Freely moving imaging

For visualizing fibers in behaving mice custom-built miniscopes (Liberti et al., 2016; Liberti et al., 2017) were positioned above the fiber bundle, focused with an XYZ manipulator, and cemented in place with Metabond (Parkell) under brief isoflurane anesthesia.

Note: Permanent implantation was used for initial experiments in this study, but baseplate attachment is equally viable.

Mice were allowed to recover for 24-48 hrs after attachment, after which imaging was performed in a custom built acrylic arena as mice foraged for Froot Loops (Kellogg’s).

The miniscope GRIN objective had an N.A. of 0.45 for high-efficiency light collection and the imaging area was approximately 800 x 600 µm, allowing visualization of a substantial fraction of the fiber bundle.

Both behavioral video and fiber signals were acquired at 30Hz using custom MATLAB acquisition software.

Neural imaging data were acquired from DAQ and CMOS PCBs from the UCLA Miniscope project (Pnevmatikakis and Giovannucci, 2017).

Illumination intensity was delivered by a 470 nm LED (Luxeon Rebel, Lumileds) at an intensity of 0.1-0.25 mW/mm2, controlled by a D-A interface (National Instruments USB-6009) and BuckPuck LED controller (LEDdynamics, Inc).

Imaging data were motion-corrected with NormCorre73, corrected for minor background light leak using spatial high-pass filtering, and converted to ∆F/F.

Intensity timeseries for each fiber were extracted using manually selected circular ROIs.

Locomotor activity was quantified using manual tracking in ImageJ.