bluuebunny/crossref_metadata_2025 huggingface dataset:
Electron Microscopy of the Sperm Tail Results Obtained with a New Fixative
Authors: Afzelius Björn
Published: May 2004

The details of a new fixation procedure using 40 per cent osmium tetroxide in carbon tetrachloride are presented. This fixative is a good general preservative, gives a higher contrast than the ordinary osmium fixatives, and may also preserve structures that are not otherwise readily revealed. Some possible reasons for the increased contrast are discussed.

Micrographs of the sea urchin spermatozoa treated with the new fixative provide more detailed information on the tail structure than has heretofore been obtainable. This information is summarized in the diagrammatic text-figure.

The sperm tail can no longer be regarded as having a bilateral symmetry, and thus, it is possible to assign an index number to each of the nine peripheral filaments.

The nine peripheral filaments have a complex morphology, each one of them seems to be composed of two subunits that have unequal diameters. The slightly larger subunits are all found in the clockwise direction with regard to the other subunit or are all found in the counter-clockwise direction in the sectioned tail. Each of the slightly larger subunits is at intervals provided with two types of projections—referred to as the arms and the spokes—that extend in respective tangential and radial direction. The arms from one filament may be in actual contact with its neighboring filament through a complex bridge-like formation. There is a quantitative difference between the nine filaments with regard to this bridge.

It is assumed that the eleven tail filaments follow straight paths. Some hypotheses on sperm movement are discussed based on this assumption and on the fact that the oscillations of an actively working sperm tail are in one plane. Probably, the nine peripheral filaments have non-equivalent functions in tail movement.

In the centriole the nine peripheral filaments characteristically appear as triplets in a whorl-like arrangement. It is suggested that the inner part of this triplet is a derivation of the arms. A structural abnormality of the tail is described that is characterized by two or three complete sets of tail filaments within one cell membrane.
Observations on Cellular Structures of Porphyridium cruentum
Authors: Brody Marcia , Vatter Albert E.
Published: May 2004

The cellular structure of Porphyridium cruentum was studied with both light and electron microscope. The photosynthetic plastid in this red alga was found to be structurally similar to that in the Chlorophyceae and higher green plants. The phycobilins, as well as the chlorophyll, seem to be associated with the lamellae of the plastid.

The pyrenoid, a region of low lamellar density, contains no tubules, and does not appear to function in synthesis or storage of reserve material. Grains of floridean starch are located in the cytoplasm, outside the plastid. Typical mitochondrial organelles were not observed.

The nucleus is eccentric, and contains a nucleolus located on the inner face of the nucleus, nearest the plastid. The schedule for staining the nucleus is given in detail. Other cell structures (sheath, dictyosomes, etc.) are described.

Growing cells in light of intensity leads to disruption of the parallel arrangement of the lamellar characteristic of cells grown in moderate light.
An Electron Microscopic Study of the Intestinal Villus
Authors: Palay Sanford L. , Karlin Leonard J.
Published: May 2004

The intestinal pathway for absorbed fat was traced in thin sections of intestinal villi from rats fed corn oil by stomach tube after a fast of 24 to 40 hours. For electron microscopy the tissues were fixed in chilled buffered osmium tetroxide and embedded in methacrylate. For light microscopy, other specimens from the same animals were fixed in formal-calcium, mordanted in K2Cr2O7, and embedded in gelatin. Frozen sections were stained with Sudan black B or Sudan IV.

About 20 minutes after feeding, small fat droplets (65 mµ maximal diameter) appear in the striated border between microvilli. At the same time fat particles are seen within pinocytotic vesicles in the immediately subjacent terminal web. In later specimens the fat droplets are generally larger (50 to 240 mµ) and lie deeper in the apical cytoplasm. All intracellular fat droplets are loosely enveloped in a thin membrane, the outer surface of which is sometimes studded with the fine particulate component of the cytoplasm. This envelope, apparently derived from the cell surface by pinocytosis, has at this stage evidently become a part of the endoplasmic reticulum. Just above the nucleus numerous fat droplets lie clustered within the dilated cisternae of the Golgi complex. As absorption progresses fat droplets appear in the intercellular spaces of the epithelium, in the interstitial connective tissue spaces of the lamina propria, and in the lumen of the lacteals. All of these extracellular fat droplets are devoid of a membranous envelope.

The picture of fat absorption as reconstructed from these studies involves a stream of fat droplets filtering through the striated border, entering the epithelial cell by pinocytosis at the bases of the intermicrovillous spaces, and coursing through the endoplasmic reticulum to be discharged at the sides of the epithelial cell into extracellular spaces. From the epithelial spaces, the droplets move into the lamina propria and thence into the lymph. If the lumen of the endoplasmic reticulum is considered as continuous with the extracellular phase, then the entire pathway of fat absorption may be regarded as extracellular. However, it is impossible to evaluate from the electron microscopic evidence thus far available the quantitative importance of particulate fat absorption by the mechanism described.
Similarity of the Structure of DNA from a Variety of Sources
Authors: Hamilton L. D. , Barclay R. K. , Wilkins M. H. F. , Brown G. L. , Wilson H. R. , Marvin D. A. , Ephrussi-Taylor H. , Simmons N. S.
Published: May 2004

