Antisedan

For stereotaxic injections, rats were anaesthetised by intramuscular administration of a medetomidine (0.25 mg/kg, Domitor, Vetoquinol) and ketamine (50 mg/kg, Narketan, Vetoquinol) mix and placed in a stereotaxic frame in the flat skull position. Stereotaxic injections were performed as described previously [17 (link)]. After surgery, the incision was closed and atipamezole (0.15 mg/kg, Antisedan, Zoetis) was administered intramuscularly and post-surgery analgesia with buprenorphine (0.05 mg/kg, Buprevet, Virbac) subcutaneously. The success of viral injections was verified at the end of each study by visualisation of the fluorescent reporter and/or qRT-PCR.

A mouse burn injury model was used to model traumatic injury in mice as previously described (24 (link)). Briefly, mice were anesthetized by intraperitoneal (IP) injection with 125 mg/kg ketamine (Fort Dodge Animal Health, Fort Dodge, IA) and 10 mg/kg xylazine (Lloyd Laboratories, Shenandoah, IA). Buprenorphine at 0.6mg/kg was injected subcutaneously at the time of injury. The mice had their dorsal fur shaved and were placed in a plastic mold exposing 20% of their total body surface area. Injury was induced by immersing the exposed part of the dorsum for 9 seconds in a 90˚ C water bath. This approach causes a full-thickness and well-demarcated anesthetic injury due to complete loss innervation. Uninjured mice underwent the same procedure but were exposed to room temperature water for 9 seconds. All animals were resuscitated with an IP injection of 1 mL of 0.9% pyrogen-free normal saline containing ANTISEDAN (atipamezole, Zoetis, US) at 1mg/kg. The mortality from this burn trauma model is less than 2%.

All Bradypus sloths were caught opportunistically by hand and equipped with tags (see below) without anaesthesia. Choloepus sloths were anaesthetised during capture using 1 mg/kg of ketamine (Ketamina 50®; Holliday-Scott, Buenos Aires, Argentina) and 0.008 mg/kg of dexmedetomidine (Dexdomitor®; Zoetis, Parsippany-Troy Hills, NJ, USA) administered intramuscularly. Anaesthesia was reversed prior to release using 0.008 mg/kg of anti-sedante (atipamezol; Antisedan®, Zoetis). Body mass measurements were taken for each individual at capture (E-PRANCE® Portable Hanging Scale (±0.01 g)).
Sloths were equipped with pre-calibrated Daily Diary (DD) data loggers (Wilson, Shepard & Liebsch, 2008 (link)), programmed to record 9 parameters (barometric pressure (mbar), external temperature (°C), relative humidity (%), tri-axial magnetometry (gauss), and tri-axial acceleration (g)) at a rate of 40 Hz. The data loggers were held within non-lubricated condoms containing 0.5 g silica gel desiccant to protect the electronics from moisture damage. DDs were combined with Very High Frequency (VHF) radio transmitters (Biotrack PIP3 VHF tag) within 3D-printed housings and attached via elastic, backpack-style harnesses, positioning the devices firmly on the upper back (Fig. S1) (Cliffe et al., 2014 (link)). The total weight of the backpack was 90 g.

At baseline (Day 0) and on Days 70, 110, 168, 240, 309, 380, and 505 PI, peripheral blood was collected for complete blood count (CBC) and serum for serology; and on Days 0, 110, 168, 240, 309, 380, and 505 PI radiographic ventrodorsal (VD) and lateral thoracic images were acquired and bronchoalveolar lavage (BAL) performed with 10 mL of lactated Ringers solution under sedation with an intramuscular (IM) dose of medetomidine (Domitor®, Zoetis, NY), butorphanol (Torbugesic®; Zoetis), ketamine (ketamine hydrochloride; Zoetis), and after the procedures an IM dose of atipamezole (Antisedan®; Zoetis) was administered. Cats were monitored daily and physical examination performed weekly.
At the termination of the study (Day 510), cats were humanely euthanized after sedation using pentobarbital sodium and phenytoin sodium solution 1 mL/10 lbs. given intraperitoneally (Euthasol®, Virbac AH). Complete necropsies were conducted with collection of lung, heart, brain, kidney, and liver for histopathology studies. Immediately post mortem a blood sample from the right ventricle was collected for serology. Right caudal lung lobes were fixed perfused with 10% formalin via the bronchi to a pressure of 14 cm H₂O. Pathologists and radiologists were blinded to the treatment groups to which cats were assigned, creating a controlled, blind study format.