Sponge cell culture: a comparative evaluation of adhesion to a native tissue extract and other culture substrates

Hexactinellids are deep water sponges that possess syncytial rather than cellular tissues. In order to investigate the syncytial character of the tissue of these unusual sponges, primary cultures were developed using a substrate of acellular tissue extract (ATE) that promotes the adhesion and spreading of sponge tissues. Primary cultures of the hexactinellid sponge Rhabdocalyptus dawsoni, grown on this substrate, form thinly spread, multinucleate, confluent tissue masses which exhibit active cytoplasmic streaming. Sponge tissue adhered equally well to commercial substrates of concanavalin A and poly-l-lysine, but did not adhere to chicken collagen. Acellular tissue extracts prepared from demosponges, which are known to be cellular, also promoted adhesion and spreading of cells from those sponges. Scanning electron microscopy showed adherent Rhabdocalyptus tissue to have an uninterrupted, smooth membrane covering the entire culture, unlike primary cultures of the cellular demosponge, Haliclona sp., which consisted of numerous individual cells. Tissue from freshly collected sponges adhered preferentially to ATE from a conspecific. However, after continued wounding, tissue adhered indescriminately to any substrate. The tissue extract congealed if added to sea water or 10 mM CaCl(2), forming a white, cloudy solid, which could be fixed and sectioned for transmission electron microscopy. Thin sections of the congealed extract showed it to contain membranes but no visible collagen fibrils.     

Ultrastructural changes in the placenta of the ewe after fetal pituitary stalk section.

Binucleate cells are a normal component of the ovine chorionic epithelium, but are usually separated from the fetal-maternal interface by a thin layer of cytoplasm derived from the principal or uni-nucleate cells of the trophoblast. They are distinguished not only by two distinct and separate nuclei, but also by conspicuous membrane-bound cytoplasmic inclusions in the form of haloed droplets. After fetal pituitary stalk section binucleate cells move up to and participate in the formation of the fetal-maternal interface; furthermore they extend clear blunt-ended pseudopodia into the maternal epithelial syncytium. These activities do not appear to be supppressed by fetal infusion of cortisol or ACTH. The apparent motility of binucleate cells, together with the presence of haloed droplets within the maternal epithelial syncytium, suggests that after fetal pituitary stalk section binucleate cells invade the uterine syncytium, lose their limiting membranes and discharge their contents into the syncytial cytoplasm. Large molecules such as ovine placental lactogen may be transported from fetal to maternal tissues by this mechanism.     

Embryogenesis in the glass sponge Oopsacas minuta: Formation of syncytia by fusion of blastomeres

Sponges (Porifera) are unusual animals whose body plans makeinterpreting phylogenetic relationships within the group andwith other basal metazoan taxa a difficult task. Although molecularapproaches have offered new insights, some questions requirea morphological approach using detailed ultrastructural or lightmicroscopical studies of developing embryos and larvae. Glasssponges (Hexactinellida) have perhaps the most unusual bodyplan within the Metazoa because the majority of the tissue ofthe adult consists of a single giant multinucleated syncytiumthat forms the inner and outer layers of the sponge and is joinedby cytoplasmic bridges to uninucleate cellular regions. Herewe have used serial section transmission and high-resolutionscanning electron microscopy to examine when syncytia firstform in the cave-dwelling glass sponge Oopsacas minuta. We confirmthat in O. minuta blastomeres are separate until the 32-cellstage; cleavage is equal but asynchronous until a hollow blastulais formed. The sixth division yields a collection of variouslysized micromeres at the surface of the embryo and large yolk-and lipid-filled macromeres lining the blastocoel. Syncytiathen form by the fusion of micromeres to form cytoplasmic bridgeswith each other and the fusion of macromeres to form the futuremultinucleated trabecular tissue of the larva and adult sponge.The multinucleated trabecular tissue envelops and forms cytoplasmicbridges with all uninucleate cells, covering the developinglarva with a continuous syncytial epithelium. Differentiationof tissues occurs very early during embryogenesis with the separationof uninucleate and multinucleate lineages, but all cells andsyncytia are joined by cytoplasmic bridges such that there iscytoplasmic continuity throughout the entire larva. Althoughglass sponges begin life as a cellular embryo, the unusual mechanismof syncytia formation at such an early stage in developmentdistinguishes this group of animals from their closest multicellularrelatives, the Demospongiae. Most important, however, thesedata lend support to the hypothesis that the original metazoanswere cellular, not syncytial.     

