Yohimbine antagonizes the anaesthetic effects of ketamine xylazine in captive Indian wild felids

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1 Veterinary Anaesthesia and Analgesia, 29, 36, doi:1.1111/j x RESEARCH PAPER Yohimbine antagonizes the anaesthetic effects of ketamine xylazine in captive Indian wild felids Sadanand D Sontakke MVSc, Govindhaswamy Umapathy MSc, PhD & Sisinthy Shivaji MSc, PhD Laboratory for the Conservation of Endangered Species (LaCONES), Annexe I of Centre for Cellular and Molecular Biology, Hyderguda, Hyderabad, India Correspondence: Sisinthy Shivaji, Centre for Cellular and Molecular Biology, Uppal Road, Hyderabad 5 7, India. shivas@ccmb.res.in Abstract Objective To determine the effectiveness of yohimbine as an antagonist of ketamine xylazine anaesthesia in captive Asiatic lions (Panthera leo persica), tigers (Panthera tigris) and leopards (Panthera pardus). Study design Prospective clinical trial. Animals Fifty-two healthy adult lions, 55 adult leopards and 16 adult male tigers. Methods Captive wild felids in Indian zoos were anaesthetized with a combination of ketamine ( mg kg )1 ) and xylazine ( mg kg )1 ) using a dart propelled from a blowpipe. Time to onset of anaesthesia, lateral recumbency and induction time were measured, and physiological variables (respiration, heart rate and rectal temperature) were recorded once after the onset of complete anaesthesia. Anaesthesia was antagonized at various time periods with an intravenous administration of either.1 or.15 mg kg )1 yohimbine. Onset of arousal and time to complete anaesthetic recovery were recorded. Results A total of 123 immobilizations were conducted between 2 and 25. Anaesthetic induction was achieved in minutes in all animals. Incidents of sudden recovery or life-threatening effects associated with immobilizations were not observed. Yohimbine effectively antagonized anaesthesia in all animals within 1 minutes without any excitatory behaviour compared to control animals. No adverse reactions/side effects to yohimbine were recorded except that a few leopards exhibited seizure-like signs for a short period immediately after yohimbine administration. The duration of anaesthesia had no significant effect on the recovery time in any of the animals. Conclusion and clinical relevance Yohimbine antagonized the xylazine portion of ketamine xylazine anaesthesia and thereby hastened recovery from anaesthesia in Asiatic lions, tigers and leopards. Keywords Asiatic lion, ketamine xylazine, leopard, reversal, tiger, wild felids, yohimbine. Introduction Chemical restraint is a valuable tool in wildlife research and management since it facilitates the handling of animals as and when required for medical procedures and experimentation (Kreeger et al. 1986). In recent years, various drug combinations, such as tiletamine zolazepam, medetomidine ketamine, ketamine xylazine and phencyclidine promazine have been used to immobilize wild carnivores (Herbst et al. 1985; Shivaji et al. 1998, 23; Grassman et al. 24; Jacquier et al. 26). Amongst them, the dissociative anaesthetic ketamine hydrochloride (ketamine), in combination with an alpha-two adrenergic agonist, xylazine hydrochloride (xylazine), has been used effectively to immobilize several wild felids including lions, leopards and tigers (Herbst et al. 1985; Seal et al. 34

2 1987; Patil et al. 1998; Shivaji et al. 1998, 23; Jayaprakash et al. 21). This anaesthetic combination produces a smooth and rapid induction of anaesthesia with the pressor and cataleptic effects of ketamine being complemented by the sedative and myorelaxing effects of xylazine, thus counteracting the adverse effects of ketamine (Amend 1972). However, this combination is known to cause prolonged sedation, which has been attributed to the xylazine (Kreeger et al. 1986). Such uncontrollable recoveries because of prolonged sedation are a serious concern in wildlife research since the sedated free-ranging animals could be easily preyed upon. A few deaths have also been recorded in tigers because of profound respiratory depression following ketamine xylazine anaesthesia (Seal et al. 1987). It is therefore very important to