Two-Photon Calcium Imaging of Spines and Dendrites
Adapted from Imaging in Neuroscience and Development (eds. Yuste and Konnerth). CSHL Press, Cold Spring Harbor, NY, USA, 2005.INTRODUCTION
This protocol describes an approach to two-photon calcium imaging in dendrites and spines of living neurons. The technique can be applied to acute slices from hippocampus, cerebellum, and neocortex, as well as to slice and neuronal cultures. It uses two fluorescence detection channels to provide quantitative estimates of calcium concentration and to minimize the required concentrations of calcium indicator. A method for estimating calcium concentrations from fractional changes in fluorescent light intensity (ΔF/F) is presented, along with two methods for loading neurons with calcium indicator.
