Comprehensive and cell-type-based characterization of the dorsal midbrain during development

The layer structure has been intensively characterized in the developing neocortex and cerebellum based on the various molecular markers. However, as to the developing dorsal midbrain, comprehensive analyses have not been intensely carried out, and thus, the name as well as the definition of each layer is not commonly shared. Here, we redefined the three layers, such as the ventricular zone, intermediate zone and marginal zone, based on various markers for proliferation and differentiation in embryonic dorsal midbrain. Biphasic Ki67 expression defines the classical VZ, in which there is clear separation of the mitotic and interphase zones. Next, we mapped the distribution of immature neurons to the defined layers, based on markers for glutamatergic and GABAergic lineage. Interestingly, Tbr2 and Neurog2 were expressed in the postmitotic neurons. We also report that active (phosphorylated) JNK is a useful marker to demarcate layers during the embryonic stage. Finally, we validated the final arrival layers of the migratory glutamatergic and GABAergic neurons. These results form a foundation for analyses of brain development, especially in the proliferation and migration of excitatory and inhibitory neurons in the dorsal midbrain.