bluuebunny/crossref_metadata_2025 huggingface dataset:
A LARGE SCALE QUASI-CRYSTALLINE LAMELLAR LATTICE IN CHLOROPLASTS OF THE GREEN ALGA ZYGNEMA
Authors: Mclean Robert J. , Pessoney George F.
Published: May 2004

A quasi-crystalline lamellar lattice was observed in chloroplasts of the filamentous green alga Zygnema. The lattice does not appear in the cells until cultures are at the end of the log phase of growth. Pseudograna are also present and become more numerous towards the middle of the log phase. The three-dimensional lattice superficially resembles the configuration of cubic prolamellar bodies but is about 10 times larger and is entirely different in internal structure. The lattice is composed of one or two appressed thylakoids in a stroma matrix which is bounded on each side by a single thylakoid membrane. This multilayered sandwich of membranes and matrix occupies a position equivalent to the single membrane of a cubic prolamellar body.
ON THE RECONSTITUTION OF THE CRYSTALLINE COMPONENTS OF THE SEA URCHIN FERTILIZATION MEMBRANE
Authors: Bryan Joseph
Published: May 2004

The characteristics of the reconstitution of a crystalline component of the sea urchin fertilization membrane are presented. The reassembly of large aggregates of cylindrical or tubular components is effected by the addition of calcium or other divalent cations. The reassembly requires a slightly alkaline pH and is little affected by increasing ionic strength. Reassembly is strongly inhibited by treatment with reducing agents such as dithiothreitol. The role of this protein in the formation of the fertilization membrane and its possible relation to the calcium-insoluble proteins of the mitotic apparatus are discussed.
DIRECT ISOLATION OF THE HYALINE LAYER PROTEIN RELEASED FROM THE CORTICAL GRANULES OF THE SEA URCHIN EGG AT FERTILIZATION
Authors: Kane R. E.
Published: May 2004

Treatment of the eggs of the sea urchin with a 1 M solution of glycerol at fertilization allows the recovery from this solution of the protein released from the cortical granules, including that which would normally give rise to the hyaline layer. The calcium-gelable protein previously extracted from whole eggs and from isolated cortical material was found to be present in the glycerol solution, confirming its localization in the cortical granules and its role in the hyaline layer. Quantitative measurements on the eggs of two Hawaiian species, Colobocentrotus atratus and Pseudoboletia indiana, which have the widest variation in the gel protein content, demonstrated that a proportionate amount of this material was released at fertilization in these species, which correlates with the thickness of the hyaline layer in the two cases. In addition, the calcium-insoluble fraction of Sakai can be extracted from these eggs after removal of the hyaline protein by glycerol, showing that this is a different material. A simple method for the separation of the hyaline protein from the calcium-insoluble fraction in solution is provided.
THE METABOLISM OF GAMMA AMINOBUTYRIC ACID IN THE LOBSTER NERVOUS SYSTEM
Authors: Hall Z. W. , Bownds M. D. , Kravitz E. A.
Published: May 2004

γ-aminobutyric acid (GABA) is the inhibitory transmitter compound at the lobster neuromuscular junction. This paper presents a comparison of the enzymes of GABA metabolism in single identified inhibitory and excitatory axons from lobster walking legs. Inhibitory axons contain more than 100 times as much glutamic decarboxylase activity as do excitatory axons. GABA-glutamic transaminase is found in both excitatory and inhibitory axons, but about 50% more enzyme is present in inhibitory axons. The kinetic and electrophoretic behavior of the transaminase activity in excitatory and inhibitory axons is similar. Succinic semialdehyde dehydrogenase is found in both axon types, as is an unknown enzyme which converts a contaminant in radioactive glutamic acid to GABA. In lobster inhibitory neurons, therefore, the ability to accumulate GABA ultimately rests on the ability of the neuron to accumulate the enzyme glutamic decarboxylase.
STRUCTURE-FUNCTION RELATIONSHIPS IN THE ADIPOSE CELL
Authors: Cushman Samuel W.
Published: May 2004

A method is described for preparing isolated rat adipose cells for electron microscopy. The ultrastructure of such cells and their production of 14CO2 from U-glucose-14C were studied simultaneously in the presence of insulin or epinephrine. Each adipose cell consists of a large lipid droplet surrounded by a thin rim of cytoplasm. In addition to typical subcellular organelles, a variety of small lipid droplets and an extensive system of membranes characterize the cell's cytoplasm. A fenestrated envelope surrounds the large, central lipid droplet. Similar envelopes surround cytoplasmic lipid droplets occurring individually or as aggregates of very small, amorphous droplets. Groups of individual droplets of smaller size also occur without envelopes. The system of membranes consists of invaginations of the cell membrane, vesicles possibly of pinocytic origin, simple and vesiculated vacuoles, vesicles deeper in the cytoplasm, flattened and vesicular smooth surfaced endoplasmic reticulum, and Golgi complexes. Neither insulin nor epinephrine produced detectable ultrastructural alterations even when cells were incubated under optimal conditions for the stimulation of 14CO2 evolution. Structural responses of the isolated adipose cell to hormones, if such occur, must, therefore, be dynamic rather than qualitative in nature; the extensive system of smooth surfaced membranes is suggestive of compartmentalized transport and metabolism.
METABOLISM OF RIBOSOMAL PRECURSOR RIBONUCLEIC ACID IN KIDNEY
Authors: Ab Geert , Malt Ronald A.
Published: May 2004

