Distinct Behaviors of Neural Stem and Progenitor Cells Underlie Cortical Neurogenesis

The human cerebral cortex consists of ≈50–60 billion neurons and glia (Pelvig et al., 2007), with each classified into numerous subtypes (Peters and Jones, 1984). Regulation of the mode of cell division is required to properly amplify cell numbers through symmetric divisions and diversify cell types through asymmetric cell divisions. In Drosophila, cleavage plane orientation plays a crucial role in determining mode of division for some, but not all, precursor cell divisions. For example, in neuroblast cells a vertically oriented cleavage within the plane of the epithelium produces symmetric daughter cells, while a horizontal cleavage orientation produces asymmetric daughter cells based on an apico-basal segregation of fate determinants (Doe, 1996). However, Drosophila sensory organ precursor cells also divide vertically within an epithelium, but these divisions produce asymmetric daughter cells based on a planar segregation of fate determinants (Mayer et al., 2005). Embryonic neuroepithelial cells also exhibit various cleavage plane angles during mitosis, and cleavage orientation is widely thought to dictate daughter cell fate in the embryonic neocortex in a manner similar to that described for Drosophila neuroblast divisions (e.g., see Langman et al., 1966; Martin, 1967; Chenn and McConnell, 1995; Haydar et al., 2003). This concept was supported by the finding that fate-determinants such as Numb display an apparent apico-basal pattern of expression that is strongest at the margin of the ventricle where cortical precursor cells divide (Zhong et al., 1996). Furthermore, examination of Numb expression in vitro shows clear segregation of the fate-determinant in dissociated cortical precursor cells prior to division (Shen et al., 2002). Thus, it was proposed that vertically oriented divisions would produce an equal partitioning of fate determinants to newborn daughter cells and, hence, a symmetric fate, while horizontally oriented divisions would produce unequal partitioning and asymmetric daughter cells (Chenn and McConnell, 1995; Zhong et al., 1996). In particular, this model specifies that horizontal divisions at the surface of the ventricle represent asymmetric neurogenic divisions: the apical daughter cell is predicted to remain proliferative in the ventricular zone (VZ), while the basal daughter is predicted to be a neuron that migrates away from the ventricle to the developing cortical plate. However, analyses of cleavage plane orientation in normal developing rodent neocortex have produced varying results (see Landrieu and Goffinet, 1979; Haydar et al., 2003), and some studies have concluded that the number of horizontal divisions at the ventricle may be insufficient to produce the large number of neurons that reside in the cerebral cortex (Sauer, 1935; Smart, 1973; Landrieu and Goffinet, 1979; Huttner and Brand, 1997). Furthermore, because of the previously unappreciated role of embryonic subventricular zone (SVZ) cells in cortical neurogenesis, the cleavage plane orientation of embryonic SVZ precursor cells has not been examined in relation to neurogenesis. This takes on added significance considering that previous studies have reported that most embryonic SVZ cells divide with a horizontal orientation (Smart, 1973; Zamenhof, 1985). We explored the relationship between cleavage plane orientation and daughter cell fate for mitotic neuroepithelial cells in the VZ (referred to as radial glial cells), and intermediate progenitor (IP) cells in the SVZ of the embryonic rat. We demonstrate that radial glial (RG) cells divide primarily with a vertical orientation throughout development, yet switch from producing symmetric daughter cells during early stages of development to producing asymmetric daughter cells at the onset of cortical neurogenesis. Thus, RG cells exhibit a shift in the mode of division that is not accompanied by a change in cleavage plane angle, as commonly thought. We show that RG cells generate IP cells, and that most IP cells divide with a horizontal orientation while producing symmetric daughter cells. We also find that IP cells intermingle with RG cells and divide in the VZ at early stages of cortical development before the SVZ has formed. Thus, the mingling of two distinct progenitor cell types, each with its own mode of division, may better explain the shifting patterns of division and cleavage plane orientation that were previously attributed to a single precursor cell type. Finally, we show in situ that RG cells are the primary form of neural stem cell in the embryonic neocortex since they undergo self-renewing divisions and produce multiple cell types. The identification of two proliferative cell populations, one of which demonstrates neural stem cell properties in the developing brain, has potential for the application of stem cell biology to disease treatment.