Abstract
In many brain areas, circuit connectivity is segregated into specific lamina or glomerula. Functional imaging in these anatomically discrete areas is particularly useful in characterizing circuit properties. Voltage-sensitive dye (VSD) imaging directly assays the spatiotemporal dynamics of neuronal activity, including the functional connectivity of the neurons involved. In spatially segregated structures, VSD imaging can define how physiology and connectivity interact, and can identify functional abnormalities in models of neurological and psychiatric disorders. In the following protocol, we describe the in vitro slice preparation, epifluorescence setup and analyses necessary for fast charge-coupled device (CCD)-based VSD imaging combined with simultaneous whole-cell patch recording. The addition of single-cell recordings validates imaging results, and can reveal the relationship between single-cell activity and the VSD-imaged population response; in synchronously activated neurons, this change in whole-cell recorded Vm can accurately represent population Vm changes driving the VSD responses. Thus, the combined VSD imaging and whole-cell patch approach provides experimental resolution spanning single-cell electrophysiology to complex local circuit responses.
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Acknowledgements
We thank Brian M. Salzberg and the reviewers for critically reading the paper and for useful comments. We also thank Chyze-Whee Ang for his part in developing these techniques. NIH NINDS Grants NS-32403 and NS38572 supported this work.
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Carlson, G., Coulter, D. In vitro functional imaging in brain slices using fast voltage-sensitive dye imaging combined with whole-cell patch recording. Nat Protoc 3, 249–255 (2008). https://doi.org/10.1038/nprot.2007.539
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DOI: https://doi.org/10.1038/nprot.2007.539
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