During the first year of this grant substantial progress has been made in setting up required neuroscientific set-ups such as 2-photon imaging (2PI) in combination with spatial light modulation. Advances in technical development specific to the Plenz lab were the following: With respect to 2-photon imaging, we implemented a robust expression and use of last generation genetically encoded calcium indicators (GECIs), specifically jGCaMP7s and YC2.0. We also upgraded our current imaging laser to include wavelengths >1000 nm for future use of red-shifted GECIs. Furthermore, we are now able to synchronize our 2PI system with optical stimulation, pupillometry and locomotion measures for routine recordings. With respect to our Spatial Light Modulation (SLM)/optical stimulation, we optimized injection and expression procedures for red-shifted opsins (C1V1, Chrimson) and demonstrated functional opsin expression in vivo in the awake mouse using wide-field stimulation of opsins with simultaneous 2PI (Abstract submitted to SfN 2019). Furthermore, we implemented and aligned a custom SLM path with our existing 2PI set-up for in vivo optical stimulation. Regarding software control, we established software control of SLM with our existing Thorlabs imaging software for stimulus location and duration control. We also established in vitro acute slice recording set-up to improve opsin screening and implemented hardware/procedures to functionally identify primary visual cortex in wake mice.
