bluuebunny/crossref_metadata_2025 huggingface dataset:
DISTRIBUTION OF LEUCINE-3H DURING AXOPLASMIC TRANSPORT WITHIN REGENERATING NEURONS AS DETERMINED BY ELECTRON-MICROSCOPE RADIOAUTOGRAPHY
Authors: Lentz Thomas L.
Published: May 2004

The distribution of leucine-3H in neurons was determined by electron-microscope radioautography after infusion of label into the spinal cord or sensory ganglia of regenerating newts. In the nerve cell bodies 3 days after infusion, the highest concentration of label per unit area occurred over the rough-surfaced endoplasmic reticulum. In the large brachial nerves, the silver grains were not distributed uniformly in the axoplasm, indicating that the labeled materials are restricted in their movement to certain regions of the axon. Almost all of the radioautographic grains observed in myelinated nerves could be accounted for by the presence of a uniformly labeled band occupying the area 1500–9000 A inside the axolemma. This region of the axon was rich in microtubules and organelles while the unlabeled central core of the axon contained mainly neurofilaments. This observation supports the hypothesis that microtubules are related to axonal transport. In small, vesicle-filled nerve terminals in the blastema, labeled material was restricted to a thin zone a short distance beneath the plasma membrane while the central region of the terminal was largely unlabeled. The peripheral pattern of labeling in the nerve endings is consistent with successive addition of newly synthesized proteins at the periphery of the growth cone and release of substances such as trophic factors at the nerve terminal.
ISOLATION AND PARTIAL CHARACTERIZATION OF MACRO- AND MICRONUCLEI FROM PARAMECIUM AURELIA
Authors: Cummings Donald J.
Published: May 2004

A method was developed for the isolation of macro- and micronuclei from Paramecium aurelia. This method utilized ionic and nonionic detergents to rupture the intact cells, calcium ions and spermidine were employed to protect the nuclei, and the nuclei were purified by centrifugation. Macronuclei consisted of 22% DNA, 10% RNA, and 68% protein. Micronuclei were composed of 9% DNA, 11% RNA, and 80% protein. DNA from both macro- and micronuclei had a density of 1.687 g/cc in CsCl and 1.417 g/cc in Cs2SO4. These values corresponded to G + C content of about 23%. The RNA of macronuclei was examined by gel electrophoresis, and two high molecular weight species were identified having molecular Weights of 1.3 x 106 and 2.8 x 106 daltons. Three syngens were studied, and in each case the conditions for isolation of the nuclei were the same and no differences were observed in the properties of the nuclei.
RESPIRATORY ENZYMES AND MITOCHONDRIAL MORPHOLOGY OF HELA AND L CELLS TREATED WITH CHLORAMPHENICOL AND ETHIDIUM BROMIDE
Authors: King Mary E. , Godman Gabriel C. , King Donald W.
Published: May 2004

Exposure of HeLa and L cells to chloramphenicol causes a progressive dose-dependent decrease in cytochrome oxidase and succinate-cytochrome c reductase activities, concomitant with an increase in the amount of cytochrome c. At 2–3 days, the specific activities of the enzymes have fallen to about one-half of control values; the mitochondria appear swollen. By day 5, enzyme activities are about one-quarter of control values; the mitochondria are more swollen, with disorientation and disintegration of cristae. By day 6–8, after three generations, growth has stopped, enzyme activities are approximately the same as on day 5, and cytochrome c content has reached 170% of control value. Mitochondria show severe changes, cristae being affected more than peripheral inner membrane. The number of profiles continues to be nearly normal. After 30 days, cytochrome oxidase activity remains low but now there are mitochondria in intermediate and condensed configuration. There is a gradual accumulation in the cytoplasm of smooth membrane elements. If chloramphenicol is removed, cells recover. Ethidium bromide treatment for up to 8 days yields results virtually identical to those obtained with chloramphenicol. Cells treated with 10-4 M KCN show a decrease in cytochrome oxidase activity to about one-third of control value and an elevated amount of cytochrome c. Only a small number of mitochondria appear damaged. Autochthonous mitochondrial syntheses appear to be essential for the organization of the cristae. When cytochrome oxidase activity is impaired, a regulatory mechanism for cytochrome biosynthesis geared to mitochondrial function may be lacking, resulting in an increase in cytochrome c content.
THE RESPONSE OF VENTRAL HORN NEURONS TO AXONAL TRANSECTION
Authors: Price Donald L. , Porter Keith R.
Published: May 2004

The morphological changes induced in the frog ventral horn neurons by axonal transection have been studied with the electron microscope. During the first 2 wk after axotomy the neuronal nucleus becomes more translucent and the nucleolus becomes enlarged and less compact. The cisternae of the granular endoplasmic reticulum vesiculate and ribosomes dissociate from membranes. Free ribosomes and polysomes are dispersed in the cytoplasmic matrix. Neurofilaments and neurotubules are increased in number. These structures appear to be important in the regeneration of the axon. It is proposed that neurotubules, neurofilaments, and axoplasmic matrix are synthesized by the free polyribosomes in the chromatolytic neuron. By the fourth postoperative week, the neurons show evidence of recovery. The cytoplasm is filled with profiles of granular endoplasmic reticulum and many intercisternal polysomes. The substances being manufactured by the newly formed granular endoplasmic reticulum are not clearly defined, but probably include elements essential to electrical and chemical conduction of impulses. The significance of these observations in respect to recent studies of axoplasmic flow is discussed.
FINE STRUCTURAL OBSERVATIONS RELATING TO THE PRODUCTION OF COLOR BY THE IRIDOPHORES OF A LIZARD, ANOLIS CAROLINENSIS
Authors: Rohrlich Susannah T. , Porter Keith R.
Published: May 2004

