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  • Cell & Developmental Biology  (419)
  • 1995-1999
  • 1980-1984  (419)
  • 1935-1939
  • 1930-1934
  • 1981  (419)
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  • 1995-1999
  • 1980-1984  (419)
  • 1935-1939
  • 1930-1934
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  • 101
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 1 (1981), S. 237-245 
    ISSN: 0886-1544
    Keywords: centrioles ; symmetry ; triplet blades ; thermal fluctuations ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The paper suggests several principles of construction of a microscopically small device for locating the directions of signal sources in microscopic dimensions. It appears that the simplest and smallest device that is compatible with the scrambling influence of thermal fluctuations as are demonstrated by Brownian motion is a pair of cylinders oriented at right angles to each other. Nine equally spaced blades run in a pitched fashion along the mantle of each cylinder. The blades have a concave cross-section and bend around the circumference of the cylinder in a certain rotational pattern. Considering the striking similarity of this hypothetical device with centrioles, the paper puts forward the conjecture that centrioles locate the direction of hypothetical signals inside cells.
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  • 102
    Electronic Resource
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 1 (1981), S. 247-260 
    ISSN: 0886-1544
    Keywords: cilia ; trachea ; ATP-reactivation ; ciliary activity ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Evidence for active sliding of microtubules during ciliary activity has been demonstrated in a number of organisms: sea urchin sperm flagella, protozoan cilia, and mollusc gill cilia. Although there is evidence that active sliding also occurs in mammalian sperm flagella, there is little or no information on whether active sliding of microtubules also occurs in the short (5-μm) cilia of the mammalian trachea or oviduct. Since these cilia are important in tracheobronchial clearance and ovum transport, respectively, it has been important to demonstrate that microtubule sliding is also involved in the activity of somatic cilia. Ciliated apical portions (cortices) and cilia were isolated from rabbit trachea and oviduct, using Triton X-100 to demembranate the cilia. Most of the ciliated cortices reactivated upon addition of ATP, whereas isolated cilia reactivated to a lesser extent. When preparations of cilia were digested with trypsin before or after ATP addition, disintegration of axonemal doublets occurred with about the same frequency as reactivation. These events were recorded using Nomarski optics and dark-field microscopy. When isolated cilia which had been digested by trypsin and exposed to ATP were also prepared for electron microscopy by negative staining, telescoping of doublet microtubules from axonemes could be shown. These results demonstrate that mammalian somatic ciliary doublet microtubules actively slide in a manner similar to that described for invertebrate cilia.
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  • 103
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 1 (1981), S. 273-273 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 104
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 1 (1981) 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 105
    Electronic Resource
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 1 (1981), S. 371-385 
    ISSN: 0886-1544
    Keywords: rotating filaments ; cytoplasmic streaming ; Nitella ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Our knowledge about the actin-containing characean filaments on the basis of light and electron microscopical investigations is reviewed. Dynamic filamentous networks, known already from isolated droplets, were detected in Nitella rhizoidal cells using light microscopical techniques. Earlier light microscopic observations in cytoplasmic droplets are confirmed and complemented by new model experiments with rotating helices. The motile phenomena occurring at the filament bundles (ring formation, wave propagation, particle translocation, net dynamics, rolling motions, formation of side arms) can, in this way, be imitated in detail. Thus, the concept of cytoplasmic streaming as a translocation along bundles of rapidly rotating helical filaments is supported. In order to explain unidirectional cytoplasmic streaming, a periodic winding up and unwinding of fine filaments is postulated by which ions are periodically bound and displaced. The formation of side arms which is favored during unwinding results in a screw-mechanical different behavior of the filaments in the two directions of rotation and therefore causes permanent particle transport in one direction.
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  • 106
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 1 (1981), S. 387-397 
    ISSN: 0886-1544
    Keywords: birefringence ; polarizing microscope ; sea urchin egg ; cortex ; mitosis ; cleavage ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Birefringence (BR) at the cell surface of fertilized eggs of the sand-dollar, Clypeaster japonicus, during mitosis and cleavage was determined with a photoelectric BR detection apparatus [Hiramoto et al, 1981a]. The cortex of about 2 μm thickness is birefringent positive with respect to the normal to the cell surface. The hyaline layer is negatively birefringent. The halo-layer consisting of a row of microvilli surrounding the egg is positively birefringent in normal Ca-free sea water, while it is negatively birefringent in Ca-free sea water with high refractive index. The BR of the cortex gradually increases over the entire surface during mitosis until the onset of cleavage. The BR of the cortex at the polar region reaches a maximum shortly after the onset of cleavage and then decreases, while the BR of the cortex at the equatorial region begins to decrease shortly before the onset of cleavage, reaches a minimum shortly after the cleavage starts, and then increases again as the cleavage furrow advances. The coefficient of birefringence of the cortex is about 2.5 × 10-5 at the maximum. The BR change of the cortex during mitosis and cleavage is interpreted as a passive deformation caused by the constriction of the contractile ring as well as an active structural change of the cortex occurring in the dividing cell.
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  • 107
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 1 (1981), S. 399-416 
    ISSN: 0886-1544
    Keywords: myosin heavy chain ; avian muscular dystrophy ; adult and embryonic fast white fibers ; slow red fiber ; rod ; subfragment-1 ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Avian muscular dystrophy is characterized by the degeneration of fast white skeletal muscle fibers, with onset during development. Using a one-dimensional peptide mapping technique, we have detected two forms of the myosin heavy chain in the fast white fibers of adult domestic chickens, one form characteristic of birds homozygous for muscular dystrophy, the other of their normal controls. Four dystrophic strains carrying the same gene for muscular dystrophy were examined.No differences were detected in the embryonic heavy chain peptide maps of normal and dystrophic chickens, consistent with the developmental onset of the condition. Differences were also absent from the peptide maps of heavy chains from slow red fibers, which are unaffected in dystrophy. No dystrophy-specific peptide map differences were detected in the three light chains. Analysis of peptide maps of rod and the heavy chain component of subfragment-1 from normal and dystrophic heavy chains indicates the presence of amino acid sequence differences in the two proteins.
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  • 108
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    Cell Motility and the Cytoskeleton 1 (1981), S. 417-431 
    ISSN: 0886-1544
    Keywords: spindle poles ; centrioles ; cell center ; scaffold ; electron microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: I have used fluorescence microscopy and antibodies to 10nm filaments and tubulin labelled with contrasting fluorochromes to compare the distribution of these proteins in endothelial cells during cell division. During interphase the two filament systems have entirely different distributions: The bulk of the 10nm filaments form a ring that surrounds the cell center and nucleus and remains parallel to the substrate, while the microtubules radiate from the cell center to the cell's border. When the mitotic spindle replaces the radial microtubule pattern in mitosis, the spindle poles remain within - and in close proximity to - the ring of 10nm filaments. This was confirmed by electron microscopy which showed the ring and centrioles in the same plane separated by a distance of 300-400 nm.
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  • 109
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    Cell Motility and the Cytoskeleton 1 (1981), S. 303-327 
    ISSN: 0886-1544
    Keywords: cilia ; microtubules ; ATPase ; vanadate ; geometry of sliding ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A dynein arm attachment cycle produces sliding between adjacent doublet microtubules (N and N + 1) of cilia. In intact axonemes, in the absence of ATP, almost all arms appear attached at both ends (rigor). When ATP is added, most arms detach from doublet N + 1. In ATP and vanadate, the arms do not return to rigor, suggesting that ATP hydrolysis is required for re-extension and reattachment of the dynein arm, but not for detachment. Using solutions containing dynein to decorate dynein-less axonemal doublets, we confirm this interpretation. In the absence of ATP, both sides of each doublet decorate with arms. Addition of ATP, ATP and vanadate or AMP-PNP causes immediate arm detachment, but only in the first instance, where extensive ATP hydrolysis can occur, does decoration eventually reappear. Dynein decorates heterologous axonemal doublets and brain microtubules, as well as homologous doublets, suggesting that this mechanochemical cycle may have general applicability in microtubule-based cell motility.
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  • 110
    ISSN: 0886-1544
    Keywords: videomicroscopy ; differential interference microscopy ; streaming ; reticulopodial motility ; Allogromia ; microtubules ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A new method called Allen Video-enhanced Contrast, Differential Interference Contrast (AVEC-DIC) microscopy is shown to be sufficiently sensitive to detect several new features of microtubule-related motility in the reticulopodial network of the foraminifer, Allogromia. The method takes advantage of the variable gain and offset features of a binary video camera to operate the DIC microscope under conditions highly favorable for video imaging, but in which the optical image is virtually invisible to the eye yet retains its full information when viewed by a suitable video camera. The improvements are made possible by setting a dé Senarmont compensator to λ/9-λ/4 at maximal working aperture of internally corrected planapochromatic objectives. Under these conditions, the offset feature of the video camera can reject so much stray light from the instrument and specimen that contrast compares favorably with that observed in high-extinction images, and polarizing rectifiers offer scarcely any advantage. Freed from the constraints of the light-limited conditions of DIC microscopy, video images can be recorded 60 times per second, or over 1,000 times the rate of photomicrographs at comparable magnifications under high-extinction conditions.Application of this method to the reticulopodial network of Allogromia has shown that cytoplasmic organelles are translocated only in contact with single microtubules or bundles of microtubules, and that these organelles fail to move when separated from microtubules. Microtubules themselves undergo both axial translatory (“sliding”) and lateral “zipping and unzipping” movements that have been suggested to occur during mitosis and other biological processes.
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  • 111
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    Cell Motility and the Cytoskeleton 1 (1981), S. 445-454 
    ISSN: 0886-1544
    Keywords: taxol ; microtubules ; polymerization ; tubulin ; mitotic inhibitor ; protein self-assembly ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Dissociated bovine brain microtubule protein has been shown to reassemble at 0°C in the presence of the drug taxol. Tubulin polymerization was monitored both by electron microscopy of the polymeric structures and by incorporation of tritiated GTP into filterable polymeric structures. Most of the labeled guanine nucleotide uptake into tubulin polymeric structures occurred in the first 30 minutes of incubation with the drug. The initial polymerization event results in the formation of protofilamentous tubulin ribbons. The first microtubules were noted after 1 hour of incubation with the drug. After 20 hours of incubation at 0°C with taxol, the bulk of the polymerized tubulin appeared to be in the form of microtubules. Cold-stable tubulin rings with a mean diameter of 34 nm were present in the reaction mixture before the addition of taxol and throughout the 20-hour incubation. Most of the rings were apparantly not involved in the taxol-induced microtubule assembly. The results are consistant with a model whereby taxol induces an initial formation of protofilamentous ribbon structures, mostly from free tubulin dimers, and a slower subsequent folding of the ribbon structures into microtubules.
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  • 112
    ISSN: 0886-1544
    Keywords: videomicroscopy ; polarization microscopy ; streaming ; reticulopodial motility ; Allogromia ; microtubules ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A new method is described for recording rapid processes of cell motility in polarized light. The Allen video-enhanced contrast (AVEC-POL) method of polarization microscopy achieves significant improvements in resolution, contrast, and the visibility of fine detail by a combination of novel adjustments to a standard (unrectified) polarizing microscope and video camera. Using the full working aperture of a high-power planapochromatic objective lens and compensator setting of λ/9-λ/4, visible images appear lacking in contrast. However, the same images viewed with an appropriate video camera equipped with an electronic offset adjustment can be made to appear with as much contrast as desired, revealing a significantly greater amount of fine detail in the image than can be seen by high extinction visual microscopy alone. At bias retardations between one-ninth and one-quarter wave, the diffraction anomaly observed near extinction disappears. Consequently, polarizing rectifiers are not required with the AVEC-POL method, and images previously requiring photographic exposures of around 20 seconds are sufficiently bright to be registered on the video monitor in 1/60 second. Using an intensity monitor, quantitative measurements of cellular birefringence can be retrieved from live or videotaped images displaying a linear relationship between contrast and phase retardation due to birefringence. The AVEC-POL method also renders accessible to polarized light analysis a number of objects that scatter or depolarize too much light to be studied by high extinction methods. The method is demonstrated on model objects and applied to the highly motile reticulopodial network of Allogromia laticollaris. Rapid motion in close association with microtubules can now be analyzed in greater detail at a significant reduction in the cost of recording.