Drosophila embryo syncytial blastoderm cellular architecture and morphogen gradient dynamics: Is there a correlation?

During embryo development in many metazoan animals, the first differentiated cell type to form is an epithelial cell. This epithelial layer is modified by developmental cues of body axes formation to give rise to various tissues. The cells that arise are mesenchymal in nature and are a source of other tissue types. This epithelial to mesenchymal transition is used for tissue type formation and also seen in diseases such as cancer. Here we discuss recent findings on the cellular architecture formation in the Drosophila embryo and how it affects the developmental program of body axes formation. In particular these studies suggest the presence of compartments around each nucleus in a common syncytium. Despite the absence of plasma membrane boundaries, each nucleus not only has its own endoplasmic reticulum and Golgi complex but also its own compartmentalized plasma membrane domain above it. This architecture is potentially essential for morphogen gradient restriction in the syncytial Drosophila embryo. We discuss various properties of the dorso-ventral and the antero-posterior morphogen gradients in the Drosophila syncytium, which are likely to depend on the syncytial architecture of the embryo.     

An ultrastructural study of alimentary tract development in the cercariae of Prosorhynchoides borealis (Digenea,Bucephalidae)

Podvyaznaya I. 2011. An ultrastructural study of alimentary tract development in the cercariae of Prosorhynchoides borealis (Digenea, Bucephalidae). —Acta Zoologica (Stockholm) 92 : 170–178. The development of digestive system in Prosorhynchoides borealis cercariae was studied using transmission electron microscopy. The foregut and caecum primordia arise in early cercarial embryos as two adjoining cellular cords. The primordial pharynx appears as a cluster of myoblasts in the mid‐part of the foregut primordium whose proximal end abuts onto the ventral embryonic tegument. Later, a lumen develops within the gut primordia and their component cells form the embryonic cellular epithelium with an essentially similar structure in the foregut and caecal regions. Subsequently, the foregut epithelial cells merge to form a syncytium. This process proceeds asynchronously and the most proximal foregut area remains cellular for the longest time. The syncytial lining of the foregut establishes syncytial connections with secretory cytons differentiating in the surrounding parenchyma. These cytons produce secretory granules, which are transported through cytoplasmic connections to the foregut syncytium. Before cercariae reach maturity, their foregut epithelium becomes anucleate and continuous with the external tegument. By the end of cercarial development, numerous short lamellae appear on the luminal surface of the caecal epithelium. The caecal cells become involved in secretory activity as indicated by the presence of Golgi‐derived secretory bodies in their cytoplasm.     

Development of the implantation chamber in the pregnant bitch.

Uteri taken from 25 bitches at various times during the early stages of pregnancy were studies cytologically to determine how the implantation chamber developed and how fetal-maternal relations were established. On day 13 after the end of estrus, knobs of trophoblastic syncytium formed and became wedged between cells of the uterine luminal epithelium. The syncytium quickly spread along the uterine lumen and into the mouths of the glands, dislodging and surrounding maternal cells. As invasion continued trophoblastic villi, consisting of cores of cytotrophoblast covered by a continuous layer of syncytium, penetrated deeper into the endometrium. The syncytium spread to surround maternal vessels and decidual cells. By day 26 the trophoblast had extended down to the large lacunae. Here syncytial trophoblast covering tips of the villi degenerated, leaving cytotrophoblast exposed to the necrotic zone. These cells possessed characteristics of absorbing cells. Hematomas were formed by focal necrosis of fetal and endometrial tissue at the poles of the implantation sites. Large pools of extravasated blood accumulated and red blood cells were phagocytized by surrounding trophoblastic cells. Therefore, the endotheliochorial relationship in the canine placenta appeared to be established by syncytial trophoblast invading a cellular endometrium. In the necrotic zone and hematomas, cellular trophoblast may have lost its syncytial covering, but elsewhere maternal vessels and decidual cells in the placenta were in direct contact only with syncytial trophoblast.     

Mutations in the cytoplasmic tail of herpes simplex virus glycoprotein H suppress cell fusion by a syncytial strain.

