Search for a specific marker in the neural membranes of scrapie mice: a freeze-fracture study
Dubois-Dalcq, M.; Rodriguez, M.; Reese, T.S.; Gibbs, C.J.; Gajdusek, D.C.
Laboratory Investigation; a Journal of Technical Methods and Pathology 36(5): 547-553
1977
ISSN/ISBN: 0023-6837 PMID: 405528 Document Number: 115508
The freeze-fracture technique was used to examine the cerebellum of scrapie-inoculated animals to determine whether this technique could uncover a recognizable scrapie virus and whether neural membranes were modified in a specific manner during scrapie infection. Mice were fixed by vascular perfusion at various times after intracerebral inoculation with normal or scrapie brain suspension. Clinical signs, high titer of virus and spongiform changes in brain were observed 20 wk after inoculation with the scrapie virus. In the cerebellum, vacuoles were preferentially located in the granular layer and readily identified in freeze-fracture replicas. No new structures likely to be the scrapie virus itself were recognized inside the cleaved membranes. In contrast, the limiting membranes of vacuoles lacked the intramembrane particles present on control membranes. The absence of intramembrane particles, which possibly represent plasmalemma proteins, was also evident on membrane fragments of various sizes and shapes floating in the vacuoles and on protrusions connected to the vacuole-limiting membrane. These changes may correspond to a loss of membrane proteins or to a movement of these proteins to other regions of the membrane bilayer. During the incubation period (11-20 wk after inoculation), no significant changes in granule cell membranes were detected in scrapie-inoculated animals vs. control animals inoculated with normal brain. In contrast, astrocytes in scrapie animals show a significant increase in the number of assemblies. These changes probably reflect astrocytic hypertrophy and start as soon as 3 mo. postinoculation when the titer of virus is already high. Thus, scrapie virus may not be integrated into the cell membrane in such a manner that it can be detected with the freeze-fracture technique.