K252a, a potent inhibitor of protein kinases, inhibits the migration of cerebellar granule cells in vitro.

In order to elucidate the cellular mechanisms of migrating neurons, we developed an assay system in vitro, using an aggregation culture of developing granule cells from the rat cerebellum. This assay system allowed us to eliminate the effects of various factors other than neurons and to examine the direct effects of individual molecules on neuronal migration. In this assay system, we examined the effects of several protein kinase inhibitors on cerebellar granule cell migration, and revealed that K252a, an inhibitor of protein kinases and of the actions of neurotrophins, inhibited the migration. Within 5 min after the addition of K252a to the culture medium, most of the migrating spindle-shaped cells changed into non-migrating large and polygonal cells, which had many microspikes. Staining with rhodamine-phalloidin revealed the appearance of actin bundles that resembled stress fibers within these large cells. On the other hand, extension of neurites was not severely inhibited by the addition of K252a. These results suggest that the migration is regulated by a different mechanism from that of neurite growth.