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PubMed1987

Sodium deprivation growth failure in the rat: alterations in tissue composition and fluid spaces.

Dietary control of sodium intake was utilized in weanling rats to study the relationships among body growth, tissue composition and extracellular fluid volume (ECFV). Forty 3-wk-old rats were divided into groups receiving 30, 150, 300, 600 or 900 mu eq sodium/d for 5 wk. The minimal daily requirement for normal growth was 300 mu eq Na, or about 60 mu eq/g of new growth. Lower doses caused dose-related growth failure associated with a reduced ECFV. Analyses of carcass, muscle and bone composition were carried out. In sodium-deprived animals there was retarded growth of protoplasm, fat and bone; the mineral composition of muscle was not altered, whereas in bone calcium concentration was reduced. Plasma concentrations of sodium, potassium and chloride remained normal. Pair-feeding indicated that sodium-deficiency growth retardation could not be attributed to starvation. Sodium-deficient animals ingested a greater amount of food per gram of weight gain, possibly reflecting an increased energy expenditure. Sodium deprivation initially permitted protoplasmic growth to proceed at a rate disproportionate to that of the ECFV. Subsequently, both continued to grow at a reduced but similar rate, suggesting that ECFV may be a controller of protoplasmic growth.

Animals↗
PubMed2000

Direct measurement of aluminum uptake and distribution in single cells of Chara corallina.

Quantitative information on the uptake and distribution of Al at the cellular level is required to understand mechanisms of Al toxicity, but direct measurement of uptake across the plasma membrane has remained elusive. We measured rates of Al transport across membranes in single cells of Chara corallina using the rare (26)Al isotope, an emerging technology (accelerator mass spectrometry), and a surgical technique for isolating subcellular compartments. Accumulation of Al in the cell wall dominated total uptake (71-318 microgram m(-2) min(-1)), although transport across the plasma membrane was detectable (71-540 ng m(-2) min(-1)) within 30 min of exposure. Transport across the tonoplast was initially negligible, but accelerated to rates approximating uptake across the plasma membrane. The avacuolate protoplasm showed signs of saturation after 60 min, but continued movement across the plasma membrane was supported by sequestration in the vacuole. Saturation of all compartments was observed after 12 to 24 h. Accumulation of Al in the cell wall reflected variation in [Al(3+)] induced by changes in Al supply or complexing ligands, but was unaffected by pH. In contrast, transport across the plasma membrane peaked at pH 4.3 and increased when [Al(3+)] was reduced by complexing ligands. Cold temperature (4 degrees C) reduced accumulation in the cell wall and protoplasm, whereas 2,4-dinitrophenol and m-chlorocarbonylcyanidephenyl hydrazone increased membrane transport by 12- to 13-fold. Our data suggest that the cell wall is the major site of Al accumulation. Nonetheless, membrane transport occurs within minutes of exposure and is supported by subsequent sequestration in the vacuole. The rapid delivery of Al to the protoplasm suggests that intracellular lesions may be possible.

2,4-Dinitrophenol↗
PubMed1972

Localization of crystals in diseased oats treated with uranyl acetate.

Uranyl acetate, a suppressor of victorin-induced electrolyte leakage in oat leaves when applied together with, or before, victorin, also suppressed victorin-induced changes in ultra-structure. Uranyl crystallized in cell walls and near the plasmalemma of vascular cells, but was excluded from the protoplasm. Fewer crystals occurred near the plasmalemma when leaves were allowed to take up uranyl and victorin simultaneously than when uranyl alone was absorbed, but deposition in cell walls was similar in the two treatments. No differences in crystal distribution were found in uranyl-treated leaves which subsequently took up either water or victorin. The most striking effect of prolonged exposure to uranyl was increased vesicular activity in the protoplasm, formation of complex concentric membranes, and tonoplast damage. Following victorin treatment, uranyl post-treatment was ineffective in suppressing electrolyte leakage or preserving normal cellular ultrastructure. More severe ultrastructural damage was found in victorintreated leaves after uranyl post-treatments than after post-treatment with water, a result of victorin-induced damage which facilitates uranyl entry into the protoplasm.

Journal Article↗
PubMed1982

Twenty-four-hour induction of freezing and drought tolerance in plumules of winter rye seedlings by desiccation stress at room temperature in the dark.

