Influence of functional glia on the electrophysiology of Purkinje cells in organotypic cerebellar cultures

Abstract

Previous studies have shown that exposure of organotypic cerebellar explants to cytosine arabinoside (Sigma) for the first five days in vitro drastically reduced the granule cell population and severely affected glial function. Myelination was absent and astrocytes failed to ensheath Purkinje cells. In the absence of astrocytic ensheathment, Purkinje cell somata became hyperinnervated by Purkinje cell recurrent axon collaterals. Recurrent axon collaterals also projected to Purkinje cell dendritic spines. In later studies, exposure of cerebellar cultures to a different formulation of cytosine arabinoside (Pfanstiehl) also affected granule cells and oligodendrocytes but did not compromise astrocyte function. The different susceptibility of astrocytes to the two preparations of cytosine arabinoside (Sigma and Pfanstiehl) has provided the opportunity to examine the electrophysiological properties of Purkinje cells in the presence and absence of functional glia. Ensheathed Purkinje cells in granuloprival cultures exhibit within two weeks in vitro similar passive membrane properties as Purkinje cells in control cultures. Their input resistance is significantly higher and their spontaneous single-unit discharge is significantly lower than that of unensheathed Purkinje cells. This effect suggests that ensheathed Purkinje cells in cytosine arabinoside (Pfanstiehl)-treated cultures are more responsive to the profuse Purkinje cell recurrent axon collateral inhibitory projection to dendritic spines. These studies also show that the presence of functional glia and/or astrocytic ensheathment can be correlated with the development of complex spike activity by Purkinje cells in vitro. Purkinje cells in cultures treated with cytosine arabinoside (Pfanstiehl), which does not compromise astrocytic ensheathment, display membrane conductances and spike activity similar to mature Purkinje cells in control cultures. By contrast, Purkinje cells in cultures treated with cytosine arabinoside (Sigma), and devoid of astrocytic ensheathment, display mainly simple spike activity reminiscent of the type of activity seen in less mature neurons.