This paper presents the results of light and electron microscopy done on iridophores in the dorsal skin of the lizard Anolis carolinensis. New fine-structural details are revealed, and their importance is discussed. Of some interest is the complex of filaments between crystalline sheets in the cell. It is proposed that this complex is involved in the arrangement of crystals into crystalline sheets, and that the crystal arrangement and spacing are critical for the production of the cells' blue-green color. Tyndall scattering and thin-film interference are discussed as possible explanations for iridophore color production in relation to the fine-structural data obtained.
ACRIFLAVIN RESISTANCE IN THE HEMOFLAGELLATE, LEISHMANIA TARENTOLAE
Authors: Strauss Phyllis R.
Published: May 2004

The accumulation, metabolism, and distribution of acriflavin (acr) in two culture strains of Leishmania tarentolae were studied. One strain, reported previously, was sensitive to the dye, i.e. became dyskinetoplastic and could not be subcultured in the presence of 470 ng/ml acr, and one was resistant. Accumulation was studied by fluorescence of the dye within cells and by uptake of acr-3H by cells. Metabolism was studied by paper chromatography of aqueous extracts from cells grown with acr-3H, and distribution was examined by fluorescence and quantitative electron microscope radioautography. Substances affecting the response to acr included hemin and an acr-sensitizing factor initially obtained from red cells but here shown to be distinct from hemoglobin. In the presence of the sensitizing factor or in the absence of hemin, the resistant strain became dyskinetoplastic and could not be subcultured. Acr fluorescence appeared in the nucleus of the resistant strain, and the percentage of radioautography grains appearing in the nucleus increased. Under these conditions the distribution of radioactivity from chromatographed extracts was altered from the normal in a similar fashion. Because sensitization of the resistant strain is associated with increased amounts of acr in the nucleus, that organelle may be implicated in the mode of action of acr. In general, the two strains behaved alike except for (a) the response to acr, (b) the arginine requirement for optimal growth, and (c) the sensitivity to cycloheximide. Thus, one cannot exclude the wider possibility that acr may act on the cytoplasm and the nucleus as well as on the mitochondrion.
A BIOCHEMICAL AND RADIOAUTOGRAPHIC ANALYSIS OF PROTEIN SECRETION BY THYROID LOBES INCUBATED IN VITRO
Authors: Feeney Lynette , Wissig Steven L.
Published: May 2004

In this study we analyzed several aspects of protein secretion by thyroid follicular cells. The study was carried out on intact thyroid lobes obtained from newborn rats and incubated in vitro. The fate of leucine-3H incorporated into protein within follicular cells of untreated and thyrotropic hormone (TSH)-treated lobes was traced by quantitative electron microscope radioautography. Our findings indicate that protein synthesized by the rough-surfaced endoplasmic reticulum during a pulse exposure to leucine-3H is released relatively slowly by this organelle. Approximately 1 hr after onset of the pulse, a peak of radioactive protein appears in the Golgi region. The significance of this peak is not clear. Newly synthesized secretory protein passes through the apex of follicular cells without being concentrated or temporarily stored there in the form of large secretory droplets. Passage probably takes place via small vesicles which are intermingled among diverse small vesicles at the apex of the cells as well as in the Golgi region. Exposure of the lobes to TSH in the incubation medium for 45 or 90 min does not stimulate incorporation of leucine-3H into protein. Acute stimulation with TSH does, however, modify the movement of secretory protein within the exocrine secretory apparatus of the follicular cell. It accelerates the arrival of the protein at the apex of follicular cells, and it accelerates the release of the protein into the follicular lumen.
EFFECTS OF α-AMANITIN ON IN VITRO LABELING OF RNA FROM DEFINED NUCLEAR COMPONENTS IN SALIVARY GLAND CELLS FROM CHIRONOMUS TENTANS
Authors: Egyházi E. , D'Monte B. , Edström J.-E.
Published: May 2004

The effect of α-amanitin on nucleoside labeling of RNA in nucleoli, chromosomes, nuclear sap, and cytoplasm from Chironomus tentans salivary gland cells was investigated by radioautography and gel electrophoresis. Preribosomal RNA formation and processing in the nucleolus was not measurably influenced by the drug, and both 28 S and 18 S ribosomal RNA were transferred to the cytoplasm. In the chromosomes the heterogeneous RNA labeling was completely inhibited for the large size range (above 45–50 S) and partially for the low range. The labeling of 4–5 S chromosomal RNA was only moderately reduced. Most of the chromosomes showed radioautographically a disappearance of the normal band pattern, but some retained a pattern of weakly labeled bands. The electrophoretic results for the nuclear sap paralleled those for the chromosomes. The effect of α-amanitin on RNA labeling in these cells is similar but not identical to that of the substituted benzimidazole 5,6-dichloro-1(ß-D-ribofuranosyl) benzimidazole (DRB).
PEROXISOMES IN ABSORPTIVE CELLS OF MAMMALIAN SMALL INTESTINE
Authors: Novikoff Phyllis M. , Novikoff Alex B.
Published: May 2004