The labile precursors of ribosomal RNA in mouse kidney are preserved when nuclei rapidly isolated after sieving through multiple screens are swollen and cleansed in the presence of an RNase inhibitor before digestion with DNase and phenol extraction. The kinetics of nucleolar labeling analyzed on polyacrylamide gels show that 36S RNA is the major intermediate product in the catabolism of the original 45S RNA precursor to 32S RNA, from which 28S RNA is derived. Each kidney nucleus contains about 200–600 molecules of 45S RNA; the turnover time of the 45S pool is about 3 ± 2 min. Compared with HeLa cells, kidney nuclei have a different major intermediate product and a much smaller and more rapidly turning-over pool of ribosomal precursor RNA.
NUCLEAR MEMBRANES FROM MAMMALIAN LIVER
Authors: Franke Werner W. , Deumling Barbara , Ermen Baerbel , Jarasch Ernst-Dieter , Kleinig Hans
Published: May 2004

Nuclear membranes were isolated from rat and pig liver by sonication of highly purified nuclear fractions and subsequent removal of adhering nucleoproteins in a high salt medium. The fractions were examined in the electron microscope by both negative staining and thin sectioning techniques and were found to consist of nuclear envelope fragments of widely varying sizes. Nuclear pore complex constituents still could frequently be recognized. The chemical composition of the nuclear membrane fractions was determined and compared with those of microsomal fractions prepared in parallel. For total nuclei as well as for nuclear membranes and microsomes, various enzyme activities were studied. The results indicate that a similarity exists between both fractions of cytomembranes, nuclear envelope, and endoplasmic reticulum, with respect to their RNA:protein ratio and their content of polar and nonpolar lipids. Both membranous fractions had many proteins in common including some membrane-bound enzymes. Activities in Mg-ATPase and the two examined cytochrome reductases were of the same order of magnitude. The content of cytochrome b5 as well as of P-450 was markedly lower in the nuclear membranes. The nuclear membranes were found to have a higher buoyant density and to be richer in protein. The glucose-6-phosphatase and Na-K-ATPase activities in the nuclear membrane fraction were very low. In the gel electrophoresis, in addition to many common protein bands, some characteristic ones for either microsomal or nuclear membranous material were detected. Significant small amounts of DNA and RNA were found to remain closely associated with the nuclear envelope fragments. Our findings indicate that nuclear and endoplasmic reticulum membranes which are known to be in morphological continuity have, besides a far-reaching similarity, some characteristic differences.
THE FINE STRUCTURE AND ELECTROPHYSIOLOGY OF HEART MUSCLE CELL INJURY
Authors: Baldwin Kate M.
Published: May 2004

Injured frog heart cells electrically uncouple from their uninjured neighbors within 30 min after injury. This uncoupling process can be shown by the disappearance of an injury potential measured between such injured and uninjured cells. In the present study, the time course of the decline of injury potentials, and thus of electrical uncoupling, in bullfrog atrial trabeculae was determined. Tissue was fixed with glutaraldehyde and osmium tetroxide at various times after injury to determine the morphological changes which accompany this uncoupling process. In some cases, ruthenium red was included in the fixatives. Normal atrial cells are long and narrow, with intercellular junctions located along the lateral surfaces of the cells. Two types of intercellular junctions have been observed: cardiac adhesion plaques (CAPs), and close junctions. Close junctions occur only infrequently. Ruthenium red penetrates all around the cells, leaving only small areas within the CAPs unstained. After injury, the cells are very dense and the myofilaments disarranged. Both types of intercellular junction remain intact, and only slight changes within CAPs are observed. The results are discussed in relation to current concepts of intercellular communication.
THE INTRARENAL DISTRIBUTION OF TRITIATED PARA-AMINOHIPPURIC ACID DETERMINED BY A MODIFIED TECHNIQUE OF SECTION FREEZE-DRY RADIOAUTOGRAPHY
Authors: Bordier Betrand , Ornstein Leonard , Wedeen Richard P.
Published: May 2004

Section freeze-dry radioautography has been used to examine the intrarenal distribution of a water-soluble organic acid (para-aminohippuric acid (PAH-3H)) under constant-infusion, steady-state conditions in mouse and rat kidney in vivo. The technique described here has the following advantages: (a) Sectioning and freeze-drying are accomplished in a closed cryostat at temperatures below -40°C; (b) Handling of the section is facilitated by mounting of the section-to-be on adhesive-coated Saran Wrap prior to cutting; (c) Unembedded freeze-dried sections are attached to photographic film at ambient temperature in the dark room; (d) Fixation follows completion of radioautographic exposure and precedes photographic development; (e) Permanent close contact is maintained between tissue and film. Morphologic preservation compared favorably with that obtained by optimal fixation techniques, which, however, permit diffusion. Cellular accumulation of PAH-3H during secretion was demonstrated in the proximal tubule under steady-state conditions in vivo. The cellular concentration of PAH-3H was uniform throughout the length of the proximal tubule in mouse and rat kidney.
EFFECTS OF SINUS NERVE STIMULATION ON CAROTID BODY GLOMUS CELLS
Authors: Yates R. D. , Chen I-Li , Duncan Donald
Published: May 2004

