Neuronal Chain Gangs: Homotypic Contacts Support Migration into the Olfactory Bulb
Authors: Nancy A O'Rourke
Journal: Neuron (1996)

A great deal of attention in the field of developmental neurobiology is presently focused on the rodent olfactory bulb, which processes signals from the nasal epithelium and relays them to the brain. The olfactory bulb is the ultimate destination of an extraordinary population of migrating neuroblasts that have been a source of intense interest for two reasons. First, because these cells are generated well into adult life, they may provide clues to the regenerative capacity of the adult brain and strategies for therapeutic approaches to neurological disorders. Second, these cells migrate in a fashion that differs from the well-characterized glial-guided mechanism employed by radially migrating neurons and the molecular basis for their migration appears to be unfolding at a particularly rapid rate. The present minireview focuses on recent studies that have unveiled the cellular and molecular details of this newly described mode of migration. While olfactory projection neurons (mitral and tufted cells) are born embryonically, the interneurons (granule and periglomerular cells) are generated during the postnatal life of the animal (9Luskin M.B Neuron. 1993; 11: 173-189Abstract Full Text PDF PubMed Scopus (1565) Google Scholar). The neurons originate in the subventricular zone of the cerebral cortex that lies adjacent to the most anterior pole of the lateral ventricle (aSVZFigure 1). Localized injections of a retrovirus containing the lacZ reporter gene into the aSVZ reveal that cells from this region migrate along a distinct rostral pathway out to the olfactory bulb (9Luskin M.B Neuron. 1993; 11: 173-189Abstract Full Text PDF PubMed Scopus (1565) Google Scholar). Once they reach the olfactory bulb, the cells turn radially, migrate outward, and settle into either the granule or periglomerular layers of the olfactory bulb where they differentiate into interneurons. The migrating cells assume the elongated bipolar morphology of migrating neurons, yet are capable of dividing as they migrate (16Rousselot P Lois C Alvarez-Buylla A J. Comp. Neurol. 1995; 351: 51-61Crossref PubMed Scopus (304) Google Scholar; Menezes et al., 1996) and are more correctly termed neuroblasts or neuronal progenitors. While the bulk of this migration occurs during early postnatal stages in the rat, newly formed progenitors continue to migrate in smaller numbers throughout the adult life of the animal (7Lois C Alvarez-Buylla A Science. 1994; 264: 1145-1147Crossref PubMed Scopus (1884) Google Scholar). A major focus of recent investigations into the olfactory interneurons is centered on the mechanism for their migration along the rostral migratory stream. In laminar structures of the brain, such as cortex, cerebellum, and tectum, the prevailing model has young neurons migrating radially along glial processes (15Rakic P Experientia. 1990; 46: 882-891Crossref PubMed Scopus (555) Google Scholar). In contrast, neuroblasts migrating toward the olfactory bulb extend leading processes perpendicular to the radial glia in their surroundings (6Kishi K Peng J.Y Kakuta S Murakami K Kuroda M Yokata S Hayakama S Kuge T Asayama T Arch. Histol. Cytol. 1990; 53: 219-226Crossref PubMed Scopus (50) Google Scholar) and are therefore unlikely to utilize a radial glial-guided mechanism. Because the cells migrate parallel to the pial surface of the brain and across the paths of radial glia, the orientation of their movements is defined as tangential. Some classes of migrating neurons in other areas of the developing brain are found in close apposition to axon tracts and appear to use an axonophilic mode of migration (15Rakic P Experientia. 1990; 46: 882-891Crossref PubMed Scopus (555) Google Scholar). Again, no such preformed axon tracts appear to guide the migrating aSVZ neuroblasts. Thus, the question becomes what substrate, if any, underlies the migration of the interneuron progenitors into the olfactory bulb? A role for the highly polysialated form of the neural cell adhesion molecule (PSA-NCAM) in the migration of olfactory interneuron progenitors was revealed serendipitously from studies of transgenic mice (18Tomasiewicz H Ono K Yee D Thompson C Goridis C Rutishauser U Magnuson T Neuron. 1993; 11: 1163-1174Abstract Full Text PDF PubMed Scopus (434) Google Scholar). NCAM is a member of the immunoglobulin superfamily that is expressed in abundance throughout the nervous system (17Rutishauser U Development (Suppl.). 