Two-Photon Imaging of Population Activity with Genetically Encoded Calcium Indicators in Living Flies
- 645 Downloads
Genetically encoded calcium indicators make it possible to track neural activity on a population-wide level. Here we describe a preparation that enables two-photon imaging of neural activity in an essentially intact fly. We present strategies to minimize motion of the brain, both in preparation technique and in apparatus design. We discuss key variables for reducing the problems of photobleaching and phototoxicity in order to collect high quality imaging data. Finally, we discuss approaches to analyze the large quantities of data that can now be readily acquired using the latest generation of genetically encoded calcium indicators.
Key wordsCalcium imaging Two-photon microscopy GCaMP Mushroom body Drosophila Olfaction Memory
K.S.H. is supported by the Crick-Clay fellowship from the Watson School of Biological Sciences and predoctoral training grant 5T32GM065094 from the National Institute of General Medical Sciences. E.G. is supported by the Elisabeth Sloan Livingston fellowship from the Watson School of Biological Sciences. This work was funded by NIH grant R01 DC010403-01A1.
- 5.Tian L, Hires SA, Mao T, Huber D, Chiappe ME, Chalasani SH, Petreanu L, Akerboom J, McKinney SA, Schreiter ER, Bargmann CI, Jayaraman V, Svoboda K, Looger LL (2009) Imaging neural activity in worms, flies and mice with improved GCaMP calcium indicators. Nat Methods 6:875–881PubMedCrossRefGoogle Scholar
- 17.Kanerva P (1988) Sparse distributed memory. MIT Press, Cambridge, MAGoogle Scholar
- 18.Marr D (1969) A theory of cerebellar cortex. J Physiol (Lond) 202:437–470Google Scholar
- 19.Yuste R (2005) Imaging in neuroscience and development: a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NYGoogle Scholar
- 26.Jayaraman V, Laurent G (2007) Evaluating a genetically encoded optical sensor of neural activity using electrophysiology in intact adult fruit flies. Front Neural Circ 1:3Google Scholar
- 28.Murthy M, Turner GC (2010) In vivo whole-cell recordings in the Drosophila brain. In: Zhang B, Waddell S, Freeman M (eds) Drosophila neurobiology methods: a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NYGoogle Scholar
- 30.Bate M (1993) The development of Drosophila melanogaster. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NYGoogle Scholar