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Genetically encoded fluorescent calcium indicators (GECIs) allow for imaging of neuronal calcium activity with high spatiotemporal resolution. However, to date only very few GECIs were developed in the near-infrared (NIR) range of 700 to 900 nm, where optical scattering and attenuation are minimal in tissue. NIR GECIs generally suffer from low brightness and weak fluorescence responses and are thus not deemed as suitable for in vivo imaging.
NIR-GECO2G is a recently developed GECI with excitation and emission maxima at 678 nm and 704 nm, respectively. Using widefield fluorescence imaging in the live the mouse brain, we demonstrate several-fold improved response magnitude compared to the original NIR-GECO1 variant. We further show several-fold increased NIR-GECO2G brightness levels in Blvra-/- mice, where high concentrations of biliverdin (BV) result from deleting the gene for the enzyme that aids in the breakdown of BV.
Our results show that NIR-GECO2G demonstrates strong in vivo responses to stimuli in mice and can be used over multiple experiments on minutes- to hours-long timescales.
Here the insertion loss of murine skull has been measured by means of a hybrid optoacoustic-ultrasound scanning microscope having a spherically focused PVDF transducer and pulsed laser excitation at 532 nm of a 20 μm diameter absorbing microsphere acting as an optoacoustic point source. Accurate modeling of the acoustic transmission through the skull is further performed using a Fourier-domain expansion of a solid-plate model, based on the simultaneously acquired pulse-echo ultrasound image providing precise information about the skull's position and its orientation relative to the optoacoustic source. Good qualitative agreement has been found between the a solid-plate model and experimental measurements.
The presented strategy might pave the way for modeling skull effects and deriving efficient correction schemes to account for acoustic distortions introduced by an adult murine skull, thus improving the spatial resolution, effective penetration depth and overall image quality of transcranial optoacoustic brain microscopy.
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