1992; 116: 99-104Google Scholar). Three forms of NCAM, 120, 140 and 180 kDa in size, are generated through alternative splicing of a single gene product. The 180 kDa form of NCAM is highly polysialated and its adhesive properties differ from the other two isoforms. The PSA residues counteract the adhesive properties of the NCAM molecules and thus serve to attenuate the degree of adhesivity between cells. When the 180 kDa form of NCAM is deleted in transgenic mice, the major defect is a diminution in the size of the olfactory bulb and an accumulation of cells in the aSVZ and rostral migratory pathway. A combined knockout of all three forms of NCAM yields the same phenotype (1Cremer H Lange R Christoph A Plomann M Vopper G Roes J Brown R Baldwin S Kraemer P Scheff S Bathels D Rajewsky K Wille W Nature. 1994; 367: 455-459Crossref PubMed Scopus (883) Google Scholar). Thus, these results suggest that the 180 kDa form of PSA-NCAM plays some role in the migration of neuroblasts into the olfactory bulb. Evidence from subsequent studies suggest that it is the high levels of polysialic acid on the NCAM that are crucial for successful migration into the olfactory bulb (12Ono K Tomasiewicz H Magnuson T Rutishauser U Neuron. 1994; 13: 595-609Abstract Full Text PDF PubMed Scopus (370) Google Scholar). The PSA moieties can be cleaved specifically by the enzyme endoneuraminidase N (endo N) without damaging the remaining NCAM molecule. The high levels of PSA present along the rostral migratory pathway in neonatal mice are removed by injection of the enzyme into the intracranial cavity. Treatment with the enzyme phenocopies the NCAM knockout in mice. As in the knockout mouse, cells accumulate in the aSVZ and there is a reduction in the size of the olfactory bulb suggesting the interneuron progenitors are unable to migrate normally. Additional experiments demonstrate that these effects are not due to either a change in the rate of proliferation or death of the neuroblasts, thus strengthening the argument that it is the process of migration that is being affected specifically. Based on these results, Ono and colleagues (1994) propose that PSA serves to weaken cell–cell interactions and allow the neuroblasts to translocate through their environment. Removal of the PSA would cause the cells to adhere to their substrate and stop in their tracks. The adhesive interactions could involve NCAM, but may also include other as yet unidentified molecules. Further clues to the mechanism of migration of the olfactory interneurons have come from careful examination of the cellular contacts of their progenitors in the rostral migratory stream in the adult rodent brain (16Rousselot P Lois C Alvarez-Buylla A J. Comp. Neurol. 1995; 351: 51-61Crossref PubMed Scopus (304) Google Scholar). As in the neonate, the olfactory neuroblasts express high levels of PSA and immunostaining with an antibody against PSA reveals their migratory route. The progenitors are closely apposed to one another and extend in long thin, interconnected “chains” reaching over distances of several millimeters from the aSVZ into the olfactory bulb. Once within the olfactory bulb, the cells turn radially and disperse, losing their contacts with other migrating neuroblasts. These observations suggest two possibilities: that the chains of cells migrate along an as yet undescribed substrate or that the neuroblasts migrate independent of a defined substrate and that the extensive contacts between them are crucial for their tangential migation. Examination of the rostral migratory stream at the electron microscopy level (8Lois C Garcia-Verdugo J.-M Alvarez-Buylla A Science. 1996; 271: 978-981Crossref PubMed Scopus (1084) Google Scholar) reveals that the chains of elongated neuroblasts are surrounded by glial processes that form a tube-like structure through which they appear to migrate (Figure 2). The progenitors form extensive contacts with each other and make specialized junctions in areas of close membrane apposition that resemble zonula adherens junctions. In contrast, the neuroblasts do not form membrane specializations with the glial processes that encircle them. The close appositions and junctions between the progenitors have led the authors to suggest that it is their mutual interactions that play a prominent role in their migration. The nature of the cellular junctions, however, has yet to be explored fully. The membrane specializations are reminiscent of junctions that have been described between granule neurons and glial processes in vitro (4Gregory W.A Edmondson J.C Hatten M.E Mason C J. Neurosci. 1988; 8: 1728-1738PubMed Google Scholar). In the culture studies, glia processes form different types of junctions with migrating granule neurons than with stationary ones, suggesting a direct correlation between junction subtypes and the migratory state of the neuron. It would be of great interest to know whether the membrane specializations in the neuroblast chains are altered either in number or morphology in NCAM knockout and endo N–treated mice. Another question is whether similar contacts occur in neonatal animals, both among neuroblasts and between neuroblasts and glia. In neonates, GFAP staining has failed to reveal a network of glial processes surrounding the migratory stream (6Kishi K Peng J.Y Kakuta S Murakami K Kuroda M Yokata S Hayakama S Kuge T Asayama T Arch. Histol. Cytol. 