determine a suitable antagonist for the speedy recovery of anaesthetized endangered wild felids. Alpha-two adrenergic antagonists such as yohimbine, tolazoline and atipamezole have been used effectively in ungulates for antagonizing the anaesthetic effects of ketamine xylazine (Kreeger et al. 1986; Sontakke et al. 27). However, to the best of our knowledge, little has been published on the antagonism of anaesthesia in wild felids (Seal et al. 1987; Miller et al. 23; Jacquier et al. 26). Studies in the domestic cat (Hsu & Lu 1984) and the Bengal tiger (Seal et al. 1987) have suggested that yohimbine could hasten the recovery of ketamine xylazine anaesthesia. The present study was undertaken as a part of ongoing research on the conservation of endangered Indian wild animals with the aim to study the ability of yohimbine to antagonize the anaesthetic effects of ketamine xylazine in three endangered Indian large felids (Asiatic lions, leopards and tigers). Materials and methods Study area and animals Captive Asiatic lions (Panthera leo persica), leopards (Panthera pardus) and tigers (Panthera tigris) were anaesthetized for collection of blood for steroid hormone assay, for collection of semen by electroejaculation and for minor surgical manipulations (vasectomy, wound dressing, etc.). Animals of 3 12 years of age were anaesthetized as a part of the present study between 2 and 25 at various zoos in India. A total of 123 immobilizations were carried out on 52 lions (22 males and 3 females), 55 leopards (32 males and 23 females) and 16 tigers (all males) during All the animals were in the breeding age-group (about 3 12 years) and were apparently healthy without any detectable illness. The mean body weight (mean ± SD) of the animals were as follows: male lions ± 9.3 kg (range kg), female lions ± 7.1 kg (range kg), male leopards 75.9 ± 9.3 kg (range 6 11 kg), female leopards 68.6 ± 7.1 kg (range kg) and male tigers ± 11.3 kg (range kg). The details of the number of animals used and their sexes are given in Table 1. All animals were housed individually in indoor concrete pens ( m) during the night and were allowed in an open exercise area (surrounded by a moat) during daytime with an exposure to a natural photoperiod. Each animal was fed with 6 1 kg of beef supplemented with calcium and vitamins once a day for 6 days week )1. The animals had free access to clean Table 1 Number of captive Asiatic lions, leopards and tigers (3 12 years) used for immobilization trials (2 25) from various zoos in India Lion Leopard Tiger Zoo Male Female Male Female Male Nehru Zoological Park, Hyderabad, Andhra Pradesh Sri Venkateswara Zoological Park, Tirupati, Andhra Pradesh Indira Gandhi Zoological Park, Vishakhapatnam, Andhra Pradesh 5 5 Sakkarbaug Zoological Garden, Junagadh, Gujarat Nandankanan Zoolgical Park, Bhubaneshwar, Orissa Bannerghatta Biological Park, Bangalore, Karnataka Ó 29 The Authors. Journal compilation Ó 29 Association of Veterinary Anaesthetists, 36,

3 drinking water ad libitum. All experimental procedures were performed in accordance with the guidelines of the Central Zoo Authority, Ministry of Environment and Forests, Government of India. Anaesthesia procedure All animals were starved for 1 day prior to the immobilization procedure and were anaesthetized between 8 and 1 hours with a combination of mg kg )1 ketamine (Ketamil, 1 mg ml )1 ; Troy Laboratories, Smithfield, NSW, Australia) and mg kg )1 xylazine (Ilium Xylazil, 1 mg ml )1 ; Troy Laboratories). Both the drugs were loaded into a 5-mL projectile plastic dart syringe equipped with a 35-mm needle (Dist Inject; Helpro Health Products and Services, Pune, India) and injected intramuscularly into the thigh or the shoulder region using a blowpipe (Dist Inject; Helpro Health Products and Services). Following darting, the animals were left undisturbed and monitored for signs of the onset of anaesthesia. The time taken for onset of anaesthesia (as judged by ataxia, sedation and pupillary dilation), lateral recumbency and the induction time were recorded. Induction time is the time that elapsed between darting to complete anaesthesia when the animals no longer responded to any physical