3x3D imaging project

Fast two-photon in vivo imaging and stimulation with simultaneous three-dimensional random-access scanning in multiple brain regions 3x3D imaging

Funder: European Comission Research & Innovation FP7

Project period: 2013.06.01.-2016.05.31.

Project description:

We developed and prototyped a novel three-dimensional two-photon laser scanning microscope which can simultaneously image different brain regions in three spatial dimensions (3x3D system). The volumes imaged could exceed cubic millimeters. Such a 3x3D system is essential in understanding distributed brain computation since it allows the simultaneous recording of neuronal activity in multiple, distant brain volumes that are functionally connected. As each site is to be scanned with sub-millisecond temporal resolution in several hundred locations, the connectivity of distant neuronal microcircuits or even neuronal compartments such as dendrites or axons will be followed.

During the years of the 3x3Dimaging project, we have finished the development of the necessary technologies and built the prototype of the 3x3D microscope product. The project started with a kick-off meeting in Budapest on 07/31/2013 and ended on the same location with a review meeting on 01/17/2017. In the third year we encountered various technical challenges and faced unexpected complications when building the 3x3D microscope prototype. To correct these delays, we asked for a six-month non-cost project extension during which we not only completed the 3x3D microscope prototype but also performed the first biological measurements with the device. Many new, unexpected methods and results were achieved and published during the project in high impact journals such as Science, Nature Communications, Neuron, and Nature Neuroscience. The collaborative scientific research embedded in the project proved to be exceptionally fruitful.

Our technology will cause long lasting changes to Science, since new principles about neuronal circuit functions across distant brain regions will be discovered. To Technology, since the 3x3D system will enable large brain regions to be scanned in 3D at a cellular resolution. To Society, since the effects of retinal vision restoration in animal models of blindness can be tested in higher visual centers at an unprecedented resolution. To Theory, since theories about distributed brain computations can now be directly tested across multiple brain regions.

Project Highlights

  • WP1 Development of new acousto-optical (AO) deflectors for the 3x3D system
  • WP2 Development of the 3x3D microscope
  • WP3 Development of ultrafast lasers for deep imaging and efficient optogenetic activation
  • WP4 Development of control hardware and 3D software
  • WP5 Neurophysiological and pharmaceutical demonstration in vitro and in vivo
  • WP6 Demonstration of simultaneous 3D measurements and photo-activation in vivo at depth
  • WP7 Development of the product prototype
  • WP8 Management
  • WP9 Dissemination, Collaboration and Exploitation



Coordinating person: Dr. Balázs Rózsa


  • Femtonics Kft.
  • Friedrich Miescher Institute, Basel
  • Max Delbrück Center for Molecular Medicine
  • University of Szeged
  • Budapest University of Technology and Economics

Project details:

  • Proposal number: 323945
  • Acronym: 3x3DImaging
  • Duration of the Project: 36 months
  • Strategic objective/theme: ICT-2013.9.2