LIGHT MICROSCOPE AND ULTRASTRUCTURAL STUDIES OF THE MALE GAMETOPHYTE IN GINKGO BILOBA: THE SPERMATOGENOUS CELL

A light microscope and ultrastructural study was made of the pollen tube of Ginkgo biloba, with special emphasis given to the spermatogenous cell that gives rise to two motile sperms. Just prior to the mitotic division that results in the formation of two sperms, the spermatogenous cell consists of a large nucleus, two blepharoplasts, two large osmiophilic globules, and a conspicuous lipo-protein body. Other organelles in the cytoplasm include numerous electron-dense proplastids (with some lamellar development), mitochondria, small vacuoles, and lipid bodies. Ribosomes are present in abundance, but endoplasmic reticulum and dictyosomes are sparse. The nucleus, prior to mitosis, is relatively Feulgen-negative, due undoubtedly to the diffuse distribution of DNA. Each blepharoplast, the main organelle of interest, is nearly spherical, measures 3.5–4.5 μm in diam, and supports about 1,000 probasal bodies. The interior of a blepharoplast consists of an electron-dense matrix and of less dense regions which appear to be infiltrated by a network of microtubules. Each probasal body is composed of a cylinder of nine separate tubules (singlets) at the basal or proximal end. The cylinder becomes elaborated distally into nine pairs of subtubules (doublets) and then into nine sets of subtubules (triplets). A central tubule is present the entire length of the probasal body. Some of the subtubules, as well as microtubules from the interior of the blepharoplast, extend into the cytoplasm and probably constitute the “astral rays” as seen with the light microscope. Comparisons are made with other published accounts of the organization of blepharoplasts in plants and of centrosomes and centrioles in animals.     

Mago nashi is essential for spermatogenesis in Marsilea

Spermatogenesis in Marsilea vestita is a rapid process that is activated by placing dry microspores into water. Nine division cycles produce seven somatic cells and 32 spermatids, where size and position define identity. Spermatids undergo de novo formation of basal bodies in a particle known as a blepharoplast. We are interested in mechanisms responsible for spermatogenous initial formation. Mago nashi (Mv-mago) is a highly conserved gene present as stored mRNA and stored protein in the microspore. Mv-mago protein increases in abundance during development and it localizes at discrete cytoplasmic foci (Mago-dots). RNA interference experiments show that new Mv-mago protein is required for development. With Mv-mago silenced, asymmetric divisions become symmetric, cell fate is disrupted, and development stops. The alpha-tubulin protein distribution, centrin translation, and Mv-PRP19 mRNA distribution are no longer restricted to the spermatogenous cells. Centrin aggregations, resembling blepharoplasts, occur in jacket cells. Mago-dots are undetectable after the silencing of Mv-mago, Mv-Y14, or Mv-eIF4AIII, three core components of the exon junction complex (EJC), suggesting that Mago-dots are either EJCs in the cytoplasm, or Mv-mago protein aggregations dependent on EJCs. Mv-mago protein and other EJC components apparently function in cell fate determination in developing male gametophytes of M. vestita.     

Immunofluorescent localization of cytoskeletal tubulin and actin during spermatogenesis inPteridium aquilinum (L.) Kuhn

