Resistance to Disruption of Multilamellar Fragments of Central Nervous System Myelin

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Abstract
Single-bilayer vesicles of myelin are desirable for studying myelin development and metabolism. A procedure for vesiculating myelin (Steck et al., 1978) was found. X-ray diffraction analysis was used to examine these putative vesicle preparations because much larger amounts of material can be surveyed by this method than by EM. The sharpness (width) of the rings in the X-ray diffraction pattern varies inversely with the number of bilayers per multilayer structure. The diffuse diffraction pattern characteristic of single bilayers was expected. Diffraction patterns were recorded from isolated rat brain myelin before and after the vesiculation procedure. Both patterns showed sharp rings, indicating numerous multilayered structures. Average values ranging from 7-10 bilayers per multilayer were calculated in both cases. This procedure did produce a small fraction of a single-bilayer structures, which were isolated by differential centrifugation; however, these accounted for only .apprx. 1% of the total myelin present. The diffraction pattern of this material showed the diffuse band typical of single-bilayer structures, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated it had the same protein composition as in normal myelin. Similar results were also obtained using either fresh or frozen bovine brain myelin. Variations of the published vesiculation procedure (incubation in 0.1 M NaCl or in buffers containing glycerol; disruption by sonication or use of a Tissumizer) also were not effective in breaking down the multilamellar fragments into thinner structures. Evidently, the multilamellar fragments of isolated CNS myelin resist disruption into single-bilayer structures.