High-salt solutions prevent reactivation of euglenoid movement in detergent-treated cell models ofEuglena gracilis

Summary Detergent-treated cell models ofEuglena gracilis showed rounded-up movement of the cell body upon addition of ATP and Ca²⁺. Reactivation of the cell models was inhibited when the cell models had been treated with solutions containing >150 mM NaCl or >300 mM KC1. When the supernatant of salt-extracted cell models was subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, two distinctive bands of 120 and 160 kDa were found to be enriched. The cell membrane and associated cytoskeleton (pellicular complex) were isolated after treatment with salt solutions, and examined by electron microscopy to identify essential cortical structures required for reactivating rounding-up cell movement. Among three regularly arranged microtubules, only one and its associated structures were selectively dissolved from the pellicular strips, while the other pellicular elements remained intact. These structures were located in the groove region where sliding between strips is believed to occur during cell shape change. These results suggest a possible involvement of microtubule 2 and its associated bridges in active sliding between adjacent pellicular strips during euglenoid movement.     

On Keelungia pulex nov. gen. et nov. sp., a heterotrophic euglenoid flagellate that lacks pellicular plates (Euglenophyceae,Euglenida)

Keelungia pulex nov. gen. et nov. sp. is described from coastal waters of NE Taiwan. The new species is heterotrophic and feeds on bacteria. Cells are oblong-ovoid, biflagellate and glide along the sides of the flask. Each cell is approximately 8–11 μm long, and one of the smallest euglenoid flagellates presently known. Keelungia lacks pellicular plates and in this respect resembles diplonemids and Symbiontida, which are thought to be among the basal groups of Euglenozoa. SEM showed the presence of 10 evenly spaced longitudinal striae in the cell surface, but the striae are difficult to see in the light microscope. TEM showed each stria to comprise a double set of very low longitudinal ridges separated by a shallow furrow, and supported by ca 5 microtubules beneath the plasmalemma, unlike the situation in diplonemids and Symbiontida. The cell surface was further subtended by an extensive system of rough cisternae of endoplasmic reticulum. Keelungia pulex is phylogenetically related to species of Ploeotia and to Lentomonas applanata, but differs in details of the feeding apparatus and in the absence of pellicular plates. Sequencing of SSU rDNA indicates that Ploeotia, Keelungia and Entosiphon form a clade near the base of the euglenoid phylogenetic tree.     

RESTING CYST FORMATION OF EUTREPTIELLA GYMNASTICA (EUGLENOPHYCEAE) IN THE NORTHERN COASTAL BALTIC SEA1

Resting cyst formation of Eutreptiella gymnastica Throndsen was observed during a mesocosm experiment, where nutrient enrichment had induced almost a unialgal bloom. Cells and resting cysts of E. gymnastica were examined in scanning (SEM) and transmission electron microscopy (TEM) and light microscopy. Mature cysts were spherical, with a smooth thick mucilaginous cover that appeared layered when observed with the TEM. Intermediate forms were spherical and lacking flagella and a mucilaginous cover; the euglenoid pellicular striation and canal opening were easily visible. The volume of these intermediate spherical cells and mature cysts was estimated to have increased threefold compared to flagellated cells and contained many paramylon grains. When the cells were grazed by zooplankton, the paramylon grains passed the gut intact and were packed into fecal pellets. Intact undigested paramylon grains were observed in SEM after the breaking up of the pellets.     

Khawkinea quartana, a Colorless Euglenoid Flagellate. I. Ultrastructure

Khawkinea quartana, a naturally occurring colorless homologue of Euglena, was examined with the electron microscope. The organism is biflagellate though only one of the 2 flagella emerges from the anterior reservoir. The pellicular strips covering the body of the organism are supported by microtubules which are continuous in part with microtubules bordering the reservoir. Additional rows of microtubules are found associated with the kinetosomes. An eyespot is located in the wall of the reservoir and, adjacent to it, the contractile vacuole. The nucleus, mitochondria, and Golgi complexes are similar to those described in other euglenoid flagellates. The food reserve is paramylon. The study supports the phylogenetic origin of Khawkinea from pigmented Euglena through the loss of chloroplasts.     

