The combination of medetomidine, azaperone, and alfaxalone has been successfully used to anesthetize captive white-tailed deer (Odocoileus virginianus). This same combination was utilized to immobilize free-ranging female mule deer (Odocoileus hemionus; MD) in urban and nonurban environments (14 urban MD, 14 nonurban MD) in British Columbia, Canada. Physiologic data were collected to assess the safety and reliability of this drug combination under field conditions. Each deer received estimated dosages of 0.15 mg/kg medetomidine, 0.2 mg/kg azaperone, and 0.5 mg/kg alfaxalone intramuscularly via a remote darting system. Inductions were calm and rapid (mean time to sternal recumbency: urban MD, 6.4±2.2 min; nonurban MD, 8.2±4.1 min). Supplemental drugs were required to induce lateral recumbency in five deer, four of which had experienced initial dart failure (mean time to lateral recumbency: urban MD, 8.5±3.8 min; nonurban MD, 18.7±16.5 min). Recoveries were smooth and uneventful (time to standing: urban MD, 12.5±3.4 min; nonurban MD, 9.0±3.5 min) for all but one debilitated nonurban MD that died shortly after atipamezole administration (at five times the medetomidine dose). The major side effects of the combination were hypoxemia and hypercapnia. The combination of medetomidine, azaperone, and alfaxalone proved suitable for the immobilization of urban and nonurban free-ranging MD.
Research in the field of wildlife conservation and population management often entails the capture and immobilization of free-ranging animals. Safe and humane handling of wildlife can be facilitated by the use of an anesthetic protocol that has a rapid onset of action, a high safety margin, minimal undesirable side effects, and is readily antagonized with reversal agents. Early drug cocktails assessed in captive and free-ranging white-tailed deer (Odocoileus virginianus; WTD) and mule deer (Odocoileus hemionus; MD) involved an ultrapotent opioid or a cyclohexamine. Beyond the technical disadvantage of controlled drugs requiring strict adherence to storage, record-keeping, and handling protocols, these substances have been associated with various side effects including unpredictable inductions, hyperthermia, hypoxemia, acidosis, poor muscle relaxation, and prolonged recoveries (Caulkett et al. 2000; Wolf et al. 2004). The suboptimal anesthesia quality, the handling hazards associated with potent opioids, and the difficulty of access to some of the drugs have rendered these protocols unpopular for anesthesia of deer.
In more-recent years, research has steered away from single-agent protocols and potent opioids and toward alpha-2 adrenergic agonist-based combinations. Medetomidine is popular for use in wildlife anesthetic protocols because it is an effective sedative, can be antagonized, and can be concentrated up to 40 mg/mL to accommodate small dart capacities. Combinations with dissociative anesthetic drugs (ketamine or tiletamine), a butyrophenone sedative (azaperone), or a mixed agonist-antagonist opioid (butorphanol) produce synergism between pharmacologic agents, allowing for dose reduction of the alpha-2 adrenergic agonist and increased reliability of immobilization (Murray et al. 2000; Mich et al. 2008). While drug combinations that include a dissociative anesthetic are amongst the most-commonly used protocols and produce a relatively stable plane of anesthesia, the recovery after antagonism of the accompanying alpha-2 adrenergic agonist may be prolonged or rough because the dissociative anesthetic cannot be antagonized. This issue was addressed by substituting the dissociative anesthetic with butorphanol and azaperone (BAM; Mich et al. 2008; Miller et al. 2009). However, the improved alpha-2 adrenergic agonist-based combination remains restricted to licensed veterinarians in some countries, and wildlife professionals are seeking a drug combination that would produce an anesthesia of quality without the use of controlled substances.
