The development of the cnidoblasts of Hydra; an electron microscope study of cell differentiation

The general histological organization of Hydra is reviewed and electron microscopic observations are presented which bear upon the nature of the mesoglea, the mode of attachment of the contractile processes of the musculo-epithelial cells, and the cytomorphosis of the cnidoblasts. Particular attention is devoted to the changes in form and distribution of the cytoplasmic organelles in the course of nematocyst formation. The undifferentiated interstitial cell is characterized by a small Golgi complex, few mitochondria, virtual absence of the endoplasmic reticulum, and a cytoplasmic matrix crowded with fine granules presumed to be ribonucleoprotein. These cytological characteristics persist through the early part of the period of interstitial cell proliferation which leads to formation of clusters of cnidoblasts. With the initiation of nematocyst formation in the cnidoblasts, numerous membrane-bounded vesicles appear in their cytoplasm. These later coalesce to form a typical endoplasmic reticulum with associated ribonucleoprotein granules. During the ensuing period of rapid growth of the nematocyst the reticulum becomes very extensive and highly organized. Finally, when the nematocyst has attained its full size, the reticulum breaks up again into isolated vesicles. The Golgi complex remains closely applied to the apical pole of the nematocyst throughout its development and apparently contributes to its enlargement by segregating formative material in vacuoles whose contents are subsequently incorporated in the nematocyst. The elaboration of this complex cell product appears to require the cooperative participation of the endoplasmic reticulum and the Golgi complex. Their respective roles in the formative process are discussed.     

The fine structure of differentiating interstitial cells in Hydra

Summary Interstitial cells of hydra are small undifferentiated cells containing an abundance of free ribosomes and few other cytoplasmic organelles. They are capable of differentiating into epitheliomuscular, digestive, glandular, nerve cells, and cnidoblasts. Developing epitheliomuscular and digestive cells acquire bundles of filaments, 50 Å in diameter, which later are incorporated into the muscular processes. Early gland cells develop an elaborate rough-surfaced endoplasmic reticulum and one or more Golgi apparatus. Secretory granules originate in the Golgi region eventually filling the apex of the cell. Neurons are recognized first by the presence of an elaborate Golgi apparatus, absence of a well-developed endoplasmic reticulum, and later the appearance of cytoplasmic processes. The most striking feature of nematocyst formation by cnidoblasts is the presence of a complex distribution system between protein synthesizing rough-surfaced endoplasmic reticulum and the nematocyst. This system consists of connections between cisternae of the endoplasmic reticulum with smooth Golgi vesicles which in turn are connected to minute tubules, 200 Å in diameter. The tubules extend from the Golgi region around the nematocyst finally entering the limiting membrane of the nematocyst. It is suggested that the interstitial cells of hydra represent a model system for the investigation of many aspects of cell differentiation.This work was supported by grants from the National Cancer Institute (TlCA-5055) and from the National Institute of Arthritis and Metabolic Diseases (AM-03688), National Institutes of Health, Department of Health, Education and Welfare.The author is indebted to Dr. Russell J. Barrnett for his guidance and interest throughout this investigation.     

The Fine Structure of Blastema Cells and Differentiating Cartilage Cells in Regenerating Limbs of Amblystoma Larvae

Regenerating forelimbs of larval salamanders, Amblystoma punctatum, were fixed in OsO₄ at various intervals after amputation and were sectioned for study with the electron microscope. The dedifferentiated cells comprising the early blastema were found to have a fine structure similar to that of other undifferentiated cells and to have lost all of the identifying morphological features of their tissues of origin. The cytoplasm of such cells is characterized by numerous free ribonucleoprotein granules and a discontinuous vesicular endoplasmic reticulum. The cells have more abundant cytoplasm and are in closer contact with each other than was previously realized. The layer of condensed ground substance investing most differentiated cell types is lacking. After a period of rapid cell division, the morphology of the blastema cell changes. Cytoplasm is now sparse and contains a high concentration of free ribonucleoprotein granules, but little endoplasmic reticulum. The differentiating cartilage cell, however, develops an extensive, highly organized endoplasmic reticulum and the Golgi apparatus also appears to become more highly differentiated and more extensive at this time. Small vesicles appear throughout the cytoplasm at the time the new cisternae originate and may contribute to their formation. These and other changes in the cytoplasmic organelles are discussed.     