We have developed a complementation assay, using transiently transfected COS cells, to facilitate a molecular analysis of the herpes simplex virus type 1 glycoprotein gH. When infected by a gH-null syncytial virus, COS cells expressing wild-type gH generate infectious progeny virions and form a syncytium with neighboring cells. By deletion and point mutagenesis, we have found particular residues in the gH cytoplasmic tail to be essential for generation of a syncytium but apparently dispensable for production of infectious virions. This study emphasizes the different requirements for cell-cell and cell-envelope fusion and demonstrates that changes in the non-syn locus UL22-gH can reverse the syncytial phenotype.     

Ultrastructure and cytochemistry of the tegument of Atriaster heterodus (Platyhelminthes: Monogenea)

The tegument of the polyopisthocotylean monogenean Atriaster heterodus Lebedev & Parukhin, 1969 was studied using transmission electron microscopy. The outer syncytial layer of the tegument is connected to the internal cell bodies by cytoplasmic extensions which interweave between the muscular fibres. The free surface of the syncytium has projections of the external membrane which are similar to microvilli. The undulating basal membrane, with numerous narrow elongate projections, is associated with the basal lamina situated between the syncytial and muscular layers. The cell bodies and syncytial layer of the tegument exhibit two types of vesicles, one with fibrous contents and one with electron-dense contents; these were analysed using two cytochemical tests, the E-PTA and alcian blue methods, used for the first time on monogeneans.     

The proteome of the developing tooth of the sea urchin, Lytechinus variegatus: mortalin is a constituent of the developing cell syncytium

Echinoderm teeth are continuously growing calcite-mineralized tissues of complex structure. Two features are of special interest: (1) cell division takes place in a restricted aboral domain, the plumula, and the cells immediately merge into multinucleated syncytial layers; (2) the major part of the heavily mineralized tooth elongates and moves towards the adoral incisal tip continuously as the syncytial cells actively expand the syncytium and intermembrane mineral phase. As the first step to understanding the nature of the mineralization processes, we have isolated the proteins of the plumula and of the mature mineralized portions of the tooth, and begun their characterization. Peptide sequences were used to screen a plumula cDNA library by polymerase chain reaction. One primer set yielded a prominent amplified product which was cloned, and sequenced. Comparison with the nucleotide and protein data banks revealed the protein to be Mortalin, a member of the hsp-70 family, with >75% of its sequences identical to that of human mortalin. Immunocytochemical localization of mortalin within the plumula, using Anti-human Grp75, showed staining of the odontoblast cytosol and matrix at the point where syncytial formation was occurring. The cytosol of the syncytial layers was weakly stained. The nuclei within the syncytia were stained at their periphery. In the mature part of the tooth, the perinuclear staining of the nuclei was more prominent. We conclude that mortalin is involved in syncytium formation and maintenance. The urchin mortalin has a distinctive aspartic acid and serine-rich C-terminal domain that may link it to the mineralization process.     

The fine structure of the chinchilla placenta.

The structure of the placental labyrinth, interlobular or "coarse" syncytium, visceral (splanchnopleuric) yolk sac, giant cells and subplacenta of the chinchilla was studied with the electron microscope. The fine structure of the interhemal membrane of the placental labyrinth was found to be hemomonochorial, consisting of a single layer of syncytial trophoblast. In this respect, the placental labyrinth was similar to that of another caviomorph rodent, the guinea pig. The labyrinthine trophoblast had pinocytotic vesicles as well as larger vaculoes and multivesicular bodies. The interlobular syncytium contained granular endoplasmic reticulum, and in one case from early in gestation there were intracisternal granules in the ER. The visceral endodermal cells of the inverted yolk sac placenta had a well-developed system of apical vesicles and tubules as well as larger cytoplasmic vacuoles. Their appearance was similar to that of endodermal cells found in other rodents which are known to absorb proteins and other substances from the uterine lumen. Towards term the giant cells were often vacuolated and contained large deposits of glycogen as well as lipid droplets. The syncytial trophoblast of the subplacenta contained numerous moderately electron-dense granules which may be secretory in function; cytotrophoblastic cells lacked these granules. The subplacental syncytium often surrounded spaces or lacunae which contained an electron-dense granular material.     