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  • 113
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 1 (1981), S. 329-347 
    ISSN: 0886-1544
    Keywords: actin ; microfilaments ; heavy meromyosin ; mammary gland ; secretion ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Cytochalasin B, a microfilament-altering drug, inhibits lactose synthesis in lactating guinea pig mammary gland [Biochim. Biophys. Acta 392:20, 1975] but not primarily by inhibiting glucose transport [Eur. J. Cell Biol. 20:150, 1979]. In order to study the possible role of microfilaments in lactose synthesis and secretion, we isolated both the alveolar (milk-secreting) and myoepithelial (contractile) cells from lactating mammary gland. Light microscopy shows that the alveolar cell fraction (viability approximately 71%) is homogenous and that the cells retain strong polarity of secretory structures in the apical region. Two proteins were extracted from the alveolar cell fraction. One (mol wt 42,000) comigrates with skeletal muscle actin on SDS-PAGE gels. The other, a high-molecular-weight (180,000) protein (HMWP) may be analogous to actin-binding protein or clathrin. An extract from the myoepithelial cell fraction also contains a protein that comigrates with actin but no HMWP. Whole tissue extract contains the 42K protein, and a 185K HMWP. Examination of the alveolar cell extract by electron microscopic (EM) negative staining revealed meshworks of multistranded, interconnecting filaments, with attached globular structures (100-200 A) (possibly the HMWP) and single filaments (40-60 A diameter) branching off. To localize these filamentous structures in situ, whole tissue was glycerinated and incubated with rabbit skeletal muscle heavy meromyosin (HMM). Masses of filaments in myoepithelial cells served as convenient standards for HMM decoration. Decorated filaments have cross-arms or projections, unlike the narrow, smooth filaments of control tissue. Decorated filaments in alveolar cells are located beneath the plasma membrane, in close association with secretory vacuoles, and near the Golgi apparatus; filaments near the latter two are often oriented perpendicular to the plasma membrane. Microvesicles are embedded in meshworks under the plasmalemma and near the Golgi apparatus. Intermediate-sized (85-115 A diameter), non-decorated filaments diverge from the meshworks of decorated filaments. Microvesicles are associated with intermediate-sized filaments as well. The association of actin-like filaments with secretory vacuoles and microvesicles and their location in areas of the cell concerned with biosynthetic activities suggest a possible function in the intracellular transport of secretory products.
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  • 114
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    Cell Motility and the Cytoskeleton 1 (1981), S. 433-443 
    ISSN: 0886-1544
    Keywords: Physarum ; acellular slime mold ; calcium ion ; calcium-ionophore ; cytoplasmic contraction ; oscillation ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Calcium is now generally thought to play a key role in regulating a variety of cellular movements. When the plasmodium of Physarum polycephalum was treated with the calcium-ionophore A23187 or the quasi-ionophore amphotericin B, Ca2+ leaked out. Ca2+ efflux into the ambient solution from the plasmodial strand segment was measured by the luminescence of a photoprotein aequorin, and the tensile force production was recorded simultaneously. Ca2+ efflux oscillated with the same period as the cycle of tension generation in the strand, but the phase of cyclic changes in Ca2+ efflux was opposite to that of tension generation. That is, Ca2+ efflux fell in the increasing tension phase and rose in the decreasing tension phase. Cyclic changes in efflux of Ca2+ are provisionally interpreted as reflecting corresponding changes in concentrations of free Ca2+ in the cytoplasm.
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  • 115
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    Cell Motility and the Cytoskeleton 1 (1981), S. 469-483 
    ISSN: 0886-1544
    Keywords: microtubules ; nucleation ; mitosis ; nocodazole ; immunocytochemistry ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The reassembly of microtubules is described in mitotic cells after release from nocodazole-induced block. The formation of microtubules was followed by light microscopic immunocytochemical staining using the PAP method, combined with to-luidine blue staining of the chromatin. The light microscopic observations on whole cells were compared with ultrastructural observations on thin sections. This step is essential to ascertain complete destruction of microtubules during the nocodazole treatment and to correlate immunocytochemical staining with the presence of microtubules.Removal of nocodazole (10 or 1 μg/ml) after a sufficiently long incubation to induce a complete disappearance of microtubules resulted in the appearance of tubulin staining specifically associated with the centromeres and with one or two isolated points in the cytoplasm. Electron microscopy confirmed that the staining was due to the massive accumulation of small microtubules at the kinetochores and centrosomes. Kinetochore nucleation was seen only in association with condensed metaphase-stage chromosomes and not with the less-condensed prophase chromosomes.In a second type of experiment cells were allowed to enter mitosis in the presence of an incompletely active concentration of nocodazole (0.1 μg/ml). The construction of the mitotic spindle was arrested; however, short microtubules were assembled at the kinetochores and centrosomes.These experiments demonstrate that in living mitotic PTK2 cells the kinetochores, as well as the centrosomes, exert a nucleating action on tubulin assembly.The further elongation of microtubules after removal of nocodazole was seen to occur preferentially along axes between the centrosomes and the kinetochores. This resulted in the construction of normal metaphases that evolved through anaphase and telophase. We have attempted to formulate a hypothesis that may explain the oriented assembly that seems to be essential in the construction of the spindle.
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  • 116
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    Journal of Morphology 167 (1981) 
    ISSN: 0362-2525
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 117
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    Journal of Morphology 167 (1981), S. 13-34 
    ISSN: 0362-2525
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The structure of the kidney of Leptonychotes weddelli was examined using corrosion casts, India ink injection, and histological methods. Some observations were made on the kidney of the crabeater seal (Lobodon carcinophagus) and the elephant seal (Mirounga leonina). The kidneys in all three species are reniculate, as in many other marine mammalian species. Features that have not been described previously in a phocid seal are a peripyramidal muscle, and venous drainage characterized by a large extrinsic system and a small intrinsic system.Examination of specialized fornices, relative medullary thickness, and the volumes of juxtamedullary relative to peripheral glomeruli (all of which relate to urine concentrating ability) revealed that each reniculus of Leptonychotes is similar to the unilobar kidney of a small mammal that produces only moderately concentrated urine. The high glomerular volume to cortical volume ratio may be related to high glomerular filtration rates after feeding observed in marine mammals.It is concluded that reniculation is more likely to be related to the large size of most marine mammals than to some factor related directly to the marine environment.
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  • 118
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    Journal of Morphology 167 (1981), S. 43-51 
    ISSN: 0362-2525
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The effects of cold-stress and hibernation on bone dynamics in the femurs of hamsters were investigated using histometric analyses. Control animals were maintained at 27° C for 90 days; experimental animals were kept at 5° C and hibernated for 7, 15, 21, 50, or 90 days. Histometric analyses of cross sections indicated that bone diameter and cortical thickness at the femoral midshaft increased after 83 days of extreme cold and 7 days of hibernation but decreased significantly after 69 days of cold stress and 21 days of hibernation. Osteoporosis was evident although the number of osteons per unit area of bone increased during hibernation. An initial decrease in the number of non-Haversian longitudinal vessels per unit area of bone was seen in experimental animals which was apparently related to a corresponding reduction in cortical thickness. Lacunar area increased in these animals, suggesting that osteocytic osteolysis may be a significant mechanism for calcium regulation during hibernation.
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  • 119
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    Journal of Morphology 167 (1981), S. 65-90 
    ISSN: 0362-2525
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Prenatal development of the eye in a microphthalmic hamster strain (“anophthalmic white”) is compared with established normal developmental periods. The mutant eye primordium is first distinguished at an average of ten gestational days (Period 6) by an incompletely invaginated optic cup, uniformly pseudostratified outer neuroepithelial layer and widely separated margins of the optic fissure. The outer layer of the mutant cup subsequently becomes abnormally thickened, especially posteriorly and midventrally, and, except in a few eyes with localized imperfect fusion, the optic fissure is unfused at twelve days (Period 9), by which time fusion is normally complete. At 13 to 15 days (Periods 10-11) the fissure is unfused or irregularly fused in regions of variable location and extent. The occurrence of fissure fusion with concomitant loss of continuity between inner and outer epithelial layers is generally restricted to expanded anterior regions in 14-16 day (Periods 11-12) eyes. The presence of presumptive neural retina in the outer layer of the cup characterizes the mutant eye; and to varying degrees, in day 13-16 eyes, the presumptive neural retina (1) provides persistent continuity between the two cup layers, (2) forms both fused and unfused margins of the optic fissure, and (3) extends into an outer position of the optic cup. As early as 13 days (Period 10), nerve fibers are present in the outer layer of the cup, and by the last prenatal and first postnatal days (Period 12), ectopic nerve fiber bundles are widely distributed.
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  • 120
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    Journal of Morphology 167 (1981), S. 139-165 
    ISSN: 0362-2525
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The new family Lobatocerebridae, Rieger, contains a group of turbellariomorph worms in the annelid line of evolution. The fine structural organization of the body wall, the digestive tract, and parts of the central and peripheral nervous system are described and the findings are discussed in light of general invertebrate cytology. The epidermis and gastrodermis contain a basal granule cell system which is structurally very similar to the neuroglia cell system of the nervous system. The continuity of the neuroglia cell system, and the epidermal basal granule cell system and the basal granule cell system in the digestive epithelia suggests the existence of a single glial-basal granule cell system, similar to the gliointerstitial cell system first recognized in the Mollusca (see Nicaise, '73). The Annelida may show a dual (ectodermal and mesodermal) origin of such a gliointerstitial cell system as suggested by similarities in the epidermal basal cell system in the Oligochaeta and of certain epidermal and gastrodermal cells in polychaete regeneration with neuroglia in the Annelida. The structural similarity of neuroglia and basal granule cells in Lobatocerebridae may be the result of similarity in the formation, maintenance, or regulation of the extracellular matrix.
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  • 121
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    Journal of Morphology 167 (1981), S. 201-209 
    ISSN: 0362-2525
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Eyes of early embryonic chicks possess 14 scleral papillae, derived from the conjuctival epithelium and present as transient structures between seven and 11 days of incubation. These papillae induce the formation of the 14 scleral ossicles, which develop in the adjacent, neural crest-derived ectomesenchyme. Each papilla undergoes a predictable series of developmental changes, divided by Murrary ('43) into six morphological stages (M stages 1-6). We have confirmed his staging, and provide a scanning electron microscopic (SEM) evaluation of papilla development. The earliest stage that can be visualized with the S.E.M. is M stage 2. We describe the initial modifications of the surface of papilla cells, the presence of large microvilli and the asymmetrical morphogenesis and growth of the papillae. Papillae are shed by a mechanism that involves elongation of the cells at the base of the papilla. Such moribund papillae consist of necrotic cells coated with fibers.
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  • 122
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    Journal of Morphology 167 (1981), S. 231-247 
    ISSN: 0362-2525
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The penetration of the sperm into the egg, and the movements of the male and female pronuclei were followed from sperm attachment through pronuclear fusion, using time-lapse video microscopy of gametes and zygotes of the sea urchin Lytechinus variegatus (23° C). The pronuclei move in four stages: I. Sperm Entry Phase, following sperm-egg fusion and a rapid radiating surface contraction (5.9 ± 1.3 μm/second) when egg microvilli engulf the sperm head, midpiece, and tail to form the fertilization cone and the sperm tail beats in the egg cytoplasm; II. Formation of the Sperm Aster, which pushes the male pronucleus centripetally at a rate of 4.9 ± 1.7 μm/minute starting 4.4 ± 0.5 minutes after sperm-egg fusion, as the male pronucleus undergoes chromatin decondensation; III. Movement of the Female Pronucleus, the greatest and fastest of the pronuclear motions at a rate of 14.6 ± 3.5 μm/minute at 6.8 ± 1.2 minute after sperm-egg fusion, which establishes the contact between the pronuclei; and IV. Centration of the Pronuclei to the egg center at a rate of 2.6 ± 0.9 μm/minute by 14.1 ± 2.6 minutes after sperm-egg fusion. Pronuclear fusion typically occurs after stage IV and proceeds rapidly starting 14.7 ± 3.6 minutes after sperm-egg fusion with the male pronucleus coalescing into the female pronucleus at a rate of 14.2 ± 2.6 μm/minute.
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  • 123
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    Journal of Morphology 167 (1981), S. 297-304 
    ISSN: 0362-2525
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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    Notes: Eggs of Chelydra serpentina were incubated at 30°C and 26°C. In addition, incubation was done at 20°C during the temperature-sensitive period for sex determination. Incubation at 20°C and 30°C resulted in females; incubation at 26°C resulted in males in 99% of the cases. The average gonadal length was less in the males. The average length of the 20°C ovaries did not vary significantly from that of the 30°C ovaries.The condition of the oviducts was correlated with histology of the gonads in hatchlings and in 3-month-old animals. When at least one of the oviducts was obvious and intact, ovaries were present. If the oviducts were absent or interrupted, testes were present. Histological characteristics of the gonads resulting from the three incubation temperatures are described. In the 26°C testes, cellular infiltrations occurred frequently. The ovaries of 20°C hatchlings tended to have a less developed germinal epithelium than that of the 30°C animals. Also, epithelial cysts occurred frequently in the 20°C ovaries. The incidence of follicles at 3 months was not differential.
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  • 124
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    Journal of Morphology 168 (1981) 
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  • 125
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    Journal of Morphology 167 (1981), S. 339-375 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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    Notes: The hemimandibles in carnivorans may be united in various ways at the symphysis menti. The symphysis may contain a readily flexible joint that permits a moderate amount of independent movement of the hemimandibles. This type of symphyseal union is primitive for and widely distributed in extant carnivorans. In other carnivorans, the symphysis is patent but allows slight or essentially no independent movement of the hemimandibles. Finally, the hemimandibles may be rigidly united by synostosis of the symphysis. The morphology, movement and, insofar as possible, function of these types of symphyses are described.
