THE FINE STRUCTURE OF CHONDROCOCCUS COLUMNARIS : I. Structure and Formation of Mesosomes

Cells of Chondrococcus columnaris were sectioned and examined in the electron microscope after fixation by two different methods. After fixation with osmium tetroxide alone, the surface layers of the cells consisted of a plasma membrane, a dense layer (mucopeptide layer), and an outer unit membrane. The outer membrane appeared distorted and was widely separated from the rest of the cell. The intracytoplasmic membranes (mesosomes) appeared as convoluted tubules packaged up within the cytoplasm by a unit membrane. The unit membrane surrounding the tubules was continuous with the plasma membrane. When the cells were fixed with glutaraldehyde prior to fixation with osmium tetroxide, the outer membrane was not distorted and separated from the rest of the cell, structural elements (peripheral fibrils) were seen situated between the outer membrane and dense layer, and the mesosomes appeared as highly organized structures produced by the invagination and proliferation of the plasma membrane. The mesosomes were made up of a series of compound membranes bounded by unit membranes. The compound membranes were formed by the union of two unit membranes along their cytoplasmic surfaces.     

Elektronenoptische Untersuchungen anEndomycopsis fibuliger auf festen Nährböden

Summary The fine structure of the filamentous and mycelial cell ofEndomycopsis fibuliger grown on the agar medium and fixed in osmium tetroxide is described. The cell wall is relatively thick. Of particular interest is the occurrence of numerous vesicles underside of the cell wall proper. They have become dilated frequently at the cell surface, probably as a result of the amylase secretion. The wall presumably is penetrated by plasmodesmata. The plasmodesmata and the intercellular connections are associated with the endoplasmic reticulum. The cytoplasmic matrix is generally granular and contains nonmembrane-limited patches of lipoid.Mitochondria are numerous and variable in shape, and have been observed in different stages of development. The mitochondria profiles are delimited from the surrounding cytoplasm by single or sometimes double electron-dense lines.The double dark lines of a unit membrane at the nuclear surface are not always clear.     

THE FINE STRUCTURE OF THE GALL BLADDER EPITHELIUM OF THE MOUSE

Sections of mouse gall bladder epithelium fixed by perfusion with buffered osmium tetroxide have been studied in the electron microscope as an example of simple columnar epithelium. The free surface presents many microvilli, each presenting a dense tip, the capitulum, and displaying a radiating corona of delicate filaments, the antennulae microvillares. Very small pit-like depressions, representing caveolae intracellulares, are encountered along the cell membrane of the microvilli. The free cell surface between microvilli shows larger cave-like depressions, likewise representing caveolae intracellulares, containing a dense material. The lateral cell borders are extensively folded into pleats, which do not interdigitate extensively with corresponding folds of the adjacent cell membrane. The terminal bars are shown to consist of thickened densities of the cell membrane itself in the region of insertion of the lateral cell wall with the free cell surface. This thickening is associated with an accumulation of dense cytoplasmic material in the immediate vicinity. The terminal bar is thus largely a cytoplasmic and cell membrane structure, rather than being primarily intercellular in nature. The basal cell membrane is relatively straight except for a conical eminence near the center of the cell, projecting slightly into the underlying tunica propria. The basal cell membrane itself is overlain by a delicate limiting membrane, which does not follow the lateral contours of the cell. Unmyelinated intercellular nerve terminals with synaptic vesicles have been encountered between the lateral walls of epithelial cells. A division of the gall bladder epithelial cell into five zones according to Ferner has been found to be convenient for this study. The following cytoplasmic components have been noted, and their distribution and appearance described: dense absorption granules, mitochondria, Golgi or agranular membranes, endoplasmic reticulum or ergastoplasm, ring figures, and irregular dense bodies, perhaps lipoid in nature. The nucleus of these cells is also described.     