Huge numbers of peroxisomes are present in guinea pig duodenum, jejunum, and ileum, and in rat duodenum. The peroxisomes have been studied by light and electron microscopy, including visualization by incubation in a newly-developed alkaline 3,3' diaminobenzidine (DAB) medium. Electron micrographs of more than 3700 guinea pig peroxisomes have been studied. The diameter of most peroxisomes ranges from 0.15 µ. to 0.25 µ. They often appear in clusters, surrounded by and continuous, in numerous places, with smooth endoplasmic reticulum (ER). The ER is extremely tortuous in these regions. Serial sectioning is valuable for studying the ER-peroxisome relationships but viewing sections at different angles, tilted with a goniometer stage, is more informative. The intimate relations of the two organelles appear the same in tissue fixed in four different fixatives. The peroxisomes may be interpreted as localized dilatations of smooth ER retaining multiple membranous continuities. This interpretation is discussed in light of the turnover data on peroxisomal proteins of rat hepatocytes reported by Poole and colleagues. The very large numbers of peroxisomes in intestinal epithelium lead to speculations concerning their functional significance. They resemble the small peroxisomes described in many other cell types. Although the distinctive relationship of these peroxisomes to the ER is probably more significant than their small size, for practical purposes we propose the term "microperoxisomes" to distinguish these peroxisomes from the better-known larger peroxisomes of liver and kidney.
COMPARATIVE ANALYSIS OF MAMMALIAN SPERM MOTILITY
Authors: Phillips David M.
Published: May 2004

Spermatozoa of several mammalian species were studied by means of high-speed cinematography and electron microscopy. Three types of motile patterns were observed in mouse spermatozoa. The first type involved an asymmetrical beat which seemed to propel the sperm in circular paths. The second type involved rotation of the sperm and appeared to allow them to maintain straight paths. In the third type of pattern, the sperm appeared to move by crawling on surfaces in a snakelike manner. Spermatozoa of rabbit and Chinese hamster also had an asymmetrical beat which sometimes caused them to swim in circles. In spite of the asymmetry of the beat, these spermatozoa were also able to swim in straight paths by rotating around a central axis as they swam. Spermatozoa of some species appeared very flexible; their flagella formed arcs with a very small radius of curvature as they beat. Spermatozoa of other species appeared very stiff, and their flagella formed arcs with a very large radius of curvature. The stiffness of the spermatozoan appeared to correlate positively with the cross-sectional area of the dense fibers. This suggests that the dense fibers may be stiff elastic elements. Opossum sperm become paired as they pass through the epididymis. Pairs of opossum spermatozoa beat in a coordinated, alternating manner.
EVIDENCE FOR A TEMPORAL INCOMPATIBILITY BETWEEN DNA REPLICATION AND DIVISION DURING THE CELL CYCLE OF TETRAHYMENA
Authors: Jeffery William R.
Published: May 2004

The mechanism of coordination between DNA replication and cell division was studied in Tetrahymena pyriformis GL-C by manipulation of the timing of these events with heat shocks and inhibition of DNA synthesis. Preliminary experiments showed that the inhibitor combination methotrexate and uridine (M + U) was an effective inhibitor of DNA synthesis. Inhibition of the progression of DNA synthesis with M + U in exponentially growing cells, in which one S period usually occurs between two successive divisions, or in heat-shocked cells, when successive S periods are known to occur between divisions, resulted in the complete suppression of the following division. In further experiments in which the division activities were reassociated with the DNA synthetic cycle by premature termination of the heat-shock treatment, it was shown that (a) the completion of one S period during the treatment was sufficient for cell division, (b) the beginning of division events suppressed the initiation of further S periods, and (c) if further S periods were initiated while the heat-shock treatment was continued, division preparations could not begin until the necessary portion of the S period was completed, even though DNA had previously been duplicated. It was concluded that a temporal incompatibility exists between DNA synthesis and division which may reflect a coupling mechanism which insures their coordination during the normal cell cycle.
THE LOCALIZATION OF ENDOGENOUS PEROXIDASE IN THE LACRIMAL GLAND OF THE RAT DURING POSTNATAL DEVELOPMENT
Authors: Herzog V. , Miller F.
Published: May 2004

The distribution of endogenous peroxidase activity in the lacrimal gland of the rat during postnatal development was investigated by electron microscope cytochemistry Peroxidase activity is first found 6 hr after birth in only a few acinar cells At this stage, reaction product fills only localized segments of the scant rough endoplasmic reticulum and of the perinuclear cisternae. Peroxidase activity thus develops asynchronously in a given cell as well as in the secretory cell population as a whole 2 days after birth, all cisternae of the rough endoplasmic reticulum of a peroxidase-positive cell contain reaction product, but the majority of the acinar cells is still negative During the next days, the number of peroxidase-positive cells and the amount of the rough endoplasmic reticulum increase rapidly. By 15 days postparturition, all secretory cells are peroxidase-positive. Reaction product is then found in all cisternae of the rough endoplasmic reticulum including the perinuclear cisternae, in smooth surface vesicles located mainly between the rough endoplasmic reticulum and the Golgi stacks, in condensing vacuoles, and in all secretory granules The Golgi cisternae rarely contain reaction product In total homogenates and in fractions of glandular tissue of adult rats, peroxidatic and catalatic activities are demonstrable. The microsomal fractions and the postmicrosomal supernatants were used to separate peroxidase from catalase by precipitation with ammonium sulfate, and the following parameters were determined: substrate (H2O2-) optimum (∼ 2.0 x 10-4M), pH-optimum (pH 6 5), temperature-optimum (42°C), and the absorption maximum (415 nm before and 425 nm after addition of H2O2) The same parameters were obtained from lacrimal fluid peroxidase. Both peroxidase from lacrimal gland and that from lacrimal fluid are almost completely inhibited by 10-3M aminotriazole and are possibly identical enzymes. Peroxidase is secreted into lacrimal fluid, which does not contain catalase.
PINOCYTOSIS IN ACANTHAMOEBA CASTELLANII
Authors: Bowers Blair , Olszewski Thomas E.
Published: May 2004

