A two-step mechanism generates the spacing pattern of the ciliated cells in the skin of Xenopus embryos.

The skin of Xenopus embryos contains a population of specialized ciliated cells that are distributed in an evenly spaced pattern. Here we describe two successive steps that govern the differentiation and the generation of the spacing pattern of these ciliated cells. The first step occurs in the inner or sensorial layer of the non-neural ectoderm where a subset of cells are chosen to differentiate into ciliated-cell precursors. This choice is under the control of lateral inhibition mediated by a Suppressor of Hairless-dependent Notch signaling pathway, in which X-Delta-1 is the putative ligand driving the selection process, and a new Enhancer-of-Split-related gene is an epidermal target of Notch signaling. Because nascent ciliated-cell precursors prevent neighboring cells from taking on the same fate, a scattered pattern of these precursors is generated within the deep layer of the non-neural ectoderm. Ciliated-cell precursors then intercalate into the outer layer of cells in the epidermis. We show that the intercalation event acts as a second step to regulate the spacing of the mature ciliated cells. We propose that the differentiation of the ciliated cells is not only regulated by Notch-mediated lateral inhibition, but is also an example where differentiation is coupled to the movement of cells from one cell layer to another.     

Radial intercalation is regulated by the Par complex and the microtubule-stabilizing protein CLAMP/Spef1

The directed movement of cells is critical for numerous developmental and disease processes. A developmentally reiterated form of migration is radial intercalation; the process by which cells move in a direction orthogonal to the plane of the tissue from an inner layer to an outer layer. We use the radial intercalation of cells into the skin of Xenopus laevis embryos as a model to study directed cell migration within an epithelial tissue. We identify a novel function for both the microtubule-binding protein CLAMP and members of the microtubule-regulating Par complex during intercalation. Specifically, we show that Par3 and aPKC promote the apical positioning of centrioles, whereas CLAMP stabilizes microtubules along the axis of migration. We propose a model in which the Par complex defines the orientation of apical migration during intercalation and in which subcellular localization of CLAMP promotes the establishment of an axis of microtubule stability required for the active migration of cells into the outer epithelium.     

Electron microscope study of the binding of Con A-gold to superficial and inner ectoderm layers ofXenopus laevis and its relation to the neural-inducing activity of this lectin

Summary Isolated competent amphibian ectoderm differentiates into neural (archencephalic) structures when treated with the plant lectin concanavalin A (Con A). While the inner ectoderm layer ofXenopus laevis forms brain structures after incubation with Con A, the outer ectoderm layer differentiates into ciliated epidermis only. This difference can be correlated with the pattern of Con A bound to the plasma membrane. With gold-labelled Con A it could be shown by transmission electron microscopy (TEM) that the outer ectoderm binds substantially less lectin than the inner layer. Furthermore we observed characteristic differences at the apical and basal surfaces of the cells of the same layer, i.e. on the apical cell surface of the superficial layer almost no Con A-gold could be found. In contrast, we observed a lot of gold particles on the basal cell side of the superficial layer. However, the number on both surfaces (apical and basal side of the cell) of the inner ectoderm layer was essentially higher, which could explain its biological reaction to the Con A stimulus and the differentiation into neural structures. The data presented in this paper indicate that early and late gastrula ectoderm bind similar amounts of Con A and support the view that the decrease in competence is not correlated with a loss of receptors for inducing factors. Furthermore, we describe the binding and the internalization of Con A via receptor-mediated endocytosis and the further fate of the Con A-gold-receptor complex inside the target cell.     

Cell rearrangement during gastrulation of Xenopus: direct observation of cultured explants.