Formation of the transverse nerve in moth embryos. II. Stereotyped growth by the axons of identified neuroendocrine neurons

We are interested in the cellular mechanisms that guide neuroendocrine axons to their neurohaemal target regions and that regulate the extent and positioning of their terminal arbor. The neurohaemal organ we have studied is the segmentally repeated transverse nerve of the moth Manduca. In the mature animal, two motor neurons and a heterogeneous set of identified neuroendocrine neurons project to this nerve; the latter release hormonal peptides from along its length. In the preceding report, we demonstrated that during embryogenesis, the position, trajectory and extent of the transverse nerve are anticipated by two sets of nonneuronal cells, the strap and the bridge. In this paper we show that four identified neuroendocrine neurons (L1 and B1-3), like the identified motor neurons before them, elaborate growth cones that use this preexisting scaffolding as a substrate for axonal elongation. Moreover, growth cone navigation by these neuroendocrine neurons is as precise and invariant as that displayed by the motor neurons. One feature that differentiates the behavior of the developing neuroendocrine cells from that of the motor neurons is a stereotyped interaction that the L1 and B1-3 axons undergo with an identified syncytial cell that lies in close proximity to the strap. Each neuroendocrine neuron specifically adheres to the syncytium by extending numerous filopodia, and an occasional large lamellopodium, over its surface. These contacts are maintained by the neuroendocrine axons after their growth cones have left the vicinity of the syncytium and proceeded into the strap/bridge complex. Adhesion to the syncytium is transient and specific to the neuroendocrine neurons: although motor neuron axons are present at this same time and place, they display no affinity for the syncytium. This distinction correlates with the fact that the neuroendocrine neurons go on to elaborate arbor within the confines of the transverse nerve, while the motor neurons do not. We suggest that the syncytium may act as a "fictive target" for these neurons to aid in the differentiation of features that are specific to their cellular phenotype.     

Three‐dimensional fate mapping of larval tissues through metamorphosis in the glass sponge Oopsacas minuta

The tissue of glass sponges (Class Hexactinellida) is unique among metazoans in being largely syncytial, a state that arises during early embryogenesis when blastomeres fuse. In addition, hexactinellids are one of only two poriferan groups that already have clearly formed flagellated chambers as larvae. The fate of the larval chambers and of other tissues during metamorphosis is unknown. One species of hexactinellid, Oopsacas minuta, is found in submarine caves in the Mediterranean and is reproductive year round, which facilitates developmental studies; however, describing metamorphosis has been a challenge because the syncytial nature of the tissue makes it difficult to trace the fates using conventional cell tracking markers. We used three‐dimensional models to map the fate of larval tissues of O. minuta through metamorphosis and provide the first detailed account of larval tissue reorganization at metamorphosis of a glass sponge larva. Larvae settle on their anterior swimming pole or on one side. The multiciliated cells that formed a belt around the larva are discarded during the first stage of metamorphosis. We found that larval flagellated chambers are retained throughout metamorphosis and become the kernels of the first pumping chambers of the juvenile sponge. As larvae of O. minuta settle, larval chambers are enlarged by syncytial tissues containing yolk inclusions. Lipid inclusions at the basal attachment site gradually became smaller during the six weeks of our study. In O. minuta, the flagellated chambers that differentiate in the larva become the post‐metamorphic flagellated chambers, which corroborate the view that internalization of these chambers during embryogenesis is a process that resembles gastrulation processes in other animals.     

Pre-gastrula expression of zebrafish extraembryonic genes

Background  

Many species form extraembryonic tissues during embryogenesis, such as the placenta of humans and other viviparous mammals. Extraembryonic tissues have various roles in protecting, nourishing and patterning embryos. Prior to gastrulation in zebrafish, the yolk syncytial layer - an extraembryonic nuclear syncytium - produces signals that induce mesoderm and endoderm formation. Mesoderm and endoderm precursor cells are situated in the embryonic margin, an external ring of cells along the embryo-yolk interface. The yolk syncytial layer initially forms below the margin, in a domain called the external yolk syncytial layer (E-YSL).     

