Excitatory Cortical Neurons with Multipolar Shape Establish Neuronal Polarity by Forming a Tangentially Oriented Axon in the Intermediate Zone

Abstract

The formation of axon-dendrite polarity is crucial for neuron to make the proper information flow within the brain. Although the processes of neuronal polarity formation have been extensively studied using neurons in dissociated culture, the corresponding developmental processes in vivo are still unclear. Here, we illuminate the initial steps of morphological polarization of excitatory cortical neurons in situ, by sparsely labeling their neuroepithelial progenitors using in utero electroporation and then examining their neuronal progeny in brain sections and in slice cultures. Morphological analysis showed that an axon-like long tangential process formed in progeny cells in the intermediate zone (IZ). Time-lapse imaging analysis using slice culture revealed that progeny cells with multipolar shape, after alternately extending and retracting their short processes for several hours, suddenly elongated a long process tangentially. These cells then transformed into a bipolar shape, extending a pia-directed leading process, and migrated radially leaving the tangential process behind, which gave rise to an "L-shaped" axon. Our findings suggest that neuronal polarity in these cells is established de novo from a nonpolarized stage in vivo and indicate that excitatory cortical neurons with multipolar shape in the IZ initiate axon outgrowth before radial migration into the cortical plate.

Actin Migration Driven by Directional Assembly and Disassembly of Membrane-Anchored Actin Filaments

“…The velocity of actin waves and axonal length were measured using Multi gauge software (Fujifilm). For live-cell imaging of cortical neurons in tissue slices, in utero electroporation was performed as described previously (Hatanaka and Yamauchi, 2013). For further details, see the Supplemental Experimental Procedures.…”

Extracellular and Intracellular Signaling for Neuronal Polarity

“…Because the intermediate progenitor cells lose their polarity, the neurons cannot inherit their polarity and thus must "establish" their polarity again to become mature neurons. First, the newly generated neurons extend multiple neurites that are identical to those in stage 2 cultured neurons, at which point they are defined as MP cells in the lower portion of the intermediate zone (IZ) (119,214,237,249,287,329). Almost 60% of MP cells initially extend a tangential trailing process, which ultimately develops into an axon, and then generate a leading process, which later develops into a dendrite (119,237).…”

“…BP cells migrate toward the cortical plate (CP) and develop into mature pyramidal neurons. The MP-to-BP transition is a critical step in neuronal polarization in vivo (119,158,233,237).…”

“…Most in vivo cells undergo the same initial step of neuronal polarization as cultured neurons: trailing process (future axon) formation (119,237). Recently, several researchers discovered a new type of basal (intermediate) progenitor cell termed basal radial glial cells, specifically in animals with gyrified brains (33,76,198,338).…”

A Phosphatidylinositol Lipids System, Lamellipodin, and Ena/VASP Regulate Dynamic Morphology of Multipolar Migrating Cells in the Developing Cerebral Cortex

“…These cells extend and retract multiple processes dynamically as their somata move slowly (multipolar migration) (Tabata and Nakajima, 2003;Noctor et al, 2004). Eventually, the multipolar cells extend an axon tangentially, transform into bipolar cells, and quickly migrate using the radial fiber as a scaffold (locomotion mode of migration) (Rakic, 1972;Nadarajah et al, 2001;Tabata and Nakajima, 2003;Hatanaka and Yamauchi, 2012). These observations suggest that the process movement of multipolar cells is a biologically important, exploratory behavior resembling that of the growth cones, which sense the microenvironment.…”