Exposure of seedlings of winter rye (Secale cereale L., cv. Puma) for 2 weeks or 24 hours to desiccation stress (40% relative humidity) at room temperature (21 degrees C) in the dark induced degrees of freezing and drought tolerance in the plumules comparable to those produced by cold conditioning for 2 weeks at 3 degrees C. The induction was associated with repression of growth and could not be produced in plumules excised from the seedlings indicating a requirement for translocation of nutrients from the endosperm. Rapid increase in osmotic pressure, soluble proteins, and phospholipids in plumules in association with the development of freezing and drought tolerance and the requirement of endosperm suggested diversion of nutrient from use in extension growth, to use in augmentation of protoplasm in plumule cells. Since cold acclimation slowed or arrested growth and is associated with augmentation of protoplasm, it is suggested that the common element in the induction of freezing tolerance by cold and drought is the necessity for producing a condition of augmented protoplasm and membranes in cells thus reinforcing a similar conclusion reached from seasonal studies on woody plants.

Journal Article↗
PubMed1994

Localization of metallothionein in aged human brain.

The localization of metallothionein (MT), a small molecular weight heavy metal binding protein in aged human brain, was investigated by immunohistochemical techniques. The amount of MT and heavy metals (Zn, Cu) were also assayed by radioimmunoassay and atomic absorption spectrophotometry, respectively. Immunohistochemically, MT was found in the pia mater, ependymal cells, protoplasmic astrocytes nand glial processes neuropil of the gray matter and fibrous astrocytes of the white matter of the telencephalon, whereas oligodendroglia and microglia did not show any positive immunostaining for MT. Cytoplasm, glial processes and some nuclei of protoplasmic and fibrous astrocytes showed strong MT immunostaining. Vascular feet and adventitia were also positive for MT immunostaining. Moreover, the pia mater, astrocytes and ependyma in the diencephalon, mesencephalon, pons, medulla oblongata and spinal cord showed the same positive immunostaining for MT as the telencephalon. In the cerebellum, Bergmann's glia, protoplasmic astrocytes of the granular layer and fibrous astrocytes of the white matter showed strongly positive immunostaining for MT. According to the radioimmunoassay, the amount of MT in the brain was relatively high at about 39.3 +/- 9.5 micrograms/g wet weight; of the heavy metals in the brain, zinc and copper were detected.

Age Factors↗
PubMed1998

Cellular events involved in survival of individual arbuscular mycorrhizal symbionts growing in the absence of the host

A survival strategy operating in the absence of the host was shown in obligately biotrophic arbuscular mycorrhizal (AM) symbionts. When no host-derived signals from the surrounding environment were perceived by germinating spores, fungal hyphae underwent a programmed growth arrest and resource reallocation, allowing long-term maintenance of viability and host infection capability. The early stages of mycelial growth of AM fungi were studied by a combination of time-lapse and video-enhanced light microscopy, image analysis, and immunodetection, with the aim of acquiring knowledge of cell events leading to the arrest of mycelial growth. The time-course of growth arrest was resolved by precisely timing the growth rate and magnitude of the mycelium originating from individual spores of Glomus caledonium. Extensive mycelial growth was observed during the first 15 days; thereafter, fungal hyphae showed retraction of protoplasm from the tips, with formation of retraction septa separating viable from empty hyphal segments. This active process involved migration of nuclei and cellular organelles and appeared to be functional in the ability of the fungus to survive in the absence of a host. Immunodetection of cytoskeletal proteins, metabolic activity, and the retention of infectivity of germinated spores confirmed the developmental data. The highest amounts of tubulins were detected when hyphal growth had ceased but when retraction of protoplasm was most active. This was consistent with the role of the cytoskeleton during protoplasm retraction. Succinate dehydrogenase activity in hyphae proximal to the mother spore was still detectable in 6-month-old mycelium, which remained viable and able to form appressoria and produce symbiotic structures.

Journal Article↗
PubMed1972

Isolation of the outer sheath of Leptospira and its immunogenic properties in hamsters.