The sinus nerve or sympathetic trunk was stimulated unilaterally in one group of adult cats or Syrian hamsters while in another group the sinus nerve or sympathetic trunk was cut unilaterally and the animals were given reserpine. In a third group, atropine was administered prior to sinus nerve stimulation. All tissues were processed for the detection of primary monoamines. The carotid bodies on the operated sides were compared with those on the unoperated sides of the same animal in order to determine if amine depletion occurred following the experimental procedures. After sinus nerve stimulation alone, the density of the granules in the glomus cells was decreased, but changes were not noted in the granules following sympathetic nerve stimulation. Sinus nerve stimulation after atropine administration resulted in no change in granule density. Sinus nerve transection followed by reserpine treatment resulted in a greater decrease in granule density on the unoperated than on the operated side. Transection of the sympathetic components to the carotid body followed by reserpine injections resulted in a decrease in granule density in the glomus cells on both the operated and unoperated sides. These results suggest that the sinus nerve must be intact for reserpine to exert an effect and that the sinus nerve may contain efferent fibers which modulate amine secretion.
NUCLEATING SITES FOR THE ASSEMBLY OF CYTOPLASMIC MICROTUBULES IN THE ECTODERMAL CELLS OF BLASTULAE OF ARBACIA PUNCTULATA
Authors: Tilney Lewis G. , Goddard Janette
Published: May 2004

In the ectodermal cells of sea urchin blastulae, the microtubules converge and appear to make contact with three distinct cytoplasmic foci or satellites associated with the basal body of the cilium. Beneath the nucleus, which lies in the apical end of the cell, the microtubules are aligned predominantly parallel to the cell's long axis and could thus make contact with the satellites as is directly suggested by observations on sections at or near the planes of the satellites. After the embryos are treated with low temperature (0°C), the microtubules disassemble; however, the satellites can still be recognized. Upon rewarming, the microtubules reappear. In early stages of reformation, when the tubules in the cell consist of short segments, tubules have only been found in the apical part of the cell. One end of each microtubule appears to make contact with its respective satellite, or is aligned so that it could contact one of the satellites, provided serial sections were cut and collected in order. After longer periods of recovery, the microtubules elongate; as before, one end of each makes contact with a satellite or is aligned so that it could attach to a satellite. Segments of microtubules seen in the basal region of the cell are aligned parallel to the long axis of the cell as in the untreated ectodermal cells and are therefore interpreted as extensions of those tubules making contact with one of the satellites. On the basis of these observations, we suggest that assembly of microtubules is initiated at the satellites. These sites, perhaps best referred to as "nucleating sites," thereby could exert considerable control over the distribution of microtubules in cells. It is hoped that this preliminary report will be followed up by a more detailed study using serial sections.
A NEW METHOD FOR THE PREPARATION OF RAT LIVER LYSOSOMES
Authors: Stahn R. , Maier K.-P. , Hannig K.
Published: May 2004

A combination of differential centrifugation and carrier-free continuous electrophoresis is introduced as a new method for the isolation of animal cell organelles. Various buffers were systematically checked in order to find the system which preserves the organelles and gives as well a good separation in the free-flow electrophoresis apparatus. Triethanolamine-acetate buffer (10 mM), pH 7.4 was used. The isolated lysosomes were pure according to marker enzymes and electron micrographs. A heterogeneity of the lysosomes in electrophoretic mobility was demonstrated with respect to the marker enzymes arylsulfatase and ß-glucuronidase. The lysosomes with higher mobility showed a maximum enrichment of 240-fold with respect to arylsulfatase. The lysosomes with lower electrophoretic mobility showed a 65-fold enrichment with respect to ß-glucuronidase. The ratio of ß-glucuronidase to arylsulfatase varied from 2:1 to 1:2 in lysosomes of different mobility. The yield amounted to approximately 1 mg of lysosomal protein per gram of liver protein. 5–8 mg of lysosomes can be obtained in one experiment. The electrophoretic separation proves to be an effective tool in obtaining pure and well preserved lysosomes.
TRANSITIONAL CARDIAC CELLS OF THE CONDUCTIVE SYSTEM OF THE DOG HEART
Authors: Martinez-Palomo A. , Alanis J. , Benitez D.
Published: May 2004

Cardiac cells with distinctive electrophysiological and morphological features were found at the junctional region between Purkinje and ventricular cells of the dog heart. The electrophysiological exploration of these "transitional" cells revealed action potentials markedly different in configuration from those generated by Purkinje or by ventricular cells. The impaled cardiac cells which generated transitional action potentials were identified in serial sections and studied with the light and the electron microscopes. The transitional cells were found to be characterized cytologically by: (a) their subendocardial location, (b) their small diameter, (c) the absence of T system and sarcoplasmic reticulum, and (d) the lack of intercalated discs under the light microscope and the sparsity of specialized intercellular junctions under the electron microscope. Purkinje, transitional, and ventricular cells were found to be joined by gap junctions permeable to lanthanum. A quantitative difference in the extent and distribution of specialized intercellular junctions may be one of the factors responsible for the slow velocity of conduction characteristic of the Purkinje-ventricular junctional region.
ANALYSIS OF SODIUM TRANSPORT IN THE AMPHIBIAN OOCYTE BY EXTRACTIVE AND RADIOAUTOGRAPHIC TECHNIQUES
Authors: Horowitz Samuel B. , Fenichel I. Robert
Published: May 2004