We have analyzed cell behavior in the organizer region of the Xenopus laevis gastrula by making high resolution time-lapse recordings of cultured explants. The dorsal marginal zone, comprising among other tissues prospective notochord and somitic mesoderm, was cut from early gastrulae and cultured in a way that permits high resolution microscopy of the deep mesodermal cells, whose organized intercalation produces the dramatic movements of convergent extension. At first, the explants extend without much convergence. This initial expansion results from rapid radial intercalation, or exchange of cells between layers. During the second half of gastrulation, the explants begin to converge strongly toward the midline while continuing to extend vigorously. This second phase of extension is driven by mediolateral cell intercalation, the rearrangement of cells within each layer to lengthen and narrow the array. Toward the end of gastrulation, fissures separate the central notochord from the somitic mesoderm on each side, and cells in both tissues elongate mediolaterally as they intercalate. A detailed analysis of the spatial and temporal pattern of these behaviors shows that both radial and mediolateral intercalation begin first in anterior tissue, demonstrating that the anterior-posterior timing gradient so evident in the mesoderm of the neurula is already forming in the gastrula. Finally, time-lapse recordings of intact embryos reveal that radial intercalation takes places primarily before involution, while mediolateral intercalation begins as the mesoderm goes around the lip. We discuss the significance of these findings to our understanding of both the mechanics of gastrulation and the patterning of the dorsal axis.     

Presence of a Ciliary Patch in Preoral Epithelium of Sea Urchin Plutei

Removal of the hyaline layer from sea urchin embryos at the pluteus stage discloses a densely ciliated region in the preoral area of the ectodermal epithelium. In four-armed plutei, this ciliary path is located between the anterolateral arms and in eight-armed plutei it becomes surrounded by preoral and anterolateral arms. The area of the patch and the number of cilia increase with age. This patch is covered by cilia of unusual morphology and orientation. There are more than two cilia per cell which are coiled together several times around a small cone at the apical end of the cell. These coiled cilia run parallel to the surface of the cell but do not extend beyond the hyaline layer. The ciliary axoneme consists of a "9+2" microtubular structure, but no outer or inner dynein arms are observed. Although the cells with coiled cilia are present in a cluster constituting a part of the epithelium, they have axons that project from their basal (inner) ends. The structural characteistics of the ciliary patch suggest that it possesses a sensory function.     

The fine structure of the planarianPolycelis tenuis ijima

Summary The fine structure of the pharynx is presented and demonstrates that the pharyngeal epithelial system is a continuous one. The epithelial lining of the pharyngeal cavity with its characteristic fibrous secretory bodies merges with the outer pharyngeal epithelium at the point of anchorage of the pharynx. A few of these cells are insunk, the nuclei occurring beneath the underlying muscular layers. The nature of the outer epithelium changes towards the free end of the pharynx; the cells become ciliated and in contents come to resemble the inner epithelium which it joins at the tip.The gut cells merge at a transitional zone with the inner pharyngeal epithelium and at this point both bear microvilli and contain rod-shaped apical bodies. Some of these cells are also insunk. Towards the mouth the epithelium shows a greater degree of insinking and exhibits microapocrine secretion. Both inner and outer epithelia bear sense receptors which are concentrated at the lip.At the point of pharyngeal insertion, the sub-epithelial tissue resembles planarian parenchyma, but is rich in gland cells. These glands open on to the outer epithelium especially towards the free end of the pharynx.This research was supported by the Scientific Research Council. Grant No. B/RG/086.     

长豇豆种皮和种脐的发育

长豇豆的胚珠具内外两层珠被,内珠被在种子发育早期退化消失,种皮仅由外珠被发育而成。外珠被的外表皮细胞径向伸长,外壁和经向壁增厚,形成约占成熟种皮厚度一半的栅栏层;亚表皮细胞发育为骨状石细胞层。第三层细胞类似于亚表皮层但细胞壁增厚不明显,其内方的多层薄壁细胞形成海绵组织。种脐具两层栅栏细胞,外栅栏层及其以外部分由珠柄组织发育而成管胞群。本文还对脐缝和管胞群的作用以及豆科种子的吸水机制进行了讨论。     

Fine structure of the excretory system of the deep posterior (Ebner's) salivary glands of the human tongue