Electron microscopy of thin sections of Candida utilis: The structure of the cell wall

Summary Details of the structure of the Candida utilis cell wall were described. Using intact cells, cell walls, about 0.02 m thick, have been resolved into three electron dense layers, each about 700 Å thick, made up of materials of very similar electron opacity. The central and the inner layers of the wall seem to be closely packed forming a slightly more compact structure of somewhat greater electron density. Laminations were observed in some of the inner layers of the sections, particularly in some areas where the cell wall had separated: The possibility of this lamination being associated with the presence of chitin in the framework of the cell wall is discussed. Interesting observations have also been made in the sections of the heat-killed cells confirming the existence of a central electron dense cell wall layer, differing from the inner and the outer ones. Underlying the cell wall is the cytoplasmic membrane, a not too well defined sinous structure with some invaginations.     

Low and high voltage electron microscopy of mitosis and cytokinesis in maize roots

The structure and distribution of cytoplasmic membranes during mitosis and cytokinesis in maize root tip meristematic cells was investigated by low and high voltage electron microscopy. The electron opacity of the nuclear envelope and endoplasmic reticulum (ER) was enhanced by staining the tissue in a mixture of zinc iodide and osmium tetroxide. Thin sections show the nuclear envelope to disassemble at prophase and become indistinguishable from the surrounding ER and polar aggregations of ER. In thick sections under the high voltage electron microscope the spindle is seen to be surrounded by a mass of tubular (TER) and cisternal (CER) endoplasmic reticulum derived from both the nuclear envelope and ER, which persists through metaphase and anaphase. At anaphase strands of TER traverse the spindle between the arms of the chromosomes. The octagonal nuclear pore complexes disappear by metaphase, but irregular-shaped pores persist in the membranes during mitosis. It is suggested that these form a template for pore-complex reformation during telophase. Phragmoplast formation is preceded by an aggregation of TER across the spindle at anaphase. Evidence is presented to suggest that the formation of the desmotubule of a plasmodesma is by the squeezing of a strand of endoplasmic reticulum between the vesicles of the cell plate.Abbreviations CER cisternal endoplasmic reticulum - ER endoplasmic reticulum - HVEM high voltage electron microscope - TER tubular endoplasmic reticulum - ZIO zinc iodide/osmium tetroxide     

An atypical crista resembling a "tight junction" in bean root mitochondria

The conformation and structure of an atypical crista found in a small percentage of the mitochondria in root tip cells of Phaseolus vulgaris L. have been studied electron microscopically in material fixed in glutaraldehyde followed by osmium tetroxide. In its transformation into an atypical crista, a normal crista elongates, broadens, and flattens, and the inner leaflets of its apposed unit membranes appear to fuse in a manner analogous to the formation of "tight junctions" between certain animal cells. The result is a large platelike, quintuple-layered structure, 240–260 A thick, whose long axis parallels that of the mitochondrion. The outer layers of the "plate," bordering on the mitochondrial matrix, are thickened and exhibit striking patterns in the micrographs. The structure of the plate is compared with that previously described for tight junctions between animal cells.     

THE FINE STRUCTURE OF DIFFERENTIATING XYLEM ELEMENTS

Differentiating xylem elements of Avena coleoptiles have been examined by light and electron microscopy. Fixation in 2 per cent phosphate-buffered osmium tetroxide and in 6 per cent glutaraldehyde, followed by 2 per cent osmium tetroxide, revealed details of the cell wall and cytoplasmic fine structure. The localized secondary wall thickening identified the xylem elements and indicated their state of differentiation. These differentiating xylem elements have dense cytoplasmic contents in which the dictyosomes and elements of rough endoplasmic reticulum are especially numerous. Vesicles are associated with the dictyosomes and are found throughout the cytoplasm. In many cases, these vesicles have electron-opaque contents. "Microtubules" are abundant in the peripheral cytoplasm and are always associated with the secondary wall thickenings. These microtubules are oriented in a direction parallel to the microfibrillar direction of the thickenings. Other tubules are frequently found between the cell wall and the plasma membrane. Our results support the view that the morphological association of the "microtubules" with developing cell wall thickenings may have a functional significance, especially with respect to the orientation of the microfibrils. Dictyosomes and endoplasmic reticulum may have a function in some way connected with the synthetic mechanism of cell wall deposition.     