The transport of Na+ in mature Eurycea oocytes was studied by quantitative radioautography of 22Na+ using techniques suitable for localization of diffusible solutes, together with conventional extractive techniques. Intracellular Na+ consisted of three kinetic fractions: a cytoplasmic fast fraction of about 8.5 µeq/ml H2O; a cytoplasmic slow fraction of about 58.7 µeq/ml H2O; and a nuclear fast fraction of about 11.1 µeq/ml H2O. A nuclear slow fraction, if it exists, does not exceed 5% of the cytoplasmic. The fast fractions represent freely diffusible Na+ in the two compartments; the nuclear solvent space is 1.3 times the cytoplasmic. The flux of both fast fractions is determined by the permeability of the cortical membrane, with neither the nuclear membrane nor diffusion in the cytoplasm detectably slowing the flux. The cytoplasmic slow fraction is interpreted to represent Na+ bound to nondiffusible constituents which are excluded from the nucleus; these may be yolk platelets, although the widespread observation of Na+ binding in other cells, and the high Na+/K+ selectivity, argues against simple ion-binding to the yolk phosphoprotein.
SEPARATION OF NUCLEI REPRESENTING DIFFERENT PHASES OF THE GROWTH CYCLE FROM UNSYNCHRONIZED MAMMALIAN CELL CULTURES
Authors: McBride O. Wesley , Peterson Elbert A.
Published: May 2004

Nuclei have been isolated from unsynchronized cultures of Chinese hamster fibroblasts after varying intervals of growth following the incorporation of thymidine -3H for 20 min. These nuclei were fractionated by unit gravity sedimentation in a stabilizing density gradient of sucrose, and fractions were analyzed for the concentration of nuclei, DNA, and radioactivity. A more rapidly sedimenting population of nuclei in the G2 phase of the cell cycle was separated from a group of nuclei in the G1 phase, and nuclei in progressive stages of DNA synthesis (S phase) were distributed between these two regions. The fractionation of intact cells by sedimentation according to their position in the cell cycle was found to be less satisfactory than the corresponding separation of nuclei. This probably results from the continuous accumulation of mass within individual cells throughout the entire cell cycle, whereas most of the mass of a nucleus is replicated during a relatively narrow interval of the total cell cycle.
THE ORGANIZATION OF CONTRACTILE FILAMENTS IN A MAMMALIAN SMOOTH MUSCLE
Authors: Rice Robert V. , Moses Joan A. , McManus G. M. , Brady Arlene C. , Blasik Lorraine M.
Published: May 2004

Ordered arrays of thin filaments (65 A diameter) along with other apparently random arrangements of thin and thick filaments (100–200 A diameter) are observed in contracted guinea pig taenia coli rapidly fixed in glutaraldehyde. The thin-filament arrays vary from a few to more than 100 filaments in each array. The arrays are scattered among isolated thin and thick filaments. Some arrays are regular such as hexagonal; other arrays tend to be circular. However, few examples of rosettes with regular arrangements of thin filaments surrounding thick filaments are seen. Optical transforms of electron micrographs of thin-filament arrays give a nearest-neighbor spacing of the thin filaments in agreement with the "actin" filament spacing from x-ray diffraction experiments. Many thick filaments are closely associated with thin-filament arrays. Some thick filaments are hollow circles, although triangular shapes are also found. Thin-filament arrays and thick filaments extend into the cell for distances of at least a micron. Partially relaxed taenia coli shows thin-filament arrays but few thick filaments. The suggestion that thick filaments aggregate prior to contraction and disaggregate during relaxation is promoted by these observations. The results suggest that a sliding filament mechanism operates in smooth muscle as well as in striated muscle.
GLUCOCORTICOID-INDUCED ALTERATION OF THE SURFACE MEMBRANE OF CULTURED HEPATOMA CELLS
Authors: Ballard Philip L. , Tomkins Gordon M.
Published: May 2004

Glucocorticoids induce an alteration of the surface of hepatoma tissue culture (HTC) cells as expressed by changes in cell electrophoretic, antigenic, and adhesive properties. The alteration is assayed by the increased adhesiveness of induced cells for a glass surface. The induction process has a lag period of about 3 hr and attains a plateau level after 24–30 hr when 50–80% of the steroid-treated cells are firmly adhered. Less than 10% of untreated cells adhere under the same conditions. Induction is inhibited by actinomycin D and cycloheximide, demonstrates both pH and temperature dependence, and responds to changes in steroid concentration and structure. By contrast, the attachment per se of preinduced cells is not affected by inhibitors of RNA and protein synthesis, fluctuations of temperature and pH, and the presence or absence of the hormone. When the induction process is reversed by removal of steroid or addition of actinomycin D, preinduced adhesiveness is lost with a half-life of 13–24 hr, but in the presence of cycloheximide the loss is accelerated (t1/2 3–5.5 hr). These results suggest that glucocorticoids induce the biosynthesis of a protein which either modifies the cell surface (an enzyme) or is incorporated into surface structures (structural protein).
AN INTERPRETATION OF LIVER CELL MEMBRANE AND JUNCTION STRUCTURE BASED ON OBSERVATION OF FREEZE-FRACTURE REPLICAS OF BOTH SIDES OF THE FRACTURE
Authors: Chalcroft J. P. , Bullivant S.
Published: May 2004