Human deep posterior lingual glands (von Ebner's glands) are located beneath the circumvallate papillae. They are formed by tubuloalveolar adenomeres, intercalated ducts and excretory ducts coming together in the main excretory duct. The tubuloalveolar cells, pyramid-shaped, show large and dense secretory granules (clear cored) throughout the cytoplasm, rare basal folds and packed cisternae of rough endoplasmic reticulum (RER) at the basal pole. The columnar cells of the intercalated ducts are arranged in a monolayer. They are characterized by dense, clear-core secretory granules (mostly in the apical cytoplasm), a basal nucleus, well-developed RER and Golgi apparatus, and thin filaments distributed in supra- and perinuclear cytoplasm. Striated ducts are absent. Excretory ducts, coming together in the main duct, are lined by a bistratified epithelium. The inner layer consists of columnar cells showing bundles of tonofilaments with scarce secretory activity. The outer layer is composed of basal cells lying on the basal lamina. The main excretory duct, which opens at the bottom of the vallum, shows a stratified epithelium. The outer side is composed of 2-3 layers of malpighian cells lying on the basal lamina. The inner side consists of a single layer of cuboidal-columnar cells with dense apical granules and well-developed organelles synthesizing and condensing secretions. These cells interpolate with goblet cells, rare mitochondria-rich cells, ciliated cells and numerous small globous cells showing a clear matrix and lacking secretory granules. The cilia show a 9 + 2 microtubular structure with basal bodies provided with striated rootlets. Myoepithelial cells surround with their processes the basal portions of the secretory cells and the intercalated ducts. The conclusions concern some comparative aspects and some hypothesis on the functional role of goblet cells, ciliated cells and epithelial cells lining the different ducts, also in relation to the final secretory product.     

Localization of actin and tubulin in developing and adult mammalian photoreceptors

Summary In order to define cytoskeletal domains of the mammalian photoreceptor, actin and tubulin were localized in adult retinae of mouse and human. For light-microscopic localization, actin was labeled using fluorescent phalloidin or monoclonal antibodies against actin, and tubulin was labeled using monoclonal antibodies against alpha- and beta-tubulin in an immunocytochemical method. Actin and tubulin were also localized by ultrastructural immunocytochemistry in the mouse. Filamentous actin was present in the retina at the outer limiting membrane and in synaptic terminals, especially of the cones, while globular actin was observed additionally in the inner segments. Müller cell cytoplasm and apical microvilli at the outer limiting membrane were also labeled for filamentous actin. Alpha- and beta-tubulin were evident throughout the photoreceptors, including the inner segments, but not in the synaptic terminals or at the outer limiting membrane. In the early postnatal retina of mouse, actin and tubulin were present at the ventricular surface. This pattern changed as photoreceptors fully elongated and as synaptogenesis occurred in the outer plexiform layer.     

Cyclic changes in the surface structure of the cervix from the ewe as revealed by scanning electron microscopy

Thirty parous ewes were divided into six groups and sacrificed on day 0 (first day of estrus), 1, 2, 10, 15 or 16 of the estrous cycle. The cervices were removed immediately and processed for examination with the scanning electron microscope. Observation of the tissues reveals that the surface of the cervix is highly convoluted, which results in the formation of numerous folds or crypts. Two forms of columnar epithelial cells, a ciliated and a non-ciliated cell with microvilli, line the luminal surface of the cerix in the day 10, luteal-phase ewes. However, on day 15, 2 days before estrus, the non-ciliated cells differentiate into two morphologically distinct types of secretory cells. One type forms when the apex of the non-ciliated cell dilates outward into the lumen of the cervix. Concurrent with apical enlargement, the microvilli are lost and the limiting cell membrane becomes smooth. The other type of cell is characterized by only a slight apical swelling. Consequently, remnants of microvilli along with secretory granules can be observed on the limiting membrane of this cell. Both cells release a particulate component, which is believed to be a precursor of mucus, into the lumen of the cerix. These particles undergo a series of morphological transformations to form a fibrillar layer, generally referred to as 'cervical mucus', that covers the epithelial surface at estrus. One to 2 days following the onset of estrus, the fibers become more closely assoicated with amorphous material that begins to coagulate, thereby revealing the underlying ciliated and non-ciliated cells that characterize the cervix of the luteal-phage ewe. The cyclical variation in secretory cells and factors that may influence that structural transformations which occur in mucus are discussed.     