Acid phosphatase activity during spore differentiation of the red algaeGigartina teedii andChondria tenuissima

The acid phosphatase activity during carposporogenesis inGigartina and tetrasporogenesis inChondria was studied using the Gomori technique. During the first steps of gonimoblast maturation ofGigartina, portions of cytoplasm are ensheathed by ER cisternae with acid phosphatase activity, giving rise to autolysosomal concentric membrane bodies. In a similar way large mucilage sacs are severed. They extrude their contents in a kind of exocytosis. Multivesicular bodies, concentrically arranged cisternae and extracytoplasmic compartments, each with acid phosphatase activity, remain in young carpospores for some time, probably as remnants of the autophagocytotic and exocytotic events. The Golgi apparatus is poorly developed in gonimoblast cells and young carpospores. It becomes a prominent cell component in maturing carpospores and then participates in cell wall formation. Only some of the dictyosomal cisternae contain acid phosphatase; these are irregularly distributed in the dictyosome. — In pre- and postmeiotic tetraspore mother cells ofChondria massive lead deposits are found in the dictyosomes and in adjacent Golgi vesicles. Finer lead precipitates occur in ER cisternae, especially in those which are sequestering starch-grain-containing portions of the cytoplasm to give rise to autolysosomes. During cell cleavage, the dictyosomes aggregate. They become devoid of acid phosphatase activity with the exception of vesicles at the trans face. Later, Golgi stacks associate and have common, Gomori positively reacting, narrow cisternae at the cis face. The Golgi apparatus derived cored vesicles do not contain lead precipitates whereas the Golgi cisternae in the final stage of tetrasporogenesis show acid phosphatase activity. Variations in acid phosphatase distribution are explained in the light of current models of membrane flow.Dedicated to Univ.-Prof. DrO. Härtel on the occasion of his 80th birthday.     

Pellicle structure in Euglena

The pellicle of Euglena has been investigated by anoptral and phase contrast light microscopy, and by electron microscopy of osmium-fixed, Epon-embedded, lead-stained sections and of carbon/platinum replicas.

Observations on living cells show that the pellicular striatiors of Euglena spirogyra trace a left-handed (S) helix in a majority of cells, and a right-handed (Z) helix in only 5 to 30% of the cells in any one culture. All cells of E. spirogyra var. fusca have a left-handed (S) helix. Ornamentation on the pellicle takes the form of rows of knobs in various patterns. The living pellicle can be dissociated into long flat strips.

Electron microscopy shows that each pellicular strip has an elaborate cross-sectional shape, features of which are accessory teeth and ribs and a continuous ridge which articulates in a groove running along the edge of the next strip. The strips can move against one another, presumably by the ridges sliding in the grooves, and it is suggested that the joints might be lubricated by mucilage supplied from the helically disposed muciferous bodies. One single and one pair of fibrils or tubes, 200–250 Å in diameter, are regularly arranged parallel to each pellicular strip. A continuous tripartite plasmalemma, 80–100 Å thick, lies externally to the strips; external to this membrane lie the pellicular knobs. Each cell has from 35 to 45 pellicular strips, reducing to a few at the posterior end of the cell by successive fusions. Similar fusions occur at the anterior end of the cell, mainly within the canal.

These observations are compared with those made on the euglenoid pellicle by previous authors, and the following problems are discussed: direction of helix; the nature and cause of ornamentation; euglenoid movement with reference to fibrils, cytoplasmic flow, pellicle flexibility and the proteinaceous nature of the pellicle; helical and bilateral symmetry in the cell; and cet growth and division.     