The uptake of radioactively labeled albumin, inulin, leucine, and glucose by Acanthamoeba castellanii (Neff strain) was measured. The uptake is linear with time and appears to be continuous under the conditions of these experiments. Uptake is abolished at 0°C. No evidence for saturation of the uptake mechanism was obtained with either albumin or leucine. Each of the four tracer molecules enters the ameba at a similar rate when the uptake is calculated as volume of fluid ingested per unit time. The data suggest that each of these molecules enters the cell by pinocytosis. The highest rate of uptake was obtained with cells in their usual culture medium containing proteose peptone, glucose, and salts but pinocytosis also continued at a reduced rate in a simple salt solution. The calculated volume of fluid taken in during pinocytosis in culture medium was about 2 µl/hr per 106 cells. The route of uptake was examined in the electron microscope using horseradish peroxidase (HRP) as a tracer. HRP activity was found exclusively within membrane profiles within the cytoplasm, confirming the pinocytotic mode of uptake. An estimate of the rate of surface membrane turnover due to pinocytosis was made using the biochemical and morphological data obtained. This estimate suggests that the plasma membrane turnover of one cell is on the order of several times an hour.
PHYSICAL STUDIES OF ISOLATED EUCARYOTIC NUCLEI
Authors: Olins Donald E. , Olins Ada L.
Published: May 2004

The degree of chromatin condensation in isolated rat liver nuclei and chicken erythrocyte nuclei was studied by phase-contrast microscopy as a function of solvent pH, K+ and Mg++ concentrations Data were represented as "phase" maps, and standard solvent conditions selected that reproducibly yield granular, slightly granular, and homogeneous nuclei Nuclei in these various states were examined by ultraviolet absorption and circular dichroism (CD) spectroscopy, low-angle X-ray diffraction, electron microscopy, and binding capacity for ethidium bromide Homogeneous nuclei exhibited absorption and CD spectra resembling those of isolated nucleohistone. Suspensions of granular nuclei showed marked turbidity and absorption flattening, and a characteristic blue-shift of a crossover wavelength in the CD spectra. In all solvent conditions studied, except pH < 2 3, low-angle X-ray reflections characteristic of the native, presumably superhelical, nucleohistone were observed from pellets of intact nuclei. Threads (100–200 A diameter) were present in the condensed and dispersed phases of nuclei fixed under the standard solvent conditions, and examined in the electron microscope after thin sectioning and staining Nuclei at neutral pH, with different degrees of chromatin condensation, exhibited similar binding capacities for ethidium bromide. These data suggest a model that views chromatin condensation as a close packing of superhelical nucleohistone threads but still permits condensed chromatin to respond rapidly to alterations in solvent environment.
VARIATIONS IN TIGHT AND GAP JUNCTIONS IN MAMMALIAN TISSUES
Authors: Friend Daniel S. , Gilula Norton B.
Published: May 2004

The fine structure and distribution of tight (zonula occludens) and gap junctions in epithelia of the rat pancreas, liver, adrenal cortex, epididymis, and duodenum, and in smooth muscle were examined in paraformaldehyde-glutaraldehyde-fixed, tracer-permeated (K-pyroantimonate and lanthanum), and freeze-fractured tissue preparations. While many pentalaminar and septilaminar foci seen in thin-section and tracer preparations can be recognized as corresponding to well-characterized freeze-fracture images of tight and gap junction membrane modifications, many others cannot be unequivocally categorized—nor can all freeze-etched aggregates of membrane particles. Generally, epithelia of exocrine glands (pancreas and liver) have moderate-sized tight junctions and large gap junctions, with many of their gap junctions basal to the junctional complex. In contrast, the adrenal cortex, a ductless gland, may not have a tight junction but does possess large gap junctions. Mucosal epithelia (epididymis and intestine) have extensive tight junctions, but their gap junctions are not as well developed as those of glandular tissue. Smooth muscle contains numerous small gap junctions The incidence, size, and configuration of the junctions we observed correlate well with the known functions of the junctions and of the tissues where they are found.
THE MOBILIZATION AND EXTRACELLULAR RELEASE OF GRANULAR ENZYMES FROM HUMAN LEUKOCYTES DURING PHAGOCYTOSIS
Authors: Wright Daniel G. , Malawista Stephen E.
Published: May 2004

The importance of granular (lysosomal) enzymes from neutrophils in producing the tissue damage of acute inflammation has been suggested by much indirect and some direct evidence. This study has investigated the kinetics of release and subsequent fate of granular enzymes from phagocytizing human leukocytes The following observations are made: (a) During phagocytosis, the granular enzyme lysozyme is released from leukocytes into the extracellular medium. (b) Release of lysozyme increases as phagocytic challenge increases, but attains a maximum. (c) Release of lysozyme accompanies phagocytosis and is not a delayed event. (d) The lack of release of a nongranular enzyme, lactic dehydrogenase, indicates that cell damage is not a necessary condition of enzyme release. (e) Like lysozyme, ß-glucuronidase is released from phagocytizing leukocytes. Acid α-naphthyl phosphatase and cathepsin also appear to be released, but are not found in appreciable amounts in the extracellular medium, in part because of their lability in solution. These results support the concept that extracellular release of granular enzymes may be a useful secretory function of inflammatory leukocytes which becomes damaging to the host in certain circumstances.
DETECTION OF DNA BY TRITIATED ACTINOMYCIN D ON ULTRATHIN FROZEN SECTIONS
Authors: Bernier Roch , Iglesias Roberto , Simard René
Published: May 2004

