Studies on the histomorphology and histochemistry of the saccus vasculosus of Cyprinion macrostomus.

The saccus vasculosus of Cyprinion macrostomus is made up of coronet and supporting cells. Neurones are not observed. The luminal surface of the coronet cells exhibits hair-like protrusions. The saccus vasculosus is mainly concerned with a secretory function. Plenty of secretory material is seen in the lumen. PAS and alcian blue tests are positive for the secretory material in the lumen and the apical protrusions of the coronet cells. Some cells also exhibit small PAS- and alcian blue-positive granules. The possibility of the elaboration of glycogen by the coronet cells is ruled out. The toluidine blue test is also positive for the secretory material in the lumen and the apical protrusions. Small metachromatic granules are also seen in some of the coronet cells. The acetone-Sudan black B and Sudan black B tests point to lipid moiety of the coronet cells, the apical protrusions and the secretory material in the lumen. The acrolein-Schiff and ninhydrin-Schiff tests point to a protein moiety of the coronet cells and the secretory material in the lumen. The performic acid-alcian blue test showed cystine to be present in small amount.     

On the presence and localization of glycogen accumulations inside coronet cells of the saccus vasculosus of the dogfish (Scylliorhinus caniculus)

Summary In several coronet cells of the saccus vasculosus of Scylliorhinus large quantities of glycogen occur, as shown by light and electron microscopy. The significance of glycogen as an energy storage necessary for a transcellular ion transport process taking place in the coronet cells is discussed.The authors thank Dr. F.C.G. van de Veerdonk, W. F. Jansen and W. F. G. Flight for reading the manuscipt and for their critical remarks. They are also indebted to Mr. H. van Kooten and his staff for their valuable photographic assistance.     

Fine structural localization of adenosine triphosphatase activities in the saccus vasculosus of the rainbow trout,Salmo gairdneri Richardson

The following characteristics of the adenosine triphosphatases (ATPase) in the saccus vasculosus were studied in Salmo gairdneri Richardson: 1) distributional pattern, 2) cytochemical properties in relation to different substrates, inhibitors, pH and bivalent metal ions, and 3) ultrastructural localization. Ultracytochemical studies using modifications of the Washstein-Meisel technique showed that within the pH range 7.1-8.0 several Mg++ or Ca++-activated ATPase are localized on the intracellular surface of membranes and in the cytoplasm of ependymal coronet cells and tanycytes ("supporting cells", "Zwischenzellen", glial cells"). The high ATPase activity at the level of the specialized luminal plasma membranes of coronet cell globules and of tanycyte microvilli is discussed in relation to phenomena of active transport and a possible resulting transfer of low-molecular weight substances into and/or from the cerebrospinal fluid (CSF). The localization of ATPase on the specialized membranes of primary vesicles is considered in connection with available structural and enzyme-cytochemical data on a possible function of these cell organelles in storage and release of substances (including Ca++ ions?). The cytoplasmic ATPase activity in coronet cells is ascribed to microtubules and/or possible existing contractile proteins/filaments, presumably concerned with internal transport or motility processes. In tanycytes ATPase activity is believed to be associated with the characteristic microfilamentous system of still unknown function. The ATPase activity in the (9 + 0) ciliary apparatus of globules could not be interpreted in terms of motility. The present study provides further support to the proposed hypothesis of the transport function of the saccus vasculosus, and an extension of the concept in the sense that not only the principal coronet cells, but also the tanycytes of this circumventricular organ are involved in CSF-homeostasis.     

Primitive Coronet Cells in the Brain of Oikopleura (Appendicularia,Tunicata)

Abstract A peculiar cell type is described from the sensory vesicle of the brain of the appendicularian tunicate Oikopleura dioica. The cells carry globular, modified cilia and resemble in several other respects the coronet cells of Saccus vasculosus in many fishes. This finding, together with earlier reports of similar cells in the sensory vesicle of ascidian tadpoles, makes it probable that a primitive form of coronet cells is a regular constituent of the tunicate brain cavity. The existence of coronet cells in the tunicate line of evolution is of interest from a phylogenetical as well as functional point of view. The author proposes that the tunicate sensory vesicle corresponds to part of the craniate third ventricle and that the tunicate coronet cells are involved in the regulation of the ventricle fluid composition.     

