Potassium chloride as a euthanasia agent in psittacine birds: Clinical aspects and consequences for histopathologic assessment

Abstract

Twelve parrots anesthetized with isoflurane were euthanized intravenously (IV) with 3 or 10 mEq/kg body weight (BW) of potassium chloride (KCl) resulting in ventricular asystole at 68.0 s and 32.8 s, respectively. Mild vocalization (1/6 birds, 3 mEq/kg BW) and involuntary muscle tremors (5/6 birds, 10 mEq/kg BW) were noted. Unlike barbiturates or T-61 no histologic artefacts resulted from this technique.

Barbituric acid derivatives, most specifically pentobarbital sodium, are the most frequently used injectable agents for humane euthanasia of mammals, and are also used for the same purpose in birds (1–4). Other injectable euthanasia agents considered acceptable by the American Veterinary Medical Association and the Canadian Council on Animal Care include intravenous T-61 (Intervet Canada, Kirkland, Quebec) and potassium chloride (KCl) injected by the intracardiac or intravenous routes under general anesthesia (1,2).

Pentobarbital sodium (Euthanyl; Bimeda-MTC Animal Health, Vétoquinol, Lavaltrie Quebec) is licensed for use in Canada by the intravenous (IV) route for euthanasia of dogs, cats, horses, cattle, mink, laboratory animals, and birds. Barbiturates cause death by their effects on the central nervous system; a rapid loss of consciousness is followed by depression of the respiratory centers and apnea, and then by cardiac arrest (1). Disadvantages of the use of barbiturate agents include the regulatory requirements associated with controlled drugs, and the potential for secondary barbiturate toxicity stemming from animals scavenging or being fed the carcasses of euthanized animals.

T-61, an injectable non-narcotic agent, is a mixture of 3 compounds: 1) embutramide, a general anaesthetic with narcotic effects that depresses the respiratory center in the brain; 2) mebenzonium iodide, a non-depolarizing neuromuscular blocking agent that paralyzes the skeletal muscles; and 3) tetracaine hydrochloride, a very potent local anaesthetic, in a mixture of water and the organic solvent dimethyl-formamide (5). In Canada, T-61 is licensed for intravenous use in dogs. This product has been reported to be advantageous over pentobarbital for euthanasia in various species, since the terminal gasping seen with pentobarbital anesthesia is not seen with T-61. However, concerns have been expressed regarding humane aspects of its use if dosage and injection recommendations are not followed (3,5,6). The issue of possible tissue residues in animals euthanized with T-61 has not been studied.

Intravenous or intracardiac injection of KCl is an acceptable method for humane euthanasia of animals, particularly livestock or wildlife, but only for animals under general anesthesia (1–3). Death occurs due to cardiotoxic effects; high levels of serum potassium lead to delayed electrical conduction between myocytes, sinoatrial and atrioventricular block, and at high enough levels, a complete block in ventricular conduction, ventricular fibrillation, and fatal asystole (7,8). General anesthesia is required prior to KCl administration to ensure lack of conciousness while cardiac arrest is being induced, particularly as seizures and muscle spasms may occur (1,3). Potassium chloride is not a controlled substance, and does not result in secondary toxicity from tissue residues.

When tissues are to be evaluated by histopathology, both barbiturate agents and T-61 are problematic in that they cause considerable tissue artefact, in a dose-related manner. Barbiturate injection results in endothelial damage and has direct caustic effects leading to congestion, edema, hemorrhage, hemolysis, emphysema, and cell destruction in various organs including lungs, kidney, heart, liver and the gastrointestinal serosa. This occurs in a variety of species including laboratory animals, sheep, dogs, cats, and nonhuman primates (9). T-61 causes dose- and time-dependent tissue toxicity including endothelial necrosis, hemolysis, pulmonary congestion, and edema in cats, dogs, monkeys, and sheep and is not recommended for euthanasia when postmortem histopathological studies are to be carried out, especially in pulmonary tissue (10).

In the collective experience of the authors, based on avian necropsy submissions through the Department of Pathobiology of the Ontario Veterinary College, University of Guelph, Canada, the use of both barbiturates and T-61 for humane euthanasia frequently causes extensive tissue damage, especially in the lungs, leading to difficulties in detailed histopathologic analysis. At the same time, it was observed that similar artefacts were never seen in the tissues from birds that had been euthanized with intravenous KCl. Hence it was decided to evaluate the clinical and histopathological aspects of the use of KCl to euthanize psittacine birds.

Twelve psittacine birds from a research colony being depopulated were used for the study. All procedures involving animals were approved by the University of Guelph Animal Care Committee in accordance with the requirements of the Animals for Research Act of Ontario, revised 1990, and the recommendations of the Canadian Council for Animal Care.

The study population was divided into 6 blocks, 5 of these consisted of 2 birds of the same species, and 1 block consisted of a white-eyed conure and a white-capped pionus (Table 1). The treatments consisted of low and high doses of KCl (Hospira, Lake Forest, Illinois, USA), 3 mEq/kg body weight (BW) and 10 mEq/kg BW, respectively, administered intravenously (IV) for euthanasia under general anesthesia. Within each block, the treatment was allocated randomly.

