Role Of Reelin And Its Signal Molecules On Neuronal Migration During Cortical Development

The mammalian cerebral cortex is a highly ordered structure composed of different classes of neurons that are arranged in a well-organized six-layered structure ranging from the pial surface to the white matter. During brain development, neurons are generated in the ventricular zone and migrate to their final destinations in the emerging cortical plate (CP). The first cohort of migrating neurons forms the innermost layer of the CP, subsequent cohorts migrating past their predecessors in the CP to take up more superficial positions. This pattern of generating cortical layers, in which the oldest neurons form the deepest layers of the CP and those arriving later are more superficial, is known as inside-out. Defects in neuronal migration are involved in many neuronal disorders such as lissencephaly (smooth brain), epilepsy, mental retardation, and severe learning disabilities. Neuronal migration is precisely orchestrated by different signaling pathways; probably the best characterized signaling pathway controlling neuronal migration is the Reelin signaling cascade. The extracellular matrix molecule Reelin, a large glycoprotein secreted by Cajal-Retzius (CR) cells located in the marginal zone (MZ) during early cortical development, is required to control proper migration and positioning of cortical neurons. Lack of Reelin expression in mice results in the reeler phenotype which is named because of its characteristic "reeling" gait. The cellular architecture of the reeler mouse brain is dramatically disrupted:the cellular layers of the cerebral cortex are disorganized and approximately inverted. The morphology and the orientation of neurons dendrites with inverted axo-dendritic polarity is severely impaired in cortical structures. The mechanisms that underly the neuronal migration, the dendritic development, the development and maintenance of this axo-dendritic polarity are largely unknown.In the present study, we successfully visualized individual migrating neurons and radial glial cells using in utero electroporation with pCAGGFP plasmid transfected at embryonic day14.5(E14.5) and E17.5, respectively. Since Reelin is required in a normotopic position to exert its function, combined with immunocytochemistry against Reelin, we demonstrated that intensively branching of migrating neurons and radial glial cells was closely correlated spatiotemporally with distribution of endogenous Reelin. To investigate the potential function of Reelin signal molecules Cdc42(cell division cycle42) and Dock9(dedicator of cytokinesis9) in the radial migration of cortical neurons, we used in utero electroporation to introduce short hairpin RNAs (shRNAs) into the cortical neurons. We demonstrated that down-regulation of Cdc42in newborn cortical neurons impeded their radial migration. Knockdown of Dock9at E14.5by in utero electroporation of the shRNA-Dock9construct coupled to EGFP mimicked the migrational defect of reeler mice.19.3%of the neurons transfected with the shRNA-Dock9construct had a ventricular orientation of the apical dendrite.