Fig 1: Lineage-tracing of cerebellar neurons arising from Olig3+ progenitor cells.(A–D) Analysis of Tomato+ (red) cells in Olig3creERT2/+;Rosa26lsl-tdT/+ mice that were recombined with tamoxifen (TAM) at E10.5 and imaged at E19. See Figure 2—figure supplement 1A and Video 1 for a description of the genetic strategy and a complete reconstruction of a recombined Olig3creERT2/+;Rosa26lsl-tdT/+ brain. (A, A’) A sagittal three-dimensional reconstruction of the cerebellum. Tomato+ cells were broadly distributed across the cerebellum and densely packed in the DCN nuclei (asterisks in A’). (B) A coronal three-dimensional reconstruction of the cerebellum. DCN nuclei are marked with asterisks. The pontine nuclei (Pn) and their axons (arrowhead), which develop from Olig3+ progenitor cells in the medulla oblongata, are labeled with Tomato. (C) A sagittal section stained against Tomato and DAPI (blue). Other known Olig3 derivatives, such as the thalamus (Th) including its projections to the cortex (Cx), pontine nuclei (Pn), inferior olive (Io) and many neurons in the medulla oblongata (Mo) are marked with Tomato. A magnification of the cerebellum is displayed with (C’) or without (C’’) DAPI. The external granule cell layer (EGL), Purkinje cell layer (PL), and DCN neurons are labeled with Tomato. (D) Left, schematic display of DCN nuclei positive for Brn2 (dentatus and interpositus) and Tbr1 (fastigii). Right, sagittal cerebellar sections stained against Tomato and Brn2 or Tbr1 (green). The boxed areas are illustrated to the bottom of the main photographs displaying individual and merged fluorescent signals. Asterisks mark double positive cells. (E–H) Analysis of cerebellar neurons with a history of Olig3 (βgal+) expression. Olig3creERT2/+;MaptnLacZ/+ mice were recombined with tamoxifen (TAM) at different embryonic stages and analyzed at E19. See Figure 2—figure supplement 1B for a description of the experiment. Sagittal cerebellar sections from these mice were stained against βgal (red) and markers for the EGL and granule cells (Pax6, blue in E), DCN neurons (Tbr1, blue in F), Purkinje cells (Foxp2, blue in G), and inhibitory interneurons (Pax2, blue in H). Double-positive (βgal+/marker+) cells were quantified at E19. The boxed areas on the micrographs are illustrated to the bottom of the main photographs displaying individual and merged fluorescent signals. The mean and SD are plotted in all graphs, and the dots represent the mean of individual animals. n = 3 mice per age. Significance was obtained using one-way ANOVA followed by post hoc Tukey’s test, see Table 2 for statistical details. Photomicrographs were acquired using the automatic tile scan modus (10% overlap between tiles) of the Zeiss LSM700 confocal microscope. The main microphotograph displayed in G was mounted on a black frame to maintain figure panel proportions.Figure 2—source data 1.Source data for Figure 2.