Ultrathin frozen sections of fresh liver tissue were floated on actinomycin D-3H. Quantitative high resolution radioautography was performed to determine the value of the method for detection of DNA by electron microscopy. A complete series of control experiments involving various treatments of frozen sections with enzymes (pronase, DNase) and 0.1 N HCl were also carried out to determine the specificity of the labeling. The results indicate the value of the method for detection of DNA directly on ultrathin frozen sections. Short treatments with pronase followed by DNase reduce the labeling to zero, whereas removal of chromosomal proteins with HCl increases the amount of radioactivity in the nucleus considerably. The results are discussed in view of the future applications opened by ultracryotomy, since radioautographic detection of various macromolecules and cellular components by labeled compound with specific affinities will now be possible.
PHYSICOCHEMICAL EFFECTS OF ALDEHYDES ON THE HUMAN ERYTHROCYTE
Authors: Vassar P. S. , Hards J. M. , Brooks D. E. , Hagenberger B. , Seaman G. V. F.
Published: May 2004

The effects of formaldehyde, acetaldehyde, and glutaraldehyde on human red blood cells were investigated. It was found that (a) The surface negative charge of the erythrocytes at pH 7 was increased 10% by glutaraldehyde, but not by the other two aldehydes. (b) The effect of incomplete fixation of the red blood cells was demonstrated by hemoglobin leakage studies The leakage of hemoglobin subsequent to formaldehyde treatment was especially pronounced Acetaldehyde-fixed cells showed some leakage of hemoglobin after an hour of exposure to the fixative, whereas glutaraldehyde-fixed cells showed no hemoglobin leakage. (c) All three aldehydes caused K+ leakage during fixation. The concentrations of K+ in the fixing solutions all reached the same level, but whereas the leakage with glutaraldehyde was immediate, that with formaldehyde was more gradual and that with acetaldehyde reached a steady state only after 24 hr. (d) The effects of the aldehydes on red cell deformability and swelling revealed that glutaraldehyde hardened the cells within 15 min, formaldehyde within 5 hr, while acetaldehyde required at least 24 hr to produce appreciable fixation. (e) The hematocrit changes accompanying the fixation process depended upon cell volume changes and loss of deformability.
LIVER SINUSOIDAL CELLS
Authors: Bissell D. Montgomery , Hammaker Lydia , Schmid Rudi
Published: May 2004

Sequestration and degradation of red blood cells (RBC) are believed to occur in part in the liver, but the magnitude and cellular localization of this process remain uncertain. This problem was studied in rats by investigating isolated parenchymal and sinusoidal cell populations of the liver. After digesting the perfused liver with pronase, hepatic sinusoidal cells were isolated free of RBC and debris. Of the isolated cells, 90% were phagocytic, as judged by their uptake of colloidal 198Au or of aggregated albumin-131I administered in vivo After administration of spherocytic (heat-treated) RBC, however, only about one quarter of the isolated cells were found to contain phagocytized RBC. This apparently distinct population of RBC-phagocytizing cells is designated as "erythrophagocytic (EP)" cells. The EP cell population was further characterized functionally by its specific phagocytosis of colloidal carbon and of 99mtechnetium-sulfur colloid and histochemically by its peroxidase activity. The role of the EP population in the catabolism of RBC-hemoglobin was studied in isolated hepatic sinusoidal cells by assay of microsomal heme oxygenase (MHO), which is the inducible enzyme system that converts heme to bilirubin. The MHO activity of individual sinusoidal isolates was related directly to their content of EP cells Assay of the MHO activity of the whole spleen and of the total EP cell population of the liver suggested that these two tissues may be of comparable importance in their ability to degrade RBC-hemoglobin.
COLCHICINE-BINDING PROTEIN AND THE SECRETION OF THYROID HORMONE
Authors: Williams John A. , Wolff J.
Published: May 2004

The role of microtubules in the thyrotropin- or adenosine 3',5' cyclic monophosphate (cyclic AMP)-stimulated accumulation of cytoplasmic colloid droplets and secretion of iodine from the mouse thyroid gland has been investigated by means of different classes of agents that affect the stability of microtubules. The onset of inhibition of secretion by colchicine, the uptake of colchicine-3H by thyroid lobes, and the binding of colchicine-3H to thyroidal soluble protein are shown to have similar time courses Colloid droplet accumulation is also inhibited and does not readily resume upon removal of colchicine from the medium. This appears to be due to the slow washout of the drug (t½ ∼ hr). Thyroids contain a soluble colchicine-binding protein that resembles microtubule proteins of other tissues with respect to apparent Km for colchicine, pH optimum, and stability characteristics Colchicine analogues inhibit iodine secretion and colchicine binding in a parallel manner and as a function of their antimitotic potencies. Microtubule-stabilizing agents such as hexylene glycol and D2O also inhibit secretion. Thus, inhibition of thyroid secretion by antimitotic agents appears to be mediated by an effect on microtubules. The inhibitory locus of colchicine inhibition occurs after the generation of cyclic AMP, since stimulation of secretion by this nucleotide is blocked by colchicine, whereas thyroid-stimulating hormone—induced accumulation of cyclic AMP is not affected. Thus, the functioning microtubule appears to play a role in the induction of colloid endocytosis.
ROLE OF THE SARCOPLASMIC RETICULUM IN GLYCOGEN METABOLISM
Authors: Wanson Jean-Claude , Drochmans Pierre
Published: May 2004