Euglenoid movement inEuglena fusca: Evidence for sliding between pellicular strips

Summary InEuglena fusca, each pellicular strip carries a row of particles on its surface. The relative displacement of particles on adjacent strips was analysed by video-microscopy and evidence was obtained that adjacent pellicular strips slide relative to each other during euglenoid movement.E. fusca shows two types of euglenoid movement, oscillatory bending and rounding-up of the cell body. During oscillatory bending, the maximum velocity of sliding was 0.4 m/s and the maximum displacement distance between adjacent strips 2.3 m about their mean position. WhenE. fusca exhibited rounding-up of the cell body, particle displacement again occurred and the angle of the pellicular strips to the long axis of the cell body increased because of pellicular sliding. As a result the distance between the cell's anterior and posterior tips was reduced. There was no change in distance either between rows of particles or between particles within the same row. The findings are incompatible with theories of euglenoid movement requiring local contraction of pellicular strips and point to the likely existence of active sliding between adjacent strips.     

Light and Electron Microscopic Description of a Colorless Euglenoid,Serpenomonas costata n. g., n. sp.1

ABSTRACT. A new genus of rigid, colorless, phagotrophic euglenoid is described from a standing pond on a salt marsh. The cell body measures 21–25 μm long, is about 18 μm wide, has a slight dorso-ventral flattening, and is marked by distinct pellicular ridges. The organism, described as Serpenomonas costata, has two unequal, antapically inserted, heterodynamic flagella. The shorter flagellum is anteriorly directed during swimming and the longer one trails posteriorly. Cells move along the substrate with a creeping motion. The ingestion apparatus is composed of separate ribs extending the length of the cell and is incorporated into the ventral pellicular ridge. The apparatus is independent of the canal and reservoir and is not protrusible. The taxonomic affinities of Serpenomonas with other euglenoids are discussed.     

The Fine Structure of Rhynchocystis pilosa (Sporozoa, Eugregarinida)

SYNOPSIS. The trophozoite of Rhynchocystis pilosa obtained from the seminal vesicles of the earthworm Lumbricus terrestris was studied by light and electron microscopy. The trophozoite's cortical organization is particularly interesting because of its unusual evaginations and associated fibrillar structures. The pellicle is formed by 2 concentric membranes elevated into 60–70 alternating primary and secondary ridges extending posteriad. Numerous long ‘hairs’ or cytopilia originate along the primary ridges and each contains a system of fibrils originating from an underlying longitudinal myoneme. Longitudinal rows of pores lie between adjacent pollicular ridges. Three systems of fibrils lie in the cortex of the trophozoite. A longitudinal myoneme consisting of 12–18 fibrils lies below each primary pellicular ridge. Circular myonemes lie below the pellicle in a parallel array along the length of the organism. Each myoneme consists of 4–8 fibrils structurally similar to those of the longitudinal myonemes. Pairs of fine filaments also lie in the inner pellicular membrane along the apex of each ridge. The trophozoite's anterior end is modified as an attachment organelle consisting of 30–35 delicate pellicular folds which originate at the base of an anterior papilla. The folds extend approximately 15 μ posteriad where they become continuous with the primary pellicular ridges. The nucleus lies in the cytoplasm near the posterior level of the attachment organelle and is surrounded by a double membrane perforated by numerous pores. The cytoplasm contains numerous small vesicles which may be found in dense aggregations. These aggregations often occur in proximity to Golgi complexes and certain membrane-bound bodies. Mitochondria are abundant in the cytoplasm as are large, ovoid paraglycogen bodies. Occasionally layers of granular membranes are arranged parallel to the surface of the paraglycogen bodies but also occur thruout the cytoplasm.     

The Fine Structure of Haemoproteus columbae Sporozoites*

SYNOPSIS. The fine structure of Haemoproteus columbae sporozoites has been studied and compared to sporozoite structure as revealed by the light microscope. The sporozoites are ultrastructurally similar to those of other Haemosporidia in that they possess a 3-layered pellicle, subpellicular microtubules, polar ring, micropore, free ribosome-like particles, micronemes, a structure resembling a Golgi complex, an irregular mitochondrion, and a large nucleus. In the anterior region of the sporozoite there are 21–22 regularly arranged longitudinal subpellicular microtubules located peripherally around the cell. In the apical region the microtubules appear thickened on 1 side. The sporozoite of H. columbae has a microneme system in which 1–3 micronemes are associated with the outer pellicular membrane at the anterior end. Micronemes are found throughout the cytoplasm, but occur in greater concentration in the anterior region of the sporozoite. A clear pellicular cavity, located between the polar ring and the termination of the inner pellicular layer, is present at the anterior end of the sporozoite. Vesicular invaginations of the inner pellicular layer have been observed in the anterior region; their function is unknown. Spherical osmophilic bodies are found throughout the cytoplasm.     