CYTOPLASMIC MICROTUBULES : I. Hydra

Small cytoplasmic tubules are present in the interstitial cells and cnidoblasts of hydra. They are referred to here as "microtubules." These tubular elements have an outside diameter of 180 A and an inside diameter of 80 A. By difference, the membranous wall is estimated to be 50 A thick. The maximum length of the microtubules cannot be determined from thin sections but is known to exceed 1.5 µ. In the interstitial cells the microtubules are found in the intercellular bridges, free in the cytoplasm and in association with the centrioles. In the cnidoblast they form a framework around the developing nematocyst and in late stages are related to the cnidocil forming a tight skein in the basal part of the cell. Especially in this cell, confluence of microtubules with small spherical vesicles of the Golgi complex has been observed. It is proposed that these tubules function in the transport of water, ions, or small molecules.     

THE NORMAL FINE STRUCTURE OF OPOSSUM TESTICULAR INTERSTITIAL CELLS

The interstitial tissue of the opossum testis includes interstitial or Leydig cells, macrophages, and small cells which morphologically resemble mesenchymal cells. The latter are thought to give rise to mature interstitial cells. The most prominent feature of the interstitial cell cytoplasm is an exceedingly abundant agranular endoplasmic reticulum. This reticulum is generally in the form of a meshwork of interconnected tubules about 300 to 450 A in diameter, but occasionally it assumes the form of flattened, fenestrated cisternae resembling those of pancreatic acinar cells, except for the lack of ribonucleoprotein particles on the surface of the membranes. The interstitial cells vary considerably in their cytoplasmic density. The majority are quite light, but some appear extremely dense, and in addition usually have a more irregular cell surface, with numerous small pseudopodia. These differences may well reflect variations in physiological state. Cytoplasmic structures previously interpreted as "crystalloids" consist of long bundles of minute parallel tubules, each about 180 A in diameter, which seem to be local differentiations of the endoplasmic reticulum. The mitochondria are rod-shaped, and contain a moderately complex internal membrane structure, and also occasional large inclusions that are spherical and homogeneous. The prominent juxtanuclear Golgi complex contains closely packed flattened sacs and small vesicles. The results of the present study, coupled with biochemical evidence from other laboratories, make it seem highly probable that the agranular endoplasmic reticulum is involved in the synthesis of the steroid hormones produced by the interstitial cell. This finding therefore constitutes one of the first functions of the agranular reticulum for which there is good morphological and biochemical evidence.     

Light and electron microscopic studies of crayfish hemocytes

Using light and electron microscopy, three hemocyte types are described in the hemolymph of the crayfish. The coagulocyte comprises 65% of the total hemocyte number and contains medium-sized cytoplasmic granules, abundant dilated rough endoplasmic reticulum, and a highly developed Golgi complex. It rapidly undergoes cytolysis in vitro and participates in coagulation by releasing the contents of its granules to the hemolymph. The granulocyte comprises 31% of the total hemocyte number and is capable of phagocytosis. It contains large, irregularly shaped cytoplasmic granules, a moderately developed Golgi complex, and moderate amounts of non-dilated rough endoplasmic reticulum. During coagulation in vitro, the cell attaches and spreads onto the substratum; this is followed by a slow intracellular granule breakdown and cytolysis. The amebocyte comprises 4% of the total hemocyte number and it is also capable of phagocytosis. It possesses small cytoplasmic granules, many vacuoles, a moderately developed Golgi complex, and large amounts of smooth endoplasmic reticulum. It is distinguished from the other two cell types by being stable and motile in vitro.     