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  • 126
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    Journal of Morphology 168 (1981), S. 5-15 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The lung volume, the morphometrically determined alveolar and capillary surface area, and the capillary volume of 27 dogs (weight 2.65-57 kg) all were linearly correlated with body weight. The thickness of the air-blood barrier increased only slightly with increasing body size. The structural diffusing capacity, containing these parameters, was used to estimate the gas exchange capabilities of the lung and was also found to scale in direct proportion to body size. This coincides with reports on physiologically estimated diffusing capacity but is obviously different from the interspecies slope for metabolism which scales to the 3/4 power of body weight.
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  • 127
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    Journal of Morphology 168 (1981), S. 1-2 
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  • 128
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    Journal of Morphology 168 (1981), S. 43-49 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Topics: Biology , Medicine
    Notes: A realistic model of the distribution of the partially orientated capillaries in skeletal muscles has been introduced for stereological analyses. Distributional parameters not previously estimated for capillary networks in muscles have been quantified. These include the lengths of capillary per unit volume of tissue (Lv) and a dimensionless index of orientation (Ω). The present study demonstrates that surgical techniques for inducing skeletal muscle hypertrophy can be an effective stimulus for the proliferation of additional capillaries. In the hypertrophic muscles studied the capillaries become more highly orientated. This suggests that the growth of new capillaries occurs preferentially along the long axis of the muscle.
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  • 129
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    Journal of Morphology 168 (1981), S. 97-108 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Notes: Eggs of a number of cockroach species are parasitized by Tetrastichus hagenowii. The ultrastructure of the sensilla on the antennae of females and males was examined by scanning and transmission electron microscopy. The females have two types of multiporous plate sensilla while the males have only one. Type 1 is found in females and males and has a relatively thin cuticular wall and many pores, while type 2 is found only in females and has a relatively thick cuticular wall and few pores. Both sexes have nonporous, thick-walled, socketed hairs; multiporous, nonsocketed hairs; multiporous, thick-walled pegs; and terminal hairs. In addition, males have multiporous, nonsocketed, long hairs. The sensilla are similar, in many respects, to the sensilla of other chalcid parasitoids. The antennal sensilla of female T. hagenowii are probably involved in ovipositional behavior. The multiporous, long hairs of the male possibly receive stimuli during mating behavior A chemoreceptive function is proposed for the multiporous plate sensilla.
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    Journal of Morphology 168 (1981), S. 137-149 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Notes: The ultrastructure of hemopoietic bone marrow of the Spanish lizard, Lacerta hispanica, has been studied for the first time. The organ consists of a stroma formed by venous sinuses and reticular cells. Erythropoiesis takes place in the lumen of blood vessels, while granulopoiesis is extravascular. Pluripotent stem cells are structurally differentiated into erythrocytes and granulocytes. Two types of granulocytes, heterophils and acidophils, have been found, and a third granular cell type is tentatively identified as granular leukocyte. Remarkably, plasmacytopoiesis occurs in the bone marrow of Lacerta hispanica. The possible functional significance of these results is discussed with emphasis on their importance for the reptilian immune system.
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    Journal of Morphology 168 (1981), S. 181-187 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Notes: The cells of mandibular organs of female Libinia emarginata exhibit changes in substructure during molting and vitellogenesis. The cytoplasm is that of a steroid-hormone producing cell. The cells do not appear to produce ecdysones.
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    Journal of Morphology 167 (1981), S. 265-276 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Notes: Morphogenesis of the eye was studied in a new strain of micro-phthalmic rat. Abnormalities were noted immediately after the formation of the optic cup. The inner layer in the central part of the optic cup was relatively thick and contained many mitotic figures, whereas that of the marginal part was thin and contained only a few. The transitional point in the inner layer between the central and the marginal parts was well marked. This is evidently due to the extreme growth inhibition of the inner layer at the marginal part.At the early developmental stage, an area of the inner layer corresponding to the transitional point protruded toward the lens because the central part of the inner layer continued to differentiate.The differentiation and the protrusion of the inner layer proceeded variably at the later stages depending on the degree of the growth inhibition. The eyes were classified into three groups: Group A-the retina was recognized as a cyst consisting of the pigment layer and the pigment-layerlike structure which originated from the inner layer; group B-the neural retina and its layered structure were inverted; group C-abnormalities, such as the destruction of the lens, were observed.Although previous authors who studied eye mutants suggested the vascular abnormality as the primary cause of the production of abnormal eyes, we feel that this is not the case in our animals.
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    Journal of Morphology 167 (1981), S. 305-312 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Notes: The fine structure of Johnston's organ in the pine sawfly, Neodiprion sertifer, was studied by electron microscopy to determine if there exists a dimorphism in this organ corresponding to the sexual dimorphism in antennal shape and surface area.The organ is made up of scolopidia that are ultrastructurally similar to those of other insects. The scolopidia, identical in both sexes, comprise three sensory cells bearing two types of sensory processes: Two are shorter and smaller in diameter than the third, which extends into the cuticle of the membrane connecting pedicel and flagellum and terminates at an epicuticular invagination. The dendrites and sensory processes are surrounded by two types of enveloping (glial) cells-a scolopale cell and an attachment cell. Other enveloping cells occur at different levels of the scolopidium.Sexual dimorphism is evident only in the numbers of scolopidial groups: Males have more groups with fewer scolopidia, but both sexes possess about the same total number of scolopidia.
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    Journal of Morphology 167 (1981), S. 333-337 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Notes: A polytrophic ovariole of the flour moth, Ephestia kuhniella, is composed of a linear series of increasingly mature egg chambers, each consisting of an oocyte, an interconnected cluster of seven nurse cells, and a covering layer of follicle cells. This study describes changes in the volume of each component as a function of the position of the egg chamber in the ovariole. Analysis of the growth curve of the Ephestia oocyte yields two possible correlations between accelerated oocyte growth and ultrastructural events enhancing the supply of yolk materials to the oocyte: the first is the initiation of yolk synthesis by the follicle cell layer and its transfer to the oocyte, and the second is the formation of channels between the follicle cells allowing hemolymph to gain access to the oocyte. An Ephestia oocyte increases in volume from approximately 2.5 × 103 μm3 to approximately 2.0 × 107 μm3 over an average series of 58 egg chambers.
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    Journal of Morphology 168 (1981), S. 3-4 
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    Journal of Morphology 168 (1981), S. 17-42 
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    Source: Wiley InterScience Backfile Collection 1832-2000
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    Notes: Embryonic development of the head of Oxyrhachis tarandus (Membracidae) has been investigated in detail to settle the controversy of head segmentation and to refute the occurrence of an intercalary segment. The head is formed from six distinct elements: the prostominal lobe, the paired cephalic lobes, the antennal segment and the three noncontroversial gnathal segments. The prostomial lobe, which possesses a neuromere and a pair of coelomic cavities, represents the first body segment, called the prostomial segment. The tritocerebral lobes of the brain and the stomatogastric nervous system, consisting of a frontal ganglion, clypeolabral nerves, and the recurrent nerve etc., develop from the neuromere of the prostomial lobe. The tritocerebrum thus belongs to the prostomial segment rather than to an imaginary intercalary segment and mainly represents the ganglionic center of the stomatogastric nervous system in the brain. Frons, clypeus, and labrum develop from the outer wall of the prostomial lobulate plate, whereas the epipharyngeal wall, including the cibarial pump, develops from its inner wall. The presence of three coelomic cavities and of three distinct neural masses in the cephalic lobes during the initial stages of development shows that they have developed by the fusion of three distinct segments during the long phylogenetic history of insects. The portion of the germ band presently considered as the intercalary segment is actually the sternal part of the antennal segment. The neural cells located in this region give rise to the deutocerebrum by shifting forward, around the stomodaeum, and always leaving a commissure behind. The intercalary segment is thus a complete illusion. The antennal segment is postoral in the beginning and bears a pair of coelomic cavities, but later on it shifts forward and its sternal part invaginates into the stomodaeum.
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    Journal of Morphology 169 (1981), S. 1-19 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Notes: Hands of two-toed sloths (Choloepus) are long, narrow, hook-like apparatuses with only two functional digits (II and III); rays I and IV are represented only by metacarpals. The proximal phalanges of digits II and III are shortened to essentially proximal and distal articulating surfaces, and all but distal interphalangeal joints of these digits are restricted by interlocking surfaces to minimal ranges of flexion and extension. Several intercarpal joints and the wrist joint, however, allow wide ranges of movement in several axes. Wide excursion at the wrist is permitted by an extremely lax joint capsule, the manner of insertion of several prime movers of the carpus, and the reduced participation of the ulna in the wrist joint. Several extrinsic digital muscles, particularly extensors, are absent and others have unusual actions. Intrinsic musculature consists primarily of mm. interossei and m. extensor digitorum brevis, although other, inconstant muscles do occur. Hands of Choloepus are used as flexible hooks on supports less than 52 mm in diameter and as fixed grapnels on larger supports. In both cases, distal phalanges (and covering claws) form the “hook” element. Whereas bare volar pads seem to be adjunctive on supports smaller than 52 mm in diameter, they are essential on those larger than 65 mm. Two-toed sloths may prefer supports 50 mm in diameter or smaller. The potential importance of vines as supports is discussed.
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    Journal of Morphology 169 (1981), S. 113-140 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Notes: A comparative study of limb morphology indicates that the osteological and myological differences between Didelphis virginiana, the Virginia opossum, and Chironectes minimus, the water opossum, may be associated in Chironectes with decreased resistance to water and increased mechanical advantage of its muscles for increased force. Limb myology is described and a synonymy of terms is applied to the musculature of these two opossums.
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    Journal of Morphology 169 (1981), S. 161-183 
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    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The larval chondrocranium of the pelobatoid anuran Pelodytes punctatus was studied, using Alcian blue-alizarin stained and cleared whole preparations, serial sections, and gross dissections. This beaked tadpole is an unspecialized pond dweller and its chondrocranium closely resembles those of other ecologically similar tadpoles of diverse systematic relationships. Detailed analysis, however, shows many differences among the chondrocrania of anuran larvae. Among other features, these include the configurations of suprarostrals, fronto-parietal fenestrae, palatoquadrate suspensoria, the ligamentum or cartilago tectum of the muscular process of the quadrate and the circumoral ligaments. The lateral circumoral ligament permits differentiation of beaked discoglossoidean and ranoidean larvae. Microhyloids conform to the ranoidean pattern in this feature. Pipoids either lack it or seem to conform to the discoglossoidean pattern. Use of these larval features as key characters enables assignment of Pelodytes to an uncertain position among pelobatoid frogs. This is totally congruent with previous assignments based on adult features and is used to support the hypothesis that anuran phylogenies based on larval characters will closely resemble, in major features at least, those based solely on adult characters.
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    Journal of Morphology 169 (1981), S. 243-251 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The site and process of urine formation in the renopericardial system of Helisoma have been investigated. Osmotic pressure and protein content of hemolymph from the heart, pericardial fluid from the pericardial cavity, prourine from the kidney sac, and urine from the ureter have been determined. Osmotic pressure is equal in hemolymph, pericardial fluid, and prourine, but less in urine. Protein content is similar in hemolymph and pericardial fluid, but much less in prourine and urine. Hemoglobin molecules are present in hemolymph and pericardial fluid but not in prourine. It is suggested that in Helisoma the kidney sac is the site of prourine formation, and prourine is an ultrafiltrate of hemolymph. The kidney epithelial cells contain 6- to 7-nm microfilaments which react with heavy meromyosin producing unidirectional arrowheads. Numerous actin filaments are present in the vicinity of the lateral cell membranes and basal processes. It is possible that the actin filaments regulate the extracellular spaces for prourine passage. It is postulated that the actin-rich kidney epithelium may generate hydrostatic pressure for ultrafiltration. Na+-K+ ATPase is located on the luminal side of the kidney epithelium, which may regulate intracellular fluid level of the kidney epithelial cells, and thereby regulate their cell volume. Thus Na+-K+ ATPase may be involved in the regulation of extracellular spaces in kidney epithelial cells. The enzyme may participate in the production of hyposmotic urine.
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    Journal of Morphology 169 (1981), S. 259-274 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The cytoarchitecture and neuromorphology of the torus semicircularis in the tokay gecko, Gekko gecko, were examined in Nissl-stained, fiber-stained, and Golgi-impregnated tissues.From a superficial position, the torus semicircularis extends rostrally under the caudal half of the optic tectum. Caudally, the two tori abut upon one another; rostrally, they diverge. The torus semicircularis consists of central, laminar, and superficial nuclei.The central nucleus consists of fusiform, spherical and triangular neurons. Their dendrites are highly branched, with numerous dendritic spines, and are oriented mediolaterally, dorsoventrally, and rostrocaudally. Fusiform and spherical neurons display two dendritic patterns: “single axis,” ramifying in one axis, and “dual axis,” exhibiting higher-order branches perpendicular to the primary dendrites. Triangular neurons exhibit a “radiate” dendritic pattern.In the rostral half of the torus semicircularis, the laminar nucleus caps the central nucleus. The laminar nucleus encircles the central nucleus in the caudal torus semicircularis. The neurons of the laminar nucleus have dendritic arrays oriented parallel to the border of the central nucleus. These dendrites exhibit a paucity of dendritic spines and higher-order branches. Fusiform and spherical neurons exhibit “single axis” and “dual axis” dendritic patterns. Triangular neurons display “radiate” patterns.The caudal superficial nucleus lies dorsal and dorsolateral to the central nucleus. The superficial nucleus is sparsely populated by small fusiform and spherical neurons with moderately branched dendrites and moderate numbers of dendritic spines. These neurons display “single axis” (fusiform neurons) as well as “dual axis” and “radiate” (spherical neurons) dendritic patterns. They are oriented either parallel to or perpendicular to the boundary of the laminar nucleus.