Alfaxalone has the potential to replace dissociative anesthetics and opioids in alpha-2 adrenergic agonist-based combinations for the anesthesia of deer. The neuroactive steroid causes central nervous system depression and muscle relaxation by binding to the gamma-aminobutyric acid type A receptors and increasing their sensitivity to the inhibitory effect of the gamma-aminobutyric acid neurotransmitters (Harrison and Simmonds 1984). A novel reformulation of alfaxalone is approved for the induction and maintenance of anesthesia in dogs and cats in Canada, the US, Australia, New Zealand, South Africa, and many Asian and European countries and is only controlled in some countries (Drug Enforcement Administration Schedule IV in the US). Alfaxalone, combined with medetomidine and azaperone (MAA), successfully anesthetized captive WTD. While hypoventilation and hypoxemia were observed, they did not significantly differ from what was observed with medetomidine-azaperone (MA) alone (Pon et al. 2016). Furthermore, the synergism between the three substances allowed for the use of a subanesthetic alfaxalone dose and a total drug volume suitable for remote delivery in large animals. While the drug combination produced a satisfactory, deep sedation and a rapid and smooth recovery (following administration of atipamezole) of deer in captivity, it is unknown how it would perform in free-ranging animals, which may experience an acute stress event in response to human presence and stalking. The stress reaction has the potential to counteract the effects of medetomidine through the activation of the sympathetic nervous system.
Our goal was to determine if the MAA combination improved the welfare of wildlife during capture and handling. We assessed the efficacy and safety of the MAA protocol in MD under field conditions and compared the quality of induction and recovery using this protocol.
MATERIALS AND METHODS
Study area and animals
Twenty-eight free-ranging adult female MD from the East Kootenay region, British Columbia, Canada, were included in this study. The deer were anesthetized in urban and nonurban environments for radiocollar fitting and biologic sample collection for ecologic studies conducted by the British Columbia Ministry of Forests, Lands and Natural Resource Operations. Captures happened over three campaigns: nine animals were immobilized in Newgate (49°03′ to 49°12′N, 115°12′ to 115°15′W, elevation [Et] 789±11 m, barometric pressure [PB] 688±3 mmHg, ambient temperature [TA] 4±2 C) in December 2014; five in Newgate (Et 856±40 m, PB 687±2 mmHg, TA 7±7 C) in April 2015; and 14 in Invermere (50°29′ to 50°30′N, 116°01′ to 116°03′W, Et 863±4 m, PB 695±3 mmHg, TA 0±3 C) in February 2016.
Capture methods, drugs, and darting equipment
All animals were located and darted from the ground. Body masses of the animals were estimated at 80 kg. Each deer received a combination of medetomidine (30 mg/mL; Bow Valley Research, Calgary, Alberta, Canada), azaperone (40 mg/mL; Elanco, Eli Lilly Canada, Guelph, Ontario, Canada), and alfaxalone (10 mg/mL; Jurox, Rutherford, New South Wales, Australia) at estimated dosages of 0.15 mg/kg, 0.2 mg/kg, and 0.5 mg/kg, respectively. Actual drug doses were later calculated using the exact weight of the animals. Drugs were delivered remotely using two different systems: urban MD were darted with 3-mL, single-barbed metal Pneu-Darts using a Pneu-Dart Model 389 rifle (Pneu-Dart, Inc., Williamsport, Pennsylvania, USA) and nonurban MD were darted with 3- or 5-mL, single-barbed plastic Dan-Inject darts using a Dan-Inject JM Special 16 rifle (Dan-Inject, Ft. Collins, Colorado, USA), with all drugs combined in a single dart. In the eventuality of failure of a dart to discharge completely, the animal was darted again with MAA at 25–100% of the initial dose, depending on the approximate residual drug volume in the first dart. If the plane of anesthesia was too light to allow for processing of the animal, a supplemental alfaxalone dose equal to the initial dose was given intravascularly in either the jugular or the medial saphenous vein. Times from injection to sternal recumbency, to head down, and to lateral recumbency were recorded. The deer were approached once they were in lateral recumbency and unresponsive to noise and immediately blindfolded. Quality of sedation was scored on a scale of 1 through 8 at postinjection (PI) times: 15, 30, 45, and 60 min (Pon et al. 2016).
Reversal of medetomidine was achieved with atipamezole hydrochloride (20 mg/mL; Bow Valley Research) administered into the gluteal or quadriceps muscles at five times the medetomidine dose (0.75 mg/kg). The deer were left undisturbed to recover and gently stimulated if still recumbent 10 min following reversal administration. The quality of recovery was scored using a five-point system (Pon et al. 2016). Times from reversal administration to head up, sternal recumbency, and standing were recorded. Total down time was calculated from recumbency to standing. Urban MD were kept in a trailer and released to the wild within 8 h of capture while nonurban MD were released to the wild upon reversal. Postanesthetic survival was assessed using the radiocollar data.
