Abstract
Over the past few years, the light-gated cation channel Channelrhodopsin-2 (ChR2) has seen a remarkable diversity of applications in neuroscience. However, commonly used wide-field illumination provides poor spatial selectivity for cell stimulation. We explored the potential of focal laser illumination to map photocurrents of individual neurons in sparsely transfected hippocampal slice cultures. Interestingly, the best spatial resolution of photocurrent induction was obtained at the lowest laser power. By adjusting the light intensity to a neuron’s spike threshold, we were able to trigger action potentials with a spatial selectivity of less than 30 μm. Experiments with dissociated hippocampal cells suggested that the main factor limiting the spatial resolution was ChR2 current density rather than scattering of the excitation light. We conclude that subcellular resolution can be achieved only in cells with a high ChR2 expression level and that future improved variants of ChR2 are likely to extend the spatial resolution of photocurrent induction to the level of single dendrites.
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Acknowledgments
We thank Daniela Gerosa-Erni for excellent technical assistance, Nunu Mchedlishvili for software development, and Georg Nagel, Karl Deisseroth and Roger Y. Tsien for essential constructs. The work was supported by the Novartis Research Foundation.
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11068_2008_9025_Fig5_ESM.gif
MOESM1 [Supplementary Fig. 1. Spatial selectivity of spike initiation breaks down at laser powers > 300 µW. A ChR2-transfected CA1 pyramidal cell was stimulated in a line pattern perpendicular to the orientation of the apical dendrite with 10 ms laser pulses at 3 different laser intensities. The soma was positioned in the middle] (GIF 325 kb)
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Schoenenberger, P., Grunditz, Å., Rose, T. et al. Optimizing the spatial resolution of Channelrhodopsin-2 activation. Brain Cell Bio 36, 119–127 (2008). https://doi.org/10.1007/s11068-008-9025-8
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DOI: https://doi.org/10.1007/s11068-008-9025-8