Spherical forms of Leptospira interrogans serotype canicola Hond Utrecht IV were induced with 1 m NaCl. Electron micrographs of these salt-altered cells (SAC) revealed that the outer envelope or sheath had pulled away from the protoplasmic cylinder. Treatment of SAC with 0.02% sodium lauryl sulfate solubilized the sheath and released the protoplasmic cylinder. Further processing of the solubilized sheath yielded a pellet which displayed a membrane structure in electron micrographs. The released protoplasmic cylinder showed loss of intracellular organization and the outer envelope present in normal cells. Immunization of hamsters with whole formalized cells, SAC, or sheath in doses as low as 10 mug/animal protected them from death upon challenge with virulent canicola 27.

Animals↗
PubMed2002

Characterization of the leptospiral outer membrane and description of three novel leptospiral membrane proteins.

The outer membrane (OM) of the mammalian pathogen Leptospira kirschneri was isolated in the form of membrane vesicles by alkaline plasmolysis and separated from the protoplasmic cylinder by sucrose density gradient ultracentrifugation. All four components of the alkaline plasmolysis buffer, including 1.0 M NaCl, 27% sucrose (wt/vol), 2 mM EDTA, and 10 mM Tris (pH 9), were required for efficient OM release, as judged by recovery of leptospiral lipopolysaccharide. Two populations of OM vesicles (OMVs) were recovered, with peak concentrations found in the sucrose gradient at densities of 1.16 and 1.18 g/ml. Transmission electron microscopy revealed that the more buoyant OMV population was smaller (<0.1 micro m in diameter) than the denser OMV population (0.2 to 0.3 micro m in diameter). The densities of both populations of OMVs were distinct from that of the protoplasmic-cylinder material, which was found in the sucrose gradient at a density of 1.20 g/ml. The OMV fractions were free of protoplasmic-cylinder material, as judged by immunoblotting with antibodies to the endoflagellar sheath protein, heat shock protein GroEL, and two novel cytoplasmic membrane proteins, lipoprotein LipL31 and transmembrane protein ImpL63. The protein components of the OMVs were characterized by one- and two-dimensional immunoblotting and found to include previously described OM proteins (OMPs), including the porin OmpL1; the lipoproteins LipL32, LipL36, and LipL41; and the peripheral membrane protein P31(LipL45). A number of less well-characterized OMPs were also identified, including those with molecular masses of 16, 21, 21.5, 22, 31, 36, 44, 48, 90, and 116 kDa. The 48-kDa OMP was identified as a novel OM lipoprotein designated LipL48. The use of membrane-specific markers in OM isolation techniques facilitates an accurate description of the leptospiral OM and its components.

Amino Acid Sequence↗
PubMed1973

Ribosomes of acid-fast bacilli: immunogenicity, serology, and in vitro correlates of delayed hypersensitivity.

Ribosomal fractions obtained from Mycobacterium bovis (BCG) and M. smegmatis (strain butyricum) were studied to determine their antigenicity, their ability to stimulate the production of soluble mediators of delayed hypersensitivity (in vitro correlates) by sensitized peritoneal exudate cells, and the antigenic relations of ribosomal antigens of BCG to BCG protoplasm and H37Rv culture filtrates. The crude ribosomes and the 50-30S ribosomal subunit pool obtained from each of the organisms induced both delayed and immediate hypersensitivity when injected in incomplete Freund adjuvant into rabbits, and skin reactions could be elicited in sensitized rabbits with those antigens. The crude ribosomes and 50-30S ribosomal subunit pool of M. smegmatis stimulated lymphocytes of guinea pigs sensitized with viable organisms to produce macrophage migration inhibition factor. Comparable ribosomal fractions from BCG bacilli caused lymphocytes of guinea pigs sensitized with viable M. bovis (BCG) to produce skin reactive factor. Immunoelectrophoretic studies showed that H37Rv culture filtrate, protoplasm, crude ribosomes, and 50-30S ribosomal subunits of BCG contain multiple precipitinogens and that many of these were shared between the different antigen systems. Comparative electrophoresis revealed that BCG protoplasm and H37Rv culture filtrate shared a major portion of their components with each other and relatively few with ribosomal systems. The ribosomal systems shared the major portion of their components with each other and relatively few with the other antigen systems.