1990; 53: 219-226Crossref PubMed Scopus (50) Google Scholar). In addition, larger numbers of olfactory interneurons are being generated and the neuronal precursors form thicker streams more akin to interwined ropes than to chains. Clearly, the cellular contacts in the neonates should be examined in more detail. The issue of whether mutual contacts between the neuronal precursors play a role in their tangential migration into the olfactory bulb has been addressed directly in transplantation studies by Hu and colleagues (1996). Reciprocal transplants of migratory cells between normal and PSA-NCAM mutant animals were performed (Figure 3). The aSVZ cells were transplanted into one of two different sites along the pathway, either into the aSVZ where the cells originate (position 1Figure 3) or into the olfactory bulb where the cells begin to migrate radially into their final laminar positions (position 2Figure 3). In a control experiment, mutant cells transplanted into the aSVZ of mutant animals do not migrate. These results again support the notion that it is the migration of neuroblasts into the bulb that is perturbed specifically in the mutant. When wild-type cells are transplanted into the aSVZ of mutant animals, they also fail to migrate (Figure 3A). In contrast, when the normal neuroblasts are transplanted into a mutant olfactory bulb, they translocate along their normal radial trajectories (Figure 3A). In reciprocal experiments, mutant cells migrate normally along the wild-type rostral migratory pathway and out into their final laminar positions in the olfactory bulb (Figure 3B). Thus, it appears that it is the environment of the cells that is affected by the mutation, not the ability of the cells to migrate. These results, along with those that demonstrate extensive contacts between migrating neuroblasts (16Rousselot P Lois C Alvarez-Buylla A J. Comp. Neurol. 1995; 351: 51-61Crossref PubMed Scopus (304) Google Scholar, 8Lois C Garcia-Verdugo J.-M Alvarez-Buylla A Science. 1996; 271: 978-981Crossref PubMed Scopus (1084) Google Scholar), suggest that cooperative interactions between the progenitors are crucial for this mode of tangential migration. Small numbers of mutant cells in a normal animal may be carried out to the olfactory bulb by the normal cells surrounding them. Conversely, normal cells in a mutant aSVZ may become trapped by neighboring mutant cells that are unable to move. The radial migration of cells within the olfactory bulb is not perturbed in any of the transplant paradigms; thus, the mechanism that underlies the radial movements is unlikely to involve PSA-mediated interactions. The most compelling evidence for homotypic interactions between progenitors playing a mechanistic role in their own migration comes from in vitro studies of aSVZ explants (5Hu H Tomasiewicz H Magnuson T Rutishauser U Neuron. 1996; 16: 735-743Abstract Full Text Full Text PDF PubMed Scopus (317) Google Scholar). When pieces of the aSVZ are cultured in a collagen gel matrix, chains of cells, resembling those described by Alvarez-Buylla and colleagues (16Rousselot P Lois C Alvarez-Buylla A J. Comp. Neurol. 1995; 351: 51-61Crossref PubMed Scopus (304) Google Scholar, 8Lois C Garcia-Verdugo J.-M Alvarez-Buylla A Science. 1996; 271: 978-981Crossref PubMed Scopus (1084) Google Scholar), are observed emanating from the edges of the explants into the matrix. The cells migrating from the explants maintain the high levels of PSA seen in vivo. Removal of the PSA with endo N greatly reduces the number of cells leaving the explants, suggesting they are utilizing a mode of migration similar to that employed by aSVZ neuroblasts in vivo. Interestingly, immunostaining reveals that no glia or axons extend from the explants into the collagen matrix. All the cells in the migrating streams express class III β-tubulin, a marker for neurons in early stages of differentiation (5Hu H Tomasiewicz H Magnuson T Rutishauser U Neuron. 1996; 16: 735-743Abstract Full Text Full Text PDF PubMed Scopus (317) Google Scholar). Thus, these results provide strong support for the notion that the neuroblasts are capable of migrating independently without the support of other cell types. In addition, the PSA residues on their surfaces appear crucial for the contacts that underlie these cooperative interactions. The role of glia in the migration of the olfactory interneuron progenitors has yet to be determined. The results of the explant culture studies (5Hu H Tomasiewicz H Magnuson T Rutishauser U Neuron. 