Light microscopic and ultrastructural study of the development of the saccus vasculosus in the rainbow trout,Oncorhynchus mykiss

This study using light and electron microscopy indicates that the saccus vasculosus is distinguishable in 9-mm embryos and grows continuously throughout embryonic development to the adult stage. In the saccus vasculosus, epithelial mitoses are observed in all stages studied. Phases of centriologenesis, ciliogenesis, and globule formation have been characterized in developing coronet cells. During the phase of centriologenesis, new centrioles appear in association with pre-existing centrioles and not on deuterosomes. After ciliogenesis, each cilium differentiates to a globule almost at the same time as the other cilia of the coronet cell. The inner membrane system of the globules seems to derive from the ciliary plasma membrane. This membrane system often produces membrane whorls during the development. The different phases of coronet cell development have been found in the same individual and in all the stages studied except the 9-mm embryo. Cerebrospinal fluid-contacting neurons are observed in the saccus epithelium from the 12-mm embryos on and are distinguishable from coronet cells in their early formative stages. The three cell types of the saccus vasculosus increase continuously in number during development. Nerve processes are found in the saccus vasculosus of embryos, whereas differentiated synapses appear later in the fry. The significance of continued coronet cell formation is discussed in relation to a putative coronet cell and/or a globule renewal cycle in the adult.     

Histophysiology of the saccus vasculosus of the Indian freshwater goby Glossogobius giuris Ham. (Teleostei)

The author studied the structure and functions of the saccus vasculosus of the Indian freshwater goby Glossogobius giuris (Ham.). The saccus is ovoid, is localized on the ventral surface of the brain and is lodged between the inferior lobes. It consist of several loculi lined with coronet cells and is bathed with blood from surrounding sinusoids. The coronet cells are variably shaped and have a conspicous central nucleus. It is suggested that the purpose of the saccus vasculosus is to act as a storage site for carbohydrates to the brain. By converting glycogen to acid mucopolysaccharides, the coronet cells are involved in glycogen metabolism.     

A histoenzymological study of the saccus vasculosus of the freshwater teleost, Clarias batrachus (L.).

The saccus vasculosus (SV) of C. batrachus is comparatively small and situated latero-dorsal to the pituitary in between the lobi inferiores. It is in open communication with the third ventricle and is made up of coronet and supporting glial cells with interspersed liquor contacting neurons. These cellular constituents are arranged in one to three layers which are not thrown into folds. The PAS positive nature of the apical part of some coronet cells and their continuation with the PAS and AF positive material present in the lumen strongly suggest their secretory role. The coronet cells exhibited strong NADH diaphorase, NADPH diaphorase, cytochrome oxidase and MAO activity. AChE activity was comparatively weak. These enzyme histochemical studies show that SV has a dual function of transport and secretion. The strong MAO activity suggests the probable aminergic control of this organ.     

The saccus vasculosus of Anguilla anguilla (L.) from larva to adult

Summary The ultrastructure of coronet cells of the saccus vasculosus has been studied in specimens of Anguilla anguilla (L.) at different stages of its life cycle. At all the stages observed coronet cells are composed of a basal and an apical part, the latter bearing globules with primary vesicles. In the larva (a marine form) and in the fully metamorphosed small eel at the time of entry into freshwater the narrow lumen and the vesicles within the apical globules are filled with electron-dense material. In forms in which adaptation to freshwater has occurred, the saccus lumen appears expanded, the apical globules are better developed, and the electron-dense material has disappeared. It is suggested that the two situations observed represent different functional states of the organ, in relation to different conditions of environmental salinity.The authors gratefully acknowledge the contribution of Dr. G. Andreoli, of the University of Parma, who provided the Atlantic larvae for this study.     

Morphogenesis of the saccus vasculosus of turbot Scophthalmus maximus: assessment of cell proliferation and distribution of parvalbumin and calretinin during ontogeny

The ontogenesis of the saccus vasculosus (SV) of turbot Scophthalmus maximus is described using histological and immunohistochemical methods to assess the general morphology, as well as the distribution of proliferative cells and several calcium‐binding proteins (CaBP). The results reveal that the SV begins to differentiate on hatching, when immature coronet cells are morphologically distinguishable. Further morphogenesis involves the formation of a tubular avascular SV, which remains until premetamorphic larval stages. Folding and vascularization of the SV occurs mostly during metamorphosis, when S. maximus settle down on the bottom. Proliferative cells were placed within the SV itself and in the neighbouring infundibular hypothalamus. Their putative relationship with the growth of the SV is discussed. The CaBPs analysed are expressed in coronet cells. Parvalbumin is expressed in these cells from the beginning of their differentiation, while calretinin expression arises in the tubular SV and becomes more widespread over time. These data emphasize the importance of calcium buffering in the function of coronet cells.     

Changes in the Saccus Vasculosus Epithelium During the Smoltification Process in Atlantic Salmon (Salmo salar)

The changes that take place in the saccus vasculosus epithelium during smoltification of Atlantic salmon (Salmo salar) are described. The major cell type of the epithelium, the coronet cells, have a number (mean = 32) of spherical protrusions connected with a non-motile cilium to the apical part of the cell. The protrusions extend into the saccus lumen. In parr the protrusions become more spherical during the spring season, while in smolt the opposite process takes place. Parr coronet cells have most of the mitochondria in the apical part of the cell, while in smolt they are concentrated basally. The relative areal fraction of heterochromatin in the nuclei increase from 23 to 29 during the smoltification process. The results are consistent with the view that the coronet cells take an active part in the smoltification process.     

