N-cadherin sustains motility and polarity of future cortical interneurons during tangential migration.

In the developing brain, cortical GABAergic interneurons migrate long distances from the medial ganglionic eminence (MGE) in which they are generated, to the cortex in which they settle. MGE cells express the cell adhesion molecule N-cadherin, a homophilic cell-cell adhesion molecule that regulates numerous steps of brain development, from neuroepithelium morphogenesis to synapse formation. N-cadherin is also expressed in embryonic territories crossed by MGE cells during their migration. In this study, we demonstrate that N-cadherin is a key player in the long-distance migration of future cortical interneurons. Using N-cadherin-coated substrate, we show that N-cadherin-dependent adhesion promotes the migration of mouse MGE cells in vitro. Conversely, mouse MGE cells electroporated with a construct interfering with cadherin function show reduced cell motility, leading process instability, and impaired polarization associated with abnormal myosin IIB dynamics. In vivo, the capability of electroporated MGE cells to invade the developing cortical plate is altered. Using genetic ablation of N-cadherin in mouse embryos, we show that N-cadherin-depleted MGEs are severely disorganized. MGE cells hardly exit the disorganized proliferative area. N-cadherin ablation at the postmitotic stage, which does not affect MGE morphogenesis, alters MGE cell motility and directionality. The tangential migration to the cortex of N-cadherin ablated MGE cells is delayed, and their radial migration within the cortical plate is perturbed. Altogether, these results identify N-cadherin as a pivotal adhesion substrate that activates cell motility in future cortical interneurons and maintains cell polarity over their long-distance migration to the developing cortex.

Dynamics of the leading process, nucleus, and Golgi apparatus of migrating cortical interneurons in living mouse embryos.

Precisely arranged cytoarchitectures such as layers and nuclei depend on neuronal migration, of which many in vitro studies have revealed the mode and underlying mechanisms. However, how neuronal migration is achieved in vivo remains unknown. Here we established an imaging system that allows direct visualization of cortical interneuron migration in living mouse embryos. We found that during nucleokinesis, translocation of the Golgi apparatus either precedes or occurs in parallel to that of the nucleus, suggesting the existence of both a Golgi/centrosome-dependent and -independent mechanism of nucleokinesis. Changes in migratory direction occur when the nucleus enters one of the leading process branches, which is accompanied by the retraction of other branches. The nucleus occasionally swings between two branches before translocating into one of them, the occurrence of which is most often preceded by Golgi apparatus translocation into that branch. These in vivo observations provide important insight into the mechanisms of neuronal migration and demonstrate the usefulness of our system for studying dynamic events in living animals.

Intrinsic and extrinsic mechanisms control the termination of cortical interneuron migration.

During development, neurons migrate from their site of origin to their final destinations. Upon reaching this destination, the termination of their migration is crucial for building functional architectures such as laminated structures and nuclei. How this termination is regulated, however, is not clear. Here, we investigated the contribution of cell-intrinsic mechanisms and extrinsic factors. Using GAD67-GFP knock-in mice and in utero electroporation cell labeling, we visualized GABAergic neurons and analyzed their motility in vitro. We find that the motility of GABAergic neurons in cortical slices gradually decreases as development proceeds and is almost abolished by the end of the first postnatal week. Consistent with this, a reduction of embryonic interneuron motility occurred in dissociated cultures. This is in part due to cell-intrinsic mechanisms, as a reduction in motility is observed during long-term culturing on glial feeder cells. Cell-intrinsic regulation is further supported by observations that interneurons labeled in early stages migrated more actively than those labeled in late stages in the same cortical explant. We found evidence suggesting that upregulation of the potassium-chloride cotransporter KCC2 underlies this intrinsic regulation. Reduced motility is also observed when embryonic interneurons are plated on postnatal cortical feeder cells, suggesting extrinsic factors derived from the postnatal cortex too contribute to termination. These factors should include secreted molecules, as cultured postnatal cortical cells could exercise this effect without directly contacting the interneuron. These findings suggest that intrinsic mechanisms and extrinsic factors coordinate to reduce the motility of migrating neurons, thereby leading to the termination of migration.

Random walk behavior of migrating cortical interneurons in the marginal zone: time-lapse analysis in flat-mount cortex.

Migrating neurons are thought to travel from their origin near the ventricle to distant territories along stereotypical pathways by detecting environmental cues in the extracellular milieu. Here, we report a novel mode of neuronal migration that challenges this view. We performed long-term, time-lapse imaging of medial ganglionic eminence (MGE)-derived cortical interneurons tangentially migrating in the marginal zone (MZ) in flat-mount cortices. We find that they exhibit a diverse range of behaviors in terms of the rate and direction of migration. Curiously, a predominant population of these neurons repeatedly changes its direction of migration in an unpredictable manner. Trajectories of migration vary from one neuron to another. The migration of individual cells lasts for long periods, sometimes up to 2 d. Theoretical analyses reveal that these behaviors can be modeled by a random walk. Furthermore, MZ cells migrate from the cortical subventricular zone to the cortical plate, transiently accumulating in the MZ. These results suggest that MGE-derived cortical interneurons, once arriving at the MZ, are released from regulation by guidance cues and initiate random walk movement, which potentially contributes to their dispersion throughout the cortex.

Intracortical multidirectional migration of cortical interneurons.

It is well documented that most cortical interneurons originate from the basal forebrain and migrate tangentially to the cortex. However, relatively little is known about their migration after their arrival at the cortex. To elucidate the route and mode of intracortical migration of the interneurons, we performed real-time analysis by utilizing glutamate decarboxylase (GAD)67/green fluorescence protein (GFP) knock-in mice and an electroporation-based gene transfer of DsRed into the ganglionic eminence (GE) of a mouse embryo. Cortical interneurons show a diverse mode of migration. In coronal slices, ventrolateral-to-dorsomedial migration predominantly occurs in the lower-intermediate zone. However, a substantial number of interneurons migrate radially either towards the pial or ventricular surface. There are also quiescent neurons. Observations of the marginal zone or the ventricular zone in flat-mounted cortex from the pial or the ventricular surface, respectively, revealed that the interneurons tangentially migrate in all directions. Medial GE-derived interneurons visualized by DsRed electroporation show similar migratory behaviours. Thus, final settlement of cortical interneurons in their destinations may be a result of successive migratory process of different modes within the cortex.