ELECTRON MICROSCOPIC OBSERVATIONS OF RAT AND MOUSE CEREBELLUM IN TISSUE CULTURE

Closely ordered stages of myelin formation in cultures of newborn rat and mouse cerebellum, selected by direct light microscopy, were studied with the electron microscope. Electron micrographs of these cultures reveal the presence of neurons, axons, neuroglia, microglia, and ependymal cells. The appearance of the neuron is identical to that previously described in vivo. The neuroglial cell has long, branching processes, and its cytoplasm is characterized by packets of long, narrow fibrils. During myelin formation, a glial cell process surrounds the axon. This process may form an internal mesaxon and may spiral for several turns around the axon. Other glial cell processes may interdigitate with or overlay the innermost process to contribute to the multilamellated structure. The glial processes flatten and the cytoplasmic surfaces of the cell membrane come into contact to form the lamellae of the myelin sheath. These adhesions may be temporarily incomplete as evidenced by sequestered islands of glial cytoplasm among the myelin lamellae. Ultimately, a compact, apparently spiral, myelin sheath is formed. These findings are discussed in relation to in vivo central myelin formation.     

Circular dichroism measurements of benzyl L-aspartate–nitrobenzyl L-aspartate copolymers and their use in detecting and characterizing preferred polymer conformations

We have examined the nature of the circular dichroism band at 330 mμ for a series of copolymers of β-p-nitrobenzyl L -aspartate with β-benzyl L -aspartate. The circular dichroism band arises from an electronic transition in the nitroaromatic group. In order to interpret the effect quantitatively, we employed a simplified statistical treatment and curve fitting for six copolymers. Both approaches gave consistent results, which indicates that the dichroism comes from pairwise interactions between two nitrobenzyl groups. We constructed a molecular model that meets the constraints and requirements of the analyses developed in this paper. In this model, it is proposed that the main chain forms a right-handed α-helix and that nitrobenzyl groups separated by four residues interact with each other.     

Inhibition of purine phosphoribosyltransferases from Ehrlich ascites-tumour cells by purine nucleotides

1. The purine bases adenine, hypoxanthine and guanine were rapidly incorporated into the nucleotide fraction of Ehrlich ascites-tumour cells in vivo. 2. The reaction of 5'-phosphoribosyl pyrophosphate with adenine phosphoribosyltransferase from ascites-tumour cells (K(m) 6.5-11.9mum) was competitively inhibited by AMP, ADP, ATP and GMP (K(i) 7.5, 21.9, 395 and 118mum respectively). Similarly the reactions of 5'-phosphoribosyl pyrophosphate with both hypoxanthine phosphoribosyltransferase and guanine phosphoribosyltransferase (K(m) 18.4-31 and 37.6-44.2mum respectively) were competitively inhibited by IMP (K(i) 52 and 63.5mum) and by GMP (K(i) 36.5 and 5.9mum). 3. The nucleotides tested as inhibitors did not appreciably compete with the purine bases in the phosphoribosyltransferase reactions. 4. It was postulated that the purine phosphoribosyltransferases of Ehrlich ascites-tumour cells may be effectively separated from the adenine nucleotide pool of these cells.     

Microexudates from Cells Grown in Tissue Culture

Cellular substrata of known molecular structure and measurable dimensions can be constructed as transferred films from Langmuir troughs or as adsorbed films. In addition, large molecules in culture media form measurable adsorbates. With the techniques of ellipsometry and surface chemistry it is possible to characterize and measure (within ± 3A) as a function of several parameters a microexudate of molecular dimensions deposited when tissue cultured cells contact certain substrata. The selective attraction of substratum and cell for microexudate has been determined, and the time course of deposition in Eagle's medium is characterized by a rapid initial accretion of material. During this period, microexudate can diffuse several cell diameters and cannot be detected in the culture medium. In Eagle's medium the cells cannot be detached from glass surfaces by versene or trypsin unless the surface of cell or substratum is coated with certain molecules. Trypsin becomes adsorbed to cell surfaces, continues to be enzymatically active on the surface, and digests protein components of microexudate and substratum. Microexudate appears to be a complex mosaic of molecules (including protein) synthesized within or on the surfaces of cells and secreted by cells or transferred from their surfaces to specific substrata. It is proposed that this mosaic plays, on the molecular level, a significant role in cell-to-cell interactions, cell locomotion and adhesion, and the selective application and spreading of cells on various surfaces.     

Production of the Milk Agent in Cultures of Mouse Mammary Carcinoma

Thin sections of tissue cultures grown from tumors of the RIII high-breast-cancer strain mice were studied in the electron microscope. These tissues contain an abundance of particles whose morphology is consistent with biophysical measurement of the milk agent. These particles, found only extracellularly in our cultures, are formed at the cell membrane. The process of formation, as reconstructed from sections, appears to include a thickening and protrusion of the cell membrane which then evolves gradually into a dense sphere and separates from the cell in much the same manner as does influenza virus. The contents of the newly formed body are later rearranged to form a nucleoid within a membranous sac.