DNA's from diverse cells of different species and from diverse tissues give the same x-ray diffraction pattern. The presently observable structure of DNA appears, then, to be the same in all cells. Thus, DNA in the resting state—the stored genetic material, from sperm of Paracentrotus lividus, Arbacia lixula, and salmon and from T2 and T7 bacteriophage—gives a pattern indistinguishable from DNA from very rapidly dividing cells, e.g., human acute leukemic leukocytes, human leukemic myeloid cells, mouse sarcoma 180, and bacteria—E. coli and pneumococci—during their logarithmic growth. The same x-ray patterns are given by DNA's from more slowly dividing tissues, e.g. calf liver, calf thymus, and human normal and leukemic lymphatic tissue. DNA from chicken erythrocytes—a DNA presumably metabolically inert—gives a similar picture. DNA's from several sources with a wide range in nitrogen base ratios, prepared independently by different workers using various methods, have given final products in varying yield; these all gave the same x-ray pattern, suggesting that all DNA is in the double-helical configuration. Finally, separation of the DNA molecule into a number of fractions with a varying adenine + thymine:guanine + cytosine ratio, but a constant adenine:thymine and guanine:cytosine ratio, each giving the same x-ray pattern as the original whole molecule, suggests that DNA cannot exist in significant amounts in forms other than the double-helix.

X-ray diffraction photographs of sperm heads, extracted nucleoprotamine, calf thymus nuclei and extracted nucleohistone, and of chicken erythrocyte nuclei, are not all as well defined as those given by extracted DNA, but it is clear from the general characteristics of the pattern that much of the DNA bound to protein in these nuclei has the usual helical configuration, and that the double-helical structure of DNA exists in the cell and is not an artifact.
Studies on the Control of Mitotic Activity in Excised Roots
Authors: Wilson G. B. , Morrison J. H. , Knobloch N.
Published: May 2004

The mitotic characteristics of excised roots of the garden pea, Pisum sativum, have been studied under conditions of controlled nutrition.

The excised root system was tested with regard to its ability to respond, mitotically, to various carbon sources. Sucrose, glucose, fructose, and DL-glyceraldehyde were found to support mitotic activity in excised roots, galactose and 2-deoxy-D-glucose were toxic, and mannose ineffective. Initiation of mitotic activity in the presence of glucose was inhibited by the respiratory poisons, KCN and malonic acid, the uncoupling agent, 2,4-dinitrophenol, but was not notably affected by the protein synthesis inhibitor, chloramphenicol. The glucose-induced response in mitotic activity was not affected by the carcinogen, urethan, and indeed, there is some evidence that the response was actually enhanced. The fact that KCN, malonic acid, and probably 2,4-dinitrophenol, in suitable concentrations inhibit the onset of cell division suggests that some level of operation of the Krebs' cycle is essential for commission of cells into mitosis. Likewise, failure to inhibit cells in the process of active mitosis by KCN and malonic acid is not inconsistent with the idea that there is a shift from reliance on aerobic to anaerobic respiration between antephase and active mitosis.
Considérations quantitatives sur des coupes ultraminces de bactéries infectées par un bactériophage
Authors: Séchaud Janine , Ryter Antoinette , Kellenberger Edouard
Published: May 2004

A quantitative relationship has been established between the number of particles, for example bacteriophages, counted in ultrathin sections of bacteria and the total number present in the whole bacterial cells. The factor F relating particles counted per section with the total number of these particles per entire bacterium could be arrived at by two methods, which proved to give results in close agreement. The first involves knowledge of the average volume of a bacterial section in proportion to the average volume of a whole bacterium; if the mean number of appearances of the same particle on consecutive sections is also known, F may then be calculated. The thickness of sections and, therefore, their volume, as well as the average number of times a single particle is sectioned could be learned by examination of serial sections. By counting the relative number of T2 phage particles which had been intersected once or twice, and relating this proportion to the known phage dimensions, the thickness of the sections was determined to be about 400 A.

The second measurement of F could be made in a particular case of late phage development where the number of particles per cell was countable or titratable directly in the bacterial lysate, this number being compared with the number seen in sections of the bacteria just before lysis.

The different sources of errors are discussed. The statistical error is under 20 per cent, while the systematic errors are higher and cannot yet be indicated precisely. After a very cautious estimation of the upper limits, we can state, however, that the counts made with this method are certainly reliable to well within a factor of two.
The Effect of Respiratory Inhibitors and Chelating Agents on the Frequencies of Chromosomal Aberrations Produced by X-Rays in Vicia
Authors: Kihlman B. A.
Published: May 2004

Nitrosophenylhydroxylamine-ammonium (cupferron), potassium cyanide, sodium azide, ethylenediaminetetraacetate (EDTA), α,α'-dipyridyl, and o-phenanthroline were tested (1) for their ability to enhance the frequencies of chromosomal aberrations produced by x-rays in the root tip cells of the broad bean, Vicia faba, and (2) for their ability to inhibit oxygen consumption of excised roots of the same plant. In all cases a close correlation was found between the inhibitory effect on respiration and the enhancement of the sensitivity to x-rays at low oxygen pressures. EDTA, dipyridyl, and o-phenanthroline did not affect respiration to any greater extent, and they were without influence on the radiosensitivity. Cyanide, azide, and cupferron, which strongly inhibited respiration, also increased the frequencies of chromosome aberrations produced by x-rays at low oxygen pressures.

The relation between oxygen concentration and radiosensitivity was determined both in the presence and the absence of the respiratory inhibitor cupferron. When cupferron was present, the radiosensitivity was influenced by oxygen concentrations 30 times lower than those effective in the absence of the inhibitor.