The Occurrence of Intercellular Bridges in Groups of Cells Exhibiting Synchronous Differentiation

A previous electron microscopic study of the cat testis revealed that spermatids derived from the same spermatogonium are joined together by intercellular bridges. The present paper records the observation of similar connections between spermatocytes and between spermatids in Hydra, fruit-fly, opossum, pigeon, rat, hamster, guinea pig, rabbit, monkey, and man. In view of these findings, it is considered likely that a syncytial relationship within groups of developing male germ cells is of general occurrence and is probably responsible for their synchronous differentiation. When clusters of spermatids, freshly isolated from the germinal epithelium are observed by phase contrast microscopy, the constrictions between the cellular units of the syncytium disappear and the whole group coalesces into a spherical multinucleate mass. The significance of this observation in relation to the occurrence of abnormal spermatozoa in semen and the prevalence of multinucleate giant cells in pathological testes is discussed. In the ectoderm of Hydra, the clusters of cnidoblasts that arise from proliferation of interstitial cells are also connected by intercellular bridges. The development of nematocysts within these groups of conjoined cells is precisely synchronized. Both in the testis of vertebrates and the ectoderm of Hydra, a syncytium results from incomplete cytokinesis in the proliferation of relatively undifferentiated cells. The intercellular bridges between daughter cells are formed when the cleavage furrow encounters the spindle remnant and is arrested by it. The subsequent dissolution of the spindle filaments establishes free communication between the cells. The discovery of intercellular bridges in the two unrelated tissues discussed here suggests that a similar syncytial relationship may be found elsewhere in nature where groups of cells of common origin differentiate synchronously.     

The occurrence of intercellular bridges in groups of cells exhibiting synchronous differentiation

A previous electron microscopic study of the cat testis revealed that spermatids derived from the same spermatogonium are joined together by intercellular bridges. The present paper records the observation of similar connections between spermatocytes and between spermatids in Hydra, fruit-fly, opossum, pigeon, rat, hamster, guinea pig, rabbit, monkey, and man. In view of these findings, it is considered likely that a syncytial relationship within groups of developing male germ cells is of general occurrence and is probably responsible for their synchronous differentiation. When clusters of spermatids, freshly isolated from the germinal epithelium are observed by phase contrast microscopy, the constrictions between the cellular units of the syncytium disappear and the whole group coalesces into a spherical multinucleate mass. The significance of this observation in relation to the occurrence of abnormal spermatozoa in semen and the prevalence of multinucleate giant cells in pathological testes is discussed. In the ectoderm of Hydra, the clusters of cnidoblasts that arise from proliferation of interstitial cells are also connected by intercellular bridges. The development of nematocysts within these groups of conjoined cells is precisely synchronized. Both in the testis of vertebrates and the ectoderm of Hydra, a syncytium results from incomplete cytokinesis in the proliferation of relatively undifferentiated cells. The intercellular bridges between daughter cells are formed when the cleavage furrow encounters the spindle remnant and is arrested by it. The subsequent dissolution of the spindle filaments establishes free communication between the cells. The discovery of intercellular bridges in the two unrelated tissues discussed here suggests that a similar syncytial relationship may be found elsewhere in nature where groups of cells of common origin differentiate synchronously.     

The embryonic development of the triclad <Emphasis Type="Italic">Schmidtea polychroa</Emphasis>

Triclad flatworms are well studied for their regenerative properties, yet little is known about their embryonic development. We here describe the embryonic development of the triclad Schmidtea polychroa, using histological and immunocytochemical analysis of whole-mount preparations and sections. During early cleavage (stage 1), yolk cells fuse and enclose the zygote into a syncytium. The zygote divides into blastomeres that dissociate and migrate into the syncytium. During stage 2, a subset of blastomeres differentiate into a transient embryonic epidermis that surrounds the yolk syncytium, and an embryonic pharynx. Other blastomeres divide as a scattered population of cells in the syncytium. During stage 3, the embryonic pharynx imbibes external yolk cells and a gastric cavity is formed in the center of the syncytium. The syncytial yolk and the blastomeres contained within it are compressed into a thin peripheral rind. From a location close to the embryonic pharynx, which defines the posterior pole, bilaterally symmetric ventral nerve cord pioneers extend forward. Stage 4 is characterized by massive proliferation of embryonic cells. Large yolk-filled cells lining the syncytium form the gastrodermis. During stage 5 the external syncytial yolk mantle is resorbed and the embryonic cells contained within differentiate into an irregular scaffold of muscle and nerve cells. Epidermal cells differentiate and replace the transient embryonic epidermis. Through stages 6–8, the embryo adopts its worm-like shape, and loosely scattered populations of differentiating cells consolidate into structurally defined organs. Our analysis reveals a picture of S. polychroa embryogenesis that resembles the morphogenetic events underlying regeneration.Edited by D. Tautz     

