Glutamate producing aspartate aminotransferase in glutamatergic perforant path terminals of the rat hippocampus

Summary The enzyme aspartate aminotransferase was demonstrated cytochemically in the rat hippocampus 4, 7, and 14 days after unilateral entorhinal cortex lesion. At the light microscopic level the enzyme showed a significant activity decrease in the ipsilateral entorhinal terminal field which was similar at all postlesion times investigated. Non-denervated areas, i.e. the inner one-third of the dentate gyrus molecular layer and the radiatum layer of CA2/3, showed an increase of aminotransferase activities. At the electron microscopic level in the entorhinal terminal field of the control (unoperated) side aspartate aminotransferase was localized preferentially in a great number of boutons, containing the cytoplasmic and mitochondrial isoenzymes. Following entorhinal lesion a significant loss of these positively reacting boutons was seen. Most of the degenerating boutons contained reaction product but a small number was negative for aspartate aminotransferase. From 4 to 14 postlesion days the positively reacting boutons of the non-denervated supragranular zone expanded outward into the denervated area according to the known terminal proliferation of the commissural and associational systems. The remaining denervated entorhinal terminal field was reinnervated predominantly by negatively reacting boutons (probably terminal proliferations of septal afferents) and by a small number of positively reacting boutons (probably terminal proliferations of the crossed temporodentate pathway). The presence of cytoplasmic aspartate aminotransferase in the terminals of a well-known glutamatergic system is discussed in relation to the possible importance of this enzyme for the production of releasable glutamate.     

An electron microscope study of the perfusion-fixed spleen

Summary By preserving the intercellular or plasmatic space, perfusion fixation provides a particularly clear picture of the histological and cytological constitution of the spleen. An analysis of documents obtained with this technique has contributed new information concerning the organization of the intermediate circulation in that organ, and on the topographical relationships and ultrastructure of the various types of cell which are usually grouped under the common denomination of reticulo-endothelial system.Currently available physiological data concerning the flow of blood through the spleen, together with the observed scarcity of sinuses, and of interstices in the latters' walls, to which must be added the narrowness of these interstices, incline us to question the existence of a circulatory system involving the passage of blood from the arterioles into the pulp cords of Billroth, and thence into the sinuses. Thus we are led to conceive of a system whereby about 97% of the blood entering the spleen would flow directly into the sinuses. Phagocytosis of worn blood cells would be carried out by a quantitatively less important side-stream, running parallel to the main stream, but nevertheless sufficiently fast to ensure that every blood cell should be shunted through the spaces of Billroth at least once every 24 hours.The unitary concept of the RES, as it appears from the bulk of existing literature, is based upon the postulated, but never proven, existence of a single cell type capable of developing distinct morphological characteristics, and of carrying out distinct functions, namely phagocytosis and the laying down of reticulin, depending on its situation in the tissue. An ultrastructural study of the various cells encountered in sections of spleen, and a comparison with their counterparts in other organs, seem to rule out this unitarian view.

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Résumé La fixation par perfusion, en conservant l'espace intercellulaire ou plasmatique, donne une image de l'organisation structurale et cytologique de la rate particulièrement claire. L'analyse des documents aínsí obtenus fournit des données nouvelles sur l'organisation de la circulation intermédiaire et sur la position et l'aspect ultrastructural des cellules habituellement réunies sous le nom de système réticulo-endothélial. Les données de physiologie sur le débit splénique, le nombre de sinus observables et le relativement petit nombre et l'étroitesse des fentes sinusiennes font douter de l'existence d'un système circulatoire faisant passer le sang des artérioles dans les cordons de Billroth et de là dans les sinus. Ceci fait envisager un schéma de circulation faisant passer environ 97% du sang directement à travers le sinus. La phagocytose des éléments à détruire est assurée par une circulation parallèle peu importante, mais suffisante pour que chaque élément sanguin ait chaque jour la possibilité de passer par les cordons de Billroth.La théorie du SRE est basée sur l'existence d'une cellule unique, différant seulement par sa position et assurant la macrophagie et la formation de la réticuline. L'étude de ces différents types de cellules au niveau de la rate et leur comparaison au niveau d'autres organes permet d'assurer qu'aucun argument de morphologie ultrastructurale ne peut l'étayer.

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This work was supported by grant No 4237 of the Swiss National Foundation for Scientific Research.

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We wish to thank Mrs. Sidler-Ansermet for her invaluable technical assistance.     

An electron microscope study of myelin figures

In the electron microscope, thin sections of OsO(4)-fixed myelin figures from the phospholipide fraction of human brain show a pattern of parallel dark lines with a repeating period of about 40 A. It is shown that the dark lines probably represent the reaction product of OsO(4) with double bonds in the fatty acid chains, thereby marking the central portion of one bimolecular lamella. The addition of globin results in dense lines 25 to 50 A wide that cover the surface of the myelin figures. When such a figure consists of only two bimolecular leaflets of lipide covered with globin, the structure shows striking similarity to the image of cell membranes in fixed tissue sections. A hypothetical schema is given of the molecular structure of the figure, and the distribution of OsO(4) in it.     