Summary Details concerning the appearance and behaviour of blepharoplasts during spermatogenesis, and the assembly of the cytoskeletal motile apparatus of spermatids were elucidated by immunofluorescence microscopy using antibodies to tubulin and actin, applied to material prepared from antheridia of the fernPteridium aquilinum (L.) Kuhn. Blepharoplast immunofluorescence with antitubulin first appears as spheres at the future spindle poles prior to the last spermatogenous division. Developing spermatids each have one blepharoplast, which gives rise to a triangular layer corresponding to the incipient microtubule ribbon. Compared to the ribbon, immunoreactivity of the multilayered structure is relatively weak. Intensely fluorescing basal bodies appear, increase in number, and become arranged in rows along two edges of the microtubule ribbon as it widens and elongates. Along the dorsal edge is a dense file of basal bodies spaced at about 0.3 m intervals, parallel to each other and oriented at 145° to the multilayered structure. This spacing and orientation is maintained throughout spermatid development. Basal bodies at the opposite edge are initially oriented at 115° to the multilayered structure but become rearranged into small groups that rotate so that the angle is reduced to 55–70° by the time the assembly of flagella commences on both sets of basal bodies. By this stage the microtubule ribbon has encircled about 2/3 of the nuclear circumference and the nucleus is assuming a crescent shape. In fully developed spermatozoids the groups of basal bodies are oriented at 25° to the multilayered structure, parallel to the long body of the now helical nucleus. Immunofluorescence using antiactin showed that towards the completion of nuclear shaping, actin forms a strip along the helical multilayered structure. Detergent-extraction of mature spermatozoids revealed that actin is associated also with the flagellar band, particularly with basal bodies.Abbreviations MLS multilayered structure - MT microtubule     

Recovery of microtubules on the blepharoplast of Ceratopteris spermatogenous cells after oryzalin treatment

Most land plants have ill-defined microtubule-organizing centers (MTOCs), consisting of sites on the nuclear envelope or even along microtubules (MTs). In contrast, the spermatogenous cells of the pteridophyte Ceratopteris richardii have a well-defined MTOC, the blepharoplast, which organizes MTs through the last two division cycles. This allows a rare opportunity to study the organization and workings of a structurally well-defined plant MTOC. In this study, antheridial plants were treated with levels of oryzalin that cause complete MT loss from the cells containing blepharoplasts. The oryzalin was then washed out and plants were allowed to recover for varying amounts of time. If the spermatogenous cells were fixed prior to washing out, the blepharoplasts had an unusual appearance. In the matrix (pericentriolar) material where MT ends are normally found, clear areas of about the diameter of MTs were seen embedded in a much deeper matrix, made more obvious in stereo pairs. Occasionally, the matrix material was highly distended, although the basal body template cylinder morphology appeared to be unaltered. The blepharoplasts often occurred as clusters of 2 or 4, indicating that blepharoplast reproduction is not affected by the lack of MTs, but that their movement to the poles is. Gamma (gamma) tubulin antibodies labeled the edge of the blepharoplast in areas where the pits are located, indicating that these might be sites for MT nucleation. After wash out, the new MTs always re-appeared on the blepharoplast and the recovery occurred within an hour of washout. MT lengths increased with increasing washout time and were indistinguishable from untreated blepharoplasts after 24 h of recovery. After washout, arrays formed in new sperm cells such as the spline and basal bodies were often malformed or present in multiple copies, as were the blepharoplasts in these cells prior to wash out. These data indicate that the blepharoplast serves as the site of MT nucleation and organization even after complete MT de-polymerization.     

Actin-Based Domains of the "Cell Periphery Complex" and their Associations with Polarized "Cell Bodies" in Higher Plants

