As indicated above, in adult mammals, neurogenesis is most prominent in the SVZ in the walls of the lateral ventricles (), a region highly related to the embryonic SVZ, which as described above is the site of significant IPC proliferation earlier in development. The SVZ contains relatively quiescent NSCs, known as B cells, which give rise to actively proliferating C cells that function as the IPCs or transit amplifying progenitors in the adult brain SVZ (Doetsch et al. 1999b
). Type C cells give rise to immature neuroblasts (A cells), which migrate in chains to the olfactory bulb (Lois & Alvarez-Buylla 1994
, Belluzzi et al. 2003
), where they differentiate into interneurons (Carleton et al. 2003
). Despite their NSC properties, type B cells display ultrastructural characteristics and markers of astroglial cells, including the expression of GFAP, GLAST, and other astroglial markers (Doetsch et al. 1997
, Colak et al. 2008
, Platel et al. 2008
). Thus type B cells are also frequently referred to as SVZ astrocytes.
Figure 5 Schematic of progenitor types and lineages in the adult brain SVZ. NSCs in the wall of the lateral ventricles of adult rodents correspond to type B cells (SVZ astrocytes). These cells retain epithelial properties, including the extension of a thin apical (more ...)
The astrocytic nature of adult SVZ NSCs has now been demonstrated in several studies. Initial evidence indicated that following a six-day treatment with cytosine-β-d-arabinofuranoside (Ara-C), which kills actively dividing type C and A cells, SVZ B cells are activated and regenerate C cells, which in turn give rise to new neuroblasts (Doetsch et al. 1999a
). A replication-competent avian leukosis (RCAS) retrovirus carrying a reporter gene has been used specifically to label mammalian SVZ astroglial cells (and their progeny) in transgenic mice (G-tva) expressing the receptor for this avian retrovirus under the control of the GFAP promoter (Holland & Varmus 1998
). RCAS-labeled SVZ astrocytes generate olfactory bulb neurons (Doetsch et al. 1999b
) (). Further evidence that B cells correspond to the adult SVZ NSCs comes from studies using transgenic mice expressing herpes-simplex virus thymidine kinase (TK) from the GFAP promoter. Ganciclovir (GCV) treatment in these mice leads to the selective death of cells expressing TK and to a severe reduction in the number of new neurons produced in the adult brain and a depletion of neurosphere-forming cells from the SVZ (Imura et al. 2003
, Morshead et al. 2003
). Together these data indicate that NSCs in the SVZ are contained within a population of cells classically considered as differentiated astrocytes.
Recent studies indicate that type B cells retain some important properties of RG. Type B cell bodies are generally located just under the ependymal cell layer but have short processes that extend through the ependymal layer with small apical endings on the ventricle (Mirzadeh et al. 2008
, Shen et al. 2008
). These apical endings form junctional complexes among themselves that are virtually identical to those that join RG earlier in development. These junctional complexes are very different from those that link apical ependymal membranes or B cells to ependymal cells (Mirzadeh et al. 2008
). The ventricular contacts of SVZ B cells can be observed in whole mount preparations of the ventricular surface as small apical surfaces containing a single primary cilium. This feature is very different from ependymal cells, which as mentioned above have very large apical surfaces and contain at least 50 long motile cilia. A novel type of ependymal cells (E2 cells) appears to contain only two long cilia (Mirzadeh et al. 2008
). The function of E2 cells remains unknown. Interestingly, the apical processes of SVZ B cells cluster together in the center of structures resembling pinwheels. The periphery of each pinwheel is formed by the large apical surfaces of ependymal cells.
The SVZ also contains blood vessels that extend a substantial extracellular matrix next to type B and C cells (Mercier et al. 2002
). However, SVZ type B and C cells are also closely associated with blood vessels (Shen et al. 2008
, Tavazoie et al. 2008
). Type B cells that contact the ventricle through their small apical endings have relatively long basal processes, frequently oriented tangentially with specialized end feet on blood vessels (Mirzadeh et al. 2008
). Proliferating SVZ cells are frequently associated with blood vessels (Tavazoie et al. 2008
) or the extracellular matrix around them (Kerever et al. 2007
), which suggests that factors derived from the vasculature may regulate both primary progenitors and IPCs in the SVZ. Adult SVZ B cells thus retain apical-basal polarity and are part of the ventricular epithelium, as were RG earlier in development.
As has been suggested for birds, the walls of the lateral ventricles where neurogenesis continues in the adult contain not only differentiated ependymal cells but also remnants of a VZ (). In the case of the rodent lateral ventricles, the VZ-like structure is present in the center of pinwheels, where adult B cells, like RG, have characteristic end feet. Thus, adult NSCs in the SVZ appear to maintain many epithelial characteristics that allow them to bridge between blood vessels underlying the SVZ and the ventricular surface. Adult stem cells likely integrate information arising at both apical and basal compartments to regulate their proliferation and differentiation, a process mirroring that of RG during development ().