Sarcoplasmic vesicles and ß-glycogen particles 30–40 mµ in diameter were isolated from perfused rabbit skeletal muscle by the differential precipitation-centrifugation method. This microsomal fraction was subjected to zonal centrifugation on buffered sucrose gradients, in a B XIV Anderson type rotor, for 15 hr at 45,000 rpm in order to separate the two cytoplasmic organelles. Zonal profiles of absorbance at 280 mµ, proteins, glycogen, and enzymatic activities (phosphorylase b kinase, phosphorylase b, and glycogen synthetase) were performed. Whereas the entire synthetase activity was found combined with the glycogen particles, 39% of phosphorylase and 53% of phosphorylase b kinase activities, present in the microsomal fraction, were recovered in the purified vesicular fraction (d = 1.175). This latter fraction consists of vesicles, derived from the sarcoplasmic reticulum, and of small particles 10–20 mµ in diameter attached to the outer surface of the membranes. These particles disappear after α-amylase treatment. Incubation of the sarcovesicular fraction with 14C-labeled glucose-1-phosphate confirms the localization of a polysaccharide synthesis at the level of the membranes. "Flash activation" of phosphorylase b, i.e. Ca "activation" of phosphorylase kinase followed by a conversion of phosphorylase b into a, was demonstrated in the purified sarcovesicular fraction. Moreover, the active enzymatic sites were detected on the membranes by electron microscopy. The presence of binding sites between the membranes of the sarcoplasmic vesicles and a glycogen-enzyme complex suggests that this association plays a role in the glycogenolysis during muscle contraction.
THE BIOSYNTHESIS AND CONTENT OF GAMMA-AMINOBUTYRIC ACID IN THE GOLDFISH RETINA
Authors: Lam Dominic M. K.
Published: May 2004

Goldfish retinas incubated with L-glutamate-14C (UL) were found to synthesize γ-aminobutyric acid-14C (GABA-14C) The accumulation of newly synthesized GABA was enhanced by physiological stimulation of the retina with flashing light; and this increase was directly proportional to the logarithm of the light intensity. The total GABA content was also higher in light-stimulated than in dark-adapted retinas, although the glutamate content remained unchanged No differences were found in the cell-free activities of glutamate decarboxylase (EC 4 1.1 15) and GABA-glutamate transaminase (EC 2.6.1.19) extracted from light-stimulated and dark-adapted retinas. These findings, together with other physiological and morphologcal evidence, suggest that GABA plays a functional role in synaptic transmission in the goldfish retina
THE THREE-DIMENSIONAL STRUCTURE OF THE BASAL BODY FROM THE RHESUS MONKEY OVIDUCT
Authors: Anderson Richard G. W.
Published: May 2004

The structure of the oviduct basal body has been reconstructed from serial, oblique, and tangential sections This composite information has been used to construct a three-dimensional scale model of the organelle The walls are composed of nine equally spaced sets of three tubules, which run from base to apex pitched to the left at a 10°–15° angle to the longitudinal axis. The transverse axis of each triplet set at its basal end intersects a tangent to the lumenal circumference of the basal body at a 40° angle (triplet angle). As the triplet set transverses from base to apex, it twists toward the lumen on the longitudinal axis of the inner A tubule; therefore, the triplet angle is 10° at the basal body-cilium junction. Strands of fibrous material extend from the basal end of each triplet to form a striated rootlet. A pyramidal basal foot projects at right angles from the midregion of the basal body. In the apex, a 175 mµ long trapezoidal sheet is attached to each triplet set. The smaller of the two parallel sides is attached to all three tubules while the longitudinal edge (one of the equidistant anti-parallel sides) is attached to the C tubule. The sheet faces counterclockwise (apex to base view) and gradually unfolds from base to apex; the outside corner merges with the cell membrane.
CHANGES IN THE ORGANIZATION OF TUBULIN DURING MEIOSIS IN THE EGGS OF THE SURF CLAM, SPISULA SOLIDISSIMA
Authors: Weisenberg Richard C.
Published: May 2004

Polymerized tubulin can be stabilized in Kane's spindle isolation medium (HGL solution), isolated by differential centrifugation and then assayed by colchicine binding activity. In the eggs of the surf clam, Spisula solidissima, the level of particulate tubulin undergoes a series of specific changes during first meiotic division. In either unactivated ("interphase") eggs or metaphase eggs the amount of particulate tubulin was about 13% of the total at 23°C. The amount of particulate tubulin decreased shortly after activation, reaching a minimum value at about 5 min, the time of nuclear membrane breakdown. The particulate tubulin concentration then rose, reaching a maximum at metaphase, and then decreased again during anaphase, reaching a minimum at first polar body formation. In HGL homogenates of unactivated eggs a structure is present which has been shown to contain the interphase particulate tubulin (IPT). This structure consists essentially of a 10–20 µ granular sphere attached to a membranous material which is probably part of the egg cortex. These particles are absent at the time of nuclear membrane breakdown, when the level of particulate tubulin is minimal and when the first signs of spindle formation are visible. Electron microscopy of these particles by negative staining indicates that they are composed of microtubules associated with a granular matrix which may be a polymorphic aggregate of tubulin.
BIOSYNTHESIS IN ISOLATED ACETABULARIA CHLOROPLASTS
Authors: Shephard David C. , Levin Wendy B.
Published: May 2004