A modification of the freeze-fracturing technique to permit observation of replicas of both sides of the fracture is described. It has been used to study mouse liver cell membrane structure. Membranes break to give two faces with three-dimensional complementarity, although there is some small-scale mismatching which is discussed. Since the two distinctive sets of membrane faces are complementary sets, they cannot be the two outside surfaces. In particular, structures (such as particles) seen on these faces are within the membrane. It is not possible from this work to say precisely where the fracture plane goes with respect to a plasma membrane, only that it must be close to the interface between membrane and cytoplasm, or at that interface. Models, consistent with the appearance of the matching replicas, are derived for three regions of the plasma membrane: (a) The nonjunctional plasma membrane, which contains many scattered particles. Except for these particles, the otherwise flat fracture face is not at variance with a bimolecular leaflet structure. (b) Gap junctions. Each of the two membranes comprising a gap junction contains a close-packed array of particles. (c) Tight junctions. Here membranes have ridges within them.
EFFECTS OF FIXATION AND SUBSTRATE PROTECTION ON THE ISOENZYMES OF ASPARTATE AMINOTRANSFERASE STUDIED IN A QUANTITATIVE CYTOCHEMICAL MODEL SYSTEM
Authors: Papadimitriou J. M. , Van Duijn P.
Published: May 2004

The cytochemical technique of Lee and Torack for the demonstration of aspartate aminotransferase activity was tested on a model system consisting of either total liver homogenate or the mitochondrial or soluble cytoplasmic fraction, incorporated in polyacrylamide film. After incubation of portions of film in a medium of α-ketoglutarate, L-aspartate, and lead nitrate, the lead oxaloacetate formed was converted to lead sulfide. The absorbance determined at 520 nm in a film spectrophotometer and expressed in terms of unit weight of film provided a measure of the contained enzymatic activity, and was directly proportional to the concentration of chemically determined oxaloacetate in the film. Both mitochondrial and "soluble" isozymes of aspartate aminotransferase reacted with the cytochemical media to a quantitatively similar degree, but were considerably inactivated after 15 min of treatment with 1% glutaraldehyde or 3.7% formaldehyde in imidazole buffer, the rate of inactivation being greater for the soluble isozyme. Application of the principle of substrate protection delayed inactivation. Thus, for both isozymes the rate of inactivation decreased if ketoglutarate was added to the fixative. Similarly, it was shown that the optimal incubation medium for the demonstration of the soluble isozyme must contain 4 mM of α-ketoglutarate and 20 mM of L-aspartate. Under these conditions the turnover-number for the cytochemical system is 70% of the value obtained from biochemical estimations. Cytochemical Km values differed for each isozyme and were in accord with values determined by biochemical techniques, indicating that the model system can be used as a link between biochemical and cytochemical data in enzymatic studies.
PROTONATION AND CHLOROPLAST MEMBRANE STRUCTURE
Authors: Murakami Satoru , Packer Lester
Published: May 2004

Light changes the structure of chloroplasts. This effect was investigated by high resolution electron microscopy, photometric methods, and chemical modification. (a) A reversible contraction of chloroplast membrane occurs upon illumination, dark titration with H+, or increasing osmolarity. These gross structural changes arise from a flattening of the thylakoids, with a corresponding decrease in the spacing between membranes. Microdensitometry showed that illumination or dark addition of H+ resulted in a 13–23% decrease in membrane thickness. Osmotically contracted chloroplasts do not show this effect. (b) Rapid glutaraldehyde fixation during actual experiments revealed that transmission changes are closely correlated with the spacing changes and therefore reflect an osmotic mechanism, whereas the light scattering changes have kinetics most similar to changes in membrane thickness or conformation. (c) Kinetic analysis of light scattering and transmission changes with the changes in fluorescence of anilinonaphthalene sulfonic acid bound to membranes revealed that fluorescence preceded light scattering or transmission changes. (d) It is concluded that the temporal sequence of events following illumination probably are protonation, changes in the environment within the membrane, change in membrane thickness, change in internal osmolarity accompanying ion movements with consequent collapse and flattening of thylakoid, change in the gross morphology of the inner chloroplast membrane system, and change in the gross morphology of whole chloroplasts.
GENESIS OF MITOCHONDRIA IN INSECT FAT BODY
Authors: Larsen W. J.
Published: May 2004

Electron microscopy and stereological methods have been used to study the time course and mechanism of mitochondrial genesis in the adult fat body of Calpodes ethlius, (Lepidoptera, Hesperiidae). Most of the larval mitochondria are destroyed during a phase of autolysis shortly before pupation, so that pupal and early adult fat body cells have few mitochondria. The number of mitochondria per cell increases rapidly at the end of the 1st day after the adult emerges. Characteristic partitioned mitochondria appear during the period when the number is rapidly increasing. This evidence, coupled with the results of morphometric analyses of mitochondrial diameter, volume, and surface area, confirms the view that the genesis of adult mitochondria involves the growth and division of mitochondria surviving from the larva.
PURIFICATION OF INTACT MICROTUBULES FROM BRAIN
Authors: Kirkpatrick Joel B. , Hyams Lyon , Thomas Virginia L. , Howley Peter M.
Published: May 2004

Hundred-fold purification of intact microtubules from homogenates of rat brain is reported. The success of purification depends on stabilizing the microtubule structure by the combined effects of hexylene glycol, acidic Ph, and low temperature. A practical, negative stain, electron microscopic assay is used to study purity and stability of microtubule fractions. The purified fractions show a major band which migrates like purified tubulin in the SDS gel electrophoresis system.
FACTORS CONTROLLING THE REASSEMBLY OF THE MICROVILLOUS BORDER OF THE SMALL INTESTINE OF THE SALAMANDER
Authors: Tilney Lewis G. , Cardell Robert R.
Published: May 2004