The localization of thiamine pyrophosphatase activity in Meckel's cartilage cells during endochondral ossification

Summary The cytochemical distribution of thiamine pyrophosphatase (TPPase) activity in Meckel's cartilage cells of the mouse embryo has been studied during the endochondral ossification. All the cartilage cells contain reaction product within the Golgi apparatus. In immature chondrocytes, at the reserve cell zone, TPPase activity is restricted to several inner cisternae of independent Golgi apparatus. In mature cells at the proliferative cell zone, several Golgi complexes form a Golgi network connecting with each other by the TPPase positive tubular stalks. Golgi cisternae, condensing vacuoles and vesicles also contain reaction product. In the hypertrophic chondrocytes located in the calcifying zone, their disorganized Golgi apparatus still retain reaction product. Some chondrocytes, even those located within calcified or opened lacunae, exhibit intact structures and normal cytochemical enzyme distribution. These data indicate the possibility that some chondrocytes may survive and contribute the formation of mandible.     

The localization of thiamine pyrophosphatase activity in Meckel's cartilage cells during endochondral ossification

The cytochemical distribution of thiamine pyrophosphatase (TPPase) activity in Meckel's cartilage cells of the mouse embryo has been studied during the endochondral ossification. All the cartilage cells contain reaction product within the Golgi apparatus. In immature chondrocytes, at the reserve cell zone, TPPase activity is restricted to several inner cisternae of independent Golgi apparatus. In mature cells at the proliferative cell zone, several Golgi complexes form a Golgi network connecting with each other by the TPPase positive tubular stalks. Golgi cisternae, condensing vacuoles and vesicles also contain reaction product. In the hypertrophic chondrocytes located in the calcifying zone, their disorganized Golgi apparatus still retain reaction product. Some chondrocytes, even those located within calcified or opened lacunae, exhibit intact structures and normal cytochemical enzyme distribution. These data indicate the possibility that some chondrocytes may survive and contribute the formation of mandible.     

蟹累枝虫的光镜及电镜观察

利用活体观察、蛋白银染色及电镜技术对蟹累枝虫(Epistylis eriocheiri)的形态学、表膜下纤维及超微结构进行了较为系统的描述,研究发现:(1)光镜下该种群体双叉分支,活体时虫体表膜柔软,完全伸展时呈长筒状;大核马蹄形,呈横位;伸缩泡单个,斜卧口围缘下方。口围缘纤维上连细密的环状纤维,下接两端较为粗壮,中间细长的纵长纤维,构成了连续的表膜下纤维系。(2)电镜下该种表面具沟、嵴、横纹结构,沟嵴相互交错;口围纤毛花瓣状;虫体表膜层呈齿状,具嵴状突起、泡间微管、表膜孔,胞质层包括内含胞器单一的致密原生质层和富含胞器的疏松原生质层。研究对该种的上述特征在虫体的收缩机制及与其他相似种之间的关系等方面进行了讨论。       

Cubic Membrane Structure in Amoeba (Chaos carolinensis) Mitochondria Determined by Electron Microscopic Tomography