Animals↗
PubMed1963

MORPHOLOGY OF TREPONEMA MICRODENTIUM AS REVEALED BY ELECTRON MICROSCOPY OF ULTRATHIN SECTIONS.

Listgarten, M. A. (Harvard School of Dental Medicine and Forsyth Dental Infirmary, Boston, Mass.), W. J. Loesche, and S. S. Socransky. Morphology of Treponema microdentium as revealed by electron microscopy of ultrathin sections. J. Bacteriol. 85:932-939. 1963.-Broth cultures of a strain of Treponema microdentium were harvested on Millipore filters, fixed in osmic acid, and sectioned for electron microscopy. The sections revealed that the spirochetes had an axial filament, made up of two fibrils approximately 150 A in diameter, which was situated between an external envelope approximately 140 A in thickness and a protoplasmic cylinder. The protoplasmic cylinder had a cross-sectional diameter of 100 to 200 mmu, and was surrounded by a double "membrane" consisting of two 40-A electron-dense structures separated by a 45-A space. Cross-sections of spirochetal "granules" revealed that the limiting membrane was continuous with the outer envelope of the spirochetes, and surrounded the protoplasmic cylinder and axial filament.

Cross-Sectional Studies↗
PubMed1964

Ultrastructure of Treponema microdentium and Borrelia vincentii.

Bladen, Howard A. (National Institute of Dental Health, Bethesda, Md.), and Edward G. Hampp. Ultrastructure of Treponema microdentium and Borrelia vincentii. J. Bacteriol. 87:1180-1191.-A small oral Treponema (FM) and Borrelia vincentii (N9) were harvested after 3 to 7 days of incubation and either embedded in Vestopal W or negatively stained with phosphotungstate. The protoplasmic cylinders of both strains were identical except for size, and had a triple-structured cell wall as well as intracellular concentric laminations. Protoplasmic cylinders of both strains were enclosed in a cell envelope which appeared amorphous in negatively stained preparations, but which had a triple-structured wall when viewed in thin sections. The cell envelope of strain FM also acted as an envelope for the terminal filament; no filament envelope was evident in strain N9. Large structures which contained variable numbers of organisms and which were representative of spirochetal granules were observed. Protoplasmic cylinders contained within such granules frequently were devoid of cell envelopes. The axial filament consisted of several individual fibers which usually terminated in small end knobs. Occasionally, a fiber of the axial filament became a fiber of the terminal filament. Fibers of the terminal filament originated in end knobs similar to, but separate from, those to which the axial filament was attached. A periodicity of 60 A was occasionally observed in the terminal filament envelope of strain FM. A microperiodicity of approximately 20 A was also observed. The fibers of the terminal filament of strain N9 were composed of a large number of fibrils approximately 15 A wide. The periodicity and fibrillar structure of the terminal filament is discussed with reference to proposed models of bacterial flagella suggested by X-ray diffraction data.

Borrelia↗
PubMed1965

ELECTRON MICROSCOPY OF LEPTOSPIRES. I. ANATOMICAL FEATURES OF LEPTOSPIRA POMONA.

Ritchie, A. E. (National Animal Disease Laboratory, Ames, Iowa), and Herman C. Ellinghausen. Electron microscopy of leptospires. I. Anatomical features of Leptospira pomona. J. Bacteriol. 89:223-233. 1965.-Anatomical features of Leptospira pomona are described. Most cells in the logarithmic phase of growth, when examined in whole cell "negative-stained" preparations, display a homogeneous finely granular protoplasmic cylinder. Some cells contain discrete or complex lamellar structures. The cell termini are attached to the protoplasmic cylinder by a membrane system. Each organism appears to have two independent axial filaments originating from opposite ends of the cell. The axial filaments are composed of a closely knit bundle of some 12 to 15 individual strands. A simple model suggesting a locomotory function for the axial filaments is presented. A superficial callouslike mass is occasionally observed in the cell's terminal region. The enveloping sheath is similar to the cytoplasmic membrane; both are triple-layered and ca. 50 A thick. The sheath preferentially forms blebs at the cell termini and midway along the protoplasmic cylinder. Septal-wall formation occurs at the mid region of the cell, and it is concluded that simple transverse binary fission is the predominant mode of reproduction.