stimuli (pedal and tail reflexes). The anaesthetized animals were then transferred on to a stretcher in lateral recumbency and their eyes were covered to avoid direct exposure to sunlight. They were removed from the cage for further procedures. An ophthalmic ointment (Neosporin; Glaxo-Smith Kline India Ltd, Mumbai, India) was applied to the eyes to prevent desiccation of the cornea and conjunctiva. Physiological parameters, such as rectal temperature, respiratory and heart rates, and the general health condition of the animal were monitored immediately after onset of anaesthesia. Respiratory rates were also recorded before and after yohimbine administration. Anaesthetic reversal Immediately after the completion of the experimental procedures, the animals were taken back to their cages and were randomly assigned into three groups: one group received an intravenous injection of.1 mg kg )1 yohimbine (Antagozil, 1 mg ml )1 ; Troy Laboratories), the second group of animals received.15 mg kg )1 yohimbine (Table 4) and the third group (i.e. control animals) were injected with 2 ml of normal saline intravenously. The recovery from anaesthesia was the time that elapsed from yohimbine/saline administration to when the animal stood. They were observed intermittently for up to 8 hours post-recovery to detect any signs of resedation. Effect of duration of immobilization on recovery To study the effect of the duration of immobilization on yohimbine-induced recovery, data from animals were categorized depending on anaesthetic duration and compared. Immobilization durations in lions and leopards were categorized in to three groups namely: 11 2 minutes, 21 3 minutes and 31 4 minutes. In tigers, we evaluated recovery only in two groups of animals, i.e minutes and 31 4 minutes. Statistical analysis All data analyses were performed using statistical software program SPSS (Ver for Windows; SPSS, Inc., Chicago, IL, USA). Data are presented as mean ± SD. The unpaired Student s t-test was applied to test significant differences, if any, between the sexes, and to compare the efficacy of two doses of the antagonist with respect to recovery time. A probability of p <.5 was considered to be the minimum level of significance. The data on respiration before and after yohimbine administration were analyzed using Box Whisker plots and unpaired Student s t-test. A nonparametric one-way analysis of variance [Kruskal Wallis test (K W test)] was used to test variation in the recovery time at various time periods in lions and leopards. Results Anaesthesia For all animals, the mean ( ± SD) dose of 2.32 ± 1.1 mg kg )1 ketamine in combination with 1.16 ±.5 mg kg )1 xylazine effectively induced a smooth, rapid and deep plane of anaesthesia with good muscle relaxation and satisfactory analgesia within minutes of darting. No deaths or serious adverse effects related to immobilization were observed in any of the animals. Following darting, the first effects of anaesthesia (such as ataxia, 36 Ó 29 The Authors. Journal compilation Ó 29 Association of Veterinary Anaesthetists, 36, 34 41

4 Table 2 Time to onset of anaesthesia, lateral recumbency and anaesthesia (minutes) in the captive Asiatic lions, leopards and tigers anaesthetized with IM administration of ketamine (2.32 ± 1.1 mg kg )1 ) and xylazine (1.16 ±.5 mg kg )1 ) Species Gender Number of animals Onset of anaesthesia Lateral recumbency Induction time Lion Male ± ± ± 5.6 Female ± ± ± 5.2 Leopard Male ± ± 3.4 a 13.7 ± 3.7 a Female ± ± 3.2 b 16.1 ± 3.7 b Tiger Male ± ± ± 3.8 Values are represented as mean ± SD. Different superscripts in a column within a species differ significantly at p <.5. in-coordination, sedation and pupillary dilation), lateral recumbency and induction time were monitored (Table 2). In lions, all three parameters did not vary significantly between the sexes. However, in leopards duration of lateral recumbency and induction time showed significant differences between the sexes (Table 2). There were no female tigers in the study (Table 2). Anaesthetized males consistently responded to electroejaculation (Shivaji et al. 23) and yielded good ejaculates. A few males were also successfully vasectomized