Intracellular transport of albumin through the secretory apparatus of rat liver parenchymal cells. An immunocytochemical study

The fine structural localization of albumin in rat liver parenchymal cells was determined by an improved immunocytochemical method and serial sectioning. Albumin in the secretory apparatus of the parenchymal cells was present in segments of the rough endoplasmic reticulum, interrupted with negative segments, in transport vesicles, Golgi saccules, finely anastomosed tubules and vesicles on the trans side of the Golgi complex, and in secretion granules. Horizontally sectioned Golgi saccules contained lipoprotein particles on one side and albumin on the other side. After transport, the vesicles that contained albumin fused with the so-called rigid lamellae on the trans-side of the Golgi complex. Ultrathin serial sections revealed no true structural continuity between the endoplasmic reticulum and the cis-aspect of the Golgi complex. We concluded that secretory proteins are transported from the endoplasmic reticulum to the Golgi complex by transport vesicles that bud from the endoplasmic reticulum and fuse with the Golgi saccules. These vesicles fuse regularly with the Golgi saccules on the cis-side and occasionally with tubular elements on the trans-aspect that may belong to the so-called GERL.     

Fine structure of nerve cells in a planarian

The fine structure of the nerve cell types in the white planarian Procotyla fluviatilis were described. Ganglion cells comprise the major portion of the brain. These cells are irregular in shape with several cytoplasmic processes and contain ribosomes, a sparse endoplasmic reticulum, microtubules, lysosomes, and a Golgi apparatus with numerous small vesicles. Granule-containing cells are situated in the peripheral regions of the brain and along the nerve cords. These cells contain ribosomes, rough-surfaced endoplasmic reticulum and a Golgi apparatus with associated dense granules. The granules occupy most of the cytoplasm and are ～ 750A in diameter with moderately dense contents, ～ 750A with opaque contents, and ～ 1000A with contents of medium density. These granules are similar to those in the nervous systems of higher animals that contain epinephrine, norepinephrine, and neurosecretory substance, respectively. Each cell contains predominantly one type of granule although there is some intermixing of granules and intermediate types between the three most abundant granules. Small clear vesicles, resembling cholinergic synaptic vesicles, and all types of dense granules occur in the neuropil and within nerve endings.     

Further observations on dividing and non-dividing cnidoblasts in the regenerating isolated gastrodermis of Hydra

Summary Cnidoblasts derived from the dedifferentiation of gland cells in the regenerating isolated gastrodermis of Hydra are capable of nuclear and cytoplasmic division. The daughter cell containing the nematocyst apparently develops normally. The fate of the other daughter cell remains obscure, but it is believed that it is also capable of developing a nematocyst. Only a single bi-nucleated cnidoblast was observed and it was in the process of degeneration. It is suggested that at least in the present system, cnidoblasts are derived not only from interstitial cells but also from pre-existing cnidoblasts.This investigation was supported by the National Science Foundation Grant No. GB 8384.With the technical assistance of Linda Bookman.     

FINE STRUCTURE OF THE NERVOUS SYSTEM OF HYDRA

Fine structural details of the cells and processes of the hydranenous system are reported in this paper. Ganglion cells aresmall bipolar or multipolar cells situated above the muscularprocesses of epitheliomusiular cells. An elaborate Colgi apparatusconsisting of parallel lamellae and small and large vesiclesis present in these cells. Some cells are poor in ribosomeswhile others contain numerous free ribosomes. In the ribosome-richcells, small membranous microtubules originating from the nuclearenvelope extend into the cytoplasm and neurites. The neuritesalso contain vesicles and mitochondria and terminate at thebases of cnidoblasts and on the muscular processes of epitheliomuscularcells. Specialized synapses were not observed. A second cell type contains many membrane-bounded dense granules,1000 A in diameter, and these are considered to be neurosecretorycells. Neurosecretory granules on cnidoblasts and epitheliomuscularcells. Sensory cells are small elongated cells originate inthe Golgi apparatus and are abundant in neurites which alsoterminate situated between the apical surfaces of epithelialand digestive cells. These cells are characterized by an apicalspecialization which appears to be a modified cilium. Neurosensorycells were also observed. The intimate connection of the nervoussystem with cnidoblasts suggests a role in nematocyst discharge.The finding of neurosecretory material supports the hypothesisthat the neural control of regeneration in hydra is regulatedby material released at nerve endings.     

