Calcium Transients In Single Dendrites And Spines Of Pyramidal Neurons In Vitro

Calcium levels within dendrites and spines rapidly change during synaptic activity and in response to action potential backpropagation following somatic firing. These calcium transients can be measured and quantified by a combination of whole-cell patch-clamp electrophysiology and two-photon calcium imaging methods using membrane-impermeable calcium-dependent indicator dyes. Such methods have been applied both in vivo in the intact brain and in vitro in acute brain slice preparations and neuronal cultures. Here, we describe a dual dye technique, using a calcium-dependent indicator in combination with a morphological calcium-independent marker dye, to measure action potential backpropagation in dendrites and spines. Calcium transients can be correlated with simultaneous electrical recordings made from the cell soma or dendrites to investigate, for example, modulation of both membrane voltage and intracellular calcium signals simultaneously at different spatial locations in the neuron or modulation by pharmacological ligands. We highlight the key advantages of this technique but also note the limitations and caveats of making dynamic measurements in small structures of living neurons.