using the present anaesthetic combination. Similarly, females could be handled safely for blood collection, wound dressing and other minor surgical procedures. The duration of anaesthesia in the present study ranged from 11 to 4 minutes in all felids. Physiological values Mean physiological values for respiration, heart rates and rectal temperature were recorded immediately after onset of anaesthesia in lions, tigers and leopards (Table 3). No significant differences in physiological parameters were recorded between sexes. Respiratory rates were also recorded just before and after yohimbine administration. A significant increase (p <.1) in respiration rate was observed in all felids following yohimbine administration (Fig. 1). Further, no cardiopulmonary irregularities, hypo- or hyperthermia were observed in any of the animals during the procedures. However, during the study, two leopards and one tiger had elevated rectal temperature, up to 42.5 C immediately after anaesthesia, which soon decreased to the normal range. Anaesthetic recovery Eighty-eight reversal trials with two doses of yohimbine (.1 and.15 mg kg )1 ) were evaluated. The onset of recovery from anaesthesia, following yohimbine administration, was characterized by tongue movement, followed by limb movement, head lifting and moving into sternal recumbency. Finally, the animals stood up unaided. The final recovery time was the time that elapsed between the administration of yohimbine and the unaided standing of the animal. All the recoveries were smooth and calm without any excitement except for a few leopards that showed seizures for a short Table 3 Respiration rate, heart rate and rectal temperature [mean ± SD, (range)] in captive Asiatic lions, leopards and tigers anaesthetized with IM administration of ketamine (2.32 ± 1.1 mg kg )1 ) and xylazine (1.16 ±.5 mg kg )1 ) Species Gender Number of animals Respiratory rate (breaths minute )1 ) Heart rate (beats minute )1 ) Rectal temperature ( C) Lion Male ± 3 (16 28) 68 ± 6 (57 8) 39.1 ±.5 (38 4.2) Female 3 19 ± 4 (12 27) 67 ± 5 (56 75) 38.9 ±.6 (38 41) Leopard Male ± 3 (18 3) 72 ± 7 (55 84) 39.9 ± 1.3 ( ) Female ± 3 (15 27) 71 ± 5 (59 8) 39.3 ±.9 ( ) Tiger Male ± 3 (12 24) 66 ± 5 (58 73) 4.4 ± 1.2 ( ) Ó 29 The Authors. Journal compilation Ó 29 Association of Veterinary Anaesthetists, 36,

5 (a) Respiration (breaths minute 1 ) (b) Respiration (breaths minute 1 ) The mean times for onset of recovery and sternal recumbency in lions, leopards and tigers following intravenous administration of two doses of yohimbine (.1 and.15 mg kg )1 of yohimbine) did not differ significantly between the doses (Table 4). However, the recovery time (time to standing) was observed to be significantly dose-dependent in lions and tigers (p <.1) but not in leopards (Table 4). Unlike the lions and the tigers, a few of the leopards anaesthetized for a short duration (11 2 minutes) showed seizures just for 2 3 minutes following yohimbine administration but no further morbidity was observed. Seizures were controlled by an intramuscular administration of.5 mg kg )1 diazepam (Calmpose; Ranbaxy India Ltd, Mumbai, India). No signs of resedation were noted for 8 hours post-recovery in any of the animals. Compared to the animals treated with yohimbine, the control animals (lions, n = 15; leopards, n =15 and tigers, n = 5), which received only normal saline following anaesthesia showed prolonged recoveries (8 12 minutes; p <.1) and the animals had a violent and excited recovery. Effect of duration of immobilization on recovery (c) Respiration (breaths minute 1 ) AA BY AY Figure 1 Box Whisker plots (using SPSS 11.1) depicting changes in respiratory rate before and after yohimbine administration in Asiatic lion (a), leopard (b) and tiger (c) immobilized with ketamine xylazine anaesthesia (AA immediately after complete anaesthesia; BY prior to yohimbine administration; AY after yohimbine administration). period. No resedation was observed. No significant difference in the recovery time was observed between the sexes; hence the data were combined for both the sexes (Table 4). In order to evaluate whether the time of administration of yohimbine