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To study the biology and repair capacities of mouse oligodendroglial cells, we established cultures of cells purified from neonatal wild-type and 9.6-kb MBP-LacZ transgenic newborn mice cerebral hemispheres as free-floating aggregates in the continuous presence of neurblastoma conditioned medium (N1-B104). In vitro analysis indicated that the initial cell preparations were enriched in oligodendrocyte pre-progenitors that expressed PSA-NCAM and GAP-43 but not GD3, O4, NF68 or glial fibrillary acidic protein (GFAP) markers. These pre-progenitors required increased concentrations of insulin and progesterone to allow their survival in vitro. With time in culture, spheres composed of oligodendrocyte pre-progenitors became oligospheres enriched in oligodendrocyte progenitors expressing GAP-43 and GD3. As well as conserving bipotentiality in vitro, these spheres were able to form myelin in vivo after transplantation into the neonatal shiverer mouse brain. Thus, the oligosphere strategy is a powerful method for generating large populations of mouse oligodendrocyte pre-progenitors and progenitors. The ability to generate oligospheres from transgenic mice will be instrumental in the further dissection of the molecular and cellular mechanisms of myelination and remyelination of the central nervous system. J. Neurosci. Res. 58:735–751, 1999. © 1999 Wiley-Liss, Inc.

QKI proteins are expressed by differentiated glia and have been implicated as regulators of myelination, but are also thought to function during early neural development. This study shows that QKI proteins are expressed in neural progenitors of the ventricular zone (vz) during murine CNS development, but that their expression is down-regulated during neuronal differentiation. By contrast, neural progenitors located in specific subdomains of the vz maintain expression of QKI proteins as they differentiate and migrate away into the emerging nervous system. These QKI+ cells have characteristics consistent with the acquisition of a glial rather than neuronal fate; they express nestin, incorporate BrdU, fail to express neuronal markers, and similar QKI+ cells are found in the postnatal subventricular zone, a known area of gliogenesis. In vitro, neural progenitor cells also down-regulate QKI expression as they differentiate into neurons, but not if they differentiate into glia. Furthermore, neural progenitors in strictly delineated subdomains of the vz dramatically up-regulate expression of the QKI-5 isoform prior to the emergence of QKI+ cells from these regions. Taken together, these data indicate that (1) glia are generated from subsets of neural progenitors found in specific, identifiable subdomains of the vz (2) QKI expression is regulated as neural progenitors undergo the neuron-glial cell fate decision and (3) QKI expression is a characteristic of glial progenitors. J. Neurosci. Res. 54:46–57, 1998. © 1998 Wiley-Liss, Inc.

While the bulk of oligodendendrocytes are generated postnatally in rodents, it is now clear that the first oligodendrocytes are born during midembryonic development. Recent studies imply that the first oligodendrocytes to appear are specified concurrently with certain neuronal subtypes. In addition, patterning molecules known to confer positional information on neural tissues during development, such as sonic hedgehog and bone morphogenetic proteins, have also been implicated in the specification of glial fate. This review discusses some of the recent advances in our knowledge of how oligodendrocytes are generated and the mechanisms by which this might occur in the developing brain and spinal cord.

Myelin basic proteins (MBPs) are major proteins of central nervous system (CNS) myelin, where they facilitate the apposition of cytoplasmic faces of myelin lamellae. Myelin-bearing oligodendrocytes transport MBP mRNA to myelin, where newly translated protein is directly inserted into the myelin sheath. An apparent absence of MBPs in oligodendrocyte perikarya has suggested that MBP localized to the soma is translationally inert. We now demonstrate by confocal immunofluorescence microscopy that not only are MBPs present in the majority of oligodendrocyte perikarya but oligodendrocytes are heterogeneous with respect to their localization of MBPs; MBPs are concentrated in some cells at the plasmalemma and distributed in others throughout the cytoplasm and, surprisingly, the nucleus. MBPs are present in the nuclei of over half of oligodendrocytes in the adult, but in almost all MBP+oligodendrocytes during myelinogenesis. Transport of MBPs into nuclei appears to be a regulated process since some cells exhibit robust MBP accumulation in their cytoplasm but exclude MBPs from their nuclei. We show that oligodendrocyte nuclei contain all four major MBP isoforms, but that in transgenic mice, the epitope-tagged 14 kD MBP isoform preferentially segregates to the plasmalemma. Our data demonstrate that oligodendrocytes are not required to exclude MBPs from their perikarya and suggest that MBPs have a specific function in the oligodendrocyte perikarya and nucleus.

Myelin sheaths develop in the central nervous system (CNS) as elaborations of the processes of oligodendrocytes. Although many details of the spiral wrapping of oligodendrocyte processes around axons and their subsequent transformation into myelin sheaths are known from thin-section electron-microscopic studies, the three-dimensional architecture of the myelin-forming cells is incompletely understood. To characterize this aspect of oligodendrocyte development, we labeled thick (100- to 300-microns) sections of developing murine CNS with oligodendrocyte marker antibodies, recorded individual cells in serial optical sections by confocal microscopy, and created whole-cell reconstructions of oligodendrocytes before and during the initiation of myelination. We distinguish three stages in the maturation of oligodendrocytes, which at all three stages are labeled by the O4, O1 and Ranscht monoclonals and by antibodies against the myelin-specific proteins CNP and myelin basic protein. Premyelinating oligodendrocytes, present before axonal ensheathment begins, emit multiple irregular processes which have predominant radial orientation. These processes, which generally terminate within 50 microns of the cell body, have a surface area 3-8 times or more that of the cell body itself and may represent a mechanism for sampling the local environment of each cell and for identifying target axons. Transitional cells have initiated one or more myelin sheaths; these cells progressively reduce the number of their radial processes as they increase the number of their myelin internodes. The radial processes of each transitional cell are most reduced in parts of the process arbor where ensheathment has begun, suggesting directional control in the elaboration or stability of the radial processes. Mature myelin-bearing oligodendrocytes entirely lack the radial processes and instead emit a few sparsely branching processes which connect cell bodies with myelin internodes. Three-dimensional analysis of the earliest stages in myelin sheath formation reveals two distinct phases. The initiating event in the formation of myelin internodes is the growth of thin unbranched processes, termed 'initiator processes', along axons. The second phase, spiral ensheathment of target axons, begins through the elaboration from each initiator process of lamellar extensions which extend circumferentially around the target axon and thereby form the first turn of its myelin sheath.