Sodium chloride (nacl)

Suvorexant (Belsomra^®; Merck, Whitehouse Station, NJ, United States of America) pills (20 mg) were crushed and dissolved in a 20% vitamin E TPGS (D-α-tocopherol polyethylene glycol 1,000 succinate; Mazuri, Richmond, IN, United States of America) solution (Cox et al., 2010 (link); Guo et al., 2013 (link); Ehlers et al., 2020 (link)) which was also used for control (i.e., 0 mg/kg SUV) group injections. Once homogenized, to maximize bioavailability of the compound, SUV was administered 30 min before the test sessions at doses of 0, 10, and 20 mg/kg (5 mL/kg, p. o.; based on previously reported doses: Simmons et al., 2017 (link); Gentile et al., 2018 (link)). Oxycodone hydrochloride (Spectrum Chemicals, St. Louis, MO, United States of America) dissolved in 0.9% sodium chloride (Hospira, United States of America) was administered intravenously (i.v., 0.15 mg/kg/0.1 mL; Wade et al., 2014 (link); Matzeu and Martin-Fardon, 2020 (link)).

Figure 1 Following the last day of extinction training, the rats that were designated for intra-pPVT OrxA prime-induced reinstatement received a sham injection (Sham) for habituation to the microinjection procedure. Twenty-four hours later, they received an intra-pPVT microinjection of 0.5 μg OrxA (Matzeu et al., 2016 (link); American Peptide, Sunnyvale, CA, United States) in 0.9% sodium chloride (Hospira, Lake Forest, IL, United States) or vehicle (i.e., 0.9% sodium chloride; Matzeu et al., 2016 (link)). The microinjections in the pPVT were performed using a microinfusion pump (Harvard 22 Syringe Pump, Holliston, MA, United States) and injectors that extended 3.5 mm beyond the guide cannula. The injections were performed at a flow rate of 0.5 μl/min over 1 min. The injectors were left in place for an additional minute to allow diffusion away from the injector tip. Following the injections, the rats were returned to their home cages for 2 min and then placed in the operant chambers under extinction conditions for 2 h. After the test, the rats were euthanized by CO₂ inhalation, and their brains were collected and snap frozen. The brains were sectioned coronally (40 μm) on a cryostat at −20°C, and injection tracks were verified (Figures 1C,D). Only rats with cannula placements that were located in the appropriate brain region were included in the data analysis.

Figure 1 Rats that were designated for cocaine self-administration were surgically prepared with jugular catheters 7–10 days before beginning cocaine self-administration training in 6 h/day (i.e., long-access [LgA]) sessions. Each session was initiated by the extension of two retractable levers into the operant conditioning chambers (29 cm × 24 cm × 19.5 cm; Med Associates, St. Albans, VT, United States). Responses at the active lever were reinforced on a fixed-ratio 1 (FR1) schedule by an intravenous (IV) infusion of cocaine (National Institute on Drug Abuse, Bethesda, MD, United States; 0.25 mg/0.1 ml/infusion) that was dissolved in 0.9% sodium chloride (Hospira, Lake Forest, IL, United States) and infused over 4 s. Each reinforced response was followed by a 20-s timeout (TO20) period that was signaled by the illumination of a cue light above the active lever. Responses at the inactive lever were recorded but had no scheduled consequences.