Electron microscope studies of the human epidermis; the cell boundaries and topography of the stratum malpighii

The thin skin of the left upper quadrant of the human abdomen has been studied by electron microscopy. Tissue removed with a high speed rotary punch was fixed in osmium tetroxide or potassium permanganate. The latter fixative in our preparations is superior to osmium for the demonstration of epidermal cell membranes and certain other membranous structures of the epidermis. The cytoplasmic membranes of basal cells and cells of the stratum granulosum have been found to be relatively straight, while those of most spinous cells are sharply scalloped. The deep cells of the stratum spinosum in the rete ridge area show cell membranes and cytoplasmic structure intermediate between true basal cells and most cells of the stratum spinosum. The extracellular material of the desmosome has been found to consist of alternate dark and light laminae similar to those described by Odland (13) and Horstmann and Knoop (7).     

Electron Microscope Studies of the Human Epidermis : The Cell Boundaries and Topography of the Stratum Malpighii

The thin skin of the left upper quadrant of the human abdomen has been studied by electron microscopy. Tissue removed with a high speed rotary punch was fixed in osmium tetroxide or potassium permanganate. The latter fixative in our preparations is superior to osmium for the demonstration of epidermal cell membranes and certain other membranous structures of the epidermis. The cytoplasmic membranes of basal cells and cells of the stratum granulosum have been found to be relatively straight, while those of most spinous cells are sharply scalloped. The deep cells of the stratum spinosum in the rete ridge area show cell membranes and cytoplasmic structure intermediate between true basal cells and most cells of the stratum spinosum. The extracellular material of the desmosome has been found to consist of alternate dark and light laminae similar to those described by Odland (13) and Horstmann and Knoop (7).     

Ultrastructure of dyads in muscle fibers of Ascaris lumbricoides

The dyads of Ascaris body muscle cells consist of flattened intracellular cisternae applied to the sarcolemma at the cell surface and along the length of T-tubules. In specimens prepared by conventional methods (glutaraldehyde fixation, osmium tetroxide postfixation, double staining of sections with uranyl acetate and lead hydroxide), both the sarcolemma and the limiting membrane of the cisterna exhibit unit membrane structure and the space between them is occupied by a layer of peg-shaped densities which is referred to as the subsarcolemmal lamina. The lumen of the cisterna contains a serrated layer of dense material referred to as the intracisternal lamina. In specimens fixed in glutaraldehyde, dehydrated, and then postfixed in phosphotungstic acid, with no exposure to osmium tetroxide or heavy metal stains, the membranous components of the dyads appear only as negative images, but the subsarcolemmal and intracisternal laminae still appear dense. Except for the lack of density in membranes and in glycogen deposits, the picture produced by the latter method is very much like that of tissue prepared by conventional methods.     

THE STRUCTURE OF SOME CYTOPLASMIC COMPONENTS OF PLANT CELLS IN RELATION TO THE BIOCHEMICAL PROPERTIES OF ISOLATED PARTICLES

