Fixation of the central nervous system for electron microscopy by aldehyde perfusion: III. Structural changes after exsanguination and delayed perfusion

Abstract

Brain tissue from aldehyde-perfused rats was analyzed with the electron microscope with respect to qualitative structural changes due to exsanguination and delayed perfusion from 5 to 60 minutes. More severe alterations were generally found after 15, 30, and 60 minutes as compared with those after 5 minutes of perfusion delay. The extracellular space was found increased after 15, 30, and 60 minutes of perfusion delay. Triple-layered membranes and myelin sheath structures were generally found resistant to the experimental procedure. Most intracellular organelles changed due to the adverse fixation situation, but changes were not at any stage ubiquitous. Nuclear material generally displayed aggregation. Nuclear membrane and α-cytomembrane cisternae widened. This was especially prominent in glial cells. Sometimes attached ribosomal particles appeared to become detached. Ribosomal aggregates showed tendency for separation into single ribosome-like particles. Golgi membrane complexes formed rounded structures, occasionally with circular continuity. Mitochondria swelled with or without retention of internal material. Some mitochondria exhibited evidence of internal autolysis, while others appeared in ring forms or partially constricted. Particularly in cerebellar Purkinje cells, stacks of small membrane-bound cisternae were associated with α-cytomembrane-bound cisternae. Both types of cisternae were also often associated with constricted mitochondria. The observed structural changes warrant the conclusion that each cell type and also individual cells differ significantly in response to disturbances enforced by the perfusion delay. Fixation criteria can be constructed for each organelle system.