Histochemische Beobachtungen am Saccus vasculosus der Forelle

Summary The saccus vasculosus of rainbow trout and brown trout, the latter caught in the wild, has been investigated by histochemical means. Isolated coronet cells and groups of them were found to be rather strongly but unspecifically stainable by alcian blue. A performic acid-aniline-aldehyde-thionine reaction demonstrated that such cells contain more disulfide groups than their nonstaining neighbours. This higher disulfide content and the stainability by alcian blue do not necessarily coincide with the presence of acid mucopolysaccharide, which was found in the cytoplasm of coronet cells in some cases. The hypothesis is discussed that cystine may be stored and used by coronet cells as a precursor of the acid mucopolysaccharide, which has been shown in the lumen of the organ.     

Ultrastructural changes in the coronet cells of the saccus vasculosus from rainbow trout,Salmo gairdneri (Richardson), kept in sea water

Summary The coronet cells of saccus vasculosus of fresh-water living and sea-water adapted rainbow trout were studied with the electron microscope, with special regard to changes in the latter group. Only quantitative differences were observed, namely a raised number of mitochondria in the apical region and the head and also a concentration of the agranular endoplasmic reticulum with a higher amount of electron-dense material and vesicles around the Golgi saccules. Together, these findings suggest a secretory function for the coronet cell. A supposed transport of vesicles from the head region of the coronet cell out into the globules is suggested. Interrelation between primary and secondary vesicles is discussed.This work was supported by grants from the Royal Physiographical Society of Lund and the Faculty of Natural Sciences at the University of Lund.—I am greatly indebted to Mrs. Lena Svenre and Miss Inger Norling for excellent technical assistance.     

Possible Ascidian Counterpart to the Vertebrate Saccus Vasculosus with Reference to Pyura tessellata (Forbes) and Boltenia echinata (L.)

The morphology of the hitherto unknown larva of Pyura tessellata (Forbes) is described and the discovery of an auxiliary brain vesicle is presented. The vesicle was also found in another pyurid species Boltenia echinata (L.). The auxiliary vesicle communicates with the sensory vesicle at the level of the statocyte and is lined with cubodial epithelial cells carrying a 2 μm globular structure projecting into the lumen. These cells very closely resemble primitive tunicate coronet cells previously reported and coronet cells typical for the saccus vasculosus found in elasmobranchs and many ganoids and teleosts.     

Relationships between the tubular system in the globules of the coronet cells of the saccus vasculosus and the cerebrospinal fluid in Gasterosteus aculeatus form leiurus (Teleostei)

Summary In the stickleback, Gasterosteus aculeatus, the globules of the coronet cells in the saccus vasculosus contain a tubular system that most probably communicates permanently with the lumen of the saccus. Only very few openings were found in random ultrathin sections. Injecting peroxidase into the cerebrospinal fluid revealed the communication between this tubular system and the cerebrospinal fluid. As early as 1 h after peroxidase injection the tracer was detected in the tubular system. This system increases the potential exchange surface between the coronet cells and the content of the saccular lumen, and may also facilitate the access of components of the cerebrospinal fluid (CSF), even of high molecular weight, into the globules. It remains to be determined whether the intratubular condensations (granules) are, as often believed, of secretory origin or are formed by accumulation of components of the CSF.With the technical collaboration of Melle A. Meunier     

Metabolic activity in the saccus vasculosus of the rainbow trout,salmo gairdneri (richardson)

Summary Labelling experiments to test mucopolysaccharide synthesis in the saccus vasculosus of the rainbow trout gave negative results and provoked analysis of succinic dehydrogenase activity in the coronet cells. Such activity was almost absent in the fresh-water living rainbow trout but was intense in sea water adapted individuals as well as in a sea water teleost, the flounder. Conversely, in sea water adapted rainbow trout which had been transferred back to fresh-water, activity had again decreased. In the sea water adapted rainbow trout high succinic dehydrogenase activity was paralleled by strongly positive staining with alcian blue, indicating the presence of mucopolysaccharides. The latter fishes also showed incorporation of N-acetyl-³H-glucosamine. Our findings indicate a role for saccus vasculosus in adaptation for osmoregulation in sea water and favour the hypothesis of a secretory function for the coronet cells.This work has been supported by grants from the Royal Physiographical Society of Lund and the Faculty of Natural Sciences at the University of Lund. We are greatly indebted to Mrs. Lena Eriksson, Mrs. Kristine Fogelström and Mrs. Lena Svenre for excellent technical assistance.     