1. Electron micrographs of thin sections of material fixed with buffered osmium tetroxide have been used for comparison of the fine structure of isolated cytoplasmic particles from silver beet petioles and roots of germinating wheat with that of the cytoplasm of the intact cells. 2. Mitochondria of wheat roots have an external double membrane and poorly oriented internal double membranes. As compared with the structures seen in situ, the isolated mitochondria showed evidence of some disorganisation of the fine internal structure, probably due to osmotic effects. The possible influence of such changes on the enzymic properties of the isolated mitochondria is discussed. 3. The isolated plant microsomes are mainly spherical vesicular structures consisting of (a) an outer membrane enclosing (b) either an homogeneous slightly dense material (wheat root microsomes) or some granular dense material (silver beet microsomes) and (c) small dense particles, mostly associated with the vesicle membranes. 4. The cytoplasm of the wheat root cells does not contain any structures similar to the isolated microsomes but has a very dense reticular network, consisting of membranes with associated small dense particles, here called the endoplasmic reticulum. The observations indicate that the isolated microsomes arise mainly by rupture and transformation of the membranes of this structure. The effects of such extensive changes in the lipoprotein membranes on the enzymic activities of the endoplasmic reticulum, as studied in isolated microsomes, is discussed. 5. Meristematic wheat root cells contain structures which consist of smooth membranes with associated vacuoles and are similar to the Golgi zones of animal cells. The membranes of these zones probably contribute to the microsomal fraction under the conditions of preparation used for the enzymic and chemical studies previously reported.     

The ultrastructure of Candida krusei

Various methods of chemical fixation and freeze-drying of Candida krusei were compared to determine the most appropriate method for the ultrastructural investigation of the thick walled organisms of this genus. Freeze-drying without chemical fixation was of little value because of insufficient variation in electron density. Potassium permanganate was able to penetrate the intact cell but failed to show cytoplasmic glycogen and lipid and some details of the cell wall. While normal glutaraldehyde, formaldehyde and osmium tetroxide treatment failed to permeate and preserve intracellular structures, several cycles of rapid freezing (–155°C) and thawing followed by glutaraldehyde fixation and osmium tetroxide post-fixation demonstrated the intracellular details of the majority of cells so treated.     

Comparative effect of osmium tetroxide and ruthenium tetroxide on Penicillium sp. hyphae and Saccharomyces cerevisiae fungal cell wall ultrastructure

The fungal cell wall viewed through the electron microscope appears transparent when fixed by the conventional osmium tetroxide method. However, ruthenium tetroxide post-fixing has revealed new details in the ultrastructure of Penicillium sp. hyphae and Saccharomyces cerevisiae yeast. Most significant was the demonstration of two or three opaque diverse electron dense layers on the cell wall of each species. Two additional features were detected. Penicillium septa presented a three-layered appearance and budding S. cerevisiae yeast cell walls showed inner filiform cell wall protrusions into the cytoplasm. The combined use of osmium tetroxide and ruthenium tetroxide is recommended for post-fixing in electron microscopy studies of fungi.     

STRUCTURAL FEATURES OF MESOSOMES (CHONDRIOIDS) OF BACILLUS SUBTILIS AFTER FREEZE-ETCHING

Freeze-etched cells of Bacillus subtilis have been studied with the electron microscope. The outer surface of the plasma membrane, i.e. the side facing the cell wall, is covered with numerous granules and short strands, each measuring approximately 50 A in diameter. These strands are occasionally seen to enter the cell wall. The inner surface of the plasma membrane, i.e. the side facing the cytoplasm, appears to be sparsely dotted with small particles measuring about 50 A. The envelope of mesosomes differs from the plasma membrane. Blunt protrusions arise from its outer surface; the inner surface appears smooth. Stalked particles, as described by other investigators after negative staining with phosphotungstic acid, were not observed on any membrane surface in our material. Preparations were also made of specimens prefixed in osmium tetroxide prior to freeze-etching. Under these conditions the bacterial membranes appeared to be surprisingly well preserved. In contrast to directly frozen, unfixed cells, some osmium tetroxide-fixed preparations showed a differentiation in cytoplasm and nucleoplasm, which made it possible to observe the close association of the mesosome with the latter.     

