Simultaneous two-photon calcium imaging at different depths with spatiotemporal multiplexing

Nat Methods. 2011 Feb;8(2):139-42. doi: 10.1038/nmeth.1552. Epub 2011 Jan 9.


In vivo two-photon calcium imaging would benefit from the use of multiple excitation beams to increase scanning speed, signal-to-noise ratio and field of view or to image different axial planes simultaneously. Using spatiotemporal multiplexing we circumvented light-scattering ambiguity inherent to deep-tissue multifocal two-photon microscopy. We demonstrate calcium imaging at multiple axial planes in the intact mouse brain to monitor network activity of ensembles of cortical neurons in three spatial dimensions.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Brain / metabolism
  • Brain Chemistry
  • Calcium / analysis*
  • Calcium / metabolism
  • Mice
  • Microscopy, Fluorescence, Multiphoton / instrumentation
  • Microscopy, Fluorescence, Multiphoton / methods*
  • Time Factors


  • Calcium