In an atmosphere of pure oxygen, an increase of radiosensitivity of about 20 per cent was obtained with cupferron, EDTA, and potassium cyanide.
THE FORMATION OF BASAL BODIES (CENTRIOLES) IN THE RHESUS MONKEY OVIDUCT
Authors: Anderson Richard G. W. , Brenner Robert M.
Published: May 2004

Basal body replication during estrogen-driven ciliogenesis in the rhesus monkey (Macaca mulatta) oviduct has been studied by stereomicroscopy, rotation photography, and serial section analysis. Two pathways for basal body production are described: acentriolar basal body formation (major pathway) where procentrioles are generated from a spherical aggregate of fibers; and centriolar basal body formation, where procentrioles are generated by the diplosomal centrioles. In both pathways, the first step in procentriole formation is the arrangement of a fibrous granule precursor into an annulus. A cartwheel structure, present within the lumen of the annulus, is composed of a central cylinder with a core, spoke components, and anchor filaments. Tubule formation consists of an initiation and a growth phase. The A tubule of each triplet set first forms within the wall material of the annulus in juxtaposition to a spoke of the cartwheel. After all nine A tubules are initiated, B and C tubules begin to form. The initiation of all three tubules occurs sequentially around the procentriole. Simultaneous with tubule initiation is a nonsequential growth of each tubule. The tubules lengthen and the procentriole is complete when it is about 200 mµ long. The procentriole increases in length and diameter during its maturation into a basal body. The addition of a basal foot, nine alar sheets, and a rootlet completes the maturation process. Fibrous granules are also closely associated with the formation of these basal body accessory structures.
FINE STRUCTURAL LOCALIZATION OF ACYLTRANSFERASES
Authors: Higgins Joan A. , Barrnett Russell J.
Published: May 2004

A study of the fine structural localization of the acyltransferases of the monoglyceride and α-glycerophosphate pathways for triglyceride synthesis in the intestinal absorptive cell is reported. Glutaraldehyde-fixed tissue was found to synthesize diglyceride and triglyceride from monopalmitin and palmityl CoA, and parallel morphological studies showed the appearance of lipid droplets in the smooth endoplasmic reticulum of the absorptive cell. Glutaraldehyde-fixed tissue also synthesized triglyceride from α-glycerophosphate, although this enzyme system was more susceptible to fixation than the monoglyceride pathway acyltransferases. Cytochemical methods for the localization of free CoA were based (a) on the formation of the insoluble lanthanium mercaptide of CoA and (b) on the reduction of ferricyanide by CoA to yield ferrocyanide which forms an insoluble precipitate with manganous ions. By these methods the monoglyceride pathway acyltransferases were found to be located mainly on the inner surface of the smooth endoplasmic reticulum. The α-glycerophosphate pathway acyltransferases were localized mainly on the rough endoplasmic reticulum. Activity limited to the outer cisternae of the Golgi membranes occurred with both pathways. The possible organization of triglyceride absorption and chylomicron synthesis is discussed in view of these results.
THE SMOOTH MUSCLE CELL
Authors: Ross Russell
Published: May 2004

Smooth muscle derived from the inner media and intima of immature guinea pig aorta were grown for up to 8 wk in cell culture. The cells maintained the morphology of smooth muscle at all phases of their growth in culture. After growing to confluency, they grew in multiple overlapping layers. By 4 wk in culture, microfibrils (110 A) appeared within the spaces between the layers of cells. Basement membrane-like material also appeared adjacent to the cells. Analysis of the microfibrils showed that they have an amino acid composition similar to that of the microfibrillar protein of the intact elastic fiber. These investigations coupled with the radioautographic observations of the ability of aortic smooth muscle to synthesize and secrete extracellular proteins demonstrate that this cell is a connective tissue synthetic cell.
PERMEABILITY OF THE OVARIAN FOLLICLE OF AEDES AEGYPTI MOSQUITOES
Authors: Anderson Winston A. , Spielman Andrew
Published: May 2004

The passage of tracers of various molecular weights into resting and vitellogenic ovarian follicles of Aedes aegypti mosquitoes was studied ultrastructurally. The outermost layer of the follicular sheath (the basement lamina) is a coarse mechanical filter. It is freely permeable to particles with molecular weights ranging from 12,000 to 500,000 (i.e. cytochrome c, peroxidase, hemoglobin, catalase, ferritin, immunoglobulin (IgG)-peroxidase, iron dextran and Thorotrast) that have dimensions less than 110 A. Molecules as large as carbon (300–500 A) are totally excluded. Whereas proteins and polysaccharide tracers permeate the basement lamina with apparent ease, certain inert particles (e.g. Thorotrast, Fellows-Testager Div., Fellows Mfg. Co., Inc., Detroit, Mich.) penetrate more slowly. With respect to the tracers tested, resting follicles are as permeable as vitellogenic follicles. The follicle epithelium of resting or vitellogenic follicles is penetrated by narrow intercellular channels. Our observations suggest that these spaces are lined with mucopolysaccharide material. After permeating the basement lamina, exogenous tracers fill these channels, while the bulk of material accumulates in the perioocytic space. Within 3 hr after imbibing blood, the pinocytotic mechanism of the oocyte is greatly augmented. Pinocytosis is not selective with regard to material in the perioocytic space, since double tracer studies show that exogenous compounds are not separated, but are incorporated into the same pinocytotic vesicle. During later stages of vitellogenesis, 36–48 hr after the blood-meal, the pinocytotic mechanism of the oocyte is diminished. Simultaneously, the intercellular channels become occluded by desmosomes, and the vitelline membrane plaques separate the oocyte and follicle epithelium.
COEXISTENCE OF GAP AND SEPTATE JUNCTIONS IN AN INVERTEBRATE EPITHELIUM
Authors: Hudspeth A. J. , Revel J. P.
Published: May 2004