An electron microscope study of lampbrush chromosomes

Lampbrush chromosomes were isolated from germinal vesicles of oocytes from Necturus maculatus, Triturus viridescens, Pseudotriton montanus and Rana pipiens. After treatment of isolated nuclei with 10 per cent sucrose, chromosomes free of nuclear sap are obtained for examination in either the light microscope or in the electron microscope. For electron microscopy the chromosomes were prepared either by Anderson's critical-point procedure or were embedded in methacrylate and sectioned. The evidence presented in favor of the view that the loops, axis, and the chromomeres of lampbrush chromosomes are formed by two chromonemata is based on the following observations: 1. Treatment of isolated chromosomes with 0.002 M KCN loosens the structure of the loops, and a more or less coiled organization is then observed in most of them with the light microscope. At the electron microscope level, each loop consists of a bundle of microfibrils. The latter are 500 A in diameter, and their complex arrangement within the loops is best studied in stereoscopic preparations. 2. Treatment of chromosomes with 0.002 M KCN also unravels the "chromomeric" regions of the axis. A fibrillar organization then becomes visible in the light microscope. In the electron microscope, wide strands are seen within some chromomeres; their diameter corresponds closely to that of the chromonemata forming the loops associated with the same chromomeres. In thin transverse sections of isolated chromosomes, no special structure is visible in the axial region except random profiles of fibrils similar to those seen in the loops of the same preparations. 3. Two strands sometimes connect adjacent chromomeres. Where gaps exist along the axis, after stretching of the chromosomes, a loop occasionally straddles the break and returns to a chromomere on each side.     

An electron microscope study on in vitro parthenogenesis in sunflower

Summary Electron microscope studies have been conducted on the parthenogenesis induced by in vitro culture of unfertilized ovules of sunflower (Helianthus annuus). In comparison with the state of the egg prior to inoculation, some eggs 5 days after culture show striking ultrastructural changes, which include, among others, nuclear migration, an increase in the number and activity of the organelles, a loss of polarity and wall formation at the chalazal end of the cell. Most of these changes are similar to those that occur normally in the zygote, indicating that parthenogenic development has been triggered in these eggs. Such eggs have been termed activated and are presumed to be capable of undergoing parthenogenesis. The parthenogenic proembryos which result share some features in common with zygotic proembryos. In addition, some parthenogenic proembryos exhibit unique properties not found in zygotic proembryos. These include embryos that consist of two parts differing markedly in density, an inversion of polarity, the frequent occurrence of autophagic vacuoles, the thickening of cell walls, a centripetal growth mode of wall formation, the appearance of an incomplete cell wall, free nuclear division, amitosis and degeneration. We believe that these ultrastructural peculiarities are the effects of in vitro culture.     

An electron microscope study of the yeast Pityrosporum ovale

Summary Cells of Pityrosporum ovale were prepared for electron microscopy by different methods of fixation and embedding, all of them causing some degree of damage to the cells. Apart from the usual organelles seen in other yeast cells, a body was found which showed an electron-dense outer layer and an electron-light centre when stained with permanganate. The cell wall showed layers of different electron-density. Buds were formed at one pole only, leaving a collar on the mother cell.     

An electron microscope study of the regenerating nerve fibers

Summary Guinea pig sciatic nerves were severed in order to obtain a regenerative process of the nerve fibers. The animals were killed at different periods of time after the severing (24 hours, 6 days and 12 days) and the specimens obtained were prepared for electronmicroscopic study.The nerve fiber growing extremities (growing cones) were specially studied. The growing cones showed the following components: a) microvesicles; b) mitochondria; c) multivesicular bodies.The microvesicles are hollow elements of about 200 to 700 Å. They constitute the main component of the growing cone. The mitochondria were seen as elongated bodies of 80 m. They were seen in many cases changing to round dense bodies which appear to break-up in irregularly-shaped fragments.The multivesicular bodies were found present in most of the growing cones. Protoneurofibrils do not exist in the growing cone but a close relationship between microvesicles and protoneurofibrils was found in the segments next to the growing cone.The above-mentioned components were found in all growing cones, disregarding the time elapsed after the nerve severing.Director of the Institute.Assistant of the Department of Neurohistology and Experimental Histology.Head of the Department of Cell Ultrastructure.     

Denervation-induced spatiotemporal upregulation of ephrin-A2 in the mouse hippocampus after transections of the perforant path

Transections of the entorhinal afferent fibers to hippocampus, perforant path (PP), result in the denervation in specific hippocampal subregions, which is followed by a series of plastic events including axon sprouting and reactive synaptogenesis. Many growth-associated molecules are thought to participate in these events. In the present study, we proved the upregulation of ephrin-A2 in the denervated areas of the ipsilateral hippocampus following PP transections. Interestingly, when the elevation of ephrin-A2 reached the maximum axon sprouting in the denervated areas almost finished, implying the possible inhibitory effect of ephrin-A2 on sprouting. In addition, ephrin-A2 expression was observed in synapses during reactive synaptogenesis, suggesting that this molecule might also be implicated in the formation and maturation of synapses in the denervated areas.