Imaging Structure Function of in the Nervous System

Imaging Structure & Function of in the Nervous System Highlights and emerging technologies August 30, 2018 1

Building microscopes… �DIY table-top confocal training kit �Cheap, hands on experience setting up light paths, aligning mirrors and learning the basics of handson imaging 2

Build a 2 -photon microscope 3

New GECIs � Have new set of green GECIs, with different properties j. GCa. MP 7 s is slower and is � very sensitive (good for analyzing synapses) j. GCa. MP 7 f is fast binding j. GCa. MP 7 c has a low baseline fluorescence j. GCa. MP 7 b has a high baseline fluorescence Currently working on j. GCa. MP 8 s Expects most improvement in performance of j. GCa. MP 7 F 4

Sn. FRs � IGlu. Sn. FR is best developed: Have fast, less sensitive version and slower more sensitive version (mimics NMDAR glutamate binding kinetics) GCa. MP 6 f i. Glu. Sn. FR-sensitive i. Glu. Sn. FR-fast � Also have ATP, Ach and GABA Sn. FRs � Lin Tian has recently developed a dopamine sensor 5 David Di. Gregorio et al.

Voltage Sensors �Michael Lin’s ASAP 3 (wants to collaberate) �Janelia’s halo linked channel-rhodopsin with dye 6

Light-sheet microscopy � Laser beam only focused in a single direction (ie. a “sheet”) is used as light source and passed though sample Limits out-of focus fluorescence since illumination is only in thin plane Limited photo-damage Samples are acquired by planes rather than individual points so potentially much faster than scanning microscopy 7

SCAPE microscopy Traditional Light-sheet microscopy has limits tied into the need to either translate the sample through the sheet or to translate both the focusing plane and sheet itself through a statically stationed sample. � SCAPE allows for volumetric imaging at high rates making it ideal for in vivo imaging. � 8

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Innovations in 2 p based optogenetics �Valentia Emiliani’s stuff �Adam Packer’s stuff 10

AOD microscopy �Stepahne Dieudonne’s stuff 11

Spatial light modulators for reducing scattering through tissue �Guide-star method would work for us Single segment illumination (Na Ji) 12

Mini. Scope 13

Suite 2 p � 14

Link to lectures on Google Drive 15