The intercellular junctions of the epithelium lining the hepatic caecum of Daphnia were examined. Electron microscope investigations involved both conventionally fixed material and tissue exposed to a lanthanum tracer of the extracellular space. Both septate junctions and gap junctions occur between the cells studied. The septate junctions lie apically and resemble those commonly discerned between cells of other invertebrates. They are atypical in that the high electron opacity of the extracellular space obscures septa in routine preparations. The gap junctions are characterized by a uniform 30 A space between apposed cell membranes. Lanthanum treatment of gap junctions reveals an array of particles of 95 A diameter and 120 A separation lying in the plane of the junction. As this pattern closely resembles that described previously in vertebrates, it appears that the gap junction is phylogenetically widespread. In view of evidence that the gap junction mediates intercellular electrotonic coupling, the assignment of a coupling role to other junctions, notably the septate junction, must be questioned wherever these junctions coexist.
STUDIES ON THE PERMEABILITY OF LYMPHATIC CAPILLARIES
Authors: Leak Lee V.
Published: May 2004

The passageway for interstitial fluids and large molecules across the connective tissue lymph interface has been investigated in dermal lymphatic capillaries in the ears of guinea pigs. Numerous endothelial cells overlap extensively at their margins and lack adhesion devices at many points. The observations suggest that these sites are free to move as a result of slight pressure changes. Immediately following interstitial injections of tracer particles (ferritin, thorium, carbon, and latex spheres), many of the overlapped endothelial cells are separated and thus passageways are provided between the interstitium and lymphatic lumen. Tracer particles also occur in plasmalemmal invaginations along both connective tissue and luminal fronts. All of the tracer particles accumulate within large autophagic-like vacuoles. Very few particles of ferritin are observed in the endothelium after 24 hr; however, the vesicles containing the nonprotein tracer particles (carbon, thorium, and latex) increase in size and content and remain within the lymphatic endothelial cells up to 6 months. The role of vesicles in the transport of large molecules and particles is discussed in relation to the accretion of tracer particles within large vesicles and autophagic-like vacuoles in the endothelial cytoplasm.
THE STRUCTURE, ORIGIN, ISOLATION, AND COMPOSITION OF THE TUBULAR MASTIGONEMES OF THE OCHROMONAS FLAGELLUM
Authors: Bouck G. Benjamin
Published: May 2004

The structure, assembly, and composition of the extracellular hairs (mastigonemes) of Ochromonas are detailed in this report. These mastigonemes form two lateral unbalanced rows, each row on opposite sides of the long anterior flagellum. Each mastigoneme consists of lateral filaments of two distinct sizes attached to a tubular shaft. The shaft is further differentiated into a basal region at one end and a group of from one to three terminal filaments at the free end. Mastigoneme ontogeny as revealed especially in deflagellated and regenerating cells appears to begin by assembly of the basal region and shaft within the perinuclear continuum. However, addition of lateral filaments to the shaft and extrusion of the mastigonemes to the cell surface is mediated by the Golgi complex. The ultimate distribution of mastigonemes on the flagellar surface seems to be the result of extrusion of mastigonemes near the base of the flagellum, and it is suggested that mastigonemes are then pulled up the flagellum as the axoneme elongates. Efforts to characterize mastigonemes biochemically after isolation and purification on cesium chloride (CsCl) followed by electrophoresis on acrylamide gels have demonstrated what appear to be a single major polypeptide and several differentially migrating carbohydrates. The polypeptide is not homologous with microtuble protein. The functionally anomalous role of mastigonemes in reversing flagellar thrust is discussed in relation to their distribution relative to flagellar anatomy and to the plane of flagellar undulations.
ISOLATION AND BIOCHEMICAL CHARACTERIZATION OF NUCLEI FROM CHICK EMBRYO LIVER
Authors: Incefy Genevieve S. , Kappas Attallah
Published: May 2004

A procedure is described for the isolation of enzymatically active nuclei from chick embryo liver. It consists of the homogenization of the pooled tissue in 0.32 M sucrose-3 mM MgCl2 followed by a slow centrifugation. The resulting nuclear pellet is then purified further in a discontinuous density gradient composed of sucrose solutions containing Mg2+ ions, the lower portion of the gradient being 2.2 M sucrose-1 mM MgCl2. Based on DNA recovery, the nuclear fraction isolated by the procedure described contained an average of 62% of the nuclei in the original filtered homogenate. Light and electron microscope examinations showed that 90% of the isolated nuclei were derived from hepatocytes. They appeared intact with well preserved nucleoplasmic and nucleolar components, nuclear envelope, and pores. The isolated nuclei were quite pure, having a very low level of cytoplasmic contamination as indicated by cytoplasmic enzyme marker activities and electron microscope studies. The nuclear fraction consisted of 19.9% DNA, 6.2% RNA, 74% protein, the average RNA/DNA ratio being 0.32. Biosynthetic activities of the two nuclear enzymes NAD-pyrophosphorylase and DNA-dependent RNA polymerase were preserved. The specific activities of these enzymes were: NAD-pyrophosphorylase, 0.049 µmoles nicotinamide adenine dinucleotide (NAD) synthesized/min per mg protein; Mg2+ activated RNA polymerase, 4.3 µµmoles UMP-2-C14 incorporated into RNA/µg DNA per 10 min; and Mn2+-(NH4)2SO4 activated RNA-polymerase, 136 µµmoles UMP-2-C14 incorporated into RNA/µg DNA per 45 min.
CYTOPLASMIC FILAMENTS IN DEVELOPING AND ADULT VERTEBRATE SMOOTH MUSCLE
Authors: Uehara Y. , Campbell G. R. , Burnstock G.
Published: May 2004

