Retrograde fluorogold labeling of retinal ganglion cells in neonatal mice

BACKGROUND: The neonatal period, especially postnatal day 10 (P10), is important for mouse retinal ganglion cells (RGCs) development, and an effective labeling technique to track neonatal RGCs is needed. Retrograde fluorogold (FG) labeling is widely used for adult mouse RGCs, but its applicability for the neonatal mouse is still unknown. This study aimed to evaluate the safety and efficiency of retrograde FG labeling in P10 mice. METHODS: The anatomic location of the superior colliculus (SC) of P10 wild-type C57/BL6J mice was clarified by histological brain section and hematoxylin and eosin (H&E) staining. Three doses of 3% FG were injected into the SC of 30 mice, and 3 days post-surgery, labeling efficiency was quantified by retinal flat-mounts, and labeling safety was evaluated by mice mortality. RESULTS: Samples of brain tissue from 2–3.5 mm posterior to the bregma, and from 0.5–2.0 mm lateral to the midline showed major SC-related structures. The FG-positive RGC density in the 0.3 µL group was 3,563.9±311.9 cells/mm(2), significantly more than in the 0.6 µL group (1,718.6±177.1 cells/mm(2)) or 1.0 µL group (2,496.8±342.2 cells/mm(2)). The mortality rate was 10% in both the 0.3 and 0.6 µL groups, but 40% in the 1.0 µL group. CONCLUSIONS: The appropriate labeling site in P10 mice was confirmed and 0.3 µL FG is an appropriate dose for retrograde labeling of RGCs.