1996; 16: 735-743Abstract Full Text Full Text PDF PubMed Scopus (317) Google Scholar) indicate that glia are not necessary for their migration. Certainly, glia do not serve as a direct migratory substrate, a role that radial glia play in other areas of the brain (15Rakic P Experientia. 1990; 46: 882-891Crossref PubMed Scopus (555) Google Scholar). In the adult olfactory bulb, however, they may contribute to these movements in several ways (8Lois C Garcia-Verdugo J.-M Alvarez-Buylla A Science. 1996; 271: 978-981Crossref PubMed Scopus (1084) Google Scholar). The glial processes that ensheath the migrating neuroblasts may provide a permissive environment for the migration through the adult brain, they may actively guide the cells to the olfactory bulb, or they may restrict them from dispersing into other regions of the brain. Migrating neuroblasts in the neonatal do not appear to be surrounded by glial processes as has been shown in the adult (6Kishi K Peng J.Y Kakuta S Murakami K Kuroda M Yokata S Hayakama S Kuge T Asayama T Arch. Histol. Cytol. 1990; 53: 219-226Crossref PubMed Scopus (50) Google Scholar). Thus, it is possible that the glia play a more active role in the tangential olfactory interneuron migration as the animals mature. An additional question is whether the mechanism described here is applicable to other types of tangential migration in the developing brain. In the tectum, young tectobulbar neurons appear to travel tangentially along axons extended by older members of the same neuronal class (3Gray G.E Sanes J.R Neuron. 1991; 6: 211-225Abstract Full Text PDF PubMed Scopus (129) Google Scholar). While homotypic contacts may support these movements, the underlying mechanism would be defined as axonophilic (15Rakic P Experientia. 1990; 46: 882-891Crossref PubMed Scopus (555) Google Scholar). Both neurons (13O'Rourke N.A Dailey M.E Smith S.J McConnell S.K Science. 1992; 258: 425-427Crossref Scopus (372) Google Scholar) and neural progenitors (Fishell et al., 1993) migrate tangentially in the main body of the developing cerebral cortex, and the mechanisms for their migration remain a mystery. Tangentially oriented neurons in the intermediate zone of the dorsal cortex extend their leading processes orthogonal to the radial glia (14O'Rourke N.A Sullivan D.P Kaznowski C.E Jacobs A.A McConnell S.K Development. 1995; 122: 2165-2176Google Scholar), yet they appear to be dispersed rather than in close contact with one another. These neurons do not travel along the length of glial processes as radially migrating neurons do, but they contact both radial glia and axons along their route and may undergo axonophilic migration. In the ventricular zone, rounded neural progenitors disperse from one another through random movements in the tangential plane of the ventricular zone (Fishell et al., 1993). Both the shapes of these progenitor cells and their mode of movement appear to distinguish them from the olfactory neuroblasts. Thus, neither of these migratory populations appear likely to employ a migratory mechanism similar to that of the aSVZ neuroblasts. A group of neurons that express class III β-tubulin also appear to migrate tangentially through the proliferative zones of the cerebral cortex (10Menezes J.R Luskin M.B J. Neurosci. 1994; 14: 5399-5416Crossref PubMed Google Scholar, 14O'Rourke N.A Sullivan D.P Kaznowski C.E Jacobs A.A McConnell S.K Development. 1995; 122: 2165-2176Google Scholar). The possibility that these cells rely on homotypic contacts for their movement has yet to be explored. Thus, while a homophilic mechanism does not appear to account for all instances of tangential migration, other regions of the brain and spinal cord should be examined closely to determine whether it could be invoked more broadly. Previous investigators have characterized populations of neurons that undergo either glial-guided or axon-guided migration (15Rakic P Experientia. 1990; 46: 882-891Crossref PubMed Scopus (555) Google Scholar). The studies on the olfactory bulb discussed here define a new category of “self-guided” migration based on homotypic interactions that are mediated, in part, by polysialic acid residues on NCAM. Many questions remain to be answered with regard to this mechanism. One wonders about the nature of the cellular behavior within the neuronal chains. Do the cells slide or “leap frog” past each other in a dynamic fashion, or do they maintain static contacts and move like a large conveyor belt? How do cells within the olfactory bulb leave the chains and migrate radially? Finally, what other molecules help drive the cells toward the olfactory bulb? Do diffusible signals attract the cells, or is the environment simply more permissive for movements in the rostral direction? Clearly, the studies in the olfactory bulb will spark further interest into the cellular and molecular cues that guide migrating cells in all regions of the brain.