The ability of chloroplasts isolated from Acetabulana mediterranea to synthesize the protein amino acids has been investigated. When this chloroplast isolate was presented with 14CO2 for periods of 6–8 hr, tracer was found in essentially all amino acid species of their hydrolyzed protein Phenylalanine labeling was not detected, probably due to technical problems, and hydroxyproline labeling was not tested for The incorporation of 14CO2 into the amino acids is driven by light and, as indicated by the amount of radioactivity lost during ninhydrin decarboxylation on the chromatograms, the amino acids appear to be uniformly labeled. The amino acid labeling pattern of the isolate is similar to that found in plastids labeled with 14CO2 in vivo. The chloroplast isolate did not utilize detectable amounts of externally supplied amino acids in light or, with added adenosine triphosphate (ATP), in darkness. It is concluded that these chloroplasts are a tight cytoplasmic compartment that is independent in supplying the amino acids used for its own protein synthesis. These results are discussed in terms of the role of contaminants in the observed synthesis, the "normalcy" of Acetabularia chloroplasts, the synthetic pathways for amino acids in plastids, and the implications of these observations for cell compartmentation and chloroplast autonomy.
PERMEABLE JUNCTIONAL COMPLEXES
Authors: Machen Terry E. , Erlij David , Wooding F. B. P.
Published: May 2004

Ionic lanthanum has been used to study transepithelial ion permeation in in vitro rabbit gallbladder and intestine (ileum) by adding 1 mM La3+ to only the mucosal bathing solution. Transepithelial fluid transport electrical potential differences (p.d.), and resistances were measured. During La3+ treatment the gallbladder's rate of active solute-coupled fluid transport remained constant, the resistance increased, and the 2:1 NaCl diffusion p.d. decreased. Mucosa-to-serosa fluxes of 140La3+ were measured and indicate a finite permeability of the gallbladder to La3+. La3+ also increased the transepithelial resistance and p d. of ileum. Electron microscopic examination of La3+-treated gallbladder showed: (a) good preservation of the fine structure, (b) electron-opaque lanthanum precipitates in almost every lateral intercellular space, most frequently near the apical end of the lateral spaces close to or within the junctional complex, (c) lanthanum among the subjacent muscle and connective tissue layers, and (d) lanthanum filling almost the entire length of so-called "tight" junctions. No observations were made which unequivocally showed the penetration of lanthanum into the gallbladder cells. Electron micrographs of similar La3+-treated ilea showed lanthanum deposits penetrating the junctional complexes. These results coupled with other physiological studies indicate that the low resistance pathway for transepithelial ion permeation in gallbladder and ileum is through the tight junctions A division of salt-transporting epithelia into two main groups, those with "leaky" junctional complexes and those with tight junctional complexes, has been proposed.
SOME PROPERTIES OF BOUND AND SOLUBLE DYNEIN FROM SEA URCHIN SPERM FLAGELLA
Authors: Gibbons I. R. , Fronk E.
Published: May 2004

Axonemes were isolated from sperm of Colobocentrotus by a procedure involving two extractions with 1% Triton X-100 and washing The isolated axonemes contained 7 x 1015 g protein per µm of their length. Treatment of the axonemes with 0 5 M KCl for 30 min extracted 50–70% of the flagellar ATPase protein, dynein, and removed preferentially the outer arms from the doublet tubules. Almost all of the dynein (85–95%) could be extracted from the axonemes by dialysis at low ionic strength. In both cases the extracted dynein sedimented through sucrose gradients at 12–14S, and no 30S form was observed The enzymic properties of dynein changed when it was extracted from the axonemes into solution. Solubilization had a particularly marked effect on the KCl- and pH-dependence of the ATPase activity. The pH-dependence of soluble dynein was fairly simple with a single peak extending from about pH 6 to pH 10. The pH-dependence of bound dynein was more complex. In 0.1 M KCl, the bound activity appeared to peak at about pH 9, and dropped off rapidly with decreasing pH, reaching almost zero at pH 7; an additional peak at pH 10 0 resulted from the breakdown of the axonemal structure and solubilization of dynein that occurred at about this pH. A similar curve was obtained in the absence of KCl, except for the presence of a further large peak at pH 8 Measurement of the kinetic parameters of soluble dynein showed that both Km and Vmax increased with increasing concentrations of KCl up to 0.5 M When bound dynein was assayed under conditions that would induce motility in reactivated sperm (0 15 M KCl with Mg++ activation), it did not obey Michaelis-Menten kinetics, although it did when assayed under other conditions. The complex enzyme-kinetic behavior of bound dynein, and the differences between its enzymic properties and those of soluble dynein, may result from its interactions with tubulin and other axonemal proteins
TUMOR ANGIOGENESIS
Authors: Cavallo Tito , Sade Robert , Folkman Judah , Cotran Ramzi S.
Published: May 2004

Walker ascites tumor cells and an extract derived from such cells (tumor angiogenesis factor, TAF) were injected into the subcutaneous tissue of rats by using a dorsal air sac technique. At intervals thereafter, thymidine-3H was injected into the air sac and the tissues were examined by autoradiography and electron microscopy. Autoradiographs of 1µ thick Epon sections showed thymidine-3H labeling in endothelial cells of small vessels 1–3 mm from the site of implantation, as early as 6–8 hr after exposure to live tumor cells At this time interval endothelial cells appeared histologically normal. DNA synthesis by endothelium subsequently increased and within 48 hr new blood vessel formation was detected. The presence of thymidine-3H-labeled endothelial nuclei, endothelial mitoses, and regenerating-type endothelium was confirmed by electron microscopy. TAF also induced neovascularization and endothelial cell DNA synthesis after 48 hr. A similar response was not evoked in saline controls. Formic acid, which elicited a more intense inflammatory response, was associated with less endothelial labeling and neovascularization at the times studied. Pericytes and other connective tissue cells were also stimulated by live tumor cells and TAF. The mechanism of new blood vessel formation induced by tumors is still unknown but our findings argue against cytoplasmic contact or nonspecific inflammation as prerequisites for tumor angiogenesis.
RADIOAUTOGRAPHIC DEMONSTRATION OF 5-HYDROXYTRYPTAMINE-3H UPTAKE BY PULMONARY ENDOTHELIAL CELLS
Authors: Strum Judy M. , Junod Alain F.
Published: May 2004