Monitoring and data collection
Heart rates, respiratory rates, and rectal temperatures were monitored with a stethoscope, by observation of chest movements, and with a digital thermometer (3M Consumer Health Care, St. Paul, Minnesota, USA) and recorded every 5–10 min. Each animal was weighed in a heavy-duty tarp (MegaMover® Portable Transport Unit, Graham Medical, Green Bay, Wisconsin, USA) with a dial hanging-scale. Arterial blood samples were collected in heparinized syringes (GASLYTE Arterial Blood Sampler, Vital Signs, Englewood, Colorado, USA) from the femoral or median saphenous arteries and immediately processed using a portable analyzer and cartridges (i-STAT® Portable Clinical Analyzer and i-STAT®1 cartridge CG4+, Abbott Laboratories, Abbott Park, Illinois, USA), corrected to the rectal temperature. The samples were analyzed for pH, partial pressure of arterial carbon dioxide (PaCO2), partial pressure of arterial oxygen (PaO2), bicarbonate, base excess, and lactate. Atmospheric pressure was measured using the blood gas analyzer, and ambient temperature was obtained from the local weather station.
The alveolar oxygen tension (PAO2) for deer at elevations of 688 to 896 m, as defined by the alveolar gas equation, ranges from approximately 95 to 96 mmHg when assuming a PaCO2 of 40 mmHg, a water vapor pressure of 47 mmHg, and a respiratory quotient of 1 (Fahlman et al. 2014). The expected PaO2 for awake deer under those environmental conditions is 80–81 mmHg, when assuming a normal alveolar-arterial oxygen tension difference [P(A-a)O2] of 15 mmHg. Anesthetized deer were considered hypoxemic if their measured PaO2 was below the expected PaO2. The degree of the ventilation-perfusion (V/Q) mismatch was quantified using the calculated P(A-a)O2 (Fahlman et al. 2012).
Data were analyzed using GraphPad Prism 7 (GraphPad Software Inc., La Jolla, California, USA). Normality of data was assessed using the D'Agostino-Pearson normality test. If data normality was shown, a parametric unpaired two-tailed t-test was used to assess difference between groups. If not, a Mann-Whitney U-test was used. A chi-square analysis was performed to investigate differences between MD groups that required physical restraint upon approach. Values from blood samples that were of potential venous origin were excluded from the statistical analysis of arterial blood gas results. The significance level was set at P<0.05.
Urban mule deer
Urban MD weighed 71.1±6.6 kg (mean±SD). Induction and recovery times are listed in Table 1. Sternal recumbency was achieved in all animals with one dart. The initial and total drug dosages are presented in Table 2. Two deer were darted twice: one arose from lateral recumbency after being darted subcutaneously and one went into lateral recumbency but remained too reactive for handling. Four deer (29%) were physically restrained upon approach, two of which required supplemental drugs to enable processing.
The first recorded heart rate, respiratory rate, and rectal temperature for urban MD were, respectively, 38±9 beats/min, 21±2 breaths/min, and 37.6±0.5 C. Comparison of vitals over time and monitoring of blood pressure were not objectives of this field study since this has already been done in a detailed cardiopulmonary study in captive WTD anesthetized with MAA (Pon et al. 2016). Quality of sedation was recorded for 14 animals (Table 3). Arterial blood samples were successfully obtained from 11 deer at 20.7±10.7 min PI (Table 4). All animals were hypoxemic (measured PaO2<81 mmHg), with a P(A−a)O2 of 51.1±10.4 mmHg.
Recovery was smooth and uneventful. Out of the 13 urban MD for which recovery scores were obtained, three had a recovery score of 4 (slight ataxia and struggling, stood at first or second attempt, no serious instability), and 10 had a recovery score of 5 (no ataxia, no struggling, stood up at first attempt as if fully conscious). All but one deer required stimulation at 10 min post atipamezole administration to rise. Ten deer went directly from lateral recumbency to standing. Time to standing was significantly longer in urban MD than in nonurban MD. Total down time was 46.9±8.7 min. Procedure duration was 39.4±9.6 min.
