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Functional imaging of hippocampal place cells at cellular resolution during virtual navigation

Abstract

Spatial navigation is often used as a behavioral task in studies of the neuronal circuits that underlie cognition, learning and memory in rodents. The combination of in vivo microscopy with genetically encoded indicators has provided an important new tool for studying neuronal circuits, but has been technically difficult to apply during navigation. Here we describe methods for imaging the activity of neurons in the CA1 region of the hippocampus with subcellular resolution in behaving mice. Neurons that expressed the genetically encoded calcium indicator GCaMP3 were imaged through a chronic hippocampal window. Head-restrained mice performed spatial behaviors in a setup combining a virtual reality system and a custom-built two-photon microscope. We optically identified populations of place cells and determined the correlation between the location of their place fields in the virtual environment and their anatomical location in the local circuit. The combination of virtual reality and high-resolution functional imaging should allow a new generation of studies to investigate neuronal circuit dynamics during behavior.

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Figure 1: Experimental setup.
Figure 2: Imaging CA1 place cells in the dorsal hippocampus.
Figure 3: Place cells differ depending on the running direction in the linear track.
Figure 4: Characterization of place cell calcium transients and place fields.
Figure 5: Place cell activity variability in place fields.
Figure 6: Spatial organization of place cells in dorsal CA1.
Figure 7: Imaging place-related activity in dendrites and putative interneurons.

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Acknowledgements

This work was supported by Princeton University, the National Institutes of Health (grant 5R01MH083686-03), The Howard Hughes Medical Institute, The Helen Hay Whitney Foundation and The Patterson Trust. We thank F. Collman for the brain motion correction algorithm.

Author information

Authors and Affiliations

Authors

Contributions

D.A.D. and D.W.T. designed the research. D.A.D. performed the imaging experiments and developed the chronic hippocampal window system and surgery/training sequences. D.W.T. designed and implemented the combined two-photon microscope and virtual reality instrumentation. D.A.D. and C.D.H. performed extracellular recording and optimized virtual reality training procedures. L.T. and L.L.L. provided AAV2/1-synapsin-1-GCaMP3. D.A.D. analyzed the data. D.A.D. and D.W.T. wrote the paper.

Corresponding authors

Correspondence to Daniel A Dombeck or David W Tank.

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Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–5 (PDF 1432 kb)

Supplementary Movie 1

Z-series movie of the hippocampal region labeled with GCaMP3 virus infection. Movie begins in the external capsule/alveus (dense plexus of fibers) and then steps ventral in 5 micron increments through stratum oriens, stratum pyramidale and then ends in stratum radiatum (350 microns below the external capsule surface). The field is 200×100 microns. (MOV 822 kb)

Supplementary Movie 2

Functional two-photon movie of a field of CA1 neurons in a mouse running back and forth along a virtual linear track. The two-photon time-series was acquired at 15fps and the movie is displayed at 30fps. The virtual linear track is shown at the bottom of the movie and a “^” indicates the position of the mouse along the linear track. This movie corresponds to the data shown in Fig. 2 (the 2 minute time period shown in Fig. 2b). The two-photon movie was made by coloring all of the pixels within a neuron's ROI a red intensity proportional to the value of the significant transient only trace corresponding to each frame (the red color was saturated at 35% changes). This red intensity was added to the still grey-scale image. (MOV 3941 kb)

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Dombeck, D., Harvey, C., Tian, L. et al. Functional imaging of hippocampal place cells at cellular resolution during virtual navigation. Nat Neurosci 13, 1433–1440 (2010). https://doi.org/10.1038/nn.2648

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