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    Journal of Morphology 169 (1981), S. 357-357 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Journal of Morphology 170 (1981), S. 43-54 
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    Notes: Major features of interest in the mature chondrocranium of Agama pallida are striking curvature of the nasal region, lack of the paranasal cartilage and concha nasalis, and presence of a cartilaginous roof over Jacobson's organ. In addition, the course of the ethmoid nerve deviates from the normal lacertilian pattern; there is no foramen epiphaniale, and the temporal region is reduced. The prefacial commissure and facial foramen lie in front of the cochlear portion of the auditory capsule, whereas the prominentia semicircularis anterior is separated from the rest of the otic capsule. Several chondrocranial characters are suggested to be unique to the agamids.
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    Journal of Cellular Physiology 106 (1981), S. 173-178 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Proteins of approximately 10,000 daltons (presumably metallothionein) and greater than 75,000 daltons bound 64Cu when this metal was added to fibroblast lysates. Treatment with either 2-mercaptoethanol or the disodium salt of ethylenediamine tetraacetic acid demonstrated that the high molecular weight copper-binding proteins in lysates prepared from both normal and Menkes fibroblasts exhibited a relatively low affinity for copper compared to the 10,000 dalton protein(s). No difference was detected in the affinity of the low molecular weight protein(s) of normal and Menkes fibroblast lysates for copper. The amount of 64Cu bound to the 10,000 dalton protein(s), however, was approximately two to three times greater in lysates prepared from Menkes fibroblasts than from normal fibroblasts. Mixing experiments indicated that the increased binding of 64Cu to the 10,000 dalton protein(s) in lysates of Menkes fibroblasts did not result from the deficiency of a factor that effects the cleavage of copper from this protein(s), from the presence of a soluble inhibitor, or from the lack of an activator. In addition, the use of lysates, rather than whole cells, demonstrated that the observed differences in copper binding between the normal and the Menkes fibroblasts were not caused by an abnormality in the membrane transport of copper in the mutant cells. Thus the findings suggest that the increased accumulation and the reduced efflux of copper previously observed in cultured Menkes fibroblasts result either from an increased amount of the 10,000 dalton copper-binding protein(s) or from an increased capacity of this molecule(s) for copper.
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    Journal of Cellular Physiology 106 (1981), S. 209-213 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Fusion of mononucleate myoblasts to form multinucleated myotubes increases when skeletal muscle cells are grown in progressively higher oxygen concentrations (5%, 20%, and 40% oxygen). At four days of growth fusion of myoblasts (as expressed by the percent of all muscle nuclei that are located in myotubes) is 57 ± 2% in 5% oxygen, 68 ± 1% in 20% oxygen, and 78 ± 2% in 40% oxygen (P〈0.001). However, at a concentration of 40%, oxygen depresses the rate of cell division and thereby affects the number of myoblasts available for fusion. Thus, oxygen concentration significantly modifies growth of skeletal muscle in vitro. Its net effect on myotube formation results from the interaction of its separate effects to enhance cell fusion and to depress cell proliferation.
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    Journal of Cellular Physiology 106 (1981), S. 225-234 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Primary cultures of bone cells and skin fibroblasts were examined for their Ca++ content, intracellular distribution and Ca++ fluxes. Kinetic analysis of 45Ca++ efflux curves indicated the presence of three exchangeable Ca++ compartments which turned over at different rates: a “very fast turnover” (S1), a “fast turnover” (S2), and a “slow turnover” Ca++ pool (S3). S1 was taken to represent extracellular membrane-bound Ca++, S2 represented cytosolic Ca++, and S3 was taken to represent Ca++ sequestered in some intracellular organelles, probably the mitochondria. Bone cells contained about twice the amount of Ca++ as compared with cultured fibroblasts. Most of this extra Ca++ was localized in the “slow turnover” intracellular Ca++ pool (S3). Serum activation caused the following changes in the amount, distribution, and fluxes of Ca++: (1) In both types of cells serum caused an increase in the amount of Ca++ in the “very fast turnover” Ca++ pool, and an increase in the rate constant of 45Ca++ efflux from this pool, indicating a decrease in the strength of Ca++ binding to ligands on cell membranes. (2) In fibroblasts, serum activation also caused a marked decrease in the content of Ca++ in the “slow turnover” Ca++ pool (S3), an increase in the rates of Ca++ efflux from the cells to the medium, and from S3 to S2, as well as a decrease in the rate of influx into S3. (3) In bone cells the amount of Ca++ in S3 remained high in “serum activated” cells, the rate of efflux from S3 to S2 increased, and the rate of influx into S3 also increased. The rate of efflux from the cells to the medium did not change. The results suggest specific properties of bone cells with regard to cell Ca++ presumably connected with their differentiation. Following serum activation we investigated the time course of changes in the amount of exchangeable Ca++ in bone cells and fibroblasts, in parallel with measurements of 3H-thymidine incorporation and cell numbers. Serum activation caused a rapid decrease in the content of cell Ca++ which was followed by a biphasic increase lasting until cell division.
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  • 148
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    Journal of Cellular Physiology 106 (1981), S. 269-277 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have shown that collagen gel can be used as a culture matrix for the cloning of granulocyte/macrophage progenitor cells (CFU-C), the production of foci of marrow stromal cells and the maintenance of stem cell proliferation, differentiation and the production of CFU-C. Since collagen is a physiological matrix and allows the simultaneous growth of a variety of cellular elements, the system should prove useful for examining the role of cell/cell interactions and regulatory molecules involved in haemopoiesis.
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  • 149
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    Journal of Cellular Physiology 106 (1981), S. 283-291 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Results of hemacytometer cell counts and of tyrosinase measurements made by the Pomerantz method demonstrate that imidazole added to the medium of cultured B 16 mouse melanoma cells can stimulate tyrosinase specific activity and inhibit cell division. These effects are greater than with adenosine 3′,5′ cyclic monophosphate (cAMP) or the cAMP-phosphodiesterase inhibitor theophylline. The effects of imidazole on cell division and tyrosinase are enhanced by theophylline and antagonized by cAMP. Cyclic AMP-phosphodiesterase activity in cell-free extracts can be inhibited by theophyllne and stimulated by imidazole. However, imidazole does not affect cAMP-phosphodiesterase specific activity in vivo, nor does it affect intracellular cAMP concentrations as determined by competitive protein-binding assays. In contrast, the specific activity of cAMP-phosphodiesterase in vivo is stimulated by cAMP and theophylline, supporting the hypothesis that cAMP and agents which increase intracellular cAMP concentrations induce the synthesis of cAMP-phosphodiesterase. Studies with actinomycin-D and cycloheximide support the hypothesis that cAMP can also mediate posttranslational activation of tyrosinase. Similar experiments suggest that imidazole, or a derivative therof, can induce the synthesis of tyrosinase at the pretranslational level of control. We hypothesize that this type of regulation (pretranslational) by imidazole may define a role for the concept of “Metabolite Gene Regulation” (MGR), in mammalian cells.
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  • 150
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    Journal of Cellular Physiology 106 (1981), S. 399-406 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The effects of valinomycin (25 pM) on the membrane potential and on initial, passive Na+ and K+ movements have been determined in Ehrlich ascites tumor cells. The membrane potential of steady-state cells in a physiologic environment was - 23.2 mV. Addition of valinomycin induced a small, significant hyperpolarization (Vm = -29.6 mV) when averaged over the population tested. However, analyses of the response of individual cells to valinomycin showed two different potential effects: (1) the majority of cells hyperpolarized after treatment; but (2) a significant fraction depolarized when exposed to valinomycin. The Vm of steady-state cells incubated in saline with K+ at concentrations of 21 mM or 75 mM was - 21.4 mV and -22.0 mV, respectively. Addition of valinomycin to these cells was without effect on Vm, thus establishing the “null point” responses. Only for cells incubated in saline with a K+ of 75 mM was there agreement between Vm and K+ equilibrium potential (Vk). Determinations of cellular Na+ and K+ showed that valinomycin induced net losses of K+ and gains of Na+ by cells incubated in either physiologic saline or saline with a K+ concentration of 21 mM. However, the celular K+ of cells incubated in saline with a K+ concentration of 75 mM was unaltered by valinomycin. There was a two- to threefold increase in K+ permeability of the cell membrane in the presence of valinomycin. These results are consistent with the existence of two null points in the membrane-potential response to valinomycin: One is established when the membrane potential corresponds to Vk; the second occurs when the effects of valinomycin on K+ loss from the cell are exactly offset by its inhibition of active Na+ + K+ transport.
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  • 151
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    Journal of Cellular Physiology 106 (1981), S. 425-434 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The mechanism of glucose entry into human vascular endothelial cells was studied in monolayer cultures of normal (primary) and virally (SV40) transformed umbilical vein endothelium. Radioisotopic uptake studies with the glucose analogues 2-deoxy-D-glucose, and 3-O-methyl-D-glucose, and the nonmetabolizable stereoisomer L-glucose, indicated the presence of a saturable, stereospecific hexose carrier mechanism in both cell types. In other experiments with D-glucose and 3-O-methyl-D-glucose, the phenomenon of countertransport was demonstrable. Hexose transport was not affected by KCN, dinitrophenol, or ouabain, but was inhibited by phloretin and phlorizin in a pattern consistent with facilitated diffusion. Kinetic constants were obtained for both 2-deoxy-D-glucose and 3-O-methyl-D-glucose uptake. Similar Km values (range, 3.3-4.7 mM) were noted with normal and transformed cells, whereas the apparent Vmax was 0.56 nmol/μ1 cytosol/minute for primary cells and 1.7-2.5 nmol/μ cytosol/minute for transformed cells. Under standard culture conditions, as well as following 18 hours of serum deprivation, insulin at concentrations up to 10-5 M did not appear to influence hexose uptake in either cell type. Metabolism of 14C(U)-D-glucose to 14CO2 also was not stimulated by insulin. The presence of an insulin-insensitive, facilitated transport system for glucose in vascular endothelium has relevance for glucose metabolism in this tissue, and potentially for the association of certain vascular diseases (e.g., diabetic microangiopathy, atherosclerosis) with altered glucose homeostasis.
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  • 152
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    Journal of Cellular Physiology 106 (1981), S. 435-444 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Activity and accuracy of chromatin-directed DNA replication have been compared in young and aged Mus musculus and Peromyscus leucopus, two murine species with contrasting maximum lifespans. Chromatin isolated from livers of mature adults of both species copied efficiently exogenous DNA templates using predominantly DNA polymerase-β. The DNA synthetic activity of liver chromatin remained constant in both species throughout their lifetimes. The fidelity of chromatin-directed poly [d(A-T)] synthesis was similar for the comparatively short-lived M. musculus and the relatively long-lived P. leucopus and remained unaltered in old animals. The fidelity of poly [d(A-T)] copying catalyzed by DNA polymerase-β dissociated from liver chromatin was comparable to that of the chromatin-directed synthesis. The dissociated enzymes did not exhibit diminished fidelity of poly [d(A-T)] synthesis with age. In all ages of both species examined, the murine liver DNA polymerase-β, both chromatin-associated and solubilized, exhibited high error frequencies; approximately one dGMP was incorporated for every 500-1,000 complementary nucleotides polymerized. The relationship of these results to the accuracy of DNA replication and repair as a determinant of aging is considered.
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  • 153
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    Journal of Cellular Physiology 106 (1981), S. 419-424 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Exposure of murine leukemia cells in culture to bis-acetyl-diaminopentane (BADP) caused erythroid maturation as measured by the accumulation of hemoglobin in treated cells. The appearance of differentiated cells in cultures exposed to BADP occurred 18 to 20 hours earlier than in those treated with dimethylsulfoxide (DMSO), a standard inducer of differentiation in this system. Studies with [3H]BADP indicated the occurrence of relatively rapid association of the inducer with cells, and subsequent linear accumulation. Fractionation of cellular components and measurement of radioactivity from BADP therein demonstrated that this agent preferentially associates with a fraction enriched for plasma membrane. In addition, [3H]BADP was capable of binding to the plasma membrane-enriched fraction isolated from murine erythroleukemia cells as measured by gel filtration. These findings support the concept that interaction of inducers of murine erythroleukemia differentiation such as BADP with components of the surface membrane may be important in the cascade of events that lead to the erythroid maturation of these leukemic cells.