Cubic membranes occur in a variety of membrane-bound organelles in many cell types. By transmission electron microscopy (TEM) these membrane systems appear to consist of highly curved periodic surfaces that fit mathematical models analogous to those used to describe lipidic cubic phases. For the first time, a naturally occurring cubic membrane system has been reconstructed in three dimensions by electron microscopic tomography, and its periodicity directly characterized. Double-tilt tomographic reconstruction of mitochondria in the amoeba, Chaos carolinensis, confirms that their cristae (inner membrane infoldings) have the cubic structure suggested by modeling studies based on thin-section TEM images. Analysis of the membrane surfaces in the reconstruction reveals the connectivity of the internal compartments within the mitochondria. In the cubic regions, the matrix is highly condensed and confined to a continuous, small space between adjacent cristal membranes. The cristae form large, undulating cisternae that communicate with the peripheral (inner membrane) compartment through narrow tubular segments as seen in other types of mitochondria. The cubic periodicity of these mitochondrial membranes provides an ideal specimen for measuring geometrical distortions in biological electron tomography. It may also prove to be a useful model system for studies of the correlation of cristae–matrix organization with mitochondrial activity.     

Fine Structure and Cytochemistry of the Hydrogenosome of Tritrichomonas foetus1

Fine structural studies of the hydrogenosomes of Tritrichomonas foetus using an improved fixative reveal that they are enclosed by two closely apposed 6 nm membranes, which separate at some regions forming a large intramembranous vacuole where Ca⁺⁺-binding sites are located. Fixation of the cells in a glutaraldehyde solution containing 5 mM CaCl₂ and postfixation in an osmium tetroxide-potassium ferrocyanide solution led to the appearance of a reaction product associated with certain regions of the membrane of the hydrogenosomes and in the cisternae of the endoplasmic reticulum, in the recurrent flagellum, and in the plasma membrane. Treatment of ultrathin sections with EGTA removed the reaction product. These results, in association with others previously described, indicate the existence of several similarities between the hydrogenosomes and the mitochondria.     

Enzyme Cytochemistry of the Alveolar Sacs and Golgian-Like Cisternae in the Ciliate Ichthyophthirius multifiliis

In the cell cortex of the parasitic ciliate Ichthyophthirius multifiliis different kinds of cisternae were observed: the alveolar sacs, thick membrane cisternae and the endoplasmic reticulum. The thick membrane cisternae possess coated dilated rims and sometimes could be observed close to the endoplasmic reticulum. Using cytochemical techniques acid phosphatase, thiamine pyrophosphatase and nucleoside diphosphatase activities were detected in the thick membrane cisternae and in the alveolar sacs of trosphozoites. In the endoplasmic reticulum acid phosphatase activity was not detected and only very small amounts of thiamine pyrophosphatase and nucleoside diphosphatase reaction product were observed. After exit from the host, a reduction in acid phosphatase activity was evident in the alveolar sacs. At theront stage acid phosphatase activity is absent from these structures. However, high thiamine pyrophosphatase and nucleoside diphosphatase activities remain in the alveolar sacs during the whole life cycle. On the other hand, acid phosphatase, thiamine pyrophosphatase and nucleoside diphosphatase activities were detected in thick membrane cisternae of theronts. Based on the morphological aspects and enzymatic content the thick membrane cisternae of the cell cortex are designated as golgian-like cisternae. The cytochemical results point out a relationship between the alveolar sacs and the Golgi complex.     

Acid phosphatase in the guinea-pig oocytes

In guinea-pig oocytes, at every developmental stage, acid phosphatase is found histochemically in cytoplasmic granules. Ultracytochemically the reaction product is located in lysosomes and is some cisternae of the rough endoplasmic reticulum, but not in cortical granules or in vesicles with a rough endoplasmic reticulum membrane which are filled with a moderately dense homogeneous substance. It is discussed whether the acid phosphatase transforms reserve material into a storable form as has been proposed for the deposition of vitellogenin in the oocytes of lower vertebrates.     

