The ultrastructural basis of alveolar-capillary membrane permeability to peroxidase used as a tracer

Abstract

The permeability of the alveolar-capillary membrane to a small molecular weight protein, horseradish peroxidase (HRP), was investigated by means of ultrastructural cytochemistry. Mice were injected intravenously with HRP and sacrificed at varying intervals. Experiments with intranasally instilled HRP were also carried out. The tissue was fixed in formaldehyde-glutaraldehyde fixative. Frozen sections were cut, incubated in Graham and Karnovsky's medium for demonstrating HRP activity, postfixed in OsO4, and processed for electron microscopy. 90 sec after injection, HRP had passed through endothelial junctions into underlying basement membranes, but was stopped from entering the alveolar space by zonulae occludentes between epithelial cells. HRP was demonstrated in pinocytotic vesicles of both endothelial and epithelial cells, but the role of these vesicles in net protein transport appeared to be minimal. Intranasally instilled HRP was similarly prevented from permeating the underlying basement membrane by epithelial zonulae occludentes. Pulmonary endothelial intercellular clefts stained with uranyl acetate appeared to contain maculae occludentes rather than zonulae occludentes. HRP did not alter the ultrastructure of these junctions.