Hydrostatic pressure, when applied to segments of the small intestine of the salamander, causes a tremendous reduction in number of microvilli and a loss of the terminal web. The intestinal epithelium strips off from its deeper layers at the level of the basement membrane. When the pressure is released and this epithelial sheet is allowed to recover, the microvilli and its terminal web reappear. Stages in the reformation of microvilli are described. In the earliest stages, foci of dense material seem to associate with the cytoplasmic surface of the apical plasma membrane. From this material, filaments appear and their regrowth is correlated with the extension of the microvilli. We suggest that the dense material nucleates the assembly of the filaments which, in turn, appear instrumental in the redevelopment of microvilli. This concept is supported by the existing literature. Further, since neither the microvilli nor the terminal web reappear on any surface but the apical surface, even though the apical and basal surfaces are bathed with the same medium, we suggest that information in the membrane itself or directly associated with the membrane dictates the distribution of the dense material which leads to the formation of the microvilli and ultimately to the polarity of the cell.
DEVELOPMENT OF THE NEUROMUSCULAR JUNCTION
Authors: Lentz Thomas L.
Published: May 2004

Following amputation of the limb of the newt, Triturus viridescens, muscle fibers dedifferentiate giving rise to mesenchymal cells. The earliest changes detected in neuromuscular junctions of dedifferentiating muscle fibers are the appearance of a few vacuoles and decrease in density of the terminal axoplasm. Later, synaptic vesicles become tightly clustered in the axon termination, and their content appears denser than normal. Then, vesicles diminish in number until few are seen in the ending. While these changes are occurring, the area of contact of nerve with muscle becomes smaller. Junctional folds persist only where the nerve maintains contact with muscle, but these are shorter than normal and appear as slight ridges on the muscle surface. Subsequently, the nerve withdraws from the muscle cell and is completely invested by Schwann cell cytoplasm, and all traces of junctional folds are lost at the former region of contact. Cholinesterase activity was localized with the thiolacetic acid-lead nitrate method. Even before marked morphological changes occur in the junction, DFP- and physostigmine-sensitive activity in the cleft between nerve and muscle is decreased in intensity. Activity continues to decrease as the area of nerve-muscle contact diminishes and junctional folds disappear. When the nerve has withdrawn from the muscle surface, only a few small deposits of lead are left in the intervening region. These results show that as muscle becomes less specialized during dedifferentiation, the neuromuscular junction also loses the cytological and cytochemical specializations associated with synaptic function.
PHYSICAL CHEMICAL STUDIES ON THE SPECIFIC INTERACTION OF AN ACRIFLAVINE-PHOSPHOTUNGSTIC ACID COMPLEX WITH DOUBLE-STRANDED NUCLEIC ACIDS
Authors: Ladoulis Charles T. , Gill Thomas J.
Published: May 2004

The detailed definition of the structure of DNA in chromosomes and in interphase chromatin is important for correlating the structure of the genetic material with various states of physiological activity. A general approach to developing specific reagents for a variety of such studies in solution and in tissues is to combine a chemically specific organic cation with the electron-opaque phosphotungstic acid (PTA) molecule. The reagent described in this paper was made from the interaction of acriflavine and phosphotungstic acid. The acriflavine-PTA complex (a) displays some unique absorption and fluorescence properties, (b) binds specifically to DNA and RNA by intercalation of the acriflavine moiety, and (c) is electron opaque. In addition, it binds to double-stranded synthetic polynucleotides, but not to a variety of proteins, nucleoproteins, or polysaccharides.
EVIDENCE FOR THE PARTICIPATION OF THE GOLGI APPARATUS IN THE INTRACELLULAR TRANSPORT OF NASCENT ALBUMIN IN THE LIVER CELL
Authors: Glaumann Hans , Ericsson Jan L. E.
Published: May 2004

A comparative biochemical and radioautographic in vivo study was performed to identify the site of synthesis and route of migration of albumin in the parenchymal liver cell after labeling with leucine-14C or leucine-3H via the portal vein. Free cytoplasmic ribosomes, membrane-bound ribosomes, rough- and smooth-surfaced microsomes, and Golgi membranes were isolated. The purity of the Golgi fraction was examined morphologically and biochemically. After administration of leucine-14C, labeled albumin was extracted, and the sequence of transport was followed from one fraction to the other. Approximately 2 min after the intravenous injection, bound ribosomes displayed a maximal rate of leucine-14C incorporation into albumin. 4 min later, a peak was reached for rough microsomes. Corresponding maximal activities for smooth microsomes were recorded at 15 min, and for the Golgi apparatus at ∼20 min. The relative amount of albumin, calculated on a membrane protein basis, was higher in the Golgi fraction than in the microsomes. By radioautography the silver grains were preferentially localized over the rough-surfaced endoplasmic reticulum at the 5 min interval. Apparent activity in the Golgi zone was noted 9 min after the injection; at 15 and 20 min, the majority of the grains were found in this location. Many of the grains associated with the Golgi apparatus were located over Golgi vacuoles containing 300–800 A electron-opaque bodies. It is concluded that albumin is synthesized on bound ribosomes, subsequently is transferred to the cavities of rough-surfaced endoplasmic reticulum, and then undergoes migration to the smooth-surfaced endoplasmic reticulum and the Golgi apparatus. In the latter organelle, albumin can be expected to be segregated together with very low density lipoprotein in vacuoles known to move toward the sinusoidal portion of the cell and release their content to the blood.
ISOLATION OF RAT LIVER PLASMA MEMBRANES
Authors: Touster Oscar , Aronson N. N. , Dulaney John T. , Hendrickson Herman
Published: May 2004