Abstract: Nascent cellulosic cell wall microfibrils and transverse (with respect of cell growth axis) arrays of cortical microtubules (MTs) beneath the plasma membrane (PM) are two well established features of the periphery of higher plant cells. Together with transmembrane synthase complexes, they represent the most characteristic form of a “cell periphery complex” of higher plant cells which determines the orientation of the diffuse (intercalary) type of their cell growth. However, there are some plant cell types having distinct cell cortex domains which are depleted of cortical MTs. These particular cell cortex domains are, instead, typically enriched with components of the actin‐based cytoskeleton. In higher plants, this feature is prominent at extending apices of two cell types displaying tip growth ‐ pollen tubes and root hairs. In the latter cell type, highly dynamic F‐actin meshworks accumulate at extending tips, and they appear to be critical for the apparently motile character of these subcellular domains. Importantly, tip growth of both root hairs and pollen tubes is immediately stopped when the most dynamic F‐actin population is depolymerized with low levels of anti‐F‐actin drugs. Intriguingly, MTs of tip‐growing plant cells are organized in the form of longitudinal arrays, throughout the cytoplasm, which interconnect the extending tips with the subapical nuclei. This suggests that actin‐rich cell cortex domains polarize plant “cell bodies” represented by nucleus‐MTs complexes. A similar polarization of “cell bodies” is typical of mitotic and cytokinetic plant cells. A further type of MT‐depleted and actomyosin‐enriched plant cell cortex domain comprises the plasmodesmata. Primary plasmodesmata are formed during cytokinesis as part of the myosin VIII‐enriched callosic cell plates, representing “juvenile” forms of the plant “cell periphery complex”. In phylogenetic terms the association between F‐actin and the PM may be considered for a more “primitive” form of cellular organization than does the association of cortical MTs with the PM. We hypothesize that the actin cytoskeleton is a natural partner of the PM in all eukaryotic cells. In most plant cells, however, it was replaced by a tubulin‐based “cell periphery apparatus” which regulates, via still unknown mechanisms, the spatial deposition of nascent cellulosic microfibrils synthesized by PM‐associated synthase complexes.     

Partial purification of centrosomes from Chinese hamster ovary cells

A method for the purification of centrosomes from cultured Chinese hamster ovary cells is described. The centrosomes produced by application of this method show good retention of their intracellular morphology: the centrioles are surrounded by an “osmiophilic halo” containing numerous pericentriolar or satellite bodies. The latter spherical structures are approx. 55 nm in diameter and possess a densely staining central core surrounded by an envelope of lighter material. The number of satellite bodies associated with the centrioles seems variable, as does their spatial disposition within the osmiophilic halo.     

ULTRASTRUCTURE OF CEPHALEUROS VIRESCENS (CHROOLEPIDACEAE; CHLOROPHYTA). III. ZOOSPORES

Transmission electron microscopic examination of Cephaleuros virescens Kunze growing on leaves of Camellia spp. and Magnolia grandiflora L. indicates that unreleased zoospores in mature zoosporangia are similar to those produced by the related genus Phycopeltis epiphyton Millardet and unlike the quadriflagellate motile cells produced by taxa in other families of Chlorophyta. The zoospores bear four smooth isokont bilaterally “keeled” flagella containing typical “9 + 2” axonemes and lacking scales. Flagellar insertion is apical and the parallel basal bodies overlap laterally at two levels. A cross section through the four basal bodies shows a trapezoidal arrangement wherein the two upper (anterior) basal bodies are closer together than are the lower (posterior) two. Serial sections indicate that diagonally opposing upper and lower basal bodies anchor flagella which emerge from the same side of the apical papilla. Each of the four basal bodies is associated with a microtubular spline which extends beneath the plasmalemma to the posterior end of the zoospore. A distinct multilayered structure is associated with each of the lower basal bodies. A nucleus, mitochondria (two of which are closely associated with the nucleus and spline microtubules), a chloroplast, and cytoplasmic haematochrome droplets are present in each zoospore. Pyrenoids and eyespots are absent. Flagellar insertion is characterized by “reversed bilateral symmetry”; and zoospores with both right-handed and left-handed arrangements are produced. The ultrastructure of the zoospores clearly indicates that: 1) the mode of flagellar insertion: 2) morphology, number, and arrangement of multilayered structures, and 3) bilaterally keeled flagella are characteristic of the Chroolepidaceae.     

The Spermatozoa of the Thysanuran Insects Petrobius brevistylis Carp. and Lepisma saccharina L.

The spermatozoa of Petrobius and Lepisma share a few general insect features (filamentous shape, two mitochondria, compact acrosome vesicle, bilateral symmetry) but differ fundamentally with regard to specializations. In Petrobius, a long coiled acrosome, a coiled nucleus, and a “body” with axonema, two mitochondria, and a pair of lateral bodies follow each other in normal sequence. In Lepisma the acrosome is a small vestige in the spoon-shaped anterior end, the centriole is dislocated anteriorly, and nucleus, two mitochondria and axonema run like parallel filaments through most of the spermatozoon. The centriole adjunct develops into a postnuclear body in Lepisma but forms a pair of complicated “lateral bodies” in Petrobius. It is concluded that ancestral forms must have had fairly primitive spermatozoa and that specialization has proceeded independently within each evolutionary line.     

