Transplantation of cultured premyelinating oligodendrocytes into normal and myelin-deficient rat brain.

Cultures of oligodendroglial cells at various stages of maturation, from progenitors to maturing oligodendrocytes, were prepared from neonatal rat brain primary cultures and then were prelabeled in the culture dish with the fluorescent dye, fast blue (FB). Single cell suspensions were grafted into normal or myelin-deficient rat brains. The normal as well as the myelin-deficient in vivo environment allowed cell survival, migration, and differentiation. The FB+ cells expressed the oligodendroglial markers, glycerol phosphate dehydrogenase, galactocerebroside, and myelin basic protein. In the normal rat transplanted cells were identifiable at all times studied up to 24 weeks. Extensive migration of FB+ cells was observed in whole-brain sagittal sections. Our results show that the plasticity of oligodendroglia differentiation, extensively studied in vitro, can now be investigated in the normal and myelin-deficient in vivo environment.

Photochemical labeling of bovine pancreatic ribonuclease A with 8-azidoadenosine 3',5'-bisphosphate.

A simple method has been developed for the preparation of 5'-32P-labeled 8-azidoadenosine 3',5'-bisphosphate (p8N3Ap) for use in photoaffinity labeling studies. Irradiation of a complex between p8N3Ap and bovine pancreatic ribonuclease A (RNase A) with light of 300-350 nm led to the covalent attachment of the nucleotide to the enzyme. RNase A could also be labeled in the dark with prephotolyzed p8N3Ap. In either case, the nucleotide reacted with the same tryptic peptide, encompassing amino acids 67-85 of the protein. The site of labeling was determined to be either Thr-78 or Thr-82, both of which are close to or at the pyrimidine binding site of the enzyme. This result is consistent with recent nuclear magnetic resonance and X-ray studies which indicate that 8-substituted adenine nucleotides interact with the pyrimidine binding site of RNase A.