In-Depth Activation of Channelrhodopsin 2-Sensitized Excitable Cells with High Spatial Resolution Using Two-Photon Excitation with a Near-Infrared Laser Microbeam
| Autor: | Samarendra K. Mohanty, Michael W. Berns, Rainer K. Reinscheid, Naoe Okamura, Tatiana B. Krasieva, Xiaobin Liu |
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| Rok vydání: | 2008 |
| Předmět: |
Infrared Rays
Confocal Biophysics Channelrhodopsin Fluorescence Cell Line law.invention Mice 03 medical and health sciences 0302 clinical medicine Optics Channelrhodopsins Two-photon excitation microscopy law Microscopy Animals Humans Organic Chemicals Photobiophysics 030304 developmental biology Neurons Photons 0303 health sciences Microscopy Confocal business.industry Chemistry Lasers Microbeam equipment and supplies Laser Mice Inbred C57BL Temporal resolution business Ion Channel Gating 030217 neurology & neurosurgery |
| Zdroj: | Biophysical Journal. 95:3916-3926 |
| ISSN: | 0006-3495 |
| DOI: | 10.1529/biophysj.108.130187 |
| Popis: | We used two-photon excitation with a near-infrared (NIR) laser microbeam to investigate activation of channelrhodopsin 2 (ChR2) in excitable cells for the first time to our knowledge. By measuring the fluorescence intensity of the calcium (Ca) indicator dye, Ca orange, at different wavelengths as a function of power of the two-photon excitation microbeam, we determined the activation potential of the NIR microbeam as a function of wavelength. The two-photon activation spectrum is found to match measurements carried out with single-photon activation. However, two-photon activation is found to increase in a nonlinear manner with the power density of the two-photon laser microbeam. This approach allowed us to activate different regions of ChR2-sensitized excitable cells with high spatial resolution. Further, in-depth activation of ChR2 in a spheroid cellular model as well as in mouse brain slices was demonstrated by the use of the two-photon NIR microbeam, which was not possible using single-photon activation. This all-optical method of identification, activation, and detection of ChR2-induced cellular activation in genetically targeted cells with high spatial and temporal resolution will provide a new method of performing minimally invasive in-depth activation of specific target areas of tissues or organisms that have been rendered photosensitive by genetic targeting of ChR2 or similar photo-excitable molecules. |
| Databáze: | OpenAIRE |
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