Buprenorphine hcl

Heroin and buprenorphine were prepared in sterile saline and briefly sonicated. In all experiments, opioids were administered intravenously using a cumulative-dosing approach. For the heroin-only condition, we administered vehicle (ie sterile saline) and then 0.27, 0.55, 1.12, and 2.24 heroin (mg/kg IV). Cumulative doses were administered every 10 minutes as illustrated in Figure 1A.
In the buprenorphine-heroin condition, we administered vehicle (ie sterile saline) and then ascending doses of buprenorphine: 0.011, 0.044, 0.177, 0.708 (Figure 1B). Continuing in 10-minutes intervals, we then administered the same doses of heroin used in the heroin-only condition. Buprenorphine HCl was purchased from Sigma-Aldrich, and heroin was provided by the NIDA drug supply program. Doses of buprenorphine were calculated as the weight of the HCl salt.

Rats were anesthetized with 2–3% isoflurane mixed with oxygen and placed into the stereotaxic apparatus (Kopf Model 902, Tujunga, CA). After incision and craniotomy, four 1/8″ self-tapping mounting screws were installed into the skull. Then, the dura was punctured with a 27-gauge needle and a 26-gauge guide cannula (PlasticsOne, Roanoke, VA), with stainless steel tubing cut to extend 3.5 mm below the 8.0 mm long pedestal, was implanted into each V2 at −5.3 mm posterior and ±3.1 mm lateral to bregma, to a depth of 0.9 mm below the skull surface. These coordinates differed only along the rostrocaudal axis from those used for PPC cannulations in Schiffino et al. (2014) (link). Cannulae were held in place with dental acrylic and fitted with obturators that were cut to match the length of the guide. Once the acrylic set, the incision was closed with surgical staples and topical antibiotic ointment was applied to the wound edges. Then, all rats received subcutaneous 0.02 mg/kg injections of sterile buprenorphine HCl (Sigma, St. Louis, MO) to ameliorate pain.

The rats were anesthetized with either 50 mg/kg pentobarbital (Duke) or 2–3% isoflurane mixed with oxygen (Johns Hopkins) and placed into a stereotaxic apparatus (Model 902, Kopf, Tujunga, CA, USA). Fourteen rats received bilateral lesions of BLA, using stereotaxic coordinates (Paxinos & Watson, 1998 ) 2.8 mm posterior of bregma and 5.0 mm from the midline, with infusions at 8.7 and 8.4 mm ventral from the skull surface at the drill site. The lesions were made using 12.5 mg/mL N-methyl-D-aspartate (NMDA, Sigma, St. Louis, MO, USA) dissolved in phosphate-buffered saline (PBS) solution, infused with a 2.0-μL microsyringe (Hamilton, Reno, NV, USA) at a rate of 0.1 μl/min; 0.2 μL at the deeper site and 0.1 μL at the shallower site. Ten sham-lesion control rats received infusions of PBS alone to these sites, in the same manner. Each rat received a subcutaneous injection (0.02 mg/kg) of buprenorphine HCl (Sigma, St Louis, MO, USA) after surgery to ameliorate pain. The rats were allowed to recover for 10–14 days before being returned to the training protocol.