Journal of Neuroscience, Vol 7, 1370-1383, Copyright © 1987 by Society for Neuroscience
Immunocytochemical and electrophysiological differentiation of rat cerebellar granule cells in explant cultures
PE Hockberger, HY Tseng and JA Connor
We have used a combination of immunocytochemical and electrophysiological
measurements to monitor the differentiation of cerebellar granule cells in
vitro. We present immunocytochemical evidence showing that several
characteristic features of developing rat cerebellar tissue were retained
in postnatal explant cultures. Most notably the cultures expressed
radiating GFAP-positive (Bergmann) glia processes, proliferating
NSE-negative neuroblasts, and migrating NSE- positive granule cells. The
latter were subdivided into 3 developmental stages--i.e., immature,
intermediate, and mature granule cells, based upon cell differences in
location from the explant, intensity of NSE staining, excitability, and the
amplitude of voltage-dependent conductances. Immature cells were
identifiable during the first week in culture and were located up to 140
micron from the explant. These cells stained lightly for NSE and displayed
conductances of insufficient magnitude to generate action potentials.
Intermediate cells were present after 1-2 weeks in culture and were located
up to 500 micron from the explant. These cells were also NSE positive and
were characterized by the presence of soma action potentials. Intermediate
cells displayed 3 large voltage-dependent conductances: a transient,
TTX-sensitive inward current; a delayed, TEA-sensitive outward current; and
a transient, 4AP-sensitive outward current. Mature cells were present after
1 month in culture and, like intermediate cells, were no more than 500
micron from the explant. However, mature cells stained more intensely for
NSE, and the somata of these cells were devoid of voltage-dependent
conductances (although axonal currents were usually present). These results
indicate that granule cells undergo a stereotypic sequence of
differentiation in postnatal explant cultures. These stages may correspond
to developmental changes in granule cells during migration into the
(internal) granular cell layer in vivo.
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