The NR2B subunit in NMDA receptors is functionally important during cerebellar granule cell migration

Migration of cerebellar granule cells (CGCs) from the external germinal cell layer (EGL) to the internal granule cell layer (IGL) within the cerebellar cortex is a crucial developmental process. Antagonists to NMDA receptors impair CGC migration significantly, but studies to determine which subunit subtypes control or affect migration have been controversial. Migrating CGCs transiently express NMDA receptor subunit subtypes NR1a plus NR2B. Grafted NR1-/- subunit knockout cells continue to migrate, indicating that the NR1 subunit is not necessary for migration. In the present study, the functional importance of the NR2B subtype in developing cerebellum was investigated using organotypic slice cultures prepared from postnatal day 8 (P8) rats. Slice cultures were labeled with bromodeoxyuridine (BrdU) during the first 20h and then continuously treated with the NR2B-subtype-specific NMDA antagonist, ifenprodil, or the non-specific NMDA antagonist, APV, for 7 days. Cultures were incubated with fluorescently tagged anti-BrdU IgG and the percent of BrdU-labeled CGCs that migrated from the EGL to the IGL during treatment was analyzed using laser confocal microscopy. Migration into the IGL was significantly impaired by treatment with 0.5 and 1.0 microM ifenprodil. Fewer cells had migrated to the IGL in 1.0 microM ifenprodil than in 0.5 microM ifenprodil; there was no significant difference between the percent impairment caused by 1.0 microM ifenprodil and 50 microM APV. Untreated controls had few, if any, CGCs in the EGL at DIV 8. The percent of CGCs remaining in the EGL following treatment with antagonists significantly increased, indicating impairment of migration. In conclusion, the NR2B subunit appears to be necessary for CGC migration.