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  • 154
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    Journal of Cellular Physiology 107 (1981) 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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  • 155
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    Journal of Cellular Physiology 107 (1981), S. 11-19 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Primary cultures of parenchymal cells isolated from adult rat liver by a collagenase perfusion procedure and maintained as a monolayer in a serum-free culture medium were used to study glucoeogenesis and the role that the glucocorticoids play in the control of this pathway. These cells carried out gluconeogenesis from three-carbon precursors (alanine and lactate) in response to glucagon and dexamethasone added alone or in combination. Maximum glucose production was observed with cells pretreated for several hours with dexamethasone and glucagon prior to addition of substrate and glucagon (8- to 12-fold increase over basal glucose production). Half-maximum stimulation of gluconeogenesis was seen with 3.6 × 10-10 M glucagon and 3.6 × 10-8 M dexamethasone. Maximum stimulation was oberved with 10-7 M glucagon and 10-6 M dexamethasone. The length of time of dexamethasone pretreatment was found to be important in demonstrating the effect of glucocorticoids on glucagon-stimulated gluconeogenesis. Treeatment of cells with dexamethasone for 2 hours did not result in an increase in glucose production over identical experimental conditions in the absence of dexamethasone, wherease pretreatment for 5 hours (1.2-fold increase) or 15 hours (1.7-fold increase) did result in an increase in glucose production. The results establish that the adult rat liver parenchymal cells in primary culture are a valid model system to study hepatic gluconeogenesis. In addition, we have established directly that the glucocorticoids amplify the glucagon stimulation of gluconeogenesis.
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  • 156
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    Journal of Cellular Physiology 107 (1981), S. 41-46 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: An intraperitoneal injection of the beta-adrenergic drug dl-isoproterenol hydrochloride (100 mg/kg body weight) into male (190-210 g) albino rats caused two cyclic AMP surges (peaking at 10 minutes and again between 8 and 12 hours) and the initiation of DNA synthesis (between 16 and 20 hours) in the parotid glands. The parotid cells in hypocalcemic thyroparathyroidectomized rats still responded to isoproterenol injection by generating the two cyclic AMP surges, but they did not initiate DNA synthesis unless a blood calcium-elevating combination of parathyroid hormone (50 USP units/100 g of body weight) and 1α,25 (OH)2 vitamin D3(200 pmoles/100 g of body weight) was injected along with the isoproterenol.
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  • 157
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    Journal of Cellular Physiology 107 (1981), S. 59-67 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The potent tumor promoter 12-O-tetradecanoyl-phorbol-13-acetate (TPA) can stimulate quiescent, nonproliferating 3T3 cells to reenter the cell cycle and divide. We have previously used a slection technique developed in our laboratory to isolate variant cell lines which no longer divide in response to epidermal growth factor. We have now utilized the same selection procedure to isolate, from 3T3 cells, two variant cell lines, TNR-2 and TNR-9, which retain growth control and divide in response to elevated serum or fibroblast growth factor, but which do not respond to TPA. The variants do not incorporate precursors into DNA in response to TPA, demonstrating that the cells do not enter the S phase of the cell cycle. The TPA nonresponsive variant TNR-2 cannot respond to epidermal growth factor; TNR-9 responds to this mitogen. TNR-2 variant cells, which do not respond to EGF, do not bind 125I-EGF. TPA can modulate 125I-EGF binding to TNR-9 cells in a manner similar to its action on parental 3T3 cells. This TPA-induced alteration of EGF binding indicates that TNR-9 cells still interact with TPA, despite their inability to mount a mitogenic response.
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  • 158
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    Journal of Cellular Physiology 107 (1981), S. 85-100 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: In an investigation of the changes that occur in cultured neoplastic cells as they outgrow their supply of nutrient, MM96 human melanoma cells were found to diminish in size and to proliferate more slowly. These changes were accompanied by a moderate increase in the proportion of cells with a G1-like DNA content. When replated under favorable conditions, many of these cells gradually resumed active proliferation. Continuing adverse culture conditions led to a continued fall in cell size, loss of reproductive viabllity, and finally to rapid cell death. Simultaneous buoyant-density and velocity-sedimentation-fractionation experiments showed that cells from exponential cultures were moderately dense and rapidly sedimenting, cells from postexponential cultures were less dense and much more slowly sedimenting, and dye-excluding cells from reproductively nonviable, late postexponential cultures were of widely variable though generally high density, and were moderately rapidly sedimenting. Although neither fractionation method resulted in significant enrichment of clonogenic cells, depletion was seen at both extremes of both types of profile. Cells fractionated by velocity were sorted according to DNA content and hence location in the cell cycle. The relationship between sedimentation rate and cell-cycle location was reflected in the continuous thymidine labeling patterns of the separated cells. Study of these patterns suggested that cycle durations lengthened as crowding increased and nutrient became depleted, and shortened upon reseeding at low density into fresh medium.
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  • 159
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    Journal of Cellular Physiology 107 (1981), S. 171-183 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The production and localization of laminin, as a function of cell density (sparse versus confluent cultures) and growth stage (actively growing versus resting cultures), has been compared on the cell surfaces of cultured vascular and corneal endothelial cells. Comparison of the abilities of the two types of cells to secrete laminin and fibronectin into their incubation medium reveals that vascular endothelial cells can secrete 20-fold as much laminin as can corneal endothelial cells. In contrast, both cell types produce comparable amounts of fibronectin. Furthermore, if one compares the secretion of laminin and fibronectin as a function of cell growth, it appears that the laminin released into the medium by either vascular or corneal endothelial cells, is a function of cell density and cell growth, since this release is most pronounced when the cells are sparse and actively growing, and decreases by 10- and 30-fold, respectively, when either vascular or corneal endothelial cell cultures become confluent. With regard to fibronectin secretion, no such variation can be seen with vascular endothelial cell cultures, regardless of whether they are sparse and actively growing or confluent and resting. Corneal endothelial cell cultures, demonstrated a twofold increase in fibronectin production when they were confluent and resting as compared to when they were sparse and actively growing. When the distribution of laminin versus fibronectin within the apical and basal cell surfaces of cultured corneal and vascular endothelial cells is compared, one can observe that unlike fibronectin, which in sparse and subconfluent cultures can be seen to be associated with both the apical cell surface. In confluent cultures, laminin can be found associated primarily with the extracellular matrix beneath the cell monolayer, where it codistributes with type IV collagen.
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  • 160
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    Journal of Cellular Physiology 107 (1981), S. 209-217 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Conditions have been established under which the antizyme of ornithine decarboxylase (E.C. 4.1.1.17, L-ornithine carboxy-lyase, ODC) a noncompetitive protein inhibitor of ODC, can be detected in cells in response to as little as 10-7 M putrescine. The maintenance of intracellular antizyme activity depends upon the continued presence of putrescine in the medium. Removal of putrescine results in a rapid decline of antizyme activity. These phenomena are unaffected by the presence of cycloheximide and are comparable to the requirement of L-asparagine for the maintenance of ODC activity.The extent to which the antizyme level is increased is inversely related to the preexisting level of intracellular ODC at the time of addition of putrescine. The time of appearance of free antizyme is delayed in cells that have high levels of ODC; the amount of free antizyme that can be assayed for in these cells, at any particular time is correspondingly less. The converse is also true. In cells that have high levels of antizyme, the delay in appearance of ODC is greater and the amount of ODC that can be assayed for is correspondingly less than in cells with low levels of antizyme.These experiments, as well as others, indicate that the ODC antizyme and ODC interact in vitro with each other to modify their respective activities.
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  • 161
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    Journal of Cellular Physiology 107 (1981), S. 243-249 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Resting Yoshida AH130 hepatoma cells, harvested at the plateau of tumor development in vivo, were recruited into the cycling state following transfer to an in vitro system whereby these cells were incubated in the autologous ascites plasma diluted with buffered saline and enriched with glucose. In this system, cell recruitment into the phase of DNA synthesis (S phase) strictly depends on the activity of the respiratory chain and is abolished by anaerobiosis as well as by antimycin A, although the intracellular levels of ATP and the rate of protein synthesis are practically unaffected by these treatments. Furthermore, 2,4-dinitrophenol, at concentrations which uncouple the respiratory phosphorylation and hence enhance both glycolysis and oxygen consumption, does not hinder cell promotion into S phase. Thus, the absolute respiration dependence of cycling resumption by resting ascites cells does not seem to rely on respiratory ATP supply, but rather is linked to the electron flow through the respiratory chain.
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  • 162
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    Journal of Cellular Physiology 107 (1981), S. 271-281 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The synthesis and turnover of sulfate-labeled glycosaminoglycans (35S-GAGs) has been investigated in diploid human embryo fibroblasts during in vitro cellular aging. With progressive subcultivation, there was a decreased incorporation of Na235SO4 into 35S-GAGs released to the medium, but not into those accumulated at the cell surface. The composition of 35S-GAGs found in extracellular medium, cell surface (removable by gentle proteolysis), and intracellular compartments of the culture after 48-hr labeling did not change significantly with progressive subcultivation. Pulse-labeled 35S-GAGs moved from intracellular to surface and extracellular compartments more slowly in late-passage cultures. Addition of 1 mM β-xyloside to both early- and late-passage cultures produced a ten-fold enhancement of extracellular 35S-GAG production without a concomitant increase in surface-associated 35S-GAG. We interpret the data of this study to mean that secreted and cell-surface glycosaminoglycans represent different pools and that cellular aging has its effect primarily upon the secreted pool of glycosaminoglycans. Late-passage fibroblasts demonstrate marked decreases in proliferation, culture density, fibronectin matrix, and gap-junction formation. Our results suggest that glycosaminoglycan synthesis and composition are not intimately related to these parameters.
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  • 163
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    Journal of Cellular Physiology 107 (1981) 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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  • 164
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    Journal of Cellular Physiology 107 (1981), S. 317-327 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Conditioned serum-free medium (CSFM) obtained from WI-38 human fibroblasts was found to contain a mitogenic factor(s) with somatomedin (SM)-like activity. Treatment of the cells with cycloheximide eliminated the SM-like activity in CSFM, suggesting that these cells produce and release the activity. Gel filtration revealed that the fibroblast SM-like activity (FSLA) had a molecular size near 45,000. Isoelectric focusing of this FSLA yielded 2 bands of SM activity with pIs of 4.7 and 6.1, and corresponding molecular sizes of ∼29,000 and 16,500, respectively. The FSLA obtained by gel filtration revealed parallel dose response curves with a basic SM in a SM radioreceptor and radioimmunoassay and stimulated: (1) 35So4 uptake by hypophysectomized rat cartilage; (2) (U-14C) glucose oxidation is isolated rat adipocytes; and (3) (3H) thymidine uptake and cell division in these same WI-38 fibroblasts. Out studies indicate that this FSLA and basic SM are similar but not identical.
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  • 165
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    Journal of Cellular Physiology 107 (1981), S. 345-358 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have shown previously (D.A. Sirbasku, 1978, Proc. Natl. Acad. Sci. U.S.A., 75:3786-3790) that an estrogen-inducible growth factor activity for rat mammary and rat pituitary tumor cells can be identified in extracts of rat uteri, although at the time of that report only a limited biochemical characterization of the activity was presented. In this report, we have evaluated the growth factor activity for lipid, steroid hormone or protein-like properties. Uterine growth factor activity was assayed by measure of the increased cell number of the MTW9/PL rat mammary tumor cell line established by this laboratory and described previously (D.A. Sirbasku, 1978, Cancer Res. 38:1154-1165). Studies showed the following characteristics of growth factor activity: destroyed by trypsin treatment; labile when heated at 80°C; partially denatured by 6 M guanidine or 8 M urea treatment or 50% aqueous solutions of organic solvents; inactivated by extremes of pH or overnight treatment with mild acid; not dialyzable at neutral pH; of apparent molecular weight of 70,000 daltons by G-100 Sephadex chromatography; possessing an isoelectric point of 4.8 to 5.2; not chloroform/methanol extractable; and not in any way identified as either a lipid or a steroid hormone. The data available suggest that the uterine growth factor activity is a protein or polypeptide of apparent high molecular weight, and that this activity does not directly correspond to other known growth factors.
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  • 166
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    Journal of Cellular Physiology 107 (1981), S. 371-378 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Chick embryo fibroblasts were maintained at confluency for up to 35 days in medium containing 0.5% or 0.75% fetal bovine serum or 2.5% or 5.0% horse serum. At weekly intervals cells were subcultured and serially propagated in medium containing 10% FBS until their replicative lifespans were completed. The results showed that the replicative lifespan of embryonic chick fibroblasts was dependent on the cumulative number of population doublings undergone by the culture and was not related to the calendar time cells were in culture. Further characterization of 0.75% FBS maintained chick cells returned to 10% FBS medium showed that cells had protein contents and incorporated 3H-thymidine into DNA at a rate that resembled that of young cells, despite an advanced chronological age.