Human retinal development: ultrastructure of the inner retinal layers

Studies of the developing human retina from 6.5 to 18 weeks' gestational age (16–156 mm) by light and electron microscopy are concerned with the morphogenesis of neuroblast cells, plexiform layers, and inner limiting membrane. The transient layer of Chievitz is formed posteriorly by 20 mm (7 weeks), inner plexiform by 48 mm (9 weeks), outer plexiform layer by 83 mm (12 weeks), identifiable cones by 83 mm, and rods by 120 mm (15 weeks). Mitotic activity continues posteriorly until 120 mm and was seen in inner layers of the retina until 103 mm (13 weeks). Outer neuroblastic differentiation is marked by diversification from a uniform cell population to one containing at least three cell types differing in their nuclear shape, chromatin pattern, and cytoplasmic characteristics. Differentiating ganglion cells accumulate polysomes, rough endoplasmic reticulum, Golgi complexes, microtubules, and dense bodies. Müller cell bodies in ganglion and inner nuclear layers extend processes between ganglion cells, and radial fibers, containing extensive smooth endoplasmic reticulum, to the vitreal surface. Synapses appear in the inner and outer plexiform layers by 83 mm (12 weeks), and by 120 mm (15 weeks) demonstrate a variety of conventional and ribbon forms similar to those found in the adult. Synaptogenesis therefore begins considerably before the development of photoreceptor outer segments.     

Fine Structure of the Female Intoshia variabili(Alexandrov & Sljusarev) (Mesozoa: Orthonectida)

The morphology and fine structure of female Intoshia variabili, new combination for Rhopalura variabiliAlexandrov & Sljusarev, 1992, were studied with transmission electron microscopy. The body surface is covered with a 3-layered cuticula, under which is a layer of ciliated + non-ciliated cells arranged in alternating rings around the body. Ciliated cells have lateral extensions that intercalate with the non-ciliated cells. The kinetosome of each cilium has two longitudinally oriented cross-striated rootlets. The outer surface of the ciliated cells is covered with small tubercles, and the cytoplasm of these cells contains granules, vacuoles, mitochondria, fibrillar structures and lamellary bodies. A band of dense fibrils passes through the upper part of each ring of cells, going from one cell junction to another, encircling the entire body. Between the layer of ciliated + non–ciliated cells and the oocytes, elongated contractile cells from 4–5 longitudinal columns and 1 ring, the latter at the level of ciliated rings 7–9. The contractile cells contain thick and thin longitudinally oriented fibrils. The oocytes contain a large nucleus, numerous mitochondria, electron–dense granules and 1–2 spherical structures. An anteriorly situated, ciliated goblet–like receptor, not described for any other orthonectids, consists of three closely apposed cells, the upper part of which contains densely packed cilia. The genital pore opens through a non–ciliated cell and is surrounded by several cells with granules.     

Gap junctions in the differentiated neural retinae of newly hatched chickens.

Gap junctions in the neural retinae of newly hatched chickens were examined in thin section and by freeze cleaving. Unusual gap junctions containing linear arrays of intramembrane particles are found between principal and accessory cones which form a double cone at the region of the outer limiting membrane. These unusual gap junctions are often continuous with macular aggregates of hexagonally packed intramembrane particles which are characteristic of a typical gap junction. Typical gap junctions are also found in both the outer and the inner plexiform layers and in the outer nuclear layer, but are not so abundant as in the outer limiting membrane region. The sizes of intramembrane particles and their centre-to-centre spacing within the macular aggregate of a gap junction in differentiated neural retinae are slightly larger than those in undifferentiated neural retinae. Tight junctions are not found in differentiated neural retinae.     

Functional and structural differences between cultured outer and inner layer cells of bovine adrenal cortex

Summary Outer and inner layer cells of bovine adrenal cortex were cultured separately to compare cellular structural characteristics and functional differences. Outer layer cells were polygonal in shape with radially distributed lipid droplets in the cytoplasm, and produced mainly aldosterone and cortisol. The aldosterone production increased upon stimulation with angiotensin II or dibutyryl-cAMP. In contrast, inner layer cells were spindle-shaped and had fine diffused lipid droplets. They produced four times as much cortisol as outer layer cells but no aldosterone. Cortisol production increased with ACTH or dibutyryl-cAMP stimulation.When stimulated by ACTH or by dibutyryl-cAMP, both types of adrenocortical cells showed cellular retraction whereby the number of cytoplasmic lipid droplets decreased and microvilli on the cellular surface increased. At the same time, the transverse distribution of actin fibers disappeared and the microtubules changed their distribution pattern from circular to radial. Stimulation by angiotensin II, on the other hand, brought no marked structural changes.These results indicate that, in functional terms, the outer layer cells and the inner layer cells in this culture system reflect zona glomerulosa and zona fasciculata-reticularis, respectively.     

