Simultaneous imaging of morphological plasticity and calcium dynamics in dendrites.

The structure and function of the nervous system are intricately connected. To investigate their relationship it is essential to image neuronal structure and function simultaneously with high spatio-temporal resolution. For this purpose, we describe here a two-step strategy. First, to visualize neurons and their entire dendritic arborization in neuronal tissue, we use ballistic delivery or single-cell electroporation of a fluorescent calcium indicator and a red fluorescent dye. Second, dual wavelength wide-field fluorescence microscopy or confocal microscopy enables imaging structural plasticity of dendrites (including filopodia and spines) and calcium dynamics together. We routinely apply this strategy to developing neurons in live tissue, but mature neurons can also be loaded and imaged as described. For labeling cells and setting up imaging equipment, approximately 2 h are required.