Nonurban mule deer
Nonurban MD weights were 70.1±9.6 kg. Induction and recovery times are presented in Table 1. One doe was chased by two bucks after darting and was excluded from the induction time analysis. All animals were induced to sternal recumbency with one dart. Time to lateral recumbency was significantly longer in nonurban MD than in urban MD. The initial and total drug dosages are presented in Table 2. Nonurban MD received significantly higher initial and total dosage of alfaxalone compared to urban MD. Three deer experienced dart failure and were darted a second time to induce lateral recumbency. Physical restraint was used on three deer (21%), which were given supplemental drugs to maintain lateral recumbency.
The first recorded heart rate, respiratory rate, and rectal temperature of nonurban MD were, respectively, 41±4 beats/min, 23±3 breaths/min, and 38.1±0.4 C. Nonurban MD had a significantly higher rectal temperature then did urban MD. Quality of sedation was recorded for nine animals (Table 3). Arterial blood samples were successfully obtained from all nonurban MD at 39.7±16.9 min PI (Table 4). All animals were hypoxemic (measured PaO2<80 mmHg), with a P(A−a)O2 of 51.3±11.1 mmHg.
Recovery was smooth and uneventful for all nonurban MD but one, which died shortly after reversal administration. Histopathology revealed that it was an animal in poor body condition with a mild, multifocal, acute to subacute bronchopneumonia with multinucleated giant cells and aspirated plant material, with concomitant Echinococcus sp. and lung worm infections. Splenic depletion, attributed to a maladaptive process, chronic inflammation, or malnutrition, was also noted. Six deer had a recovery score of 4 and seven had a recovery score of 5. All but three MD required stimulation at 10 min post atipamezole administration to rise, and all but one MD went directly from lateral recumbency to standing. Total down time was 54.9±17.1 min. Procedure duration was 56.8±19.7 min.
Resedation and postanesthetic survival
All urban MD appeared bright and alert upon unloading from the trailer 3–8 h postcapture. Resedation could not be assessed in nonurban MD, as they were returned to the wild upon recovery. The death of two nonurban MD, which occurred between 1 wk and 1 mo postanesthesia, resulted from collisions with motor vehicles and were not attributed to residual effects of anesthesia.
Free-ranging MD were successfully immobilized with MAA, in both urban and nonurban environments. The drug combination produced a deep sedation to light anesthesia adequate for safe handling and minor procedures. Induction with MAA was satisfactory in both speed and quality. It was faster compared to MA alone and comparable to other commonly used protocols used in cervids (Caulkett et al. 2000; Mich et al. 2008; Caulkett et al. 2014). Overall, induction took longer in free-ranging MD than in captive WTD anesthetized with the same protocol (Pon et al. 2016). The longer induction in free-ranging deer when compared to captive deer may be attributed to an acute stress response triggered by the unusual presence of stalking humans during darting. This hypothesis is supported by the significantly higher rectal temperature in nonurban MD compared to urban MD. Based on the rapid induction to an anesthetic plane adequate for minimally invasive procedures, as well as the absence of injury, hyperthermia, and lactic acidosis, the quality of induction was deemed satisfactory for all animals.
Quick recovery of immobilized wildlife is desired to minimize the risk of injury or death following an anesthetic event. Quality of recovery in this study was good to excellent (recovery scores of 4 or 5) for all animals, and times to standing for nonurban MD (9.0±3.5 min) were comparable to those of BAM and MA protocols (Mich et al. 2008; Miller et al. 2009; Caulkett et al. 2014). However, similar to the study of MAA in captive WTD, comparison of results to other studies is complication by the fact that deer were encouraged to stand at 10 min postinjection of atipamezole (Pon et al. 2016). The longer recovery time recorded in urban MD (12.8±3.6 min) may be attributed to the shorter duration of the procedure. Early reversal of medetomidine (39.4±9.6 min PI) in urban MD may result in alfaxalone lingering, thus prolonging recovery. A time lapse of 56±24 min from injection to head lift upon recovery was reported in cats given alfaxalone intramuscularly at a 5 mg/kg dose (Rodrigo-Mochilí et al. 2016). Alfaxalone was shown to prolong recovery when added to MA (9.1±2 min; MAA, 12.2±2.6 min; Pon et al. 2016). Additionally, urban MD were allowed to recover in a darkened trailer with conspecifics whereas the nonurban MD woke up outside by themselves. The reduced visual stimulation in the trailer and close presence of group members may have kept the urban MD calmer upon awakening and less motivated to rise.