The rat pituitary cleft: A correlated study by scanning and transmission electron microscopy

Summary SEM reveals that the inner surface of the pituitary cleft is lined by a continuous layer of marginal cells possessing microvillous and ciliated apical surfaces. The ciliated cells are more numerous on the posterior side (toward the pars intermedia) than on the anterior side of the cleft (toward the pars distalis). In contrast small infoldings (crypts) were occasionally noted only on the marginal layer covering the distal part of the hypophysis. In some areas of the cleft the surface features of the marginal cells are rather similar to the epithelial cells populating the upper parts of the respiratory tract in their topography and distribution. In other regions they also show striking similarities with the ependymal cells (tanycytes) lining the lateral recesses of the 3rd ventricle and the infundibular process with which the pituitary cleft has a very close topographical relationship.The parenchymal cells of the pars distalis are closely related to the flattened marginal cells of the cleft. The intercellular spaces of the pars distalis form a three-dimensional labyrinthic series of cavities continuous with the submarginal spaces of the cleft. Further SEM and TEM results demonstrate that the majority of the microvillous marginal cells lining both sides of the cleft possess surface features such as bulbous protrusions, laminar evaginations and large cytoplasmatic vacuoles, which are very likely the expression of an active transport of fluids.On the basis of these results it is concluded that the fluid-like material (colloid) present in the pituitary cleft is mainly derived from the fluids contained in the lacunar spaces of the pars distalis. Thus, marginal cells by absorbing fluids from the cleft by active endocytosis, may transport to the pars intermedia material (or hormones) produced in the distal part of the gland and vice versa.The cilia present on many marginal cells, based on their 9+2 tubular pattern, possess a kynetic role. This is very similar to that shown by the ciliated cells of the ependyma lining the brain ventricles. The occurrence of ciliated cells within the pituitary parenchyma (mainly in the follicles) suggests that they probably arise from the ciliated cells populating the marginal layer of the cleft and with which the parenchyma cells are closely related.     

Radioimmunocytochemical localization of retinal S-antigen with monoclonal antibodies

Two monoclonal antibodies (RSA1/83 and RSA2/83) were developed against a homogeneous preparation of bovine retinal S-antigen. The two hybridomas produced by mouse X mouse hybrid myeloma cells secrete immunoglobulin G. Indirect autoradiography on glutaraldehyde-fixed preparations of bovine explants was used to locate the antigenic site. Antibody RSA1/83 recognizes the antigen primarily in the apical region of the rod outer segment, while antibody RSA2/83 located the antigen both in the outer and inner segments of the rod photoreceptor cells. A distinct band of silver grains also appeared along the inner limiting membrane with both antibodies. Control explants showed no specific labeling pattern over the various retinal compartments.     

Photoreceptor fine structure in the southern fiddler ray (Trygonorhina fasciata).

The fine structure of the retinal photoreceptors has been studied by light and electron microscopy in the southern fiddler ray or guitarfish (Trygonorhina fasciata). The duplex retina of this species contains only rods and single cones in a ratio of about 40:1. No multiple receptors (double cones), no repeating pattern or mosaic of photoreceptors and no retinomotor movements of these photoreceptors were noted. The rods are cylindrical cells with inner and outer segments of the same diameter. Cones are shorter, stouter cells with a conical outer segment and a wider inner segment. Rod outer segment discs display several irregular incisures to give a scalloped outline to the discs while cone outer segment discs have only a single incisure. In all photoreceptors a non-motile cilium joins the inner and outer segments. The inner segment is the synthetic centre of photoreceptors and in this compartment is located an accumulation of mitochondria (the ellipsoid), profiles of both rough and smooth endoplasmic reticulum, prominent Golgi zones and frequent autophagic vacuoles. The nuclei of rods and cones have much the same chromatin pattern but cone nuclei are invariably located against or particularly through the external limiting membrane (ELM). Numerous Landolt's clubs which are ciliated dendrites of bipolar cells as well as Müller cell processes project through the ELM, which is composed of a series of zonulae adherentes between these cells and the photoreceptors. The synaptic region of both rods (spherules) and cones (pedicles) display both invaginated (ribbon) synapses and superficial (conventional) synapses with cones showing more sites than the rods.     