Nucleotide pyrophosphatase and phosphodiesterase I of rat liver have been found to be localized primarily in cell particulates highly enriched with respect to the most commonly accepted plasma membrane marker, 5'-nucleotidase, and therefore should themselves be assigned a plasma membrane localization. The observation that plasma membranes sediment in isotonic sucrose with both nuclear and microsomal fractions was exploited to obtain plasma membrane preparations from each fraction. Both preparations are similar in chemical and enzymic composition. Moreover, the preparative method developed in this study appears to give the best combination of yield, purity, and reproducibility available. The question of the possible identity of nucleotide pyrophosphatase and phosphodiesterase I is considered, and evidence is presented suggesting that these activities may be manifestations of the same enzyme.
STUDIES ON NUCLEAR STRUCTURE AND FUNCTION IN TETRAHYMENA PYRIFORMIS
Authors: Gorovsky Martin A.
Published: May 2004

Histones were extracted from isolated macro- and micronuclear fractions and from nucleohistone fibers which were prepared from the isolated macronuclear fraction. Analysis of these histones by polyacrylamide gel electrophoresis indicated that there are electrophoretic differences between the histones of macro- and micronuclei.
A CYTOLOGICAL STUDY OF THE CENTRIFUGED WHOLE, HALF, AND QUARTER EGGS OF THE SEA URCHIN ARBACIA PUNCTULATA
Authors: Anderson Everett
Published: May 2004

While the ooplasmic components of centrifuged eggs of Arbacia punctulata do not stratify in homogeneous layers, we have obtained the following strata beginning with the centripetal end: lipid droplets, pronucleus, clear zone, mitochondria, yolk, and pigment. Whereas mitochondria may be found mingled with yolk bodies, we have never observed lipid droplets nor pigment bodies among any of the other inclusions. The so-called clear zone contains a heterogeneous population of inclusions: annulate lamellae, heavy bodies, Golgi complexes, and rod-containing vacuoles. The peripheral cortical granules of immature (germinal vesicle stage) and of mature eggs are not dislodged from the cortical ooplasm with the centrifugal force utilized. When the eggs are treated with urethane, prior to centrifugation, the cortical granules of mature eggs abandon their peripheral position.

Further centrifugation of the initially stratified eggs produces nucleated and nonnucleated halves and the centrifugation of the halves results in quarters. The cytology of the halves and quarters is discussed. The halves and quarters have been activated with either sperm or hypertonic sea water. With the exception of the nucleated halves, we were unable to obtain plutei larvae from the other fractions (red halves and quarters). We believe that the lack of development of the various fragments is a function of the balance of particular inclusions necessary for differentiation.
RNA SYNTHESIS IN HELA CELLS
Authors: Pederson Thoru , Robbins Elliott
Published: May 2004

Interphase HeLa cells manifest a stepwise shutoff of RNA synthesis when the tonicity of the extracellular medium is gradually increased. Synthesis of heterogeneous nuclear RNA is most sensitive and is selectively inhibited at 1.5 times isotonicity (450 milliosmols/liter), while 45S ribosomal RNA synthesis is not affected significantly below 2.0 times isotonicity. Transfer RNA synthesis is least sensitive to increased osmolarity and is not completely inhibited until the electrolyte concentration of the medium is elevated to 2.8 times isotonicity. Although the transcription and methylation of 45S ribosomal precursor is unaffected at 1.5 times isotonicity, there is pronounced impairment of its processing into 32S and 18S RNA. Using a refined cell synchronization technique, we have been able to compare these effects of hypertonicity with the shutoff of RNA synthesis which occurs during the G2-prophase interval of the cell division cycle. In this case, as with random cells in hypertonic medium, a selective inhibition of heterogeneous nuclear RNA synthesis and slowed processing of 45S ribosomal RNA were found, whereas synthesis of 45S and transfer RNA continued unabated throughout G2-prophase. While it is known that RNA synthesis essentially ceases during metaphase, we have noted that transfer RNA synthesis continues in metaphase at 10–15% of the interphase rate, which is of particular interest in view of the relative resistance of this species to hypertonicity. The close correlation between the patterns of cessation of RNA synthesis at mitosis and during exposure to hypertonic medium supports our earlier contention that alteration of intracellular electrolyte levels provides a useful model for studying the mechanism of mitosis.
FERRITIN IN THE FUNGUS PHYCOMYCES
Authors: David Charles N. , Easterbrook Kenneth
Published: May 2004

The iron-protein ferritin has been purified from mycelium, sporangiophores, and spores of the fungus Phycomyces blakesleeanus. It has a protein-to-iron ratio of 5, a sedimentation coefficient of 55S, a buoyant density in CsCl of 1.82 g/cm3, and the characteristic morphology of ferritin in the electron microscope. Apoferritin prepared from Phycomyces ferritin has a sedimentation coefficient of 18S and consists of subunits of molecular weight 25,000. In the cytoplasm of Phycomyces, ferritin is located on the surface of lipid droplets (0.5–2.0 µ in diameter) where it forms crystalline monolayers which are conspicuous in electron micrographs of sporangiophore thin-sections. Ferritin is found in all developmental stages of Phycomyces but is concentrated in spores. The level of ferritin iron is regulated by the iron level in the growth medium, a 50-fold increase occurring on iron-supplemented medium.
FACTORS CONTROLLING DEVELOPMENT OF CHICK EMBRYO LIVER CELLS DURING ORGAN CULTURE
Authors: Benzo Camillo A. , Nemeth Andrew M.
Published: May 2004