An Electron Microscopic Study of the Ductuli Efferentes and Rete Testis of the Guinea Pig

The ductuli efferentes and rete testis of the guinea pig were isolated by micro dissection, fixed in cold buffered osmium tetroxide, and sectioned for examination with the light and electron microscopes. Proximal and distal segments of the ductuli efferentes were identified and their respective cytological organizations characterized. The cytological components of the rete testis are briefly described and figured. Non-ciliated and ciliated cells are found in both segments of the ductuli efferentes. The non-ciliated cells have a microvillous border, mitochondria, a Golgi complex, an ubiquitous endoplasmic reticulum, and numerous cytoplasmic vacuoles. The ciliated cells contain more mitochondria, an endoplasmic reticulum with a relatively sparse distribution, and few, if any, cytoplasmic vacuoles. A regional difference exists in proximal and distal segments based on the distribution, size, number, and electron opacity of the cytoplasmic vacuoles. Attention was paid to the disposition of the endoplasmic reticulum and its relation to the system of cytoplasmic vacuoles. These findings are interpreted as suggesting that the continuity of the vacuolar system with elements of the endoplasmic reticulum represents a pathway for transfer of large quantities of fluid, an activity which has long been ascribed to the epithelium of the ductuli efferentes. Periductular capillaries possess pore-like apertures in their endothelia similar to those in other tissues known to engage in fluid transfer.     

The fine structure of elongate gametocytes of Leucocytozoon ziemanni (Laveran).

In an effort to establish comparative data within the genus Leucocytozoon, elongate gametocytes of L. ziemanni from naturally infected great horned owls (Bubo virginianus) were examined by electron microscopy. Micro- and macrogametocytes proved to be easily distinguishable at the electron microscopic level due to dramatic dimorphism at maturity and cytoplasmic and nuclear morphology. The parasite membrane architecture, number and type of cytoplasmic ribosomes of both micro- and macrogametocytes, presence and arrangement of osmiophilic bodies and electron dense spheres, mitochondrial morphology, endoplasmic reticulum cisternae morphology, mitochondria containing pocket infoldings of the nuclear membrane of the microgametocytes, and cytostome and food vacuole formation compare favorably with available information on L. simondi and L. smithi. Comparative variations exist only in that L. ziemanni gametocytes apparently lack compartmentalization of the cytoplasm by aligned unit membranes and parasite induced separations of the host cell nucleus as reported for L. simondi.     

Ultrastructural and ultrahistochemical studies on ventricular endocardial cells in teleosts (Pisces)

The ultrastructure of the ventricular endocardial cells in 17 bony-fish species representing eight families, is described. In species of Characidae, Cobitidae, Cyprinidae, and Gyrinocheilidae these cells are flat (1–2 µm at nucleus) and contain numerous ribosomes, some few bristle-coated vesicles (BCV) and small (0.243 pm) electron dense inclusion bodies. However, in species of Cichlidae, Gadidae, and Poeciliidae most endocardial cells appear relatively thicker (2–5 µm at nucleus) and contain numerous BCV, tubules of agranular endoplasmic reticulum, and large (0.5-1.5 µm) moderately electron dense bodies (MDB). In the MDB occur a number of small (20–150 nm) electron dense granules. Within the family Anabantidae, most endocardial cells seem to be of the first type in Colisa laliu and Trichogaster leeri. whereas in Helosioma iemmincki there are numerous cells of the second type. When glutaraldehyde/tannic acid fixed heart tissue of Pollachius virens is treated with ferrous chloride, ferric chloride, or osmium tetroxide, and grid stained by uranyl and lead solutions, damaged endocardial cells appear highly electron dense, whereas undamaged ones are electron lucent. Further, when glutaraldehyde fixed tissue is treated with osmium tetroxide/potassium ferrocyanide the subendocardial space is filled by a highly electron dense material. The methods described in this study make it possible to distinguish between those endocardial vacuoles having structural contact with the cell membrane, and those lacking such contact, and also to determine whether the lumen of the former is continuous with the subendocardial space, or with the intertrabecular lumen.     

