In vivo calcium imaging of brain activity in Drosophila by transgenic cameleon expression

Various genetically encoded fluorescent sensors that monitor changes in intracellular calcium concentration have been developed over the last few years. The ability to target these calcium indicators to cells and structures of interest makes them valuable tools for diverse applications and gives them distinct advantages over conventional fluorescent dyes in transgenically tractable organisms. In particular, the cameleon calcium sensors have been used successfully in a number of applications. For example, we use cameleon-2.1 to monitor in vivo brain activity in Drosophila. However, using cameleons to image intracellular calcium concentration changes in vivo is still evolving and is by no means a standard technique. Experimental details and "tricks" for dealing with equipment, techniques, and data evaluation are still restricted to a few laboratories. In this protocol for calcium imaging in Drosophila brain using cameleon-2.1, we provide guidelines to the basic principles of this novel technique in Drosophila neuroscience and, more generally, to the broad field of signal transduction research.