Investigation on Fertilization of Cephalotaxus

The structure of spermatogenous cell of Cephalotaxus is unique among the gymnosperms. While towards the mature stage, its nucleus is close to one side of the spermatogenous cell, and on the other side there is abundant and prominent . cytoplasm, which contain a group of the aggregate cytoplasms of radial arrangement similar to blepharoplast of spermatogenous cell of Ginkgo. But, there are two opposite blepharoplasts at either side of the nucleus in Ginkgo, and while there is only one blephareplast at one side in Cephalotaxus. This is one feature of the sexual process in Cephalotaxus. When the pollen tubes approach the top of the archegonia, the division of the spermatogenous cell takes place and there are two almost similar sperm cells both in size and morphology. It is interesting to note that the cytoplasm of the sperm cell contains certain granules of nucleolus-like structure, which appears to be a rare phenomenon among the gymnosperms. This is another feature of the sexual process in Cephalotaxus. These two features are the important characters of Cephalotaxaceae. The egg morphology of Cephalotaxus is also unique among the conifers, its outline looks like a carrot. The upper part of the egg is rather wide and is about 85 to 108 μm in width. On the other hand, the opposite end is gradually becoming narrow and about 910 to 1100 μm in total length. So the ratio of the length and width in the egg of Cephalotaxus is about 10:1. The structure of the egg in Celhalotaxus fortunei and C. oliveri have the following common feature: 1. When their eggs mature the cytoplasm of the egg at lower part of the nucleus possesses deep- staining and fine granules of 2 to 3 groups of aggregate cytoplasm. 2. During maturation of the egg, some of the granules of nucleolus-like structure are scattered in the cytoplasm. As fer- tilization takes place the number of these granules reaches the peak. This condition has been encountered in the egg of Amentotaxus argotaenia. Therefore we could conclude that they are closely related between Cephloraxaceae and Taxaceae. The fertilization of Cephalotaxus fortunei occured on May 10 to 24 (1983), and that of C. oliveri took place on May 28 to June 13 (1983). The fertilization of the genus belong to the type of undergoing mitosis prior to complete fuse of both male and female nuclei. This type of fertilization has been found only in Pinaceae and Cephalotaxaceae. After fertilization the structure of fertilized egg becomes prominent in polar organization. In other words, the cytop- lasm at upper part of the fertilized egg becomes highly vacuolated and that at lower portion, conversely, is rich in abundant proteinous vacuoles and certain granules of nucleolus-like structure dispersed in the cytoplasm. Because the division and differentiation of the proembryo are proceeding at the base of the archegonium, the large inclusions and the nucleolus-like granules may be involved in the nourishing and development of the proembryo.     

Ultrastructural pattern of the nucleus of Entamoeba coli

The ultrastructural organization model of the nucleus of Entamoeba coli, as outlined on the basis of our electron microscope research, shows the following characteristics: (1) karyosome located eccentrically in the nucleoplasm (in the equatorial sections of the nucleus), the heterochromatin laid out in rough, tangled sinuous bands; (2) “ball of thread-like” formations at the level of the heterochromatin border; and (3) circular dark bodies falling into two morphological types on account of their size, the “larger inclusions” type being morphologically similar to the “button bodies” of Entamoeba histolytica according to Ludwik &; Shipstone (1970). Characteristic organized clumps of these structures have not been described before and are especially evident in the cystic nuclei. These findings are discussed in relation to the nuclear organization pictures given by E. histolytica, E. moshkovskii, and E. invadens under the electron microscope.     