post-immobilization had any influence on the recovery of the animals, two doses of yohimbine (.1 and.15 mg kg )1 body weight) were administered to lions, leopards and tigers at 11 2, 21 3 and 31 4 minutes of immobilization. There were insufficient tigers anaesthetized for 11 2 minutes for further analysis. It was observed that irrespective of the dose of yohimbine used, immobilization duration had no significant effect on the recovery time in lions (K W test; v 2 =.81, p =.65) and leopards (K W test; v 2 = 3.1, p =.2) (Fig. 2). Discussion The present study documents, to our knowledge, the first report on the effectiveness of yohimbine for antagonizing the anaesthetic effects of ketamine xylazine, which has been used to immobilize lions, tigers and leopards in India and other countries (Herbst et al. 1985; Seal et al. 1987; Patil et al. 1998; Shivaji et al. 1998, 23; Jayaprakash et al. 21; Grassman et al. 24). Further, the ketamine and xylazine doses (2.32 ± 1.1 mg kg )1 ketamine with 1.16 ±.5 mg kg )1 xylazine) 38 Ó 29 The Authors. Journal compilation Ó 29 Association of Veterinary Anaesthetists, 36, 34 41

6 Table 4 Stages of yohimbine-induced recovery of ketamine (2.32 ± 1.1 mg kg )1 ) and xylazine (1.16 ±.5 mg kg )1 ) induced anaesthesia in the captive Asiatic lions, leopards and tigers Stages of yohimbine-induced recovery Species Yohimbine (mg kg )1 body weight) Number of animals Onset of arousal (minutes) Sternal recumbency (minutes) Standing/recovery time (minutes) Lion Saline ± ± ± 15.5 a ± ± ± 1. b ± ± ± 1. c Leopard Saline ± ± ± 12.5 a ± ± ± 3.2 b ± ± ± 3.5 b Tiger Saline ± ± ± 2.2 a ± ± ± 1.5 c ± ± ± 1.6 d Values are represented as mean ± SD. Different superscripts in a column within a species differ significantly at b,c p <.1 and c,d p <.5. employed in this study were shown to effectively induce smooth and rapid anaesthesia within minutes with good muscle relaxation and profound analgesia (Patil et al. 1998; Shivaji et al. 1998; Jayaprakash et al. 21). Therefore, in the present study, attempts were not made to evaluate different doses of ketamine and xylazine as this would be stressful for the animals and was not acceptable by the animal ethics committee of the zoos in India. The anaesthetic induction time in lions, leopards and tigers in the present study was consistent with that reported for the free-ranging African lions and tigers (Herbst et al. 1985; Seal et al. 1987), despite lower doses used in our study. During the course of the study, we did not observe any sudden recovery or life threatening drug-effects in any of the animals. Animals remained anaesthetized for up to an hour with satisfactory analgesia and were laterally recumbent for about 8 12 minutes. Thus, our observations with a large sample size (123 animals) demonstrated the effectiveness and usefulness of this drug combination for anaesthetizing wild felids in captivity either for research purposes or for routine veterinary manipulation procedures lasting up to 1 hour. The physiological parameters in all species recorded immediately after anaesthesia in the present study were consistent with previous reports (Herbst et al. 1985; Seal et al. 1987; Miller et al. 23; Jacquier et al. 26). However, two leopards and one tiger had elevated rectal temperatures (up to 42.5 C) because of excess struggling and over excitement of the animals during darting but it decreased gradually and no morbidity occurred. Although we did not evaluate the serial recordings of physiological parameters, as a result of the experimental procedures involved, none of the animals showed any signs of cardiopulmonary or thermoregulatory dysfunction throughout the immobilization period. Yohimbine is a known potent alpha-two adrenergic antagonist and has been used as an antagonist for xylazine-induced sedation and also in ketamine xylazine anaesthesia in wild ungulates (Jessup et al. 1983; Sontakke et al. 27). Studies in the domestic cat (Hsu & Lu 1984) and the Bengal tiger (Seal et al. 1987) suggested that yohimbine could hasten the recovery of ketamine xylazine induced anaesthesia. In the present study, two different doses of yohimbine (.1 and.15 mg kg )1 body weight) were used in these