An extensive study of adult and developing smooth muscle has revealed the widespread occurrence of a distinct filament with an average diameter of about 100 A (termed the 100 A filament). Unlike that of myofilaments, their appearance in longitudinal section is uniform, but in transverse section they have a round profile, occasionally exhibiting a less electron-opaque core. The 100 A filaments are almost invariably preserved under a variety of fixation procedures, whereas myofilaments, particularly the thicker filaments, are preserved inconsistently. The 100 A filaments appear to be randomly oriented throughout the cytoplasm, either singly or in small groups, although they are sometimes concentrated in the juxtanuclear region of the smooth muscle cells. The intimate association of 100 A filaments with dark bodies, in both developing and adult smooth muscle cells, may indicate that these filaments either play a role in dark body formation or, at least, constitute a part of the dark body.

The 100 A filaments are conspicuous in developing smooth muscle cells and occasionally form networks or clusters; they appear to decrease in relative number as maturation proceeds, but considerable numbers are still present in adult tissue.
DIFFERENTIATION OF MONOCYTES
Authors: Nichols Barbara A. , Bainton Dorothy Ford , Farquhar Marilyn G.
Published: May 2004

The origin, content, and fate of azurophil granules of blood monocytes were investigated in several species (rabbit, guinea pig, human) by electron microscopy and cytochemistry. The life cycle of monocytes consists of maturation in bone marrow, transit in blood, and migration into tissues where they function as macrophages. Cells were examined from all three phases. It was found that: azurophil granules originate in the Golgi complex of the developing monocyte of bone marrow and blood, and ultimately fuse with phagosomes during phagocytosis upon arrival of monocytes in the tissues. They contain lysosomal enzymes in all species studied and peroxidase in the guinea pig and human. These enzymes are produced by the same pathway as other secretory products (i.e., they are segregated in the rough ER and packaged into granules in the Golgi complex). The findings demonstrate that the azurophil granules of monocytes are primary lysosomes or storage granules comparable to the azurophils of polymorphonuclear leukocytes and the specific granules of eosinophils. Macrophages from peritoneal exudates (72–96 hr after endotoxin injection) contain large quantities of lysosomal enzymes throughout the secretory apparatus (rough ER and Golgi complex), in digestive vacuoles, and in numerous coated vesicles; however, they lack forming or mature azurophil granules. Hence it appears that the monocyte produces two types of primary lysosomes during different phases of its life cycle—azurophil granules made by developing monocytes in bone marrow or blood, and coated vesicles made by macrophages in tissues and body cavities.
MORPHOLOGICAL AND BIOCHEMICAL STUDIES OF B CELLS OF FETAL RAT ENDOCRINE PANCREAS IN ORGAN CULTURE
Authors: Orci Lelio , Lambert Andre E. , Kanazawa Yasunori , Amherdt Mylene , Rouiller Charles , Renold Albert E.
Published: May 2004

Fetal rat pancreases explanted on the 18th day of gestation and maintained in organ culture for 1–10 days were utilized for this series of studies. Ultrastructurally, at the time of explantation, the majority of fetal B cells was sparsely granulated and characterized by numerous free ribosomes and undeveloped rough endoplasmic reticulum (RER) and Golgi complexes. During the culture period, extensive development of the RER and Golgi complexes preceded an increasing accumulation of ß-granules. This later increase in the number of ß-granules and in the concentration of immunoreactive insulin was paralleled by a reduction of RER and Golgi complex activity. High resolution radioautographic studies of pulse-chase experiment over a 1 hr period demonstrated the shift of silver grains from the elements of the RER, through the Golgi region, and finally to the ß-granules. Incubation with 14C-labeled leucine demonstrated the incorporation of radioactivity into molecules possessing the immunological and electrophoretic properties of insulin. These studies indicate that de novo synthesis of (pro)insulin occurs also during culture of fetal rat pancreas explanted relatively late in gestation.
ASSEMBLY OF LIPIDS INTO MEMBRANES IN ACANTHAMOEBA PALESTINENSIS
Authors: Chlapowski Francis J. , Band R. Neal
Published: May 2004