Unlike ependymal cells that line most of the ventricular surface and extend a large number of long motile cilia, B cells have short, single primary cilia extending into the ventricle (Doetsch et al. 1999c
, Mirzadeh et al. 2008
). The function of this organelle in NSCs remains unknown, but as discussed below, recent work indicates that primary cilia are important sites for signal reception, particularly sonic hedgehog (Shh) (Huangfu & Anderson 2005
, Singla & Reiter 2006
). Shh is a critical morphogen and growth factor during development. Shh binding to receptor Patch
results in disinhibition of Smoothened
), an effector of Hh signaling. Conditional ablation of Smo using NestinCre
or an inducible NestinCreERT2
results in postnatal depletion of SVZ B and C cells (Machold et al. 2003
), which suggests that Shh signaling is important for the postnatal maintenance of SVZ B cells and possibly for the regulation of proliferation in type C cells. As described below in more detail, conditional deletion of primary cilia in GFAP-expressing progenitors of the dentate gyrus also results in the postnatal depletion of NSCs. These animals also display decreased proliferation in the SVZ (Y.-G. Han & A. Alvarez-Buylla, unpublished observation), which is consistent with aforementioned observations that Shh plays a critical role in SVZ stem cell maintenance.
No single molecular marker is available to identify adult SVZ primary progenitors. Although SVZ B cells, like embryonic RG, express Nestin and Sox2, they also express GFAP and GLAST, which are more frequently associated with astrocytes. Other cells both within and outside the SVZ also contain some of these alleged NSC markers. Initial experiments suggested that neurospheres, which self-renew in vitro and can generate oligodendrocytes, astrocytes, and neurons (Weiss et al. 1996
), are derived from the SVZ primary progenitors (Morshead et al. 1994
), presumably corresponding to the NSCs in vivo. However, neurosphere-forming ability is not restricted to primary progenitors; both SVZ type B and C cells can generate neurospheres (Doetsch et al. 2002
). Similarly, progenitors for oligodendrocytes can, upon stimulation with growth factors, make multipotent neurospheres (Kondo & Raff 2000
). Lewis X (LeX) is an extracellular carbohydrate expressed by embryonic pluripotent stem cells that is also enriched in a subpopulation of GFAP-expressing SVZ cells (Capela & Temple 2002
). LeX has been used to enrich the population of SVZ cells that can form neurospheres in vitro and may be a possible SVZ stem cell marker. Marker combination has also been employed to isolate SVZ cells with the potential to form neurospheres. For example, a subpopulation of adult VZ cells that is positive for CD133 but negative for CD24 generated neurospheres in vitro (Coskun et al. 2008
). A previous study suggested that multiciliated ependymal cells could function as NSCs (Johansson et al. 1999
). However, other studies indicated that ependymal cells do not generate neurospheres with NSC properties in vitro (Chiasson et al. 1999
, Capela & Temple 2002
) and that ependymal cells become postmitotic during development (Spassky et al. 2005
). Ependymal cells can be distinguished by their large apical surface, long cilia with 9+2 microtubule structure, and S100β staining. In whole mount preparations of the ventricular wall, all cells with a large apical surface stained positive for CD24, and these cells costained with S100β (Mirzadeh et al. 2008
). None of these cells stained positively for proliferative markers or incorporated BrdU. Although ependymal cells are also CD133+, the only cellular profiles in the ventricular wall that are CD133+/CD24− are cells in the center of pinwheels that correspond to the small apical end feet of type B cells. Therefore, CD133+/CD24− cells in the ventricular walls are likely to be type B cells. The overall evidence indicates that ependymal cells are post-mitotic differentiated cells that originate from RG during perinatal development (Spassky et al. 2005
). With age, these cells die. Recent observations indicate that in aging animals some SVZ astrocytes can develop multiple cilia similar to those observed in ependymal cells (Luo et al. 2008
), which further suggests that cells with multiple cilia do not divide.
Better markers may be emerging to define the subpopulation of cells in the adult VZ that function as stem cells. One promising candidate is the nuclear orphan receptor tailless,
which appears to be expressed in SVZ type B cells and to be essential for neurogenesis (Liu et al. 2008
). Markers are also required to identify NSCs better in the adult human brain. It remains controversial whether neurogenesis and migration of neuroblasts to the olfactory bulb continue in adult humans (Curtis et al. 2007
, Sanai et al. 2007
). A prominent band of SVZ astrocytes exists in the adult human brain, in an abventricular location in the walls of the anterior lateral ventricles (Sanai et al. 2004
, Quinoñes-Hinojosa et al. 2006
, Oldham et al. 2008
). Some of these cells can function in vitro as stem cells. Transcriptional analysis suggests that periventricular astrocytes in humans are distinct from parenchymal astrocytes (Oldham et al. 2008
), which may lead to a better definition of the astrocytes in the human SVZ that, like B cells in rodents, continue to function as stem cells in the adult. Although both parenchymal and SVZ astrocytes appear to be directly derived from RG, only SVZ B cells retain expression of critical determinants for their function as NSCs.