Perfusion fixation of lungs for structure-function analysis: Credits and limitations

The quality of tissue preservation in lungs fixed by vascular perfusion has been reevaluated. Excised rabbit lungs inflated to 60% of total lung capacity were perfused (zone III conditions) with different but widely used fixatives. The effects of the perfusates on pertinent physiological variables have been assessed by a continuous monitoring, the effects on the pulmonary microstructure by qualitative and morphometric analysis of electron micrographs. Important results include the following. 1) Perfusions with isotonic glutaraldehyde at flow rates within the physiological range produce large increases of perfusion pressure and lung weight that reflect intracellular, interstitial, and intra-alveolar edema. 2) No edema occurs if glutaraldehyde is added to isotonic buffer solutions (total osmolarity 510 mosM). 3) Glutaraldehyde as sole perfusate does not fully eliminate the retractive force of lung tissue. Upon release of transpulmonary pressure the lungs retract by an indeterminable amount. 4) Satisfactory results can be obtained by sequential perfusion with osmium tetroxide and uranyl acetate or glutaraldehyde (510 mosM) followed by osmium tetroxide and uranyl acetate. The latter combination yields optimal preparations to study the alveolar and capillary architecture but causes a hyperosmotic volume loss of lung cells (cell shrinkage).