A cytochemical study on the effects of energy deprivation on autophagocytosis in Ehrlich ascites tumor cells

The effect of energy deprivation on autophagocytosis in Ehrlich ascites tumor cells was studied using cytochemical techniques. Autophagocytosis was induced with vinblastine incubation (0.1 mM) and the cellular ATP-level was lowered with 2-deoxy-D-glucose (0.35 mM). Acid phosphatase was used as a marker for lysosomal enzymes and imidazole-buffered osmium tetroxide impregnation in order to study the effects of energy deprivation on the maturation of autophagic vacuole (AV) membranes. Control and vinblastine treated cells maintained their ATP-levels throughout the incubation period tested (120 min). 2-Deoxy-D-glucose alone and with vinblastine decreased the intracellular ATP-level significantly after only 3 min incubation. Most of the AV's in control and vinblastine treated cells contained degraded material and acid phosphatase activity. Their membranes were stained only slightly or not at all with imidazole-buffered osmium tetroxide. 2-Deoxy-D-glucose alone as well as with vinblastine induced in particular an accumulation of early stages of AV's. These vacuoles contained undegraded cytoplasmic material and no acid phosphatase activity and their membranes were stained usually partly with imidazole-buffered osmium tetroxide. The membranes of some early AV's resembled endoplasmic reticulum and still had attached ribosomes. It was concluded that the inhibition of cellular energy production used in the present study did not inhibit autophagic sequestration but retarded the maturation of AV membranes and impaired the functioning of lysosomal hydrolases.     

A cytochemical study on the effects of energy deprivation on autophagocytosis in Ehrlich ascites tumor cells

Summary The effect of energy deprivation on autophagocytosis in Ehrlich ascites tumor cells was studied using cytochemical techniques. Autophagocytosis was induced with vinblastine incubation (0.1 mM) and the cellular ATP-level was lowered with 2-deoxy-d-glucose (0.35 mM). Acid phosphatase was used as a marker for lysosomal enzymes and imidazole-buffered osmium tetroxide impregnation in order to study the effects of energy deprivation on the maturation of autophagic vacuole (AV) membranes.Control and vinblastine treated cells maintained their ATP-levels throughout the incubation period tested (120 min). 2-Deoxy-d-glucose alone and with vinblastine decreased the intracellular ATP-level significantly after only 3 min incubation. Most of the AV's in control and vinblastine treated cells contained degraded material and acid phosphatase activity. Their membranes were stained only slightly or not at all with imidazole-buffered osmium tetroxide. 2-Deoxy-d-glucose alone as well as with vinblastine induced in particular an accumulation of early stages of AV's. These vacuoles contained undegraded cytoplasmic material and no acid phosphatase activity and their membranes were stained usually partly with imidazole-buffered osmium tetroxide. The membranes of some early AV's resembled endoplasmic reticulum and still had attached ribosomes.It was concluded that the inhibition of cellular energy production used in the present study did not inhibit autophagic sequestration but retarded the maturation of AV membranes and impaired the functioning of lysosomal hydrolases.     

THE MICROSTRUCTURE OF THE COMPOUND EYES OF INSECTS

The apposition eyes of two diurnal insects, Sarcophaga bullata (Diptera) and Anax junius (Odonata), have been examined with the electron microscope. In the latter case only the rhabdom is described. The rhabdom of the fly consists of a central matrix and seven rhabdomeres, one for each retinula cell. The rhabdomeres show an ordered internal structure built up of transverse tubes, hexagonal in cross-section. These slender compartments running the width of the rhabdomere are 370 A in diameter. After fixation with osmium tetroxide the walls of the compartments are more electron dense than the interiors. The retinula cells contain mitochondria, and pigment granules smaller than those found in the pigment cells. These granules tend to cluster close behind the membranes which separate the retinula cells from their rhabdomeres. The rhabdom of the dragonfly is a single structure which appears to be composed of three fused "rhabdomeres," each similar to a rhabdomere of Sarcophaga. Reasons are given for believing that the rhabdom may be the site of photoreception, as well as the organ for analyzing plane-polarized light, as suggested by other workers.