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Main Menu - Block
- Overview
- Anatomy and Histology
- Cryo-Electron Microscopy
- Electron Microscopy
- Flow Cytometry
- Gene Targeting and Transgenics
- Immortalized Cell Line Culture
- Integrative Imaging
- Invertebrate Shared Resource
- Janelia Experimental Technology
- Mass Spectrometry
- Media Prep
- Molecular Genomics
- Primary & iPS Cell Culture
- Project Pipeline Support
- Project Technical Resources
- Quantitative Genomics
- Scientific Computing Software
- Scientific Computing Systems
- Viral Tools
- Vivarium
Abstract
Two-photon imaging and optogenetic stimulation rely on high illumination powers, particularly for state-of-the-art applications that target deeper structures, achieve faster measurements, or probe larger brain areas. However, little information is available on heating and resulting damage induced by high-power illumination in the brain. Here we used thermocouple probes and quantum dot nanothermometers to measure temperature changes induced by two-photon microscopy in the neocortex of awake and anaesthetized mice. We characterized heating as a function of wavelength, exposure time, and distance from the center of illumination. Although total power is highest near the surface of the brain, heating was most severe hundreds of microns below the focal plane, due to heat dissipation through the cranial window. Continuous illumination of a 1mm2 area produced a peak temperature increase of approximately 1.8°C/100mW. Continuous illumination with powers above 250 mW induced lasting damage, detected with immunohistochemistry against Iba1, GFAP, heat shock proteins, and activated Caspase-3. Higher powers were usable in experiments with limited duty ratios, suggesting an approach to mitigate damage in high-power microscopy experiments.