Manipulating Adsorption–Insertion Mechanisms in Nanostructured Carbon Materials for High‐Efficiency Sodium Ion Storage

Hard carbon is one of the most promising anode materials for sodium‐ion batteries, but the low Coulombic efficiency is still a key barrier. In this paper, a series of nanostructured hard carbon materials with controlled architectures is synthesized. Using a combination of in situ X‐ray diffraction mapping, ex situ nuclear magnetic resonance (NMR), electron paramagnetic resonance, electrochemical techniques, and simulations, an “adsorption–intercalation” mechanism is established for Na ion storage. During the initial stages of Na insertion, Na ions adsorb on the defect sites of hard carbon with a wide adsorption energy distribution, producing a sloping voltage profile. In the second stage, Na ions intercalate into graphitic layers with suitable spacing to form NaC _x compounds similar to the Li ion intercalation process in graphite, producing a flat low voltage plateau. The cation intercalation with a flat voltage plateau should be enhanced and the sloping region should be avoided. Guided by this knowledge, nonporous hard carbon material has been developed which has achieved high reversible capacity and Coulombic efficiency to fulfill practical application.     

Ultrastructural Architecture and Organization of the Spore Envelope during Development in Thelohania bracteata (Strickland, 1913) after Freeze-Etching*

An ultrastructural study was made of the spore envelope during development in the microsporidan, Thelohania bracteata. The frozen-etched outer (convex) face of the relatively thin spore coat in the earliest immature stage of development has a granular structure in regular array. The inner (concave) face bears particles as well as depressions arranged in a net-like pattern. The mature spore coat has a substructure of numerous microfibers, ～8 nm in diameter, arranged in a matrix and forming thin layers which run parallel to the spore surface. The mature spore coat possesses both outer and inner limiting layers. The outer (convex) face of the outer limiting layer is granular. The convex face of inner limiting layer bears many particles as well as many long, narrow depressions. The concave face of the inner limiting layer carries many stud-like projections, ～40 nm long and 30 nm high, which are complementary to the depressions observed on the convex face. In addition, the concave face has subunits ～15 nm in diameter, apparently arranged in a hexagonal pattern with a center to center distance of ～18 nm. The change in size of these projections, depressions, and subunits presumably is related to spore maturation.     

Müller cell zinc transporter-3 labeling suggests a role in outer retina zinc homeostasis

Labeling for zinc transporter protein-3 (ZnT-3), which can be found localized to glutamatergic vesicles elsewhere in the nervous system, has revealed an unexpectedly high concentration of this transporter protein in the outer limiting membrane region of the murine retina, a region that contains the mitochondria-rich portion of photoreceptor inner segments and is not involved with vesicle release. Having suggested the possibility that Müller cell apical villi forming the outer limiting membrane may be associated with the labeling observed, we used immunohistochemical techniques to look for ZnT-3 labeling of Müller cells isolated from rat and mouse retinas. With DAB labeling, rat Müller cell apical villi, soma, and endfeet exhibited ZnT-3 reactivity. FITC label and confocal analysis revealed that ZnT-3 protein appeared throughout the length of mouse Müller cells. We conclude from these observations that the dense labeling for ZnT-3 in the photoreceptor inner segment region of murine retinal slices is due to labeling of ZnT-3 protein associated with Müller cell apical villi. Based on these findings we suggest that Müller cells utilize ZnT-3 to regulate retinal zinc homeostasis and that this role is important to mitochondrial function in the photoreceptor inner segments.