Microglial motility in the rat facial nucleus following peripheral axotomy

Microglial motility was studied in living mammalian brain tissue using infrared gradient contrast microscopy in combination with video contrast enhancement and time lapse video recording. The infrared gradient contrast allows the visualization of living cells up to a depth of 60 μm in brain slices, in regions where cell bodies remain largely uninjured by the tissue preparation and are visible in their natural environment. In contrast to other techniques, including confocal microscopy, this procedure does not require any staining or labeling of cell membranes and thus guarantees the investigation of tissue which has not been altered, apart from during preparation. Microglial cells are activated and increase in number in the facial nucleus following peripheral axotomy. Thus we established the preparation of longitudinal rat brainstem slices containing the axotomized facial nucleus as a source of activated microglial cells. During prolonged video time lapse recordings, two different types of microglial cell motility could be observed. Microglial cells which had accumulated at the surface of the slice remained stationary but showed activity of the cell soma, developing pseudopods of different shape and size which undulated and which were used for phagocytosis of cell debris. Microglial phagocytosis of bacteria could be documented for the first time in situ. In contrast, ameboid microglia which did not display pseudopods but showed migratory capacity, could be observed exclusively in the depth of the tissue. Some of these cells maintained a close contact to neurons and appeared to move along their dendrites, a finding that may be relevant to the role of microglia in “synaptic stripping”, the displacement of synapses following axotomy. This approach provides a valuable opportunity to investigate the interactions between activated microglial cells and the surrounding cellular and extracellular structures in the absence of staining or labeling, thus opening a wide field for the analysis of the cellular mechanisms involved in numerous pathologies of the CNS.     

SPORANGIAL STRUCTURE AND ZOOSPOROGENESIS IN CHORDA TOMENTOSA (LAMINARIALES)1,2,3

Members of the genus Chorda represent the simplest form of sporophyte in the order Laminariales. The present study deals with reproduction in Chorda tomentosa, involving the initiation, growth, and structure of the sporangium and the process of zoosporogenesis. The simple tube-like sporophyte of Chorda tomentosa grows in diameter by means of repeated anticlinal divisions in a superficial meristematic layer known as the meristoderm. The onset of reproduction is marked by the conversion of the meristoderm from contributing cells to the vegetative plant body to producing 2 new cell types: paraphyses and sporangial mother cells. At the time of initiation, sporangial mother cells are crescent shaped and possess a densely staining cytoplasm. Sporangial mother cells increase in size, become ellipsoid, decrease in staining density, and undergo meiosis. After meiosis, sporangia increase in size while their nuclei undergo successive cycles of synchronous mitotic divisions. Sporangia increase to a maximum length of 120 μ;m at which time they possess the characteristic “cap” found in all members of the order studied thus far. At this stage the protoplast of the sporangium is organized into a peripheral layer of nucleus-chloroplast pairs and a central region of vacuoles. Cleavage furrows begin to form at the cell membrane and are met by furrows developing in the interior of the cytoplasm resulting in the division of the entire protoplast into separate units. Each unit is an individual zoospore. The biflagellate zoospores contain a nucleus, one chloroplast with eyespot, perinuclear Golgi, and several bodies of presumed storage carbohydrate. The occurrence of a small population of early developing sporangia is described. In essential details, the origin, development, and structure of sporangia in Chorda tomentosa are identical to all earlier observations in the Laminariales.     

The spermatogenous body cell of the coniferPicea abies (Norway spruce) contains actin microfilaments

Summary In conifer pollen, the generative cell divides into a sterile stalk cell and a body cell, which subsequently divides to produce two sperm. InPicea abies (Norway spruce, Pinaceae) this spermatogenous body cell contains actin microfilaments. Microfilament bundles follow the spherical contour of the body cell within the cell cortex, and also traverse the cytoplasm and enmesh amyloplasts and other organelles. In addition, microfilaments are associated with the surface of the body cell nucleus. The sterile stalk cell also contains microfilament bundles in the cytoplasm, around organelles, and along the nuclear surface. Within the pollen grain, microfilament bundles traverse the vegetative-cell cytoplasm and are enriched in a webbed cage which surrounds the body cell. Microfilaments were identified with rhodamine-phalloidin and with indirect immunofluo-rescence using a monoclonal antibody to actin. The majority of evidence in literature suggests that the spermatogenous generative cell in angiosperms does not contain actin microfilaments, so the presence of microfilaments within the spermatogenous body cell inP. abies appears to be a fundamental difference in sexual reproduction between conifers and angiosperms.     