Ultrastructure of somatic tissues in the ovaries of a chaetognath (Ferosagitta hispida)

Transmission electron microscopy reveals that the ovaries of Ferosagitta hispida contain four somatic tissues. A myoepithelial ovary wall, continuous with a thin layer of peritoneocytes lining the coelomic cavity, encloses a fluid-filled ovarian space in which oocytes develop. Lamellar extensions of a “follicular reticulum” branch throughout the ovarian space and ensheath developing oocytes. This tissue has been overlooked in most previous studies of chaetognath ovaries. A bipartite oviductal complex extends the length of each ovary just within the lateral ovary wall. It consists of a flattened, blindly ending cellular tube, herein referred to as the cellular sheath, and an enclosed syncytium. Sheath cells secrete an electron-dense product into the ovarian space. Those sheath cells directly bordering the syncytium are contractile and are joined to the to the syncytium by gap junctions and microvillar interdigitations. The syncytium contains a complex of membrane-bounded lumina. The latter sometimes enclose sperm received during mating or ovulated eggs. Thus the syncytium serves both as a seminal receptacle and as a duct for passage of eggs to the outside. Contrary to several classical reports, the cellular sheath and syncytium of the oviductal complex do not separate at ovulation to form a temporary oviductal lumen.     

Effects of serotonin (5HT) and complement C3 on the synthesis of the surface membrane precursors of adult Schistosoma mansoni

The syncytial surface epithelium of Schistosoma mansoni plays an important role in immune evasion. This syncytium is covered by an unusual double-membrane complex consisting of an apical plasma membrane (APM) and an overlying envelope (En) that have been shown to have different rates of synthesis and turnover. It has been suggested that discoid bodies (DBs) and multilamellar bodies (MLBs), the major syncytial inclusion bodies of schistosomes, may be the precursors of the APM and En, respectively. In this ultrastructural study, we examined the effects of serotonin (5HT) and complement C3, which have been shown to stimulate synthesis and turnover of the APM and En, respectively, on the synthesis of DBs and MLBs in vitro. With short-time incubations (20 or 40 min), 5HT stimulated the synthesis of the DBs by 2-fold, whereas C3 accelerated synthesis of the MLBs by 2-fold. Furthermore, when microtubules within the cytoplasmic connections between the syncytium and the underlying cell bodies (the site of membrane synthesis) were disrupted with colchicine, the DBs and MLBs synthesized in response to 5HT or C3 accumulated in the cell bodies. This suggests that the transport of the organelles to the syncytium is dependent upon the microtubules but not the signaling mechanism in response to 5HT or C3. These observations also support the suggestion that the DBs and MLBs are synthesized in subsyncytial cell bodies and serve as precursors of the APM and En, respectively. The rapid synthetic response to 5HT and C3 is also consistent with rapid synthesis and turnover of the APM/En, as suggested by previous studies.     

ON THE ROLE OF MICROTUBULES IN MOVEMENT AND ALIGNMENT OF NUCLEI IN VIRUS-INDUCED SYNCYTIA

Infection of baby hamster kidney (BHK21-F) cells with the parainfluenza virus SV5 causes rapid and extensive cell fusion. Time-lapse cinematography shows that when cells fuse, their nuclei migrate straight to the center of the syncytium at rates of 1–2 µ/min. Nuclei are often arranged in long, tightly packed, parallel rows in syncytia derived from the fibroblastic BHK21-F cells. Polarization microscopy shows birefringent material between and parallel to these rows of nuclei, and electron microscopy shows bundles of cytoplasmic microtubules, ~250 A in diameter, and filaments, ~80 A in diameter, parallel to and between the rows of nuclei. Colchicine treatment causes disappearance of microtubules from BHK21-F cells and an apparent increase in the number of 80-A filaments. Although colchicine-treated, SV5-infected cells fuse, their nuclei do not migrate or form rows but remain randomly scattered through the syncytial cytoplasm. Incubation at 4°C does not disrupt microtubules in BHK21-F cells. Rows of nuclei have been isolated from SV5-induced syncytia, and the nuclei in them have been found to be intimately associated with microtubules but not with other cytoplasmic structures. These results suggest that microtubules demarcate cytoplasmic channels through which nuclei migrate and that they may also be involved in the mechanism of nuclear movement.