The cytology of the testaceous rhizopod Lesquereusia spiralis (Ehrenberg) Penard. I. Ultrastructure and shell formation

Ultrastructure and shell formation in the testaceous ameba, Lesquereusia spiralis, were investigated with both scanning and transmission electron microscopy and X-ray microanalysis. The nucleus, surrounded by a fibrous lamina, contains multiple nucleoli. The cytoplasm, containing a well developed granular endoplasmic reticulum, also contains remnants of starch granules in stages of digestion. Spherical aggregates of ribosome-like particles may be seen. Golgi complexes seem to produce both a nonordered fibrous material and an electron dense vesicle. Only the latter appears to bleb off from the Golgi complex. X-ray microanalysis demonstration of silicon in Golgi vesicles and in some dense vesicles suggests that the fibrous component of the cisternae may take up and concentrate silica to form the electron-dense component of the vesicles. Membrane-bound siliceous crystals are often seen adjacent to the Golgi, suggesting either a Golgi origin or platelet formation in vesicles after release from the Golgi complex. Both electron-dense bodies and siliceous platelets are released from the cell by a process similar to apocrine secretion and may be seen outside the cell in route to the shell during shell morphogenesis. Shell development involves fusion of electron-dense bodies to form a matrix, positioning of siliceous platelets in this matrix parallel to the shell surface, and development of a system of matrix chambers. A particulate glycoconjugate is released to the shell surface upon rupture of the matrix chamber.     

LIVER PARENCHYMAL CELL INJURY : I. Initial Alterations of the Cell Following Poisoning with Carbon Tetrachloride

The structure of the endoplasmic reticulum, plasma membrane, mitochondria, and Golgi apparatus of the liver parenchymal cell is strikingly altered within 1 hour following the administration of a single oral dose of carbon tetrachloride to rats. Progressive loss of glucose-6-phosphatase activity accompanies dispersal of the ergastoplasm. Electron microscopy reveals that these changes are associated with vacuolization of the cisternae of the granular endoplasmic reticulum, degranulation of its membranes, and the appearance of increased number of free ribosomes in the adjacent cytoplasmic matrix. Concomitantly, calcium enters the liver parenchymal cell and is sequestered by mitochondria. First increased at 30 minutes, calcium content is maximal at 1 hour and returns to normal at 2 hours. Although succinic and glutamic dehydrogenase activity patterns within the liver lobule are unaffected, liver cell mitochondria enlarge and some appear to fuse or assume cup-like configurations. Microvilli lining the space of Disse become irregularly indistinct and increasingly pleomorphic by 30 minutes when the plasma membrane becomes increasingly permeable to calcium. Golgi vesicles swell and discharge their granules during the period of poisoning studied. Although all the changes observed may be the result of direct interaction of carbon tetrachloride with the membranes of the cytoplasmic constituents of the liver parenchymal cell, the possibility that the irreversible changes observed in the granular endoplasmic reticulum may be due to the chemical interaction between the poison and this system is discussed.     

THE FINE STRUCTURE OF TESTICULAR INTERSTITIAL CELLS IN GUINEA PIGS

In guinea pig testes perfused with either glutaraldehyde or osmium tetroxide fixative, the cytoplasm of the interstitial cells contains an exceptionally abundant agranular endoplasmic reticulum. The reticulum in central regions of the cell is a network of interconnected tubules, but in extensive peripheral areas the reticulum is commonly organized into closely packed, flattened cisternae which are fenestrated. Occasional small patches of the granular reticulum occur in the cytoplasm and connect freely with the agranular reticulum. The mitochondria have a dense matrix and contain cristae and some tubules. The Golgi complex is disperse and shows no evidence of secretory material. The cytoplasm also contains lipid droplets. Lipofuscin pigment granules are probably polymorphic residual bodies and contain three components: (1) a dense material which at high magnification shows a 75-A periodicity; (2) a medium-sized lipid droplet; and (3) a cap-like structure. In glutaraldehyde-perfused testis the interstitial cell cytoplasm appears to have the same density from cell to cell, and the agranular reticulum is tubular or cisternal but not in the form of empty vesicles. Thus the "dark" and "light" cells and the vesicular agranular reticulum sometimes encountered in other fixations may be artifacts. Biochemical results from other laboratories, correlated with the present findings, indicate that the membranes of the agranular endoplasmic reticulum in guinea pig interstitial cells are the site of at least two enzymes of androgen biosynthesis, the 17-hydroxylase and the 17-desmolase.     