Structure of the abdominal receptor responsive to internally applied pressure in the blood-feeding insect,Rhodnius prolixus

Summary The sensory receptor responsive to pressure applied internally to the ventral abdominal body wall of the blood-feeding insects, Rhodnius prolixus, is a single sense cell containing, at its distal end, a cilium enclosed within a scolopale, a densely staining structure characteristic of insect scolopidial sensilla. A small spherical structure lies within a dilation near the midregion of the cilium, and contains nine heavily staining bodies, the position of each corresponding to a pair of microtubules in the cilium. Proximal to the dilation, the microtubules are organized in a ring of nine pairs with one microtubule of each pair associated with dyneinlike arms. Dastal to the dilation a central pair of microtubules is present, but dyneinlike arms are absent. The scolopale cell, which gives risc to the scolopale, has cytoplasmic invaginations that form an elaborate array of extracellular compartments surrounding the body wall of the sense cell. These compartments may serve to dampen high frequency vibrations permitting the receptor to respond to pressure exerted by touch, an attribute in keeping with the receptor's proposed function of detecting abdominal distension related to the size and movement of the stomach.     

Ultrastructure of spermatogenesis and mature spermatozoa in the flatworm Prosthiostomum siphunculus (Polycladida,Cotylea)

This is the first study investigating spermatogenesis and spermatozoan ultrastructure in the polyclad flatworm Prosthiostomum siphunculus. The testes are numerous and scattered as follicles ventrally between the digestive ramifications. Each follicle contains the different stages of sperm differentiation. Spermatocytes and spermatids derive from a spermatogonium and the spermatids remain connected by intercellular bridges. Chromatoid bodies are present in the cytoplasm of spermatogonia up to spermatids. During early spermiogenesis, a differentiation zone appears in the distal part of spermatids. A ring of microtubules extends along the entire sperm shaft just beneath the cell membrane. An intercentriolar body is present and gives rise to two axonemes, each with a 9 + “1” micro‐tubular pattern. Development of the spermatid leads to cell elongation and formation of a filiform, mature spermatozoon with two free flagella and with cortical microtubules along the sperm shaft. The flagella exit the sperm shaft at different levels, a finding common for acotyleans, but so far unique for cotylean polyclads. The Golgi complex produces numerous electron‐dense bodies of two types and of different sizes. These bodies are located around a perinuclear row of mitochondria. The elongated nucleus extends almost along the entire sperm body. The nucleus is wide in the proximal part and becomes narrow going towards the distal end. Thread‐like chromatin mixed with electron‐dense intranuclear spindle‐shaped bodies are present throughout nucleus. The general sperm ultrastructure, the presence of intranuclear bodies and a second type of cytoplasmic electron‐dense bodies may provide characters useful for phylogenetic analysis.     

Neuronal organization of the accessory olfactory bulb of the lizard Podarcis hispanica: Golgi study

