Spatio-temporal 3D Mapping of Mouse Cerebellar Vascularization during Embryonic Development

Despite major advances in the study of cerebellar neurogenesis, cerebellar angiogenesis during embryogenesis remains poorly described. Recent advances in tissue clearing, light-sheet microscopy, and artificial intelligence have increasingly enabled detailed 3D modelling of cerebellar vasculature at early developmental stages. Here, vascular networks in mouse embryos from E11 to birth (P0) were labelled with podocalyxin, aSMA, and PECAM-1 antibodies together with the nuclear marker TO-PRO-3 iodide, cleared, imaged by light-sheet microscopy, and finally modelled and quantitatively analyzed using Imaris and VesselVio. Our mapping reveals that the three main paired cerebellar arteries, the superior (SCA), anterior inferior (AICA), and posterior inferior (PICA) cerebellar arteries, emerge sequentially between E11 and E13 and display significant topographical variability comparable to that observed in humans. Morphometric analysis demonstrates distinct developmental dynamics, with SCA growth proportional to cerebellar expansion, whereas the AICA and PICA exhibit accelerated extension during later embryonic stages. Interestingly, the PICA does not reach the cerebellum before birth, highlighting the question of its contribution to embryonic cerebellar vascularization. The intrinsic vascular network evolves from a rudimentary bilayer at E11 into a highly branched architecture organized around radial penetrating vessels, giving rise to collaterals that progressively colonized the cerebellar parenchyma during foliation and lobulation. These vascular changes temporally coincided with the successive stages of cerebellar neurogenesis, supporting an interplay between vascular and neuronal development. Together, our findings provide the first spatio-temporal three-dimensional atlas of cerebellar vascularization during mouse embryogenesis, establishing a reference framework for investigating cerebellar angiogenesis in developmental and pathological conditions.