Am 3000

A recovery time of 1 week was maintained prior to the first recording session. Cats were trained over 2–3 days to remain in head-restrained position for 2–4 h and cycle through periods of quiet wakefulness, NREM, and REM sleep. Recordings were obtained over two weeks following the post-operative recovery period. All recordings were conducted within a Faraday chamber using AM 3000 amplifiers (A-M Systems), bandpassed 0.1 Hz to 10 kHz with a 1 k gain. Respiration was recorded using a thermocouple (TAC80B-K, Omega) placed near the animal’s nostrils. Exhalation and inhalation phases were determined according to temperature changes produced near the nostril. Exhalation was recorded by the thermocouple as a temperature-dependent voltage increase and inhalation as a decrease. All signals were sampled at 20 kHz and digitized with PowerLab (ADInstruments—Data Acquisition Systems for Life Science, RRID:SCR_001620).

The EEG/EMG signals were amplified (gain ×1000) and filtered (EEG:1–300 Hz, EMG:10–300 Hz) using a DC/AC differential amplifier (AM-3000, AM systems, Sequim, WA, USA). The input was then received via an input module (NI-9215, National Instruments, Austin, TX, USA), digitized at a sampling rate of 1000 Hz using a data acquisition module (cDAQ-9171, National Instruments, Austin, TX, USA), and recorded using a custom-made LabVIEW program (National Instruments, Austin, TX, USA). We habituated the 24-hour EEG/EMG recordings more than three times; when REM sleep (see vigilance state assessment) was consistently observed, the experiment was initiated.

The EEG/EMG signals were amplified (gain 1000×) and filtered (EEG: 1–300 Hz, EMG: 10–300 Hz) using a DC/AC differential amplifier (AM-3000, AM Systems). The input was received via an input module (NI-9215, National Instruments), digitized at a sampling rate of 1000 Hz using a data acquisition module (cDAQ-9174, National Instruments), and recorded using a custom-made LabVIEW program (National Instruments). We habituated the 24 h EEG/EMG recordings more than three times, and REM sleep (see the vigilance state assessment) was often observed when we started the experiment. Three days before the control recordings on DOX-off day −1 (DOX-off day −4), the mice were placed in a soundproof box and connected to the EEG/EMG cable for the 24 h EEG/EMG recording of D2-DTA mice. They remained continuously tethered to the cable in the same soundproof box until day 11 of the DOX-off experiment. Cages and food were not changed during the stay in the soundproof box, except changing from DOX-containing chow to normal chow on Dox-off day 0.