felids, based on earlier studies in the domestic cat (.1 mg of yohimbine kg )1 body weight) (Hsu & Lu 1984) and in ungulates (.15 mg of yohimbine kg )1 body weight) (Jessup et al. 1983; Sontakke et al. 27). Intravenous administration of either dose of yohimbine was found to be effective in hastening the recovery from ketamine xylazine anaesthesia in a dose-dependent manner. Yohimbine significantly shortened the recovery from anaesthesia time period of 8 12 minutes in control animals that did not receive yohimbine to 5 15 minutes following yohimbine injection and eliminated the excitatory behaviour as Ó 29 The Authors. Journal compilation Ó 29 Association of Veterinary Anaesthetists, 36,

7 Recovery time (minutes) Recovery time (minutes) Recovery time (minutes) Asiatic lion Leopard Tiger.1 mg kg 1.15 mg kg 1 Yohimbine Figure 2 Effect of duration of ketamine xylazine anaesthesia (n 11 2 minutes, h 21 3 minutes and n 31 4 minutes) on the yohimbine-induced anaesthetic recovery in the captive Asiatic lions, leopards and tigers. recorded in control animals. However, a few leopards exhibited seizures for a very short period (2 3 minutes) immediately after yohimbine administration but without any life-threatening incidents. The reason for these adverse reactions could probably be an individual sensitivity to a particular drug or could be because of continuing ketamine effects since these animals had received yohimbine after only minutes of anaesthesia. Respiratory rates in all animals increased significantly (p <.1) following yohimbine injection and the pattern became more regular with a marked increase in thoracic breathing. The shortest recovery following yohimbine-treatment was observed in lions (4.7 ± 1. minutes) followed by tigers (7.7 ± 1.6 minutes) and leopards (1.8 ± 3.5 minutes) and was correlated significantly (p <.1) with yohimbine doses in lions and tigers but not in leopards. The rapid antagonism of ketamine xylazine anaesthesia by yohimbine has been attributed to the ability of yohimbine to antagonize xylazine effects (Hsu 1983). However, yohimbine may have a stimulant effect which shortened ketamine-induced anaesthesia (Hatch et al. 1983). Such a speedy recovery is always advantageous in wild felids, especially in freeranging animals, and for research purposes. A similar recovery period (4 8 minutes) was recorded in tigers administered with yohimbine (Seal et al. 1987). Atipamezole has also been used as an antagonist in lions and tigers but the recovery period was long and inconsistent. In tigers, recovery was achieved in 6 27 minutes after atipamezole administration (Miller et al. 23), whereas in lions, atipamezole-induced recovery varied from 14 to 338 minutes (Jacquier et al. 26). In fact, atipamezole is a more potent, selective and specific alpha-two adrenergic antagonist compared to yohimbine. We did not use atipamezole because of its lack of availability in India. However, the results of the present study and also the study by Seal et al. (1987) suggested that yohimbine was more effective in antagonizing/hastening the anaesthetic recovery in these felids. In the present study, we observed a variation in response to yohimbine-induced recovery between the species. This could be due to differences in the adrenergic receptor population or due to variation in the peripheral metabolism of the drug (Kreeger et al. 1986). But the precise reason for these differences could not be determined and thus further pharmacological studies are warranted. The species-specific response to yohimbine in these felids also supports our finding in an antelope, Black buck (Antelope cervicapra), in which yohimbine completely failed, whereas tolazoline proved to be very effective, in antagonizing ketamine xylazine anaesthesia (Sontakke SD, Umapathy G, Patil MS, Shivaji S, unpublished data). Moderate to deep resedation may occur if the dose of antagonist or the duration of anaesthesia is insufficient (Tyler et al. 199). However, in the present study, no resedation effects were noticed, thereby indicating the doses of yohimbine used in our study were sufficient to antagonize the drug effects. Antagonism of anaesthesia might uncover residual effects of ketamine if the antagonist is administered too early (<3 minutes) (Kreeger et al. 22). However, no such side effects or excitation 4 Ó 29 The Authors. Journal compilation Ó 29 Association of Veterinary Anaesthetists, 36, 34 41