Elektronenmikroskopische Untersuchungen von Differenzierungsvorgängen bei Moosen

Summary In meristematic cells of the gemma of Riella helicophylla and in young bud cells from the protonema of Funaria hygrometrica the cell plate is formed by fusion of small vesicles originating from the Golgi apparatus. These spherical vesicles of about 0.1 m diameter have an electron dense centre, probably consisting of pectic substances or their precursors. The endoplasmic reticulum producing multivesicular bodies participate in cell plate formation too. Another cytoplasmic component forming the cell plate are coated vesicles, the origin of which is the Golgi apparatus and perhaps also the endoplasmic reticulum. In view of these observations the question of whether the endoplasmic reticulum or the Golgi apparatus forms the cell plate must be answered in this way: both endoplasmic reticulum and Golgi apparatus supply material for growth of the cell plate. Multivesicular bodies, coated vesicles and other small vesicles of unknown nature participate in the formation of the primary wall.

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Zum Teil finanziert mit Sondermitteln des Landes Niedersachsen an Prof. Dr. M. Bopp.     

Quantitative aspects of membrane shifts in rat parathyroid cells initiated by decrease in serum calcium

The secretory activity of parathyroid glands in rats was stimulated by decreasing the serum Ca++ concentration through constant intravenous infusion of EGTA. The morphometric analysis of the nuclear and cytoplasmic volume and of the surface area of the rough endoplasmic reticulum, Golgi complex, secretory granules and plasma membrane revealed a membrane shift from secretory granules and Golgi complex to the plasma membrane within 1 hr of calcium depression. Subsequently, between 1 and 3 hr of calcium depression, the membrane shift was from the plasma membrane to the Golgi complex. It is considered likely that these membrane shifts are related to a rise in release of parathyroid hormone by exocytosis and a subsequent increase in retrieval of plasma membrane by endocytosis—probably through the compartment of coated pits and coated and uncoated vesicles.     

Cytochemical characterization of the endomembranous system during the oocyte primary growth in Serrasalmus spilopleura (Teleostei, Characiformes, Characidae)

The morphophysiological changes that occur during oocyte primary growth in Serrasalmus spilopleura were studied using ultrastructural cytochemical techniques. In the previtellogenic oocytes endoplasmic reticulum components, Golgi complex cisternae and vesicles, lysosomes, multivesicular bodies and some electron-dense vesicles react to acid phosphatase (AcPase) detection. The endoplasmic reticulum components, Golgi complex cisternae and vesicles also react to osmium tetroxide and potassium iodide impregnation (KI). These structures, except for the Golgi complex cisternae, are strongly contrasted by osmium tetroxide and zinc iodide impregnation (ZIO). Some electron-dense vesicles are ZIO-stained, while microvesicles in the multivesicular bodies and other large isolated cytoplasmic vesicles are contrasted by KI. At primary oocyte growth, the activity of the endomembranous system and the proliferation of membranous organelles are intense. The biosynthetic pathway of the lysosomal proteins such as acid phosphatase, involves the endoplasmic reticulum, Golgi complex, vesicles with inactive hydrolytic enzymes and, finally, the lysosomes. The oocyte endomembranous system have reduction capacity and are involved in the metabolism of rich in SH groups.     

Transition vesicle formationin vitro

Summary Isolated fractions enriched in transition elements derived from part rough—part smooth regions of endoplasmic reticulum of rat liver respondin vitro to ATP plus a concentrated fraction of cytoplasmic proteins by formation of ca. 60 nm vesicles with nap-like coats resembling those of transition vesicles of the intact cell. Similar vesicles are normally considered to function in the transfer of materials from endoplasmic reticulum to cis elements of the Golgi apparatus.     