The 5 day chick embryo liver cell still lacks many of the ultrastructural features of the adult liver cell. During organ culture on rafts over Eagle's medium, it develops electron-opaque mitochondria with granules, biliary microvilli, and compact Golgi complexes containing what appears to be secretory material. Rough ER proliferates and free ribosomes become bound to membrane. Thus, the 5 day cell, exposed only to simple nutrients (glucose, amino acids, vitamins) develops the general appearance of the adult liver cell except for the continued absence of smooth ER and glycogen. The significance of this incomplete differentiation and the factors controlling development are discussed in the light of accompanying metabolic changes.
5-AMINOURACIL TREATMENT
Authors: Socher S. H. , Davidson D.
Published: May 2004

Treatment of Vicia faba lateral roots with a range of concentrations of 5-aminouracil (5-AU) indicate that cells are stopped at a particular point in interphase. The timing of the fall in mitotic index suggests that cells are held at the S - G2 transition. When cells are held at this point, treatments with 5-AU can be used to estimate the duration of G2 + mitosis/2 of proliferating cells. Treatment with 5-AU can also be used to demonstrate the presence of subpopulations of dividing cells that differ in their G2 duration. Using this method, 5-AU-induced inhibition, we have confirmed that in V. faba lateral roots there are two populations of dividing cells: (a) a fast-dividing population, which makes up ∼85% of the proliferating cell population and has a G2 + mitosis/2 duration of 3.3 hr, and (b) a slow-dividing population, which makes up ∼15% of dividing cells and has a G2 duration in excess of 12 hr. These estimates are similar to those obtained from percentage labeled mitosis (PLM) curves after incorporation of thymidine-3H.
THE MORPHOLOGY OF THE SWELLING PROCESS IN RAT LIVER MITOCHONDRIA
Authors: Myron D. R. , Connelly J. L.
Published: May 2004

Through the use of combined spectrophotometric and electron microscope techniques, large amplitude swelling of rat liver mitochondria has been described as an ordered sequence of ultrastructural transitions. Prior to the actual swelling, mitochondria undergo two major conformational changes: condensed to twisted form and twisted to orthodox form. This sequence is independent of (a) the nature of swelling agents and (b) the time of onset of swelling. Agents that delay the onset of swelling act to increase the duration of the twisted conformation. Agents that prevent extensive swelling hold mitochondria in intermediate conformations. Gross swelling, immediately preceded by a decrease in electron opacity of the matrix, involves the rupture of the outer membrane and expansion of the inner compartment of the mitochondrion.
SELECTIVE EXTRACTION OF ISOLATED MITOTIC APPARATUS
Authors: Bibring Thomas , Baxandall Jane
Published: May 2004

Mitotic apparatus isolated from sea urchin eggs has been treated with meralluride sodium under conditions otherwise resembling those of its isolation. The treatment causes a selective morphological disappearance of microtubules while extracting a major protein fraction, probably consisting of two closely related proteins, which constitutes about 10% of mitotic apparatus protein. Extraction of other cell particulates under similar conditions yields much less of this protein. The extracted protein closely resembles outer doublet microtubule protein from sea urchin sperm tail in properties considered typical of microtubule proteins: precipitation by calcium ion and vinblastine, electrophoretic mobility in both acid and basic polyacrylamide gels, sedimentation coefficient, molecular weight, and, according to a preliminary determination, amino acid composition. An antiserum against a preparation of sperm tail outer doublet microtubules cross-reacts with the extract from mitotic apparatus. On the basis of these findings it appears that microtubule protein is selectively extracted from isolated mitotic apparatus by treatment with meralluride, and is a typical microtubule protein.
THE GEOMETRY OF PERIPHERAL MYELIN SHEATHS DURING THEIR FORMATION AND GROWTH IN RAT SCIATIC NERVES
Authors: Webster Henry deF.
Published: May 2004

In rat sciatic nerves, a small bundle of fibers was identified in which myelin sheaths were absent at birth, appeared within 3 days, and grew rapidly for 2 wk. During this interval, nerves were removed from littermates and were sectioned serially in the transverse plane. Alternating sets of thin and thick sections were used to prepare electron micrograph montages in which single myelinating axons could be identified and traced distally. During the formation of the first spiral turn, the mesaxon's length and configuration varied when it was studied at different levels in the same Schwann cell. The position of the mesaxon's termination shifted while its origin, at the Schwann cell surface, remained relatively constant. Along myelin internodes composed of two to six spiral turns, there were many variations in the number of lamellae and their contour. Near the mesaxon's origin, longitudinal strips of cytoplasm separated the myelin layers. Thicker sheaths were larger in circumference, more circular in transverse sections, and more uniform at different levels. Irregularities were confined to the paranodal region, and separation of lamellae by cytoplasm occurred at Schmidt-Lantermann clefts. Approximate dimensions of the bundle, its largest fibers, and their myelin sheaths were measured and calculated. The myelin membrane's transverse length and area increased exponentially with time; the growth rate increased rapidly during the formation of the first four to six spiral layers and remained relatively constant during the subsequent enlargement of the compact sheath.