In order to determine the feasibility of using radioactive precursors as markers for membrane phospholipids in Acanthamoeba palestinensis, the characteristics of phospholipids labeled with choline-14C and glycerol-3H were examined. Choline-14C was found to be a specific label for phosphatidyl choline. There was a turnover of the radioactive moiety of phosphatidyl choline at a rate that varied with the concentration of nonradioactive choline added to the growth medium. Radioactivity was lost from labeled phosphatidyl choline into the acid-soluble intracellular pool and from the pool into the extracellular medium. This loss of radioactivity from cells leveled off and an equilibrium was reached between the label in the cells and in the medium. Radioactive choline was incorporated into phosphatidyl choline by cell-free microsomal suspensions. This incorporation leveled off with the attainment of an equilibrium between the choline-14C in the reaction mixture and the choline-14C moiety of phosphatidyl choline in the microsomal membranes. Therefore, a choline exchange reaction may occur in cell-free membranes, as well as living A. palestinensis. In contrast to choline-14C, the apparent turnover of glycerol-3H-labeled phospholipids was not affected by large concentrations of nonradioactive choline or glycerol in the medium. The radioactivity in lipids labeled with glycerol-3H consisted of 33% neutral lipids and 67% phospholipids. Phospholipids labeled with glycerol-3H turned over slowly, with a concomitant increase in the percentage of label in neutral lipids, indicating a conversion of phospholipids to neutral lipids. Because most (∼96%) of the glycerol-3H recovered from microsomal membranes was in phospholipids, whereas only a minor component (∼2%) of the glycerol-3H was in the phospholipids isolated from nonmembrane lipids, glycerol-3H was judged to be a specific marker for membrane phospholipids.
ASSEMBLY OF LIPIDS INTO MEMBRANES IN ACANTHAMOEBA PALESTINENSIS
Authors: Chlapowski Francis J. , Band R. Neal
Published: May 2004

The membranes of Acanthamoeba palestinensis were studied by examination in fixed cells, and then by following the movements of glycerol-3H-labeled phospholipids by cell fractionation. Two previously undescribed structures were observed: collapsed cytoplasmic vesicles of cup shape, and plaques in food vacuole and plasma membrane similar in size to the collapsed vesicles. It appeared that the plaques formed by insertion of collapsed vesicles into membranes and/or that collapsed vesicles formed by pinching off of plaques. Fractions were isolated, enriched with nuclei, rough endoplasmic reticulum (RER), plasma membrane, Golgi-like membranes, and collapsed vesicles. The changes in specific activity of glycerol-3H-labeled phospholipids in these membranes during incorporation, turnover, and after pulse-labeling indicated an ordered sequence of appearances of newly synthesized phospholipids, first in nuclei and RER, then successively in Golgi membranes, collapsed vesicles, and finally, plasma membrane. In previous work we had found no large nonmembranous phospholipid pool in A. palestinensis. These observations are consistent with the hypothesis that membrane phospholipids are synthesized, perhaps as integral parts of membranes, in RER and nuclei. Subsequently, some of the newly synthesized phospholipids are transported to the Golgi complex to become integrated into the membranes of collapsed vesicles, which are precursors of the plasma membrane. Collapsed vesicles from the plasma membrane by inserting into it as plaques. When portions of the plasmalemma from food vacuoles, collapsed vesicles pinch off from their membranes and are recycled back to the cell surface.
PRIMITIVE ERYTHROPOIESIS IN EARLY CHICK EMBRYOGENESIS
Authors: Weintraub Harold , Campbell Graham Le M. , Holtzer Howard
Published: May 2004

The primitive line of embryonic chick blood cells develop as a relatively homogeneous cohort of cells. Using an analysis based on the continuous uptake of thymidine-3H, we have established the generation time, G1, S, and G2 for progressively more mature generations of these immature erythroblasts. The data indicate that after the initiation of hemoglobin synthesis, the average cell will yield six generations of hemoglobin producing erythroblasts. The older generations of erythroblasts exhibit a longer generation time, G1, S, and G2 than the earlier generations of erythroblasts. Other methods of analysis corroborated these findings. One of these methods, an estimate of total erythrocyte productivity from the primitive stem cells (hematocytoblasts), led to the conclusion that the erythroblast cell lineage might be initiated as early as the sixth or seventh division following fertilization. In addition, primitive erythroblasts characterized by one set of cell cycle parameters, when grown in serum associated with erythroblasts of different parameters, showed no alteration in mitotic behavior. These results suggest the presence of programmed cell division not immediately cued by extracellular influence.
PRIMITIVE ERYTHROPOIESIS IN EARLY CHICK EMBRYOGENESIS
Authors: Campbell G. Le M. , Weintraub H. , Mayall B. H. , Holtzer H.
Published: May 2004

Primitive erythroblasts in the circulating blood of the chick embryo continue to divide while synthesizing hemoglobin (Hb). Hb measurements on successive generations of erythroblasts show that there is a progressive increase in the Hb content of both interphase and metaphase cells. Furthermore, for any given embryo the Hb content of metaphase cells is always significantly greater than that of interphase cells. The distribution of Hb values for metaphase cells suggests that there are six Hb classes corresponding to the number of cell cycles in the proliferative phase. The location of erythroblasts in the cell cycle was determined by combining Feulgen cytophotometry with thymidine radioautography on the same cells. Measurements of the Hb content for erythroblasts in different compartments of the cell cycle (G1, S, G2, and M) show a progressive increase through the cycle. Thus, the amount of Hb per cell is a function of the number of cell divisions since the initiation of Hb synthesis and, to a lesser degree, the stage of the cell cycle. Earlier generations of erythroblasts synthesize Hb at a faster rate than the terminal generation. Several models have been proposed to explain these findings.
MITOCHONDRIAL BIOGENESIS IN NEUROSPORA CRASSA
Authors: Howell Neil , Zuiches Carol A. , Munkres Kenneth D.
Published: May 2004

