Mvl7000

Eye and body movements were monitored by illuminating the subject with infrared light (830 nm, Mightex SLS-0208-A). The right eye was monitored with a camera (The Imaging Source, DMK 23U618) fitted with zoom lens (Thorlabs MVL7000) and long-pass filter (Thorlabs FEL0750), recording at 100 Hz. Body movements (face, ears, front paws and part of the back) were monitored with another camera (same model but with a different lens, Thorlabs MVL16M23) situated above the central screen, recording at 40 Hz for the experiments in V1 and HPF (Figs. 1 and 2) and 60 Hz for the transectomy experiments (Fig. 3). Video and stimulus time were aligned using the strobe pulses generated by the cameras, recorded alongside the output of a screen-monitoring photodiode and the input to the speakers, all sampled at 2,500 Hz. Video data was acquired on the computer using mmmGUI (https://github.com/cortex-lab/mmmGUI). To compute the singular value decompositions of the face movie and to fit pupil area and position, we used the facemap algorithm^{13 (link)} (www.github.com/MouseLand/facemap, MATLAB version).

Head-fixed pupillometry was recorded as described by Gao et al 2020 (link) (Gao et al., 2020 (link)). Briefly, mice were first acclimated to a custom-built head fixation device for 5 days, 30 min per day. A monochromatic CMOS camera equipped with a macro zoom lens (MVL7000, ThorLabs) and an infrared LED lamp (LIU850A, ThorLabs) were used to collect images from the left pupil at 5 frames per second. The start of camera recording and the delivery of mild aversive stimulus were triggered by TTL signal controlled by Clampex. 5 consecutive air puffs (100 ms, 60 psi) or tail shocks (1s, ~250 μA) were started 5 min after the baseline recording, with the interval between stimulation range from 30-60s. The 29G tubing for air puff was placed 2mm under the tip of the snout so that the air puff won’t cause eye blink of the mouse. The recorded videos were subsequently analyzed using Bonsai 2.4, and changes in pupil size (area) were plotted.

An inverted microscope, Eclipse Ti (Nikon), equipped with a confocal A1R‐MP system (Nikon) was used for the microscopic optical measurements. For the characterization of the light backscattered by the CDD material, a polarized Ar ion laser (wavelength, λ = 488 nm) was used as the light source, while the backscattered signal was collected by a 10 × objective (numerical aperture, N.A. = 0.25) and measured by a photomultiplier. At the beginning of the measurement, the objective is positioned in order to have the focal plane at the surface of the sample (air/sample interface), and this objective position is fixed throughout the overall measurement. Upon sublimation from the sample, the light intensity that is backscattered by the air/sample interface and measured by the detector of the confocal microscope is to decrease because of the increasing spatial gap between the air/sample interface and the fixed focal plane. The sublimated thickness was estimated by the focal depth of the confocal systems: Δz = 2λ/(N.A.)².^[17] The bright field imaging time‐lapse was collected by a DS‐Ri1 color charged‐coupled‐device (CCD) camera (Nikon) in cross‐polarization mode. Analysis was performed by the ImageJ software. QR‐codes were imaged by using either a smartphone camera or a CCD camera (Leica) coupled to a long working‐distance optical system (MVL7000, Thorlabs).