C3334

FOXN1^−/− rats were anaesthetized with isoflurane and stereotactically implanted with stainless steel wires (791400, A-M Systems) over the bilateral somatosensory cortices and the bilateral motor cortices. A reference wire was positioned over the cerebellum, and implants were secured with dental cement (Metabond, S399, S371 and S398; Jet Set4 Liquid, Lang Dental, 3802X6). The following stereotactic coordinates were used, relative to Bregma: primary somatosensory cortex (S1BF), anterior–posterior −1.3 mm and lateral 3.3 mm; and primary motor cortex (M1), anterior–posterior +2.5 mm, lateral 2.5 mm. The wires of the implant were secured onto custom-made Mill-Max headpiece adapters (ED90267-ND, Digi-Key Electronics). To initiate the recording, the adapters were connected to the head stage, consisting of a digitizer and amplifier board (C3334, Intan Technologies). Awake, freely behaving animals were tethered to an acquisition board (Open Ephys) with lightweight SPI interface cables (C3206, Intan Technologies). Continuous real-time EEG was recorded with Open Ephys software (version 0.4.4.1; https://open-ephys.org). Data were sampled at 2 kHz and bandpass filtered between 1 and 300 Hz.

Mice and rats were stereotactically implanted with wires (A-M Systems, 791400) soldered to screws (J.I. Morris, FF00CE125) over the bilateral somatosensory cortices as well as a reference wire over cerebellum, and implants were secured with dental cement (Metabond, S399, S371 and S398; also Jet Set4 Liquid, Lang Dental, 3802X6). The following stereotactic coordinates were used, relative to bregma: primary somatosensory cortex (S1BF), AP −1.3 mm and lateral 3.3 mm; primary visual cortex (V1), AP −3.3 mm, lateral 2.5 mm. Implanted wires were integrated into custom-made Mill-Max headpieces (Digi-Key Electronics, ED90267-ND) that could be connected to a head stage, consisting of a digitizer and amplifier board (Intan Technologies, C3334). Awake and freely behaving animals were tethered to an acquisition board (Open Ephys) with lightweight SPI interface cables (Intan Technologies, C3206). Continuous real-time EEG was recorded with Open Ephys software (https://open-ephys.org, version 0.4.4.1). Data were sampled at 2 kHz and bandpass filtered between 1 Hz and 300 Hz. All animals underwent 3–4 hours of continuous EEG recording between the hours of 10:00 and 18:00. Longitudinal data were collected whenever possible (for example, when not precluded by EEG implant loss or failure).

Recordings were made via a 16-channel digitising headstage (C3334, Intan Technologies, USA) connected to a flexible tether cable (12-pin RHD SPI, Intan Technologies, USA), custom built commutator, and Open Ephys acquisition board (OEPS, Portugal). LFP signals were bandpass-filtered from 0.1 to 600 Hz and sampled at 2 kHz in Open Ephys software. Rats implanted with LFP electrodes underwent auditory fear conditioning as described above. However, a tone habituation session of three 30 s tones (5 kHz, 75 dB, 1-min intervals) was also added before conditioning, in order to observe if ERPs were present to an unconditioned tone (NB. LFPs were only recorded during tone habituation in subset of animals (WT n = 5, KO n = 7)). Video recordings were made using Freeze Frame software (15 frames per second, Actimetrics) synchronised with electrophysiological signals using TTL pulses.