Animals↗
PubMed1968

Fine structure of Spirochaeta stenostrepta, a free-living, anaerobic spirochete.

The fine structure of Spirochaeta stenostrepta strain Z1, a free-living anaerobic spirochete, was studied by electron microscopy. The organism possessed a coiled protoplasmic cylinder, an axial filament inserted subterminally, and a loosely fitting sheath which enclosed both the protoplasmic cylinder and the axial filament. The axial filament consisted of two fibrils partially overlapping in a 1-2-1 arrangement. The axial fibrils appeared to possess a sheath surrounding an inner core. Both inner core and sheath were apparently enclosed in a cross-striated tubular structure, which was itself surrounded by an outer sheath. The axial filament exhibited a basal hook. A disc- or mushroom-shaped structure, possibly consisting in part of cytoplasmic membrane, was observed at the insertion end of isolated filaments. The protoplasmic cylinder had a distinctive surface structure consisting of an array of tightly packed, longitudinally arranged helices measuring 2.0 to 2.5 nm in diameter. This layer of helices lay below the outer cell sheath and the axial filament. Ballistic disintegration loosened the helical array, causing individual helices or segments of helices to become separated from the cell. The function of this layer of helices is still obscure.

Cytoplasm↗
PubMedDate not supplied

Metallothionein in developing human brain.

Localization of metallothionein (MT) in the developing human brain was investigated by immunohistochemical techniques. Fetal brain at 21 weeks showed no MT expression. In 35-week-old fetuses, glial cells in the gray matter showed MT expression in the nucleus and perinuclear cytoplasm, but glial cells in the white matter showed MT not only in the nucleus and perinuclear cytoplasm but also in glial processes. At 40 weeks, glial cells in gray and white matter expressed immunoreactive MT in the nucleus, cytoplasm and glial processes. Blood vessels were positive for MT. In the infant brain, there were clear differences in glial cells between protoplasmic and fibrous astrocytes; their nuclei, cytoplasm and processes were positive for MT, but the MT-positive glial processes of protoplasmic astrocytes were fine. In the child, typical protoplasmic astrocytes in gray matter and fibrous astrocytes in white matter were observed and their nuclei, cytoplasm and glial processes were positive for MT. Pia mater and blood vessels expressed MT in infants and children.

Adolescent↗
PubMed1975

Elemental balances during intravenous hyperalimentation of underweight adult subjects.

Intravenous hyperalimentation was done in 11 underweight adults whose body weight (body wt) was less than 85 percent of ideal. For the first 6 days, "complete formula" was infused furnishing per kilogram ideal body wt per day: 15 g glucose, 0.40 g N, 0.018 g P, 2.4 meq K, 3.0 meq Na, 2.3 meq C1, 0.5 meq Mg, 0.45 meq Ca, and 50 ml H20. Patients gained weight at an average rate of 9.0 g/kg ideal body wt/day and showed average balances/kilogram ideal body wt/day as follows: plus 0.14 g N; plus 0.012 g P; plus 0.43 meq K; plus 0.49 meq Na; plus 0.37 meq Cl; and plus 0.085 meq Ca. Application of standard equations to the elemental balances indicated weight gain consisted of 35-50 percent protoplasm, 35-50 percent extracellular fluid, 5-25 percent adipose tissus, and less than 1 percent bone. Withdrawas of N, P, Na, or K impaired or abolished retention of other elements. Removal of N halted retention P, K, Na and C1; withdrawal of K stopped retention of N and P; and removal of Na or P interrupted retention of all other elements. Weight gain continued at a rate of 1.4-3.1 g/kg ideal body wt/day despite zero or negative elemental balances of N, K, P, and sometimes Na and C1. Calculations showed that weight gain during infusion of fluids lacking N, P, K, or Na consisted largely of adipose tissue, with little or no contribution by protoplasm or extracellular fluid. Data show that repletion of protoplasm and extracellular fluid of wasted adults by intravenous hyperalimentation is retarded or abolished if N, P, Na, or K is lacking. Repletion of bone mineral does not occur in absence of Na or P but proceeds in absence of N, P, K, or Na. Thus, quality of weight gained by underfed adult patients during hyperalimentation depends on elemental composition of the infusate.

