acids. Cells were ovoid to cylindrical in shape, budded from a broad base, and budded and divided at the same location. Thus, the isolate was identified as M. pachydermatis. We compared this very rare isolate from a marine mammal with four strains of M. pachydermatis using the freeze-etching electron-microscopy technique. The cells showed the same characteristic ring-swellings on the protoplasmic membrane on the neck site between the mother and the daughter parts, and the same accumulation of circumvallate bulgings in a small area near the straight sections of spiral grooves as four reference strains. Thus, in terms of morphology and ultrastructure, the isolate could be regarded as a typical M. pachydermatis.

Animals↗
PubMed2004

Differential expression of apolipoprotein d in human astroglial and oligodendroglial cells.

Apolipoprotein D (Apo D) is a secreted lipocalin in the nervous system that may be related to processes of reinnervation and regeneration. Under normal conditions, Apo D is present in the central nervous system in oligodendrocytes, astrocytes, and some scattered neurons. To elucidate the regional and cellular distribution of Apo D in normal human brain, we performed double immunohistochemistry for glial fibrillary acidic protein (GFAP) and Apo D in samples of postmortem human cerebral and cerebellar cortices. Most of the GFAP-positive cells in the gray matter had features of protoplasmic astrocytes and were mainly Apo D-positive. Apo D staining was mostly confined to the cell soma and proximal processes, whereas GFAP extended to a rich and extensive array of processes. The fibrous astrocytes in the white matter were immunoreactive for GFAP but not for Apo D. In the white matter, Apo D was mainly detected in oligodendrocytes and extracellularly in the neuropil. The results of the present study support a specific behavior for each astrocyte type. These findings suggest that Apo D expression may be cell-specific, depending on the particular tissue physiology at the time of examination.

Adult↗
PubMed2003

Research of the entry of rare earth elements Eu3+ and La3+ into plant cell.

Whether rare earth elements can enter into plant cells remains controversial. This article discusses the ultracellular structural localization of lanthanum (La(3+)) and europium (Eu(3+)) in the intact plant cells fed by rare earth elements Eu(3+) and La(3+). Eu-TTA fluorescence analysis of the plasmalemma, cytoplast, and mitochondria showed that Eu(3+) fluorescence intensities in such structures significantly increased. Eu(3+) can directly enter or be carried by the artificial ion carrier A23187 into plant cells through the calcium ion (Ca(2+)) channel and then partially resume the synthesis of amaranthin in the Amaranthus caudatus growing in the dark. Locations of rare earth elements La(3+) and Eu(3+) in all kinds of components of cytoplasmatic organelles were determined with transmission electron microscope, scanning electron microscope, and energy-dispersive X-ray microanalysis. The results of energy-dispersive X-ray microanalysis indicated that Eu(3+) and La(3+) can be absorbed into plant cells and bind to the membranes of protoplasm, chloroplast, mitochondrion, cytoplast, and karyon. These results provide experimental evidence that rare earth elements can be absorbed into plant cells, which would be the basis for interpreting physiological and biochemical effects of rare earth elements on plant cells.

Cations↗
PubMed1976

Ultrastructural studies of frozen-thawed 8-cell mouse embryos.

Survival of frozen 8-cell mouse embryos transferred directly upon thawing to the uteri of Day 3 pseudopregnant foster mothers was significantly lower (26%) than the survival of unfrozen 8-cell embryos transferred immediately after collection (73%). When frozen-thawed 8-cell embryos were cultured for 20-24 hr before transfer survival was similar to that of unfrozen 8-cell embryos transferred after 20-24 hr inculture (65% and 73%, respectively). Ultrastructural examination of the frozen-thawed 8-cell embryos revealed no obvious damage to protoplasmic components.

Animals↗
PubMed1983

Spermiogram and sperm reserves in hybrid Bos indicus X Bos taurus bulls after scrotal insulation.

Scrotal insulation for 48 h raised subcutaneous scrotal temperature by 4 degrees C in hybrid Bos indicus X Bos taurus bulls. The incidence of decapitated spermatozoa in the ejaculate increased significantly between 6 and 14 days and that of protoplasmic droplets and tail abnormalities between 20 and 23 days after insulation, respectively. Simultaneously, the percentages of spermatozoa with lost and damaged acrosomes increased significantly 12-17 days after insulation. At slaughter 23 days after scrotal insulation sperm production rates and gonadal reserves had not been affected by insulation, but epididymal reserves were markedly reduced, particularly in the cauda. Elevated testicular temperature therefore had an effect on immature spermatozoa in the caput epididymidis and on spermatids, but it is suggested that selective sperm resorption in the rete testis and excurrent ducts may prevent some of these changes being expressed in the ejaculate.

Animals↗