Effect of colchicine on the Golgi complex of rat pancreatic acinar cells

Colchicine administered to adult rats at a dosage of 0.5 mg/100 g of body weight effected a disorganization of the Golgi apparatus in pancreatic acinar cells. The results obtained after various periods of treatment (10 min to 6 h) showed (a) changes in all components of the Golgi complex, and (b) occurrence of large vacuoles that predominated in cytoplasmic areas outside the Golgi region. The alterations in Golgi stacks concerned elements of the proximal and distal side: (a) accumulation of transport vesicles, (b) formation of small, polymorphic secretion granules, and (c) alterations in the cytochemical localization of enzymes and reaction product after osmification. Transport vesicles accumulated and accompanied short, dilated cisternae, which lack mostly the reaction products of thiamine pyrophosphatase, inosine diphosphatase, and acid phosphatase, and osmium deposits after prolonged osmification. After 4 to 6 h of treatment, accumulated transport vesicles occupied extensive cellular areas; stacked cisternae were not demonstrable in these regions. The changes on the distal Golgi side included GERL elements: condensing vacuoles were diminished; they were substituted by small, polymorphic zymogen granules, which appeared to be formed by distal Golgi cisternae and by rigid lamellae. Unusually extended coated regions covered condensing vacuoles, rigid lamellae, and polymorphic secretion granules. A cytochemical distinction between Golgi components and GERL was possible neither in controls nor after colchicine treatment. The cytochemical alterations in Golgi components were demonstrable 20-30 min following administration of colchicine; at 45 min, initial morphological changes--augmentation of transport vesicles and formation of polymorphic zymogen granules--became apparent. 20 min after administration of colchicine, conspicuous groups of large vacuoles occurred. They were located mostly in distinct fields between cisternae of the endoplasmic reticulum, and were accompanied by small osmium--reactive vesicles. Stacked cisternae were not demonstrable in these fields. Vacuoles and vesicles were devoid of reaction products of thiamine pyrophosphatase, inosine diphosphatase, and acid phosphatase. The results provide evidence that formation of stacked Golgi cisternae is impaired after colchicine treatment. The colchicine--induced disintegration of the Golgi complex suggests a regulatory function of microtubules in the organization of the Golgi apparatus.     

Acid phosphatase in the guinea-pig oocytes

Summary In guinea-pig oocytes, at every developmental stage, acid phosphatase is found histochemically in cytoplasmic granules, Ultracytochemically the reaction product is located in lysosomes and in some cisternae of the rough endoplasmic reticulum, but not in cortical granules or in vesicles with a rough endoplasmic reticulum membrane which are filled with a moderately dense homogenous substance. It is discussed whether the acid phosphatase transforms reserve material into a storable form as has been proposed for the deposition of vitellogenin in the oocytes of lower vertebrates.This investigation was supported by the Deutsche Forschungsgemeinschaft (SFB 105)     

Fine structure of the corpus Luteum of the raccoon during pregnancy

Summary Ultrastructure of the granulosa lutein cells of the raccoon from throughout pregnancy has been described. The lutein cells often from epithelial cords which are separated by the connective tissues, capillaries and lymphatics. Based on the arrangements and modifications of the cytoplasmic organelles and inclusions, three types of lutein cells have been recognized. The type I lutein cells predominantly contain tubular, agranular endoplasmic reticulum, juxtanuclear Golgi complexes, a few round to rod-shaped mitochondria, some free ribosomes, and occasional lipid droplets. Occasionally the tubular cristae of mitochondria and tubular smooth endoplasmic reticulum appear contiguous. The type II cells contain abundant lace-like and/or stacked fenestrated endoplasmic reticulum cisternae that frequently form membranous whorls, some tubular, agranular endoplasmic reticulum, mitochondria, and lipid droplets. Mitochondria are usually small, but unusual large ones also occur. The small, rod-to round-shaped mitochondria usually have tubular cristae; but the large, oval, elongate, and cup shaped mitochondria possess tubular, lamellar, plate like, and whorl-like cristae. The plasma membranes of the cells are complexly elaborated and folded, especially when apposing each other. In favorable sections, strands of fenestrated cisternae appose the folds of the plasma membranes. In general, the amount of cytoplasmic organelles and inclusions vary greatly in the cells. The type III cells predominantly contain lipid droplets and sparse cytoplasmic organelles. The type I and II cells are found throughout pregnancy, but the type III cells are observed from mid gestation to term. The cytological features of type I and II cells suggest that they probably secrete most of the steroids, whereas the type III cells primarily store lipids.This research was supported by UPSHS grant AM-11376 and NIH contract 69-2136.