Cerebellar granule cell precursors can extend processes, undergo short migratory movements and express postmitotic markers before mitosis in the chick EGL

Cerebellar granule cell precursors (GCPs) form a secondary germinative epithelium, the external germinal layer (EGL) where they proliferate extensively to produce the most numerous cell type in the brain. The morphological sequence of events that characterizes the differentiation of GCPs in the EGL is well established. However, morphologies of individual GCP and their differentiation status have never been correlated. Here, we examine the morphological features and transitions of GCPs in the chicken cerebellum by labelling a subset of GCPs with a stable genomic expression of a GFP transgene and following their development within the EGL in fixed tissue and using time-lapse imaging. We use immunohistochemistry to observe cellular morphologies of mitotic and differentiating GCPs to better understand their differentiation dynamics. Results reveal that mitotic activities of GCPs are more complex and dynamic than currently appreciated. While most GCPs divide in the outer and middle EGL, some are capable of division in the inner EGL. Some GCPs remain mitotically active during process extension and tangential migration and retract their processes prior to each cell division. The mitotically active precursors can also express differentiation markers such as TAG1 and NeuroD1. Further, we explore the result of misexpression of NeuroD1 on granule cell development. When misexpressed in GCPs, NeuroD1 leads to premature differentiation, defects in migration and reduced cerebellar size and foliation. Overall, we provide the first characterisation of individual morphologies of mitotically active cerebellar GCPs in ovo and reaffirm the role of NeuroD1 as a differentiation factor in the development of cerebellar granule cells.