Endothelial plasmalemmal vesicles as elements in a system of branching invaginations from the cell surface.

Abstract

In electron microscopy studies of the endothelial vesicles in frog mesenteric capillaries, an accidental observation was made concerning vesicular organization. When tannic acid was added to already fixed tissue, the mordant reached apparently free vesicles in the cytoplasm under conditions in which vesicular movement was excluded and in which the impermeability of the cell membranes was preserved. This indicates a spatial continuity between the vesicles and the cell exterior. It is proposed that cytoplasmic vesicles in endothelial cells are elements of branching, permanent or semipermanent invaginations of the plasmalemma.Palade discovered numerous vesicles in the capillary endothelium and suggested that they ferry small amounts of material between blood and the pericapillary space (1). This picture implies that the vesicles go through a cycle. They are temporarily attached to the cell surface, where they appear as invaginations of the plasmalemma. In this state the vesicular content equilibrates with the extracellular fluid. Subsequently, the vesicles pinch off from the surface and move through the cytoplasm to the opposite cell front where the vesicular membrane fuses with the plasmalemma and the content is discharged to the cell exterior (2). The process is assumed to be bidirectional. Subsequent observations have been interpreted in support of the above hypothesis of "vesicular transport" (3-1). Although some authors have expressed doubts about the hypothesis (12-16), it nevertheless stands as a generally accepted idea, explaining, in particular, macromolecular transport in capillaries.During an ultrastructural study of endothelial vesicles in frog mesenteric capillaries we made an accidental observation that indicates that almost all vesicles are members of a system of invaginations of the plasmalemma. By using tannic acid after aldehyde and osmium fixation to increase membrane contrast, we observed in many cases a connection between vesicles deep in the cytoplasm and the surface. This observation invites the proposal that the cytoplasmic, plasmalemmal membranes in endothelial cells, rather than being organized as freely moving vesicles, represent elements in an organized system of a more permanent or semipermanent nature.The picture we propose reconciles several conflicting interpretations of the behavior of macromolecular tracers in endothelia.MATERIALS AND METHODS The experiments were carried out on mesenteric capillaries of eight frogs (Rana temporaria), each weighing approximately 40 g. The animals, which had been kept for 6-8 months at 40C, were anesthetized by exposure to 5% urethane for about 5 min until mouth respiration had stopped.Preparation for Electron Microscopy. The frog mesenteries were fixed in situ by injecting 8-10 ml of 1% formaldehyde and 1.25% glutaraldehyde in 0.1 M sodium cacodylate buffer (pH 7.3) into the abdominal cavity. After 10 min the mesenteries were excised and left for 20 hr in the above fixative. After a buffer rinse, the tissue was postfixed in 2% OS04 in 0.1 M ...

Visualizing caveolin-1 and HDL in cholesterol-loaded aortic endothelial cells

“…Alternatively, a pathway of endosomal internalization and subsequent retroendocytosis of the HDL particle after enrichment with cholesterol has been suggested in rat hepatoma cells and macrophages by several investigators (5, 30); however, direct visualization of the distribution of HDL in cholesterol-loaded cells has been lacking. The vesicular structure of the endothelial cells and their role in vesicular transport have been questioned for many years (31). People argue that at least some of the vesicles revealed under the electron microscope appeared to be surface membrane invaginations instead of free vesicles; however, due to the limitation of microscopic techniques, there was no consensus until the invention of confocal microscopy in recent years.…”

Endothelial plasmalemmal vesicles have a characteristic striped bipolar surface structure.

“…Chains of two or three vesicles were rare (arrows) and complexes that could be dendritic structures of the type previously described (10)(11)(12) were only occasionally encountered (double alTOWS).…”

Evidence of a Tubular System for Transendothelial Transport in Arterial Capillaries of the Rete Mirabile

“…In support of the existence of tubular systems are results obtained from cryofixation-cryosubstitution experiments in which cellular structures are rapidly immobilized (Frokjaer-Jensen et al 1988). In such tissues, as well as in those fixed in the presence of tannic acid (Bundgaard et al 1979), virtually all the apparent free plasmalemmal vesicular profiles present in endothelial cell cytoplasm were reported to be interconnected, forming an integral part of channel-like structures (Bundgaard et al 1979;Frokjaer-Jensen et al 1988). In addition, in permeability studies it was found that tracer proteins of different molecular weights display differences in permeability coefficients, indicating that transport is compatible with diffusion processes rather than with vesicular bulk quanta movement from one cell front to the other (Bundgaard 1980;Bendayan and Rasio 1996a,b).…”