The lung is able to rapidly remove 5-hydroxytryptamme (5-HT) from the circulation by a Na+-dependent transport mechanism. In order to identify the sites of uptake, radioautographic studies were done on rat lungs which had been isolated and perfused with 5-HT-3H and 0 5 mM iproniazid, a monoamine oxidase inhibitor. In control experiments 10-4 M imipramine was added to the perfusate to inhibit the membrane transport of 5-HT At the light microscope level, silver grains were seen concentrated near capillaries and in the endothelium of large vessels From electron microscope radioautographs a semiquantitative grain count was made and 90% of the silver grains were observed over capillary endothelial cells. The grains were found over the nucleus and cytoplasm of the cell and shewed no preferential association with any particular cytoplasmic inclusion bodies, organelles, or vesicles Other cell types were unlabeled except for a few mast cells, certain vascular smooth muscle cells, and one nerve ending. This radioautographic demonstration of the cell type responsible for the rapid removal of 5-HT from the lung circulation clearly establishes the existence of a new metabolic role for pulmonary endothelial cells.
MYONEURAL JUNCTIONS OF TWO ULTRASTRUCTURALLY DISTINCT TYPES IN EARTHWORM BODY WALL MUSCLE
Authors: Rosenbluth Jack
Published: May 2004

The longitudinal muscle of the earthworm body wall is innervated by nerve bundles containing axons of two types which form two corresponding types of myoneural junction with the muscle fibers Type I junctions resemble cholinergic neuromuscular junctions of vertebrate skeletal muscle and are characterized by three features: (a) The nerve terminals contain large numbers of spherical, clear, ∼500 A vesicles plus a small number of larger dense-cored vesicles (b) The junctional gap is relatively wide (∼900 A), and it contains a basement membrane-like material, (c) The postjunctional membrane, although not folded, displays prominent specializations on both its external and internal surfaces The cytoplasmic surface is covered by a dense matrix ∼200 A thick which appears to be the site of insertion of fine obliquely oriented cytoplasmic filaments The external surface exhibits rows of projections ∼200 A long whose bases consist of hexagonally arrayed granules seated in the outer dense layer of the plasma membrane The concentration of these hexagonally disposed elements corresponds to the estimated concentration of both receptor sites and acetylcholinesterase sites at cholinergic junctions elsewhere. Type II junctions resemble the adrenergic junctions in vertebrate smooth muscle and exhibit the following structural characteristics: (a) The nerve fibers contain predominantly dense-cored vesicles ∼1000 A in diameter (b) The junctional gap is relatively narrow (∼150 A) and contains no basement membrane-like material, (c) Postjunctional membrane specialization is minimal. It is proposed that the structural differences between the two types of myoneural junction reflect differences in the respective transmitters and corresponding differences in the mechanisms of transmitter action and/or inactivation.
ELECTRON MICROSCOPE STUDIES OF SPERMATOZOA OF RHYNCHOSCIARA SP
Authors: Shay Jerry W.
Published: May 2004

The flagellar complex of the unusual motile spermatozoon of the fungus gnat, Rhynchosciara sp, does not conform to the usual "9 + 2" filament pattern but rather consists of over 350 pairs of filaments (doublet microtubules) distributed in a spiral array. Experiments were designed to disrupt and extract flagellar microtubular components from spermatozoa of the fungus gnat. Pepsin, chymotrypsin, potassium iodide, urea, and heat were used to extract specific portions of microtubule walls Such experiments provide information on the composition of the wall and the existence of wall sites selectively sensitive to various treatments Results obtained include: (a) doublet microtubules are comprised at least in part of protein, and all subunits are probably not identical; (b) a portion of the B subfiber is apparently more sensitive to disruption than other portions of the doublet microtubule; and (c) the ac cessory singlet microtubules may be chemically different from the doublet microtubules
COLLAGEN SUBSTRATA FOR STUDIES ON CELL BEHAVIOR
Authors: Elsdale Tom , Bard Jonathan
Published: May 2004

A simple technique is described for the preparation of collagen substrata containing 0 1% of collagen by weight, in the form of native bundles with a 640 A period, the substrata are similar in these respects to soft-tissue matrices These substrate are hydrated collagen lattices (HCLs) in which the watery milieu is held within a fibrous collagen net mainly by capillary forces. HCLs have been characterized in terms of the course of collagen precipitation and aggregation, ultrastructure, and their stability under various conditions. The ways in which HCLs can be employed as both two- and three-dimensional substrata in cell behavioral studies are illustrated with some preliminary observations on the form, motility, adhesion, and growth of human diploid cells and two lines of malignant cells.
FRACTIONATION OF THE NUCLEUS BY DIVALENT CATIONS
Authors: Monneron Ariane , Blobel Günter , Palade George E.
Published: May 2004

0.3–0.5 M MgCl2 was used to disassemble nuclei and to isolate by a single centrifugation in less than 3 hr a nuclear envelope fraction in 55–60% yield as assessed by phospholipid recovery. Its gross chemical composition was determined and its morphology was studied electron microscopically by sectioning, freeze etching, and negative staining procedures.