NP Ultra & Psychedelics Stimuli and Optotagging protocol

Spontaneous Activity

Each session includes at least three epochs of spontaneous activity. During this epoch the monitor is held at mean luminance gray and there is no patterned stimulus presented. This provides a valuable time that can be used as a baseline comparison for visually evoked activity. Epochs 0 - 2 are each 20 minutes long. During saline injection experiments, two additional spontaneous epochs of 10 minutes each are included at the end.

Receptive Field Mapping with Gabor patches

The gabor patch stimulus consists of sinuisoidal gratings that are spatially restricted. Key parameters include orientation (in degrees, eight orientations), temporal frequency (4 Hz), spatial frequency (0.08 cycles/deg), contrast (in percent), patch size (20 degrees), and patch location (in degrees, 24 locations). Each trial is presented for 0.25 seconds. Epochs 0 - 2 are each 20 minutes long. During saline injection experiments, one additional receptive field mapping epoch of 10 minutes in length include at the end between two 10 minute spontaneous activity epochs.

Optotagging

Due to the nature of recurrent connections in the cortex, photo-excitation of glutamatergic neurons may lead to feedforward excitation of post-synaptic partners that may make precise identification of optotagged neurons difficult. To avoid this, the NP Ultra & Psychedelics optotagging protocol utilized glavonometer-controlled X/Y mirrors to precisely focus and target 470 nm (Sim1-Cre;Ai32) or 630 nm for (Tlx3-Cre;Ai167) laser light to the cortical surface.

Laser stimuli and power consisted of spot on the cortex of ~15 mW with a 200 \(\mu\)m circular cross-section. The laser was directed at random to predefined locations that corresponded to probe insertion sites to activate neurons near the Neuropixel probes.

Two different laser stimuli were used to asses photoactivation in targeted neurons: a 10 ms and 200 ms square-wave pulse. Intervals between stimuli were drawn from at random from distribution with a mean of 1.0 seconds.