Cell-free systems to study vesicular transport along the secretory and endocytic pathways

Proteins bound for the cell surface, lysosomes, and secretory storage granules share a common pathway of intracellular transport. After their synthesis and translocation into the endoplasmic reticulum, these proteins traverse the secretory pathway by a series of vesicular transfers. Similarly, nutrient and signaling molecules enter cells by endocytosis, and move through the endocytic pathway by passage from one membrane-bound compartment to another. Little is known about the mechanisms by which proteins are collected into transport vesicles, or how these vesicles form, identify their targets, and subsequently fuse with their target membranes. An important advance toward our understanding these processes has come from the establishment of cell-free systems that reconstitute vesicular transfers in vitro. It is now possible to measure, in vitro, the transport of proteins from the endoplasmic reticulum to the Golgi, between Golgi cisternae, and the formation of transport vesicles en route from the trans Golgi network to the cell surface. Along the endocytic pathway, cell-free systems are available to study clathrin-coated vesicle formation, early endosome fusion, and the fusion of late endosomes with lysosomes. Moreover, the selective movement of receptors between late endosomes and the trans Golgi network has also been reconstituted. The molecular mechanisms of vesicular transport are now amenable to elucidation.     

Ultrastructural studies on the formation of yolk granules in the statoblast of a fresh-water bryozoan,Pectinatella gelatinosa

The formation of protein-carbohydrate yolk in the statoblast of a fresh-water bryozoan, Pectinatella gelatinosa, was studied by electron microscopy. Two types (I and II) of yolk cells were distinguished. The type I yolk cells are mononucleate and comprise a large majority of the yolk cells. The type II yolk cells are small in number; they become multinucleate by fusion of cells at an early stage of vitellogenesis. In both types of yolk cells, electron-dense granules (dense bodies) are formed in Golgi or condensing vacuoles, which are then called yolk granules. For the formation of yolk granules, the following processes are considered: 1. Yolk protein is synthesized in the rough-surfaced endoplasmic reticulum (RER) of the yolk cells. 2. The synthesized protein condenses in the cisternal space of the RER and is packaged into small oval swellings, which are then released from the RER as small vesicles (Golgi vesicles, 300-600 A in diameter). 3. The small vesicles fuse with one another to form condensing vacuoles, or with pre-existing growing yolk granules. 4. In the matrix of the condensing vacuoles or growing yolk granules, electron-dense fibers are fabricated and then arranged in a paracrystalline pattern to form the dense body. 5. After the dense body reaches its full size, excess membrane is removed and eventually the yolk granules come to mature. Toward the end of vitellogenesis of the yolk cells, the cytoplasmic organelles are ingested by autophagosomes derived from multivesicular bodies and disappear.     

Immunocytochemical localization of renin in juxtaglomerular cells

The involvement of various organelles in the synthesis, transport, and packaging of renin in the juxtaglomerular cells of newborn mice has been investigated by immunocytochemistry with the protein A-gold technique. Highly specific rabbit antibodies against mouse submandibular renin were used. Mild fixation and embedding in glycol methacrylate allowed enough sensitivity to identify a steep gradient of labeling from rough endoplasmic reticulum to Golgi complex to secretory granules. Routine fixation and embedding in Epon produced labeling differentials that allowed delineation of hitherto ill-defined types of secretory granules and vacuoles. The classical pattern of synthesis, transport, and packaging of secretory proteins involves the rough endoplasmic reticulum and Golgi complex and seems to apply to renin secretion. Immunoreactive renin is packaged as rhomboid crystals at the trans face of the Golgi complex. The limiting membrane of these rhomboids fuses to form coalescing protogranules where the crystals eventually yield their individuality maturing into secretory granules. Vacuoles containing a flocculent material, with or without a dense core, show significant immunocytochemical labeling. These vacuoles are not associated with the Golgi complex but occupy cytoplasmic areas well endowed with rough endoplasmic reticulum. As judged from their morphological features and their immunoreactivity, the vacuoles do not seem to follow the sequence of events typical of protogranules and coalescing protogranules. They possibly represent a parallel pathway of renin synthesis and transport, involving the nuclear envelope and bypassing the Golgi complex.