Cerebellar Purkinje cells from the lurcher mutant and wild-type mouse grown in vitro

Lurcher is an autosomal dominant murine mutation. Lurcher heterozygotes (+/Lc) lose all their cerebellar Purkinje cells by adulthood. Cerebellar explants from 2 days postnatal (P2) wild-type (+/+) and +/Lc mutant mice were grown in vitro to investigate the role of local neuronal environment and afferent input in the degenerating +/Lc Purkinje cell. No significant difference was observed between the numbers of Purkinje cells in +/+ and +/Lc explants grown for 10-20 days in vitro (10-20 DIV), as revealed by calbindin-D (CaBP) immunocytochemistry. Quantitative analysis of the morphology of the CaBP immunostained Purkinje cells revealed significantly inferior dendritic development in the cells in the +/Lc explants (15-25 DIV). Wild-type and +/Lc explants were examined in the electron microscope after 10-25 DIV. No difference in ultrastructure was observed between +/+ and +/Lc Purkinje cells grown in vitro. Many features similar to normal Purkinje cell development in vivo were present, including basket and/or stellate cell axon synapses with the dendrites and somata of Purkinje cells, and parallel fibre synapses with the Purkinje cell dendritic spines. Immunocytochemical labelling of parvalbumin and γ-aminobutyric acid (GABA) confirms the presence of basket, stellate and Golgi cell inhibitory interneurons in +/+ and +/Lc cerebellar explants grown in vitro (15 DIV). Electron microscopical analysis of the GABA immunolabelled +/+ and +/Lc explants revealed a complex distribution of the immunostain and not all the basket and/or stellate cell-like structures were immunolabelled. It is proposed that the altered synaptic investment of +/Lc Purkinje cells in culture, and the loss of the climbing fibre input in particular, is responsible for their survival in vitro. Experiments conducted to investigate the influence of the abnormal climbing fibre input on the developing +/Lc Purkinje cell neither confirmed, nor ruled out, a role for this afferent in the degeneration of the cell in vivo.