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    Journal of Cellular Physiology 106 (1981), S. 309-319 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We describe the isolation and characterization of a Chinese hamster ovary cell line selected for resistance to N-carbamoyloxyurea. Using the mammalian cell permeabilization assay developed in our laboratory, a detailed analysis of the target enzyme, ribonucleotide reductase (EC 1.17.4.1), was carried out. Both drug-resistant and parental wild-type cells required the same optimum conditions for enzyme activity. The Ki values for N-carbamoyloxyurea inhibition of CDP reduction were 2.0 mM for NCR-30A cells and 2.3 mM for wild-type cells, while the Ki value for ADP reduction was 2.3 mM for both cell lines. Although the Ki values remained essentially unchanged, the Vmax values for NCR-30A cells were 1.01 nmoles dCDP formed/5 × 106 cells/hour and 1.83 nmoles dADP/5 × 106 cells/hour, while those for the wild-type cells were 0.49 nmoles dCDP produced/5 × 106 cells/hour and 1.00 nmoles dADP/5 × 106 cells/hour. This approximate twofold increase in reductase activity at least partially accounts for a 2.6-fold increase in D10 value for cellular resistance to N-carbamoyloxyurea exhibited by NCR-30A cells. The NCR-30A cell line was also cross-resistant to the antitumor agents, hydroxyurea and guanazole. No differences in Ki values for inhibition of CDP and ADP reduction by these two drugs were detected and cellular resistance could be entirely accounted for by the elevation in activity of the reductase in the NCR-30A cell line. The properties of N-carbamoyloxyurea-resistance cells indicate they should be useful for further investigations into the regulation of mammalian enzyme activity.
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  • 168
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    Journal of Cellular Physiology 106 (1981), S. 349-360 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: L5178Y/TK+/- cells treated with methyl methanesulfonate (MMS) were allowed to recover for 0,48,96,144, or 240 hours, and were then plated in soft-agar medium containing trifluorothymidine (TFT). Dose-dependent and consistent increases in the frequency of TFTR cells were observed after each of the 48-240-hour expression periods through the counting of predominantly large, mutant colonies. Size distributions of soft-agar colonies from either MMS-treated or control cells were bimodal in the presence, and unimodal in the absence, of TFT. An increase of small, presumptive TFTR colonies with either increasing MMS concentration or decreasing recovery time was probably a manifestation of chemical toxicity, for a similar increase in small-colony number was observed in the absence of TFT when cells were cloned immediately after MMS treatment, when no induced mutants were yet detectable. Recloning experiments with 22 small-colony-derived cell lines revealed that, with one exception, small-colony morphology was not a heritable trait. While all large- and some small-colony-derived stocks from MMS-treated cells were of the phenotypically stable TK-/- type; spontaneous small TFTR colonies generally were not, their occurrence being directly correlated with serum concentration. No aneuploidy was evident in MMS-treated cell lines several generations after isolation as small TFTR colonies. These results suggest that delayed MMS cytotoxicity in TK+/- cells can temporarily produce increased physiological resistance to TFT in some cells, giving rise to secondary populations of small-colony TFTR variants.
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  • 169
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Topics: Biology , Medicine
    Notes: We have examined the effect of alpha-methylmannoside (α-MM) addition to concanavalin A (con A)-stimulated peripheral human lymphocytes. With a previously established kinetic model, we have, from the time course of proliferation, extracted the responding clone size, rate of entry of this clone into S phase, and the length of the lag period. We have studied the effect of con A dose and time of addition of α-MM to optimally stimulated cells on these kinetic parameters. We show that neither a low dose of con A nor an early addition of --MM to optimally stimulated cells results in a change in the responding clone size. That is, all of the potentially responsive cells appear to become “committed” to enter the cell cycle regardless of the presence of α-MM early in the culture or in the presence of suboptimal stimulation. However, the rate at which these committed cells enter the first S phase is a function of the dose of con A and time of addition of α-MM and varies over a wide range. It is the variation in this parameter that accounts for virtually all of the diminished response previously interpreted as a time-dependent irreversible commitment of mitogen-stimulated cells. The previous work using only fixed time points for measuring thymidine uptake and the concept of commitment must be reevaluated in light of the kinetic evidence presented.
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  • 170
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    Journal of Cellular Physiology 106 (1981), S. 407-418 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The effects of altered external sodium and potassium concentrations on steady state, active Na+ + K+ transport in Ehrlich ascites tumor cells have been investigated. Membrane permeability to Na+ and K+, intracellular [Na+] and [K+], and membrane potential were measured. Active cation fluxes were calculated as equal and membrane potential were measured. Active cation fluxes were calculated as equal and opposite to the net, diffusional leak fluxes. Elevation of external K+ (6-60 Mm)by equivalent replacement of Na+ (154-91 mM) inhibits both active Na+ and K+ fluxes, but not proportionally. This results in a decrease of the coupling ratio (rp = -Jkp/JpNa) as external K+ is increased. Elevation of external K+ (3-68 mM) at constant Na+ (92mM) inbibits Jpk, but is without effect on JpNa. The coupling ratio declines from 1.01 ± 0.14 to 0.07 ± 0.05, a 14-fold alteration. Reduction of external Na+ (154-25 mM) at constant K+ (6mM) depresses JpNa, but is without effect on Jpk. The coupling ratio increases from 0.63 ± 0.04 at 154 mM Na+ to 4.5 ± 2.04 at 25 mM Na+. The results of this investigation are consistent with the independent regulation of active cation fluxes by the transported species. Kinetic analysis of the data indicates that elevation of external sodium stimulates active sodium efflux by interacting at “modifier sites” at the outer cell surface. Similarly, external potassium inhibits active potassium influx by interaction at separate modifier sites.
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  • 171
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    Journal of Cellular Physiology 106 (1981), S. 445-450 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Tumor-promoting phorbol esters, such as 12-0-tetradecanoyl-phorbol-13-acetate (TPA), stimulate mouse peritoneal exudate macrophages to undergo DNA synthesis without added macrophage growth factor (MGF). Resident peritoneal macrophages do not respond in this manner. Plasminogen activator (PA) levels of both exudate and resident peritoneal macrophages are elevated by TPA. Thus the phorbol esters appear to mimic the action(s) of MGF and may be useful in understanding the events involved in macrophage and precursor cell growth responsiveness.
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  • 172
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    Journal of Cellular Physiology 107 (1981), S. 21-29 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Intracellular recordings of cultured human peritoneal exudate cells reveal that cells within the culture exhibit an active depolarizing response to injected currents which can reach positive potentials and resemble slow spikes. The cells exhibiting spikes are similar to the reticular cells described by Stuart and Davidson (1971a,b) in that they are esterase(+), acid phosphatase(+), and internalize colloidal carbon but not opsonized red blood cells. The active depolarizing response is unaffected by either decreasing the external sodium concentration or by adding tetrodotoxin (3 × 10-5 M), whereas increasing the external calcium concentration increases both the spike amplitude and rate of rise, and the addition of cobalt (3 mM) blocks the response. Addition of barium increases the duration and amplitude of the spikes but reduces the afterhyperpolarization. The data indicate that cultued human reticular cells from the peritoneal cavity exhibit a calcium spike.
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  • 173
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    Journal of Cellular Physiology 107 (1981), S. 47-57 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Human NHIK 3025 cells growing exponentially in 30% or 3% serum had population doubling times of 19.1 and 27.6 hours, respectively. These values were equal to the calculated protein doubling times (17.6 and 26.5 hours, respectively), showing that the cells were in balanced growth at both serum concentrations. Stepdown from 30% to 3% serum reduced the rate of protein synthesis within 1-2 hours, from 5.7% hour to 4.3% hour, while the rate of protein degradation was unchanged (1.7%/hour). In cells synchronized by mitotic selection from an exponentially growing population, the median cell cycle durations in 30% and 3% serum were 17.2 and 23.6 hours, respectively, which were also in good agreement with the protein doubling times. The median G1 durations were 7.1 and 9.6 hours, respectively. Thus the duration of G1 relative to the total cell cycle duration was the same in the two cases. Complete removal of serum for a period of 3 hours resulted in a 3-hour prolongation of the cell cycle regardless of the time after mitotic selection at which the serum was removed. For synchronized cells, the rate of entry into both the S phase and into the subsequent cell cycle were reduced in 3% serum as compared to 30% serum, the former rate being significantly greater than the latter at both serum concentrations. Our results thus indicate that these cells are continuously dependent upon serum throughout the entire cell cycle.
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  • 174
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Topics: Biology , Medicine
    Notes: Cation transport and membrane potential were studied during the cell cycle of neuroblastoma cells (clone Neuro-2A) to investigate the role of these parameters in growth regulation. The cells were synchronized by selective detachment of mitotic cells. The membrane potential and intracellular K+ activity were measured with conventional and K+-selective microelectrodes respectively. Both the membrane potential and K+ activity were high in mitosis, decreased to half maximal in G1 phase, and rose again during S phase.K+ efflux across the plasma membrane was studied with 42K+ as a radioactive tracer using a washing method for cells grown in monolayer and a continuous efflux method for mitotic cells in suspension. The intracellular K+ content and unidirectional K+ efflux rate obtained from these measurements showed modulations during the cell cycle similar to those of the membrane potential. Using equations of electrodiffusion theory the membrane permeabilities to K+ and Na+ were calculated. These permeabilities were high in mitosis, decreased rapidly in G1 phase and increased during S phase, followed by a transient decrease in G2 phase. A rapid increase was observed between G2 phase and the next mitosis. A similar pattern was obtained for the K+ conductance. K+ resistance changes during the cell cycle were similar to changes in the specific membrane resistance, measured by microelectrodes, except for the early cell cycle phases (mitosis and G1).These studies clearly demonstrate large modulations of the passive membrane permeability properties during the cell cycle. These modulations can be correlated with physicochemical membrane variations during the cell cycle, such as membrane fluidity and lateral mobility of lipids.
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  • 175
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    Journal of Cellular Physiology 107 (1981), S. 115-122 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Flow cytometry indicated that significant amounts of dsRNA were accumulated in HeLa S3 cells blocked at or near G1/S boundary by hydroxyurea (HU) or excess thymidine (TdR). The dsRNA/DNA ratio increased in these cells in a manner characteristic of unbalanced cell growth. In HU-treated cells, dsRNA content was maximal 16 hours after addition of the drug and did not change significantly during the next 24 hours. The DNA content in blocked cells increased by 10%. Cell viability assessed by colony formation in soft agar decreased exponentially in HU-treated cultures after 16 hours of incubation. Correlation between loss of cell viability and rate of cell proliferation after removal of HU was observed, as determined by cell count and analysis of cell cycle progression. In TdR-treated cultures cells slowly progressed into mid S-phase during 40 hours and dsRNA accumulation continued during this period. Cell viability was not significantly affected by treatment with excess TdR, indicating that unbalanced growth per se, as measured by dsRNA accumulation, is not lethal for the cells. After reversal of DNA synthesis inhibition by removal of the drug, cells treated with HU for 16 hours or TdR for 16-24 hours promptly progressed through the cell cycle. This progression was accompanied by accumulation of significant amounts of dsRNA. As a result, cells in G2 phase had a very high dsRNA content leading to retention of the unbalanced condition (increased dsRNA/DNA ratio) in the daughter cells. It is suggested that dsRNA accumulation in the cell is controlled to a certain degree by cell progression through the S phase. This type of control, evidently, was reflected in limited dsRNA accumulation in the cells blocked at or near G1/S border, in continuous dsRNA accumulation in the cells slowly progressing through S phase, and in accumulation of large amounts of dsRNA after renewal of progression through the S phase.
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  • 176
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    Journal of Cellular Physiology 107 (1981), S. 139-145 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Using the pulsed nuclear magnetic resonance (NMR) spectroscopy, the spin-lattice (T1) and the spin-spin (T2) relaxations times of water protons from samples of pectoralis major muscles of normal (line 412) and homozygous dystrophic (line 413) chickens were measured. Both the T1 and T2 were significantly increased (P 〈 0.05) in the dystrophic muscles. The mean values of the relaxation times are given ± S.D. The T1 values were 654 ± 22 msec in normal and 692 ± 41 msec in dystrophic muscles. The T2 values for normal and dystrophic muscles were 39 ± 4 msec and 52 ± 7 msec, respectively. Although the water content of dystrophic muscles (78.9 ± 0.6%) determined by gravimetric methods was significantly higher than normal muscles (74.9 ± 1.1%), this difference in tissue hydration could not explain quantitatively the increase of T1 and T2 values in the dystrophic muscles. The results of the measurements of the relaxation times seem to suggest that there are changes in the composition and/or conformational state of the proteins.
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  • 177
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    Journal of Cellular Physiology 107 (1981), S. 165-170 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have quantified the effect of EGTA on K exodus and uptake in human blood lymphocytes. When lymphocytes were exposed to a medium containing an EGTA concentration that resulted in an ionized Calcium (Ca) of less than 10 μM, K exodus began to increase. This increase reached nearly threefold that of the control rate in a medium containing sufficient EGTA to reduce the ionized Ca concentration below 0.1 μM. When K exodus was increased, K uptake increased proportionately. This increase in K uptake represented active transport and was associated with an 80% increase in intracellular Na concentration from 15 to 27 mM. The addition of Ca to a medium containing EGTA reversed to normal the increased K exodus and uptake. Histidine, a potent chelator of divalent cations other than Ca, had no effect on K transport. These data indicate that extracellular Ca chelation leads to an increase in lymphocyte membrane permeability and cation leak. This increased leak is associated with an elevation of the cell Na and an increase in transport to a rate equivalent to that of the exodus rate. The compensatory increase in active transport maintains the cell monovalent cation concentration within 10 to 15 mM of unperturbed levels.
