Previous work has shown that endogenous chemical mediators, of which histamine is the prototype, increase the permeability of blood vessels by causing gaps to appear between endothelial cells . In the present paper, morphologic and statistical evidence is presented, to suggest that endothelial cells contract under the influence of mediators, and that this contraction causes the formation of intercellular gaps . Histamine, serotonin, and bradykinin were injected subcutaneously into the scrotum of the rat, and the vessels of the underlying cremaster muscle were examined by electron microscopy . To eliminate the vascular collapse induced by routine fixation, in one series of animals (including controls) the root of the cremaster was constricted for 2-4 min prior to sacrifice, and the tissues were fixed under conditions of mild venous congestion . Electron micrographs were taken of 599 nuclei from the endothelium of small blood vessels representing the various experimental situations . Nuclear deformations were classified into four types of increasing tightness (notches, foldsl closing folds, and pinches. In the latter the apposed surfaces of the nuclear membrane are in contact) . It was found that : (1) venous congestion tends to straighten the nuclei in al groups ; (2) mediators cause a highly significant increase in the number of pinches (P < 0 .001), also if the vessels are distended by venous congestion ; (3) fixation without venous congestion causes vascular collapse . The degree of endothelial recoil, as measured by nuclear pinches, is very different from that caused by mediators (P < 0 .001) . (4) Pinched nuclei are more frequent in leaking vessels, and in cells adjacent to gaps (P < 0 .001) ; (5) mediators also induce, in the endothelium, cytoplasmic changes suggestive of contraction, and similar to those of contracted smooth muscle ; (6) there is no evidence of pericyte contraction under the conditions tested . Occasional pericytes appeared to receive fine nerve endings. Various hypotheses to explain nuclear pinching are discussed ; the only satisfactory explanation is that which requires endothelial contraction .

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Among the techniques which have been reported to stain the surface coat of cells, for electron microscopy, is lanthanum staining en bloc . Similarly, the presence of the cationic dye, Alcian blue 8GX, in a primary glutaraldehyde fixative has been reported to improve the preservation of the surface coat of cells of many types ; however, the preserved coat is not very electron opaque unless thin sections are counterstained . The present paper shows that for several rat tissues lanthanum staining en bloc is an effective electron stain for the cell surface, giving excellent contrast, if combined sequentially with prefixation in an aldehyde fixative containing Alcian blue . The cationic substance cetylpyridinium chloride was found to have a similar effect to that of Alcian blue in enhancing the lanthanum staining of the surface coat material of the brush border of intestinal epithelial cells . The patterns of lanthanum staining obtained for the tissues studied strikingly resemble those reported in the literature where tissues are stained by several standard methods for demonstrating mucosubstances at the ultrastructural level . This fact and the reproduction of the effect of Alcian blue by cetylpyridinium chloride constitute a persuasive empirical argument that the material visualized is a mucopolysaccharide or mucopolysaccharide-protein complex .

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