The isolation of a new class of mutants permitting facultative anaerobiosis in Neurospora crassa is described. Backcross analyses to the obligate aerobe prototroph (An-) indicate single nuclear gene inheritance (An-/An+). An+ and An- are indistinguishable in morphology and growth rates under aerobic conditions. Anaerobic growth requires nutritional supplements that are dispensable for aerobic growth. Conidiogenesis, conidial germination, and vegetative growth rate are suppressed by anaerobiosis. An+ mutants produce substantial quantities of ethanol under anaerobic conditions. Anaerobiosis and chloramphenicol both affect mitochondrial enzyme activity and morphology. Chloramphenicol inhibition leads to reduction in cytochrome oxidase and swollen mitochondria with few cristae. Anaerobiosis leads to reduction in both cytochrome oxidase and malate dehydrogenase activities, enlarged mitochondria with fewer cristae, enlarged nuclei, and other alterations in cellular morphology. The fine structure of anaerobically grown cells changes with the time of anaerobic growth. We conclude that either inhibition of mitochondrial membrane synthesis or inhibition of respiration might lead to the observed alterations in mitochondria.
ELECTRICAL COUPLING BETWEEN FAT CELLS IN NEWT FAT BODY AND MOUSE BROWN FAT
Authors: Sheridan Judson D.
Published: May 2004

White fat from the newt, Triturus pyrrhogaster, fat body, and brown fat from the interscapular fat pad of newborn mice have been tested for the presence of low-resistance intercellular junctions. 42 pairs of amphibian fat cells and 15 pairs of mammalian brown fat cells were found to be "electrically coupled." In most of these cases intracellular deposition of a dye, Niagara Sky Blue: 6B, was used to supplement and confirm direct observations of impalements. Coupling was often difficult to find in both preparations, but the mechanical disturbance of the tissue during the preparative procedures may have uncoupled many cells. The fact that, in both types of fat, coupling was observed between cells separated by one or more other cells suggests that coupling may be more widespread in vivo. Electron microscopy (provided by Dr. J. -P. Revel and Mrs. K. Wolken) of the brown fat revealed frequent intercellular junctions resembling "gap junctions" but possibly lacking the substructure usually visible with colloidal lanthanum infiltration. The results are discussed in relation to current ideas about the exchange of regulatory molecules via low-resistance junctions and about the control of brown fat by hormones and nerves.
GOLGI APPARATUS, GERL, AND LYSOSOMES OF NEURONS IN RAT DORSAL ROOT GANGLIA, STUDIED BY THICK SECTION AND THIN SECTION CYTOCHEMISTRY
Authors: Novikoff Phyllis M. , Novikoff Alex B. , Quintana Nelson , Hauw Jean-Jacques
Published: May 2004

New insights into the ultrastructure and phosphatase localizations of Golgi apparatus and GERL, and into the probable origin of lysosomes in the neurons of fetal dorsal root ganglia and the small neurons of adult ganglia have come from studying thick (0.5–1.0 µ) as well as thin (up to 500 A) sections by conventional electron microscopy. Tilting the thick specimens, by a goniometer stage, has helped to increase our understanding of the three-dimensional aspects of the Golgi apparatus and GERL. One Golgi element, situated at the inner aspect of the Golgi stack, displays thiamine pyrophosphatase and nucleoside diphosphatase activities. This element exhibits regular geometric arrays (hexagons) of interconnected tubules without evidence of a flattened portion (saccule or cisterna). In contrast, GERL shows acid phosphatase activity and possesses small cisternal portions and anastomosing tubules. Lysosomes appear to bud from GERL. Osmium deposits, following prolonged osmication, are found in the outer Golgi element. Serial 0.5-µ and thin sections of thiamine pyrophosphatase-incubated material demonstrate that, in the neurons studied, the Golgi apparatus is a continuous network coursing through the cytoplasm. Serial thick sections of acid phosphatase-incubated tissue suggest that GERL is also a continuous structure throughout the cytoplasm. Tubules of smooth endoplasmic reticulum, possibly part of GERL, extend into the polygonal compartments of the inner Golgi element. The possible physiological significance of a polygonal arrangement of a phosphatase-rich Golgi element in proximity to smooth ER is considered. A tentative diagram of the Golgi stack and associated endoplasmic reticulum in these neurons has been drawn.
THE EFFECT OF ETHIDIUM BROMIDE ON MITOCHONDRIAL DNA SYNTHESIS AND MITOCHONDRIAL DNA STRUCTURE IN HELA CELLS
Authors: Leibowitz Robert D.
Published: May 2004

The synthesis of mitochondrial DNA (mDNA) in HeLa cells is selectively inhibited by relatively low concentrations of ethidium bromide. After exposure of cells to strongly inhibitory concentrations of the drug, the apparent superhelix density of mDNA is rapidly increased, as judged by its buoyant density in CsCl in the presence of ethidium bromide. Mitochondrial DNA synthesized in the presence of partially inhibitory concentrations of ethidium bromide is also altered in its buoyant density in the presence of the dye, but is more heterogeneous in this respect. However, the change in buoyant density of newly synthesized mDNA may be explained by changes in structure other than a change in superhelix density, as indicated by its increased resistance to digestion by pancreatic DNase.
APPEARANCE AND FUNCTION OF ENDOGENOUS PEROXIDASE IN FETAL RAT THYROID
Authors: Strum Judy M. , Wicken Janice , Stanbury John R. , Karnovsky Morris J.
Published: May 2004