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  • 178
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    Journal of Cellular Physiology 107 (1981), S. 195-207 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The effect of rat submaxillary extract on the growth of rat C6 glioma cells in serum-free culture has been examined. Extracts (10-15 μg/ml) of submaxillary glands from both male and female rats markedly enhanced the growth of serum-deprived C6 cells and, in combination with insulin, transferrin, and NIH-LH (a source of fibroblast growth factor), were able to stimulate C6 cell growth to an extent comparable to that achieved with an optimal amount of fetal calf serum. The mitogenic activity of rat submaxillary extracts was found to be heat-labile, acid-stable, and partially inactivated by protease and 2-mercaptoethanol. Under our assay conditions, biologically active preparations of purified mouse submaxillary gland epidermal growth factor (EGF) or nerve growth factor (NGF) were not mitogenic for C6 cells, nor was the mitogenic activity of rat submaxillary extracts inhibited by antiserum to these mouse submaxillary gland growth factors. These results suggest that the active component(s) of rat submaxillary extracts is unrelated to either EGF or NGF. The growth-enhancing effect also appears unrelated to esteropeptidase activity present in these extracts since the mitogenic activity was unaffected by several protease inhibitors. Moreover, two purified mouse submaxillary gland arginylesteropeptidases, EGF-binding protein and γ-subunit of 7 S NGF, were unable to elicit a comparable growth response even when added to cell culture medium at unreasonably high concentrations.The C6 cell mitogenic activity of crude submaxillary extracts could be separated into two biologically similar components by either gel filtration on Sephadex G-100, preparative isoelectric focusing in a pH gradient of 3-10, or adsorption to DEAE-cellulose followed by elution with a sodium chloride gradient. One of the active components was acidic in nature and had an apparent molecular weight of 40,000, while the other was near neutral in charge and possessed a molecular weight of approximately 20,000. The relationship between these two C6 cell mitogenic components and the rat submaxillary gland component responsible for stimulating Balb/c-3T3 cell growth in serum-free, factor supplemented medium (McClure et al., 1979, J. Cell Biol. 83: 96a) is also discussed.
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  • 179
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    Journal of Cellular Physiology 107 (1981), S. 231-236 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A bovine calf lens epithelial cell line (CLE-1) that synthesizes crystallin has been established in culture and some of its transport properties have been characterized using both cells and membrane vesicles derived from them. The membrane vesicles fractionate with high recovery of plasma membrane markers, showing a 40-fold purification of 5′-AMPase and a 20-fold decrease in the specific activity of the mitochondrial marker enzyme succinic dehydrogenase relative to a cell homogenate. Transport sites demonstrated higher specific activity than has been seen in vesicles from cell lines studied previously. The uptake of α-amino isobutyric acid (AIB) (an alanine analog) by CLE-1 cells is stimulated four- to fivefold by Na+ and exhibits a Km of 5.4 mM with a Vmax of 50 pmoles/min μg of cell protein. The uptake of leucine was not Na+ stimulatable. The uptake of AIB by the cells was reduced by 43% at confluence. Thus, the cell density dependent behavior of the uptake of the alanine amino acid family in CLE-1 is similar to that of various fibroblast cells.The Na+ caused a threefold stimulation of AIB uptake in the membrane vesicles, while vesicular uptake of leucine was unaffected by Na+. The uptake of adenine, guanine, uridine, and guanosine was also tested in these vesicles. The substrates were rapidly accumulated, came to a steady state distribution within 1-2 minutes, and were recovered as the unaltered compounds after uptake.
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  • 180
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    Journal of Cellular Physiology 107 (1981), S. 251-254 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The quantity of aspartate, but not of glutamate, synthesized by human diploid fibroblasts (HDF) was inversely proportional to the glucose concentration. Forty hours after confluent cells were refed with media containing 1.5 mM or 55 μM glucose, the aspartate concentration was 8 μM in medium high in glucose and 28 μM in medium low in glucose; glutamate was 350 μM at both glucose levels. The label incorporation from [U-14C] glutamine (0.8 C/ml) into aspartate after 40 hours in 1.5 mM glucose medium was less than 1000 DPM/ml and into glutamate 540,000 DPM/ml. The respective label incorporation into aspartate in 55 μM glucose medium was 118,000 DPM/ml and into glutamate 450,000 DPM/ml. In media with 1.5 mM glucose, label incorporation into both aspartate and glutamate was observed from [6-14C] glucose, [3-14C] pyruvate, and [1-14C] pyruvate. Since [1-14C] pyruvate labeled aspartate in 1.5 mM glucose medium, whereas [U-14C] glutamine did not, the observations support a cystolic pathway of aspartate synthesis from glycolytic intermediates in the presence of glucose. However, when the glucose concentration is decreased, glutamine appears to be the primary carbon source for aspartate synthesis. Therefore, both the quantity and the pathway of aspartate synthesis in HDF are functions of the glucose concentration. It is postulated that increased accumulation of aspartate in the media of HDF at confluency may be explained by decreased availability of acetyl CoA or of NADH.
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  • 181
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    Journal of Cellular Physiology 107 (1981), S. 329-334 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The objectives of this investigation were to produce a reliable, senstive probe to measure intracellular PO2 with a high degree of resolution and to apply this technique to biological systems. A fluorescent molecule, pyrene dissolved in paraffin oil, was encapsulated in polyacrylamide to form a probe of nanometer dimensions. The quantitative and microscopic oxygen values were determined by analyzing the quenching of the fluorescence of the probe by oxygen, as displayed on a television monitor by a silicon-intensified-target camera. The nanocapsules had a sensitivity of approximately 1 mm PO2, a spatial resolution of 0.5 μm, and a temporal resolution of milliseconds. Calibrated nanocapsules within nonrespiring Amoeba proteus responded to ambient partial pressures of oxygen. At two different ambient partial pressures, nanocapsules engulfed by respiring amoebas indicated an intracellular PO2 28 mm Hg less than extracellular PO2. The capsules retained their sensitivity to oxygen for at least 8 months.
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  • 182
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    Journal of Cellular Physiology 107 (1981), S. 359-369 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Human fibroblasts that have been serum deprived for 4 hours have a digitoxin-insensitive Na influx of 9.5 ± 1.0 (n = 4) μmol/g prot/min which is not significantly different from the influx of 9.4 ± 0.6 (n = 3) μmol/g prot/min measured in cells arrested in the G1/G0 state by serum-deprivation for a period of four days. The Na influx in serum-deprived cells is rapidly stimulated (within one minute) simply by assaying the cells in medium containing 10% fetal bovine serum (FBS). The digitoxin-insensitive Na influx for cells in the presence of 10% FBS is 22.9 ± 1.1 (n = 6) μmol/g prot/min. the stimulation of Na influx in serumdeprived cells can also be achieved by the addition of the purified mitogen, epidermal growth factor (EGF). Addition of EGF to serum-deprived cells gives a maximal stimulation of Na influx of approximately 1.6-fold, with the concentration for half-maximal stimulation being 7.5 ng/ml. The stimulation of Na influx results from the activation of an amiloride-sensitive pathway, which appears to be minimally active in serum-deprived cells. Kinetic analysis of Na influx experiments in the presence of 10% FBS and varying concentrations of amiloride indicate that at infinite concentrations of amiloride the Na flux would be reduced to 8.9 μmol/g prot/min, which is comparable to the level of Na flux measured in serum-deprived cells in the presence of 5 mM amiloride. Thus, amiloride can totally inhibit the serum-stimulated component of Na influx while inhibiting less than 10% of the Na influx in serum-deprived cells. The Na influx in serum-deprived cells can also be stimulated 2.5-fold by preincubating cells in the presence of the Ca+ ionophore A23187 to elevate the intracellular Ca content. This stimulation of Na influx by intracellular Ca+2 can be virtually eliminated by adding 1 mM amiloride.
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  • 183
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    Journal of Cellular Physiology 107 (1981), S. 379-384 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Murine multipotential hematopoietic stem cells (CFU-s) bear an antigen (SC-1) which is recognized by heterologous antisera to mouse brain. We have found that cloned Thy-1 negative variants of the T-cell lymphoma RL ♂1 are sensitive to complement-mediated cytolysis by anti-brain serum and can absorb the anti-stem cell activity from the antiserum. We have isolated several subclones derived from a primary Thy-1 negative variant which are not susceptible to anti-brain serum. The surface of the resistant lines has little or no antigen capable of binding anti-mouse brain antibodies as measured by either immunofluorescence or a radioimmunoassay. These lines are also unable to absorb the anti-bodies responsible for the cytotoxic effect of rabbit anti-mouse brain serum against CFU-s. We conclude that the predominant antigen, serologically detectable on Thy-1 negative variants of RL ♂1, is SC-1.
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  • 184
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    Journal of Cellular Physiology 107 (1981), S. 399-412 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Vole cells transformed by avian sarcoma virus carrying the src gene lose their fibroblastic morphology, the organized cytoskeletal system of the normal fibroblastic cell, the typical fibronectin deposit around the cell membrane, and the ability to shut off multiplication when suspended in liquid medium. All of these transformation characteristics are reversed by treatment with cAMP derivatives. Moreover, the cAMP treatment does not cause loss of activity of the src gene product. These data imply that cAMP exerts its effect at or after the point in the metabolic pathway affected by the src gene product, pp60src. Presumably, the decision to adopt the transformed or the normal state is determined by the degree to which the src gene or cAMP-mediated kinase activities respectively predominate in the cell. The development of all four transformation characteristics as a result of introduction of the src gene, and their coordinate reversal by cAMP derivatives, supports the previous thesis that in the normal vole or CHO fibroblast all four properties are part of a common regulatory system.
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  • 185
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    Journal of Cellular Physiology 107 (1981), S. 413-426 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Chinese hamster ovary (CHO) cells in culture were limited for polyamines through the use of α-methylornithine (αMO), a competitive inhibitor of ornithine decarboxylase. Initial exposure of the cells to the inhibitor caused growth rate and intracellular polyamine content to decline continuously. Reseeding the αMO-treated cells into medium containing the inhibitor resulted in steady-state (exponential) growth at cell densities below 5 × 103 cells/cm2, at a rate approximately twofold slower than untreated cells. Under these conditions, putrescine and spermidine were undetectable and spermine remained relatively constant at a level approximately half that found in untreated cells. Addition of exogenous putrescine elevated the polyamine content and stimulated the growth of αMO-treated cultures. Thus, growth rate correlated with polyamine content in the αMO-treated cells.The growth of reseeded. αMO-treated cells became nonexponential at a density (5 × 103 cells/cm2) far below that at which untreated cells departed from exponential growth (1 × 105 cells/cm2). Medium obtained from high density, αMO-treated cultures inhibited the growth of cells at low density in the presence of αMO. Doubling the concentration of the defined components of conditioned medium did not markedly affect its capacity to inhibit growth. However, dialysis completely removed the inhibitory activity from conditioned medium. The results imply that a low molecular weight inhibitor of growth is produced by polyamine-limited cells. This is a variable that must be controlled in studies with polyamine-limited animal cells.Morphological studies indicated that subcellular organelles, including mitochondria, were largely unaffected by treatment with αMO. The maintenance of mitochondrial integrity in the presence of αMO demonstrates that the swelling of mitochondria observed previously in cells treated with methylglyoxal bis(guanylhydrazone) was not due to polyamine limitation. αMO-treated cells did, however, accumulate numerous cytoplasmic vacuoles. The identity of these vacuoles and their relationship to cellular physiology is not yet understood.
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  • 186
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    Journal of Cellular Physiology 108 (1981), S. 475-482 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The effect of decreasing cellular sterol content on neurite outgrowth in C1300 (Neuro 2A) neuroblastoma cells in serum-free medium has been studied. Sterol-depleted, undifferentiated neuroblastoma cells were obtained by growing cells for 24 h in medium containing lipoprotein-poor serum and 25-hydroxy-cholesterol (25-OHC). Under these conditions the activity of 3-hydroxy-3-methyl-glutaryl-CoA reductase and the incorporation of [14C] acetate into sterols were almost completely suppressed, and the sterol/phospholipid ratio of the cells declined to 60% of that in cultures grown without 25-OHC. The sterol-depleted cells were viable and exhibited rates of DNA, RNA, protein and fatty acid synthesis comparable to those measured in control cultures.Sterol depletion had no detectable effect on the number of cells that were able to undergo morphological differentiation within 3 h after removal of serum from the medium. However, by 24 h most of the sterol-depleted cells had retracted their neurites. The observation that addition of low-density lipoprotein was able to restore neurite outgrowth in cultures treated with 25-OHC indicates that the inability of sterol-depleted cells to maintain their neurites is related specifically to the decline in the sterol content rather than to a general cytotoxic effect of 25-OHC. Our findings suggest that incorporation of cholesterol into the cell membrane is important for long-term maintenance and elongation of neuroblastoma neurites, but that the initial morphological change (i.e., within 3 h after removal of serum) is apparently a separate and distinct event, not dependent on the availability of cholesterol.
