Delayed genesis of interneurons in the rabbit cerebellum after the end of granule cell genesis

The granule cell population of the cerebellum is generated postnatally from the external germinal layer (EGL). In mammals, the EGL is thought to disappear at different postnatal stages, e.g. after 2 months in rabbits. We have recently shown that a neurogenetic layer called subpial layer (SPL) persists in rabbits between the second and the fifth month of life (Ponti et al., in press). The SPL is structurally different from the EGL, acquiring features proper of adult neurogenetic zones, such as the presence of PSA-NCAM+ chains. Here we describe a population of newly generated cells detectable in the cerebellar cortical layers during the period of SPL existence. Most of these cells co-express PSA-NCAM and doublecortin, and fall into two main morphological types: bipolar and neuronal-like cells, the presence of the latter being restricted to the molecular layer. Using bromodeoxyuridine (BrdU) systemic administration, and BrdU/PSA-NCAM double stainings in confocal microscopy at different survival times (5, 10, 15 days), we show that the majority of early newly generated cells (after 5 days) are bipolar, whereas they are neuronal-like after 15 days. This suggests the existence of a neurogenetic sequence in the cerebellar cortex. To further analyse the fate and the origin of the newlyborn elements, immunocytochemistry for GABA and Pax-2 was performed. Most of neuronal-like cells were also stained for GABA, suggesting their identity as interneurons, although their morphology does not fit with any of the known cerebellar interneuronal cell populations. Accordingly, most of these cells also express Pax-2, a marker considered specific for GABAergic interneurons of neuroepithelial origin (Maricich et al., 1999). These results suggest that the rabbit SPL is linked to neurogenetic processes different from granule cell genesis,