The neuronal organization of the accessory olfactory bulb (AOB), which receives sensory information from the vomeronasal organ, was described in a squamate reptile (Podarcis hispanica) by means of light microscopy. Using the Golgi-impregnation method, seven neuronal types could be distinguished: Periglomerular cells constitute a morphologically heterogeneous population of small neurons located between and around the glomeruli. The mitral cells are diffusely distributed in the AOB. Their cell bodies are usually located within the mitral cell layer, but some of them could be also observed in the plexiform layers. Mitral cells were classified into three subgroups on the basis of their sizes and dendritic tree morphologies. Thus, the “outer mitral cells” have the biggest cell bodies, and their distal secondary dendrites are mainly distributed rostrocaudally in the external plexiform layer. The “inner mitral cells” have large cell bodies, and their secondary dendrites are distributed dorsoventrally and are located deeper than those of the other two subgroups. The third type, the “small mitral cells,” is the smallest one among mitral cells in the AOB, and from their cell bodies, only two main dendritic trunks arise. The granule cells are composed of several categories based on their different cell body locations and dendritic tree morphologies. Thus, the “superficial granule cells” are located exclusively in the external plexiform layer and have small dendritic fields. The “middle granule cells” have fusiform cell bodies—situated in the internal plexiform layer—and present a wide dendritic projection area. Finally, the “deep granule cells” are distributed throughout the granule cell layer and include a great variety of dendritic tree morphologies. The distribution and morphological features of all neuronal types constituting the AOB of Podarcis were compared with those reported on other vertebrates. The results suggest that the lamination pattern and neuronal organization of the AOB in lizards are more similar to that of mammals than to that of the remaining vertebrates.     

MITOSIS IN THE ALGA VACUOLARIA VIRESCENS

Details of mitosis in the chloromonadophycean alga Vacuolaria virescens Cienk. have been studied with the light microscope. The chromosomes are relatively large (up to μ in length at metaphase) and so mitotic stages are readily distinguishable. Chromosomes can be recognized in interphase nuclei as fine strands of chromatin. Contraction of these chromosomes marks the beginning of mitosis and continues progressively until the transition from metaphase to anaphase. Disintegration of nucleoli is complete by late prophase and nucleolar reformation begins in telophase. Some chromosomes exhibit less densely stained regions; centromeres are also present as indicated by their differential staining and by the behavior of chromosomes at metaphase and anaphase. At anaphase progeny chromosomes move apart parallel to the division axis of the nucleus. As anaphase progresses the chromosomes fuse at the polar surface of the progeny chromosome groups. This process continues in telophase and the chromosome groups become more spherical. By the end of telophase nucleolar reformation has begun and the chromosomes have relaxed to their interphase condition.     

Structural and immunocytochemical characterization of microtubule organizing centers in pteridophyte spermatogenous cells

Summary During the development of the spermatogenous cells, the pteridophyteCeratopteris richardii produces three structurally well-defined microtubule organizing centers (MTOCs). The blepharoplast, a spherical body that occurs during the last two spermatogenous divisions, organizes two microtubule (MT) arrays, one associated with a nuclear indentation and the other that organizes the spindle apparatus for the final divisions. After the last spermatogenous division, the blepharoplast reorganizes to produce two new putative MTOCs: the lamellar strip (LS) of the multilayered structure (MLS), which apparently organizes the spline microtubule array, and an amorphous zone (AM), that connects the basal bodies. Thin and semi-thin sections of this tissue were probed with antisera which recognize MTOCs in lower eukaryotes and animals to determine if any of these structures contain MTOC-associated proteins or epitopes recognized by monoclonal antisera. Gamma tubulin antibodies, which recognizeonly the minus ends of MTs in mammalian cells, label along the MT in all arrays found in the pteridophyte spermatogenous cells. Kinetochore MTs are unlabelled near the kinetochore, however. The monoclonal antibodies MPM-2 and C-9, that recognize centrosomal and nuclear epitopes in mammalian cells, label the interphase nucleus, the cytoplasm of mitotic cells, and the blepharoplast during both nuclear indentation and spindle formation. Double labelling of the blepharoplast-containing cells with anti-tubulin and either MPM-2 or C-9 reveals that the blepharoplast-associated fluorescence is the focus of the tubulin arrays. Centrin labels the reorganizing blepharoplast, the MLS, the AM, and a stellate pattern in the transition region of the flagella. These data indicate the usefulness of the structurally well-recognized MTOCs in pteridophyte spermatogenous cells in investigation of land plant MTOCs.     