Iodination within the thyroid follicle is intimately associated with a thyroid peroxidase. In order to locate the in vivo site of iodination, the initial cytochemical appearance of this enzyme has been determined in fetal rat thyroid and its presence correlated with the onset of iodinated thyroglobulin synthesis. Peroxidase first appears in follicular cells during the 18th day of gestation. It is seen first in the perinuclear cisternae, the cisternae of the endoplasmic reticulum, and within the inner few Golgi lamellae. These organelles presumably represent sites of peroxidase synthesis. During the 19th and 20th days of gestation, there is a tremendous increase in peroxidase activity. In addition to the stained sites described, there are now many peroxidase-positive apical vesicles in the follicular cells. Newly forming follicles stain most conspicuously for peroxidase, the reaction product being heavily concentrated at the external surfaces of apical microvilli and in the adjacent colloid. Iodinated thyroglobulin becomes biochemically detectable in thyroids during the 19th day of gestation and increases greatly during the 20th day. The parallel rise in peroxidase staining that just precedes, and overlaps, the rise in iodinated thyroglobulin, suggests that apical vesicles and the apical cell membrane are the major sites of iodination within the thyroid follicle.
PLASMA AND PHAGOSOME MEMBRANES OF ACANTHAMOEBA CASTELLANII
Authors: Ulsamer Andrew G. , Wright Paddy L. , Wetzel Mary G. , Korn Edward D.
Published: May 2004

Plasma membranes were isolated from the ameba Acanthamoeba castellanii by low-speed velocity centrifugation followed by equilibrium centrifugation in a sucrose gradient. The isolated membranes had a high ratio of sterol to phospholipid (0.98 moles/mole) and of phospholipid to protein (0.43 mg/mg). The plasma membranes had very low concentrations of DNA, RNA, lipid inositol, and glycerides. Glycolipids and glycoproteins were enriched in the plasma membranes relative to their concentrations in the whole cell. The plasma membranes were also judged to be of high purity by the absence, or very low level, of enzymatic activities considered to be indicative of other cell membranes, and by electron microscope examination. Alkaline phosphatase and 5'-nucleotidase activities were enriched in the plasma membranes 13-fold relative to the whole homogenate and had higher specific activities in the plasma membranes than in any other cell fractions. A Mg++ adenosine triphosphatase (ATPase) was enriched sixfold in the plasma membranes relative to the whole homogenate. The phospholipids of the plasma membranes contained more phosphatidylethanolamine and phosphatidylserine and less phosphatidylcholine than did the phospholipids of the whole cells. There were differences in the fatty acid compositions of corresponding phospholipids in the plasma membranes and whole cells but no difference in the ratios of total saturated to unsaturated fatty acids. The membranes of phagosomes isolated from amebae that had ingested polystyrene latex had essentially the same phospholipid, sterol, and enzymatic composition as plasma membranes.
ELECTRON MICROSCOPE HISTOCHEMICAL EVIDENCE FOR A PARTIAL OR TOTAL BLOCK OF THE TRICARBOXYLIC ACID CYCLE IN THE MITOCHONDRIA OF PRESYNAPTIC AXON TERMINALS
Authors: Hajós Ferenc , Kerpel-Fronius Sándor
Published: May 2004

Respiration-linked, massive accumulation of Sr2+ is used to reveal the coupled oxidation of pyruvate, α-oxoglutarate, succinate, and malate by in situ mitochondria. All of these substrates were actively oxidized in the dendritic and perikaryal mitochondria, but no α-oxoglutarate or succinate utilization could be demonstrated in the mitochondria of the presynaptic axon terminals. A block at an early step of α-oxoglutarate and succinate oxidation is proposed to account for the negative histochemical results, since the positive reaction with pyruvate and malate proves that these mitochondria possess an intact respiratory chain and energy-coupling mechanism essential for Sr2+ accumulation. This indicates that the mitochondria in the axon terminals would be able to generate energy for synaptic function with at least some of the respiratory substrates. With regard to the block in the tricarboxylic acid cycle, the oxaloacetate necessary for citrate formation is suggested to be provided by fixation of CO2 into some of the pyruvate.
ISOLATION OF GERM CELL GOLGI APPARATUS FROM SEMINIFEROUS TUBULES OF RAT TESTES
Authors: Cunningham W. P. , Mollenhauer H. H. , Nyquist S. E.
Published: May 2004

Intact Golgi apparatus have been isolated with good purity from rat testis by a simplified sucrose gradient technique. The procedure is inherently selective in that most of the Golgi apparatus in the isolate are from germ cells in late spermatocyte or early spermatid development. No Sertoli cell Golgi apparatus are present in the fraction. Biochemical analyses showed that the enzyme N-acetylglucosamine galactosyltransferase is enhanced in the band containing Golgi apparatus. Because they are easy to isolate, and because they are involved with the formation of a specific internal cellular component (the acrosome), these Golgi apparatus will be useful objects for comparison with other kinds of Golgi apparatus and for other studies leading to a better understanding of the basic functioning of Golgi apparatus.
ACTINOMYCIN-D ENHANCEMENT OF GLUTAMINE SYNTHETASE ACTIVITY IN CHICK EMBRYO RETINAS CULTURED IN THE PRESENCE OF CORTISOL
Authors: Beif-Lehrer Liane
Published: May 2004

Cortisol can prematurely induce glutamine synthetase activity in the chick embryo retina. Under appropriate conditions, this effect can be enhanced by addition of low levels of actinomycin D; this enhancement is reversibly inhibited by cycloheximide. The magnitude of the effect is a function of time of exposure to hormone as well as antibiotic and is also a function of the age of the embryo; within the limits of the present study it did not appear to be a function of actinomycin-D concentration. The data are discussed in terms of current ideas of possible control mechanisms in animal cells.