Hypoxemia may contribute to anesthesia-related mortalities in wildlife and should be minimized by providing supplemental oxygen intranasally, when possible (Spraker 1993; Fahlman et al. 2014). Immobilization with MAA in the current study induced hypoxemia in all animals. The decreased PaO2 was likely a consequence of both hypoventilation and intrapulmonary changes. Hypoventilation, as indicated by hypercarbia (CO2>45 mmHg), likely resulted from the dose-dependent depression of the central respiratory center by both alfaxalone and medetomidine (Celly et al. 1997; Moll et al. 2013). Atelectasis secondary to prolonged recumbency or subclinical lung diseases may have contributed to hypoxemia in the deer anesthetized with MAA (Read 2003). Intrapulmonary changes, leading to physiologic shunting or diffusion impairment, may result in a low V/Q ratio and a subsequent increased venous admixture. In the present study, V/Q mismatch occurred in all animals, as indicated by an increased P(A-a)O2 (>15 mmHg; Fahlman et al. 2012). The degree of hypoxemia reported here compares to what was observed in captive WTD deer immobilized with MA, MAA, or BAM (PaO2, MA, 54±7 mmHg; MAA, 54±9 mmHg at 15 min PI; BAM, 42.4±7.8 mmHg at 15 min PI), but was more severe than in free-ranging WTD immobilized with BAM (PaO2>60 mmHg; Mich et al. 2008; Caulkett et al. 2014; Pon et al. 2016).
Retention of CO2 secondary to hypoventilation resulted in a mild primary respiratory acidosis in all but one deer. Blood gas values were compared to values in awake sheep (Ovis aries) and tranquilized wapiti (Cervus elaphus), as reference values could not be found for awake MD (Read et al. 2000; Onmaz et al. 2009). Furthermore, urban MD had a significantly lower blood pH than did nonurban MD, and their lactate levels were marginally higher. Although the latter observation is not statistically significant, it is likely of physiologic importance. The increase in lactate may be attributable to the fact that more urban MD were physically restrained upon approach. Deer were more easily tracked in a town and were approached more quickly after becoming laterally recumbent. Excitement and struggle upon approach ultimately resulted in an increased anaerobic metabolism and accumulation of lactic acid. Furthermore, some nonurban deer received a higher dose of alfaxalone, as it was used to top-up the larger darts, and these were more deeply sedated upon approach.
Wildlife anesthesia is often conducted without prior knowledge of patients' health status, and visual examination can, at best, approximate condition. This presents challenges because drug protocols cannot be adjusted to the patient. In this study, one nonurban MD died in December, and only after the fact was it discovered it had pre-existing health issues, including bronchopneumonia complicated with Echinococcus sp. and lung worm infections. Anesthesia likely exacerbated pulmonary dysfunction, leading to agonal regurgitation/aspiration and death. It is unknown whether this animal would have died if anesthetized with a lower dose of MAA or with a different drug combination or whether these postmortem findings are reflective of subclinical respiratory diseases in this deer population. In the present study, MAA was used at the reported doses throughout winter and spring, in deer of varying body conditions and weights, and proved adequate for most animals.
The use of MAA proved suitable for the immobilization of urban and nonurban free-ranging MD. Anesthesia was characterized by a rapid induction, the maintenance of adequate anesthetic plane and muscle relaxation for performance of minimally invasive procedures, and a rapid, smooth recovery following administration of atipamezole. Supplemental oxygen is recommended to treat the hypoxemia when using MAA in the field.
We thank Aaron Reid and Ian Adams for accommodating this research in their deer collaring projects and for sharing survival data. We had the assistance of the following people: Jenny Coleshill, Becky Phillips, Kylie Pon, Irene Teske, Larry Ingham, David Lewis, Holger Bohm, Brian MacBeth, Joan Caulkett, and the many volunteers who helped with deer captures. This project was funded by the University of Calgary Clinical Research Fund.