ULTRASTRUCTURE OF CEPHALEUROS VIRESCENS (CHROOLEPIDACEAE; CHLOROPHYTA). II. GAMETES

Transmission electron microscopic examination of Cephaleuros virescens Kunze growing on leaves of Camellia sp. indicates that gametes are similar to those of Trentepohlia aurea. The gametes bear two, smooth isokont “keeled” flagella containing typical “9 + 2” axonemes and lacking scales. Flagellar insertion is apical and the parallel basal bodies overlap laterally. Each basal body is associated with a separate multilayered structure and component microtubular spline. The latter extends posteriorly beneath the plasmalemma. A nucleus, mitochondria, chloroplasts, and cytoplasmic haematochrome droplets are present. Pyrenoids and eyespots are absent. The subcellular components of C. virescens gametes are comparable to those found in gametes of T. aurea; however, the arrangement of basal bodies and multilayered structures differs slightly from that in T. aurea. Comparison of the fine structure of gametes from Cephaleuros, Phycopeltis, and Trentepohlia clearly indicates that the (1) mode of flagellar insertion, (2) morphology, number, and arrangement of multilayered structures, and (3) keeled flagella are common to these three genera and, thus far, unique among the green algae. Although flagellar insertion is apical, it is not bilaterally symmetrical (sensu stricto), nor is it asymmetrical (cf. Chara and Nitella sperms). The arrangement may be termed “reversed bilateral symmetry” and standardization of the terminology is recommended.     

An Analysis of the Formation of Ciliary Primordia in the Hypotrich Ciliate Urostyla weissei*

SYNOPSIS. The protargol technic was used in a study of the development of oral, cirral, and dorsal primordia of Urostyla weissei fixed during division, reorganization, and regeneration following transection at different levels. While the course of development is similar in all situations, differences were observed in the way in which some primordia are initiaily formed. The primordium of the new AZM always appears posterior to the old AZM. It develops into an entire new membranellar band in dividing cells and in opimers (posterior fragments from equatorial transections), while it eventually joins with a portion of the old AZM in reorganizers, promers (anterior fragments from equatorial transections) and “large opimers” (cells whose anterior tip has been cut off). The UM-primordium of proters is derived from disaggregation of the kinetosomes of the 2 old UM's, that of opisthes and opimers is formed “de novo” to the right of the AZM-primordium, while the UM-primordium of reorganizers, promers, and “large opimers” is of composite origin, partly “de novo” and partly from the old UM's. The UM primordium differentiates into the new UM's and the 1st frontal cirrus. The primordia of the remaining frontal, ventral, transversal (F-V-T) and marginal cirri originate as “streaks” of cilia, most of which are derived from re-alignment of the constituent cilia of certain pre-existing cirri. New cirri differendiate from the streaks, and replace the remaining old cirri. The streaks are formed similarly in all developmental situations, except for the 1st 3 F-V-T streaks. In proters, reorganizers, and promers, these originate from the posterior 3 frontal cirri, while in opisthes and opimers they are formed “de novo” to the right of the UM-primordium. In the “large opimers” these streaks are formed “de novo” behind the 1st 3 frontal cirri, in spite of the continued presence of these cirri at the anterior tip of the fragments. The site of formation of these streaks thus appears to be determined by an anteriorposterior gradient, rather than by any preformed cortical structure. The new dorsal bristle rows I to III develop from the proliferation of portions of the old rows, while rows IV and V originate from short kineties formed “de novo” on the right margin. New caudal cirri differentiate at the posterior ends of the new rows I to III. The numbers of ventral cirral rows and transversal cirri are variable; these variations are correlated, and related to variations in numbers of developing streaks. A survey of hypotrich developmental patterns revealed extensive parallels, especially in the sites of appearance of primordia. The primordium site appears to be a more constant feature of cortical development than is the “source” of ciliary units. It is concluded that sites of primordia are determined by cellular gradients, with competent preformed structures being utilized if they are appropriately positioned within these gradients.