Fluorescence Proteins and Time-Lapse Imaging of the Cytoskeleton
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Visualization of cytoskeletal dynamics in real time is of paramount interest in cell biological research. With the aid of
fluorescent-cytoskeletal fusion proteins and enhancement of confocal laser scanning microscopes with high-end objectives and
cell-incubation chambers, high-resolution time-lapse imaging is nowadays possible for long time periods. However, most of
the cytoskeletal dynamics can be detected during short observation periods.
In this chapter, we provide a detailed description for time-lapse imaging of microtubules, neurofilaments, and microfilaments
within primary neurons. We use two primary neuronal cell culture systems for the analysis of different aspects of cytoskeletal
motion and organization: (1) dissociated dorsal root ganglia, which are highly practical to study cytoskeletal dynamics along
their neurites or within the growth cone, and (2) cerebellar slice cultures, which are characterized by their organotypic
morphology even after 30 days in vitro. In particular in these slice cultures Purkinje cells exhibit highly dynamic dendritic
spines within a functional neuronal network.
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