Adolescent↗
PubMed1996

Topographical distribution of phospholipids in boar sperm plasma and intracellular membranes as revealed by freeze-fracture cytochemistry.

We used fracture-label and label-fracture cytochemistry in conjunction with the phospholipase A2-colloidal gold (PLA2-CG) technique to study the distribution of phospholipids in ejaculated boar spermatozoa. These techniques provide visualization of the topographical distribution of phospholipids in freeze-fractured sperm membranes in a three-dimensional view. In various freeze-fractured boar sperm membranes and crossfractured cytoplasmic structures, quantitative analysis revealed that the nuclear envelope membranes and the nuclear content possessed the highest labeling density of PLA2-CG. Moderate labeling was detected over acrosomal membranes, especially the inner acrosomal membrane. Replicas of both protoplasmic and exoplasmic fracture faces of the plasma membrane of boar sperm head showed a relatively low density of PLA2-CG labeling. Moreover, a differential distribution of phospholipids was seen over the protoplasmic face of the plasma membrane domains of the sperm head, which showed the highest concentration of gold particles in the postacrosomal region, followed by the equatorial segment and the anterior acrosome region. The PLA2-CG labeling densities over the post-acrosomal region and the equatorial segment were significantly higher than that over the anterior acrosome region. In the flagellum, an intense labeling was also seen over crossfractured mitochondria, dense fibers, and fibrous sheath. The protoplasmic fracture face of the plasma membrane over the middle piece, the annulus, and the principal piece was moderately labeled by PLA2-CG. No significant difference in mean labeling density of PLA2-CG was detected among the three membrane domains. In label-fracture preparations, exoplasmic halves of the plasma membrane of the head and the middle piece of the tail were uniformly labeled with PLA2-CG. However, the annulus and principal piece were devoid of PLA2-CG binding sites. These results indicate that differential distribution of phospholipids associated with the boar sperm membranes may reflect phospholipid composition of membrane domains characteristic of special physiological functions.

Animals↗
PubMed1986

The intramembrane structure of septate junctions based on direct freezing.

Smooth septate junctions from the midgut of the cricket, Acheta, and the horseshoe crab, Limulus, as well as Hydra-type septate junctions from the epidermis of Hydra have been studied by freeze-fracture after direct freezing using the liquid helium-cooled copper block/slam freezing method. The exoplasmic fracture face at both types of septate junction exhibits rows of closely packed but irregularly shaped intramembrane particles. Complementary to these particle rows, on the protoplasmic fracture face, are sharply defined grooves with a periodic variation in depth and width that was conspicuous in Hydra but less well defined in arthropods. The closely packed, irregular particles on the exoplasmic faces could represent plastically deformed portions of transmembrane proteins pulled through the bilayer during freeze-fracture. On the basis of this interpretation, the grooves on the protoplasmic faces represent a confluence of the bilayer disruptions occurring during fracturing. The structures observed here are different from those reported in replicas of glutaraldehyde-fixed and glycerol-cryoprotected tissue, in which the intramembrane junctional components partition with the protoplasmic face and often assume the appearance of continuous cylinders. This comparison illustrates some of the artifacts associated with freeze-fracturing and shadowing. On the basis of a comparison of freeze-fracture replicas and sections of lanthanum-infiltrated tissues, the relationship between intramembrane junctional components and intercellular septal elements is analysed.

Animals↗
PubMed1986

Freeze-fracture analysis of thylakoid membranes and photosystem I and II enriched fractions from Phormidium laminosum.

Thylakoid membranes of the thermophilic cyanobacterium Phormidium laminosum have been fractionated into photosystem II and photosystem I particles. These fractions have been characterized by their partial electron transport activities, and biochemical and spectral properties. Exoplasmic fracture face and protoplasmic fracture face particles in the unfractionated thylakoid membranes were shown to correspond in size to particles in freeze-fractured photosystem II and photosystem I fractions, respectively. Differences between the histograms of the thylakoid membrane protoplasmic fracture face particles and the isolated photosystem I particles suggest that in addition to photosystem I complexes some of the particles on the thylakoid protoplasmic fracture face may be related to cytochrome b/f complexes, the hydrophobic component of the coupling factor, or respiratory complexes.

Cell Membrane↗