Late embryonic and postnatal cerebellar folial surface area expansion promotes cerebellar cortical cytoarchitectural lamination. We developed a streamlined sampling scheme to generate unbiased estimates of murine cerebellar surface area and volume using stereological principles. We demonstrate that during the proliferative phase of the external granule layer (EGL) and folial surface area expansion, EGL thickness does not change and thus is a topological proxy for progenitor self-renewal. The topological constraints indicate that during proliferative phases, migration out of the EGL is balanced by self-renewal. Progenitor self-renewal must, therefore, include mitotic events yielding either 2 cells in the same layer to increase surface area (β-events) and mitotic events yielding 2 cells, with 1 cell in a superficial layer and 1 cell in a deeper layer (α-events). As the cerebellum grows, therefore, β-events lie upstream of α-events. Using a mathematical model constrained by the measurements of volume and surface area, we could quantify inter-mitotic times for β-events on a per-cell basis in post-natal mouse cerebellum. Furthermore, we found that loss of CCNA2, which decreases EGL proliferation and secondarily induces cerebellar cortical dyslamination, shows preserved α-type events. Thus, CCNA2-null cerebellar granule progenitor cells are capable of self-renewal of the EGL stem cell niche; this is concordant with prior findings of extensive apoptosis in CCNA2-null mice. Similar methodologies may provide another layer of depth to the interpretation of results from stereological studies.

Postnatal cerebellar folial surface area expansion promotes cerebellar cortical cytoarchitectural lamination. We developed a stream‐lined sampling scheme/work‐flow to generate objective/unbiased estimates of cerebellar surface area and volume using stereological principles. Our data demonstrate that during the proliferative phase of EGL and folial surface area expansion, EGL thickness does not change and thus is a topological proxy for progenitor self‐renewal. These topological constraints indicate that during proliferative phases, migration out of the EGL is balanced by self‐renewal. Progenitor self‐renewal must therefore include mitotic events yielding either two cells in the same layer to increase surface area (b‐events) and mitotic events yielding two cells, with one cell in a superficial layer and one cell in a deeper layer (a‐events). Using a mathematical model constrained by these macroscopic observations, we were able to relate easily‐obtained global measurements of volume and surface area to the underlying cellular physiology. We used this overall method to quantify the lengthening of inter‐mitotic times for b‐events on a per‐cell basis. Similar methodology may prove useful by providing another layer of depth to the interpretation of results from stereological studies.