Über Aktivitätsphasen im Epithel des Saccus vasculosus von Etmopterus spinax

Summary The coronet cells in the saccus vasculosus of Etmopterus spinax L. were investigated histologically and histochemically. Inclusions of granula in the coronet cells were observed in 8 specimens. The granules react positivly as lipid and protein containing substances. They are attached to the Einschlußkörper in the cell base and accumulate around the nuclei or in the inner fibre apparatus of the cell apex. The diameter of the granules reaches 2,5 in the cell base and diminishes to less than 1 in the apical structures. These granula are interpreted as a sign of secretion into the third ventricle.This secretory activity is high in gravid females (31,4–54,7 % of the cells) and low in juvenile females or specimens in which no ovulation could be stated (0–4,7 %). Gravid females show differences in the distribution pattern of the granula within the cells, thus indicating a phasic activity.The coronet cells in the saccus vasculosus of Etmopterus seem to be involved in the secretory phenomena related to gravidity.

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Die Untersuchungen wurden mit dankenswerter Unterstützung durch die Deutsche Forschungsgemeinschaft durchgeführt.     

Elektronenmikroskopische Beobachtungen am Saccus vasculosus einiger Knorpelfische

Zusammenfassung Das Epithel des Saccus vasculosus zweier Arten von Selachiern und von einem Holocephalen wurde, nach Perfusionsfixierung der Tiere mit Glutaraldehyd, elektronenmikroskopisch untersucht.Krönchenzellen werden von kleineren Stützzellen unterschieden. Beide Zellarten sitzen der Basalmembran auf und begrenzen apikal das Lumen des III. Ventrikels. Das gut entwickelte agranuläre endoplasmatische Reticulum der Krönchenzellen deutet auf eine hohe biochemische Aktivität hin. Feinstruktur, Gestalt und Lage der Krönchenzellen im Epithel sprechen für ihre Rezeptorennatur. Neben Krönchenzellen ragen einzelne birnenförmige Zellen in das Lumen des Saccus vasculosus der Haie. Zahlreiche Vesikel im apikalen Cytoplasma weisen ebenso wie ein großer Golgi-Apparat auf sekretorische Funktion dieser Zellart. Basisnah splittern Bündel markloser Axone in Einzelfasern auf. Synaptische Verbindungen zwischen Krönchenzellen und solchen Axonen wurden nicht beobachtet.

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Electronmicroscopical observations on the Saccus vasculosus of some Chondrichthyes

Summary The Saccus vasculosus of two species of Selachians and one species of Holocephala was investigated with the electron-microscope after perfusion of the animals with glutaraldehyde.Coronet cells are distinguished from smaller polymorphic supporting-cells. Both types of cells extend from the basement membrane to the lumen of the IIIrd. ventricle. Smooth endoplasmic reticulum is well developed in the coronet cells, and indicates a high biochemical activity. Ultrastructure, shape and position of the coronet cells in the epithelium speak in favour of the receptor hypothesis of these elements. Besides coronet cells occur pear shaped cells which project into the lumen of the Saccus vasculosus of the two species of sharks. Their submicroscopical structure points out secretory function of these cells. Bundles of nonmyelinated nerve fibers split into individual fibers near the basement lamina. Synaptic connections between these axons and the coronet cells have not been observed.

Herrn Prof. Dr. H. Leonhardt danke ich für Themenstellung und zahlreiche Anregungen, Herrn Prof. Dr. Dr. h. c. W. Bargmann für einen Arbeitsplatz im Anatomischen Institut Kiel.     

Seahorse fingerprints: a new individual identification technique

Individual identification is particularly important for ethological studies and sampling design. Previous studies have developed various types of marking by tags and chemical marks, but these methods involve significant manipulation of the individuals. Other studies have reported natural marks as an efficient method for individual recognition. Our study aims to elucidate a new method for natural mark identification of seahorses, which we tested with the endangered Brazilian seahorse Hippocampus reidi. We avoid pseudoreplication by recognizing individuals. Seahorses have a hard bony structure on the top of their heads called the coronet, which has a different shape for each individual, corresponding to a fingerprint. We tagged seahorses in the field with collar tags and photographed their coronets. After two days, we identified seahorses by their photographs and verified their identification with the collar tags. We correctly identified all individuals by fingerprint identification. Although this method was only tested with adults, we suggest that it applies to seahorses in general, as all species possess the coronet structure.     

Sperm ultrastructure in Kellia suborbicularis (Bivalvia: Galeommatoidea: Kellidae)

The sperm cells of Kellia suborbicularis are narrow with a short bullet‐shaped acrosome, a 5.0–5.5 µm long and 0.4–0.6 µm broad nucleus, and a short midpiece with a ring of five mitochondria. The disposition of the subacrosomal substance into a coronet‐like formation is unique, and the sperm structure offers no clue to the relationship between Kellia and other galeommatoidean genera. The possible significance of narrow elongate sperm for their entry into the brood pouch is discussed.