8 signs were observed during recovery in the present study. Similar findings were also recorded in our earlier study in Axis deer (Sontakke et al. 27). In conclusion, we recommend ketamine xylazine anaesthesia for immobilization of captive Indian wild felids and yohimbine to hasten recovery from anaesthesia. Acknowledgements The authors thank the Department of Biotechnology, Government of India, the Department of Biotechnology, Government of Andhra Pradesh and the Central Zoo Authority of India, Ministry of Environment and Forests, Government of India for financial support. They would also like to thank the Chief Wildlife Wardens of Andhra Pradesh, Gujarat, Karnataka and Orissa for kindly providing the animals for the above studies. Their sincere thanks are due to Dr Kholkute for his constant suggestions and encouragement. References Amend JF (1972) Premedication with xylazine to eliminate muscular hypertonicity in cats during ketamine anaesthesia. Vet Med Small Anim Clin 67, Grassman LI Jr, Austin SC, Tewes ME et al. (24) Comparative immobilization of wild felids in Thailand. J Wildl Dis 4, Hatch RC, Booth NH, Kitzman JV et al. (1983) Antagonism of ketamine anaesthesia in cats by 4-aminopyridine and yohimbine. Am J Vet Res 44, Herbst LH, Packer C, Seal US (1985) Immobilization of free-ranging African lions (Panthera leo) with a combination of xylazine hydrochloride and ketamine hydrochloride. J Wildl Dis 21, Hsu WH (1983) Antagonism of xylazine-induced CNS depression by yohimbine in cats. Calif Vet 37, Hsu WH, Lu ZX (1984) Effects of yohimbine on xylazineketamine anaesthesia in cats. J Am Vet Med Assoc 185, Jacquier M, Aarhaug P, Arnemo JM et al. (26) Reversible immobilization of free-ranging African lions (Panthera leo) with medetomidine-tiletamine-zolazepam and atipamezole. J Wildl Dis 42, Jayaprakash D, Patil SB, Kumar MN et al. (21) Semen characteristics of the captive Indian leopard (Panthera pardus). J Androl 22, Jessup DA, Clark WE, Gullet PA (1983) Immobilization of mule deer with ketamine and xylazine and reversal with yohimbine. J Am Vet Med Assoc 183, Kreeger TJ, Del Giudice GD, Seal US et al. (1986) Immobilization of white-tailed deer with xylazine hydrochloride and ketamine hydrochloride and antagonism by tolazoline hydrochloride. J Wildl Dis 22, Kreeger TJ, Arnemo JM, Raath JP (22) Aplha-2-adrenergic antagonists. In: Handbook of Wildlife Chemical Immobilization. Kreeger TJ, Arnemo JM, Raath JP (eds). Wildlife Pharmaceuticals, Inc., Fort Collins, Colorado. pp Miller M, Weber M, Neiffer D et al. (23) Anaesthetic induction of captive tigers (Panthera tigris) using a medetomidine-ketamine combination. J Zoo Wildl Med 34, Patil SB, Jayaprakash D, Shivaji S (1998) Cryopreservation of semen of tigers and lions: computerized analysis of the motility parameters of spermatozoa. Curr Sci 75, Seal US, Armstrong DL, Simmons LG (1987) Yohimbine hydrochloride reversal of ketamine hydrochloride and xylazine hydrochloride immobilization of Bengal tigers and effects on hematology and serum biochemistries. J Wildl Dis 23, Shivaji S, Jayaprakash D, Patil SB (1998) Assessment of inbreeding depression in big cats: testosterone levels and semen analysis. Curr Sci 75, Shivaji S, Kholkute SD, Verma SK et al. (23) Conservation of wild animals by assisted reproduction and molecular marker technology a review. Indian J Exp Biol 41, Sontakke SD, Reddy A, Umapathy G et al. (27) Ketamine-xylazine induced anaesthesia in captive Indian Axis deer (Axis axis) and its reversal with yohimbine hydrochloride. Am J Vet Res 68, Tyler NJC, Hotvedt R, Bix AS et al. (199) Immobilization of Norwegian reindeer (Rangiferus tarandus tarandus) and Svalbard reindeer (R.t. platyrhynchus) with medetomidine and medetomidine-ketamine and reversal of immobilization with atipamezole. Acta Vet Scand 31, Received 27 February 27; accepted 7 August 27. Ó 29 The Authors. Journal compilation Ó 29 Association of Veterinary Anaesthetists, 36,