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  • 187
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    Journal of Cellular Physiology 109 (1981) 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 188
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    Journal of Cellular Physiology 108 (1981), S. 47-54 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The ionic basis of volume regulation by human peripheral blood lymphocytes in hypotonic Tyrode's medium has been studied. The intracellular water space of lymphocytes increased to a maximum after 1 min in 0.68 × isotonic Tyrode's but returned to the isotonic value by 20 min at 37°C. During this phase of volume regulation (1-20 min) both 42K+ efflux and 42K+ influx were stimulated severalfold, but the increase in 42K+ efflux exceeded the influx, resulting in a net loss of 20% of the lymphocyte K+. The increase in 42K+ efflux during the phase of cell shrinkage was unaffected by ouabain or by quinidine. Hypotonicity increased both the ouabain-sensitive (active) and ouabain-insensitive components of 42K+ influx by 76% and 123% respectively. Hypotonic shock stimulated 22Na+ influx by only 25%, but cell Na+ content was unchanged at 1 min and even decreased after 20 min. Thus active K+ influx and Na+ extrusion is increased by hypotonicity, but greater pumping cannot explain the net decrease in cell cations that leads to volume regulation. The 45Ca2+ uptake was not significantly changed by hypotonicity. Although volume regulation was abolished in a hypotonic high K medium, 42K+ efflux was still stimulated 2-fold by the reduction in tonicity. These findings support the hypothesis that volume regulation in hypotonic media occurs largely by a passive loss of cell K+, which results from a selective increase in membrane permeability to this ion. The increase in K+ permeability in hypotonic media is observed even in the absence of volume regulation by the cell.
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  • 189
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    Journal of Cellular Physiology 108 (1981), S. 77-82 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Kinetics of glucose transport in K-562 cells was studied using 3-0-methylglucose, a nonmetabolizable analog of glucose. A Km of 3.7 mM and Vmax of 32.0 nmoles/minute/106 cells was found for the process. D-Glucose, phloretin, and phlorizin competitively inhibit the transport of 3-0-methylglucose with Ki values of 4.1 mM, 4.1 μM and 225 μM, respectively, whereas L-glucose did not inhibit transport at all. The results indicate that K-562 cells, which are known to have erythropoietic characteristics, possess a glucose carrier system similar to the one in adult human erythrocytes. However, the Vmax data suggest that more copies of the carrier are present in the malignant cell, presumably to support the high rate of anaerobic glycolysis.
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  • 190
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    Journal of Cellular Physiology 108 (1981), S. 91-97 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The specific activity of HMG-CoA reductase, the major rate-limiting enzyme in the sterol biosynthetic pathway, declined linearly with increasing cell density in four different lines of mammalian cell cultures. As expected, this caused the rates of sterol synthesis from [14C]acetate to decline in a parallel manner. The decrease in reductase activity in the dense cultures was also correlated with decreased incorporation of [14C]acetate into fatty acids and [3H]thymidine into DNA. In contrast, the activities of two enzymes, NADH dehydrogenase and 5′-nucleotidase, which are not involved in lipid synthesis, were independent of changes in cell density. The simplest explanation for these data is that HMG-CoA reductase and the synthesis of sterol and fatty acids are regulated in concordance with the rate of cell growth and proliferation.
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  • 191
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    Journal of Cellular Physiology 108 (1981), S. 115-122 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: “Fertilization acid” is released from sea urchin eggs upon fertilization and decreases the pH of the surrounding seawater. In bicarbonate-free artificial seawater flushed with nitrogen gas, the pH shift still occurs but returns to the original value in a few minutes, suggesting that the released acid is volatile. A likely candidate for a volatile acid is carbon dioxide released from the eggs. Therefore, the total CO2 content of seawater was measured pre- and post-fertilization and was found to be correlated stoichiometrically with released proton equivalents, leading to the conclusion that fertilization acid is largely carbon dioxide. Manometric analysis of cell extracts and ashed eggs suggest that the carbon dioxide may be stored in the unfertilized egg as an inorganic carbonate.
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  • 192
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Epidermal growth factor (EGF) stimulates the initiation of DNA synthesis in Swiss 3T3 cells after a constant prereplicative period of 14-15 hours. The final rate of initiation follows apparent first-order kinetics and can thus be quantified by a rate constant k. The value of k can be changed by later additions during the prereplicative period: When cells stimulated by a very low concentration of EGF, alone or with insulin, which results in a relatively low value of k, receive a saturating amount of EGF at 15 hours, then k is markedly increased after 4-6 hours. Insulin alone (up to 200 ng/ml) is unable to set the lag phase, but does have a synergistic effect on the value of k given by EGF. When added at 15 hours, insulin also increases k, but after a delay of 4-6 hours. In contrast, both hydrocortisone and prostaglandin E1 (PGE1) inhibit the stimulation of DNA synthesis by EGF only during the first 8 hours of the prereplicative period of decreasing the value of k.Prostaglandin F2α (PGF2α), which stimulates DNA synthesis in a similar mode as EGF, when added with EGF has a synergistic effect on DNA synthesis. This suggests that EGF and PGF2α, nevertheless, act through different regulatory events.
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  • 193
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    Journal of Cellular Physiology 108 (1981), S. 163-173 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Inhibition of the gap-filling, polymerizing step of excision repair by 1-β-D-arabinofuranosylcytosine (ara-C) after irradiation with ultraviolet light in human diploid fibroblasts resulted in the formation of persistent DNA strand breaks in G1, G2, and plateau phase cells, but not in S phase cells. Addition of hydroxyurea to ara-C resulted in partial inhibition of repair in S phase cells. These observations can be explained either in terms of changing roles in repair for different DNA polymerases throughout the cell cycle or by the presence of a pool of deoxycytidine nucleotides during S phase equivalent to an external source of deoxycytidine at 50 μM concentration. A similar concentration dependence on ara-C was observed for inhibition of repair in normal human, xeroderma pigmentosum (XP) variant, and Cockayne's syndrome cells. Ara-C produced a similar number of breaks in normal and Cockayne's syndrome cells but slightly more in XP variant cells. Exonuclease III and S1 nuclease independently both degraded about 50% of the 3H-thymidine incorporated into repaired regions in the presence of ara-C. Sequential digestion with both enzymes degraded nearly 90% of the repaired regions. These observations can be explained if excision repair proceeds by displacing the damaged strand so that both the 3H-labeled patch and the damaged region are still ligated to high molecular weight DNA and compete for the same complementary strand during in vitro incubation with the nucleases. The amount of 3H-thymidine incorporated in DNA by repair decreased with increasing concentrations of ara-C and hydroxyurea, suggesting that the incomplete patches became shorter under these conditions. Extrapolation of the digestion kinetics with exonuclease III permits an estimate of the normal patch size of about 100 nucleotides, consistent with previous estimates.
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  • 194
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    Journal of Cellular Physiology 108 (1981), S. 195-211 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Colchicine and vinblastine inhibited endothelial cell migration but had no effect on the stimulation of replication seen at wound edges in cultures of endothelium at stationary density. This is in contrast to the effects of cyto-chalasins which inhibit both migration and replication at wound edges. Moreover, colchicine and vinblastine stimulated cell replication in the unwounded, confluent monolayer. This effect has kinetics similar to the stimulation of replication at a wound edge and is associated with an initial retraction of cell borders, leaving gaps between cells. Cytochalasin D inhibited the growth response to microtubule disrupting agents but did not prevent cell retraction. Stimulation of replication by microtubule disrupting agents was not dependent on serum but was synergistic with serum in cultures rinsed repeatedly with serum-free medium. The replication occurred prior to any cell loss. When, however, cells were allowed to complete mitosis, about one-half of the daughter cells detached from the monolayer so that there was no increase in cell density. We conclude that microtubule disrupting agents are the first agents found to be effective in stimumating growth of vascular endothelium at saturation density. These data further suggest that colchicine and vinblastine stimulate cell growth in a manner similar to wounding, where cell movement is a prerequisite to cell replication.
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  • 195
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    Journal of Cellular Physiology 108 (1981), S. 221-230 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Two spontaneously arising variant clones were selected from the N18 neuroblastoma cell line solely on the basis of their flattened morphology and tight adherence to the culture flask. Two other clones having the round loosely adherent morphology typical of the parent line were also selected, and flat variants were shown to arise in them upon prolonged cultivation. The flat variant clones have slower growth rates in culture, lower cloning efficiencies in suspension, and reduced acetylcholinesterase inducibility when compared with either the parent N18 line or the round cell clones. Cells of both morphologic types have high levels of plasminogen activator and are tumorigenic, although the variants have a slower growth rate in vivo, consistent with their slower growth rate in culture. SDS-polyacrylamide gel electrophoresis of total protein from the two cell types shows that the flat variants have increased amounts of a 200,000 molecular weight polypeptide that has tentatively been identified as the heavy chain of myosin. Round morphological revertants from one of the flat variant clones exhibited growth characteristics typical of the parent N18 line, but their content of myosin heavy chain, although reduced, was not so low as that in the round cell clones originally isolated. The possibility of a causal relationship between flat morphology, reduced suspension cloning efficiency, and increased content of myosin heavy chain is discussed.
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  • 196
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    Journal of Cellular Physiology 108 (1981) 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 197
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    Journal of Cellular Physiology 108 (1981), S. 291-298 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Addition of H1 histone or polylysine (10 μg/ml) to cultured Friend erythroleukemia cells or to two mouse lymphoma cell lines (el-4 and S-49) increased levels of cell division in these cultures. There is a stimulation of incorporation of labeled thymidine into DNA in cultures containing H1 histone and polylysine. DNA fiber autoradiographic experiments revealed that replicon size is decreased in the cells cultured with H1 histone and polylysine at later periods of culture.
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  • 198
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    Journal of Cellular Physiology 108 (1981), S. 299-307 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Trifluoperazine (TFP) blocks spreading and migration of cultured mammalian cells. These are calcium-dependent and microfilament-mediated processes. Calmodulin, a regulator of many calcium-dependent processes in cells, is selectively inhibited by TFP. Cell spreading on a plastic- or collagen-coated substratum was reversibly inhibited by 10 μM TFP. The drug blocks cell spreading even in the presence of 1 mM cAMP. TFP is as effective as cytochalasin B (CB), an inhibitor of microfilament function, in blocking cell spreading. All cell lines tested, whether “normal” or virally transformed, failed to spread in TFP. The drug, at a concentration sufficient to inhibit spreading, does not interfere with the initial attachment of a cell to a plastic surface. Cells plated in the presence of 10 μM TFP attach at a rate and to an extent equal to untreated controls. TFP added to already spread cells results in a reversible cell rounding. Detection of fibronectin by indirect immunofluorescence suggests TFP-induced cell rounding is not due to shedding of fibronectin from the cell surface. TFP reversibly blocks cell migration into a wound edge almost as effectively as CB. We suggest that TFP interferes with these microfilament-mediated functions by direct action on the microfilaments or indirect action by inactivating calmodulin.
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  • 199
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    Journal of Cellular Physiology 108 (1981), S. 327-335 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The action of procaine on the terminal erythroid differentiation of murine erythroleukemia (MEL) cells has been investigated at the level of individual cells. At concentrations (7 × 10-4 M) which had no inhibitory effect on cell growth, pretreatment of these cells with procaine for 12-24 hr caused a pronounced inhibition (〉 90%) of commitment to terminal erythroid differentiation of dimethyl sulfoxide (DMSO)-treated cells. Simultaneous treatment of MEL cells with DMSO and procaine, however, resulted to only slight inhibition (〈 20%) of commitment. Blockade of commitment by procaine pretreatment appears to be general since it was observed in cells treated with other inducers (6-thioguanine, dimethylformamide). Procaine pretreatment did not abolish the ability of MEL cells to complete the “latent period” and commit upon the removal of the block. Reversal of procaine inhibition of commitment was obtained by the addition of either CaCl2 (1.0 mM), calcium ionophore A23817 (1 μg/ml), but not of MgCl2 (1.0 mM). From these data we conclude that procaine inhibits the terminal erythroid differentiation of MEL cells by blocking an event or process required for commitment which occurs prior to commitment itself. Our results suggest that this process involves calcium metabolism.
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  • 200
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    Journal of Cellular Physiology 108 (1981), S. 365-373 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Several studies indicate that glutamine is a critical requirement for cell growth in vitro. Growing and quiescent (serum-starved) 3T3-fibroblasts were exposed to media (Dulbecco's modified Eagle's minimal essential medium) in which the concentration of the 13 essential amino acids had been lowered to 1/100 or 1/1,000 of that in DMEM - either all together or one by one. The effects on DNA synthesis were measured by autoradiographic determinations of the percentage of labeled cells after 24 hours exposure to 3H-thymidine. A reduction of all 13 essential amino acids to 1/100 or 1/1,000 of the normal concentration in the medium resulted only in a minor growth inhibitory effect during the first cell cycle. A similar growth inhibitory effect was caused by the depletion of one of the 13 essential amino acids (except glutamine) from the medium. However, a depletion of glutamine from the medium resulted in a marked inhibition of growth. Conversely, a relative excess of glutamine, when the other 12 amino acids were lowered to 1/1,000 of the normal concentration, counteracted the growth inhibitory effect of serum starvation. It was even possible to stimulate quiescent cells to undergo DNA synthesis by exposing them to a serum-depleted (0.5% serum) medium with a relative excess of glutamine.
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