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 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 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 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.     

The use of backscattered electrons to examine selectively stained nerve fibers in the scanning electron microscope

Selective staining of neuronal tissues using standard light microscopic techniques has been combined with backscattered electron scanning electron microscopy. This technique allows neurons to be readily distinguished from their surrounding tissues and examined at high resolution. The technique overcomes some of the problems involved in scanning electron microscopy of nervous tissue in situ.     

An electron microscope study of sunflower crown gall tumor

Summary The inception and developmental phases of sunflower (Helianthus annuus) crown gall tumor growth induced byAgrobacterium tumefaciens were studied submicroscopically. Bacteria were found only in the inception phase. Chloroplasts from tumor cells of both phases showed several kinds of structural variation. Other cytoplasmic organelles remained more or less normal.This investigation was supported in part by an American Cancer Society Institutional Grant.