Table 1.

Dose of potassium chloride administered IV for euthanasia and the time taken to reach electrical ventricular asystole in 12 psittacine birds

Bird	Species	Sex	IV dose of KCl (mEq/kg BW)	Time to reach electrical ventricular asystole (s)
1	Moluccan cockatoo (Cacatua moluccensis)	Female	10	14
2	Moluccan cockatoo (Cacatua moluccensis)	Male	3	22
3	Sun conure (Aratinga solstitialis)	Female	3	Not examined
4	Sun conure (Aratinga solstitialis)	Male	10	14
5	Blue fronted Amazon (Amazona aestiva)	Female	3	69
6	Blue fronted Amazon (Amazona aestiva)	Male	10	11
7	African grey (Psittacus erithacus timneh)	Male	10	23
8	African grey (Psittacus erithacus timneh)	Female	3	32
9	Mealy Amazon (Amazona farinosa)	Male	3	84
10	Mealy Amazon (Amazona farinosa)	Male	10	45
11	White eyed conure (Aratinga leucophthalma)	Male	10	90
12	White capped pionus (Pionus senilis)	Male	3	137

After manual restraint with a towel, each bird was mask induced with 5% isoflurane (Forane; Baxter, Deerfield, Illinois, USA) for general anesthesia. Once the anesthetic plane deepened and wing tone was lost, the birds were intubated with uncuffed endotracheal tubes of appropriate sizes and were maintained at 2.5% isoflurane concentration. A sample of whole blood was collected from each bird from the right jugular vein for a complete blood (cell) count and a serum biochemical profile. A 24-gauge IV catheter (BD Insyte-W; Singapore) was placed in the medial ulnar vein for the purpose of administration of KCl. A portable ECG machine with ECG recording facility (MAC5500; GE Healthcare, Milwaukee, Wisconsin, USA) was used for monitoring the electrical cardiac activity during the procedure. Standard ECG leads were connected to both the wing webs and near the proximal attachment of the gastrocnemius muscle on both legs (11). A thin piece of gauze dipped in alcohol was placed between the alligator clips of the leads and the skin to enhance the electrical conduction as well as to soften the grip of the clips. An ECG strip was run for a few seconds at speeds of 25 mm/s and 50 mm/s alternately, prior to the administration of the KCl to assess the normal ECG for each bird. Continuous ECG monitoring was started immediately prior to the administration of the IV bolus of KCl with the speed of the ECG output set at 50 mm/s. The point of time at which the IV bolus administration was started was recorded. Cardiac activity was also monitored by auscultation, and any other patient activity, including vocalization, muscle tremors, and muscle contraction, was also recorded. The endpoint was defined as the time at which the heart beats could not be auscultated and, on ECG, electrical ventricular asystole lasted more than 3 s. The time taken from administration of the assigned dose of KCl to the endpoint was recorded. The euthanasia procedure was videotaped from the point where the birds attained a surgical anesthetic plane to the point the birds were declared dead in order to more closely correlate ECG changes with any patient activity.

Sections of formalin-fixed, paraffin-embedded tissues, including cerebrum, cerebellum, brain stem, spinal cord, brachial nerve, vagus, heart, adrenal gland, crop, proventriculus, ventriculus, duodenum, kidney, lungs, pancreas, ovary/testes, and thyroid and parathyroid glands from each bird were stained with hematoxylin and eosin. A pathologist blinded to treatment subjectively evaluated the slides for histopathologic quality of the sections and presence of tissue artefacts. Histopathological sections of the lungs (the most common site of euthanasia induced artefacts in various species) and the heart (the site of action of KCl for euthanasia) were considered particularly relevant.

There were no abnormalities observed on pre-euthanasia ECGs of any bird comparied to published normal values for similar species (11–14). The ECG data (after administration of the agent) could not be monitored in 1 patient (Bird 3) due to technical reasons. The time taken for cessation of the cardiac electrical activity after administration of KCl for each bird is shown in Table 1. The average time to reach endpoint for birds administered the lower dose of KCl was 68.80 s (1.15 min), and for the higher dose was 32.83 s (0.55 min). Within each block, the time to death was shorter for the higher dose compared to the lower dose.

One bird (Bird 5) vocalized mildly for a few seconds immediately after administration of the lower dose of KCl. Birds 1, 4, 6, 7, and 10 (all receiving the higher dose) exhibited mild muscle tremors of the wings and involuntary contraction of the skeletal muscles, most notably the leg muscles, immediately after cessation of the heartbeat. Muscle contractions were not noted in any bird receiving the lower dose of KCl.

There was no evidence, based on clinical pathology, the pre-euthanasia ECGs, and the histopathology of the heart, of an underlying electrolyte abnormality or cardiac condition in any bird that might have influenced the outcome of this trial.

Both doses of KCl administered intravenously (3 mEq/kg and 10 mEq/kg BW) resulted in rapid death, although a faster euthanasia was achieved with the higher dose. Both these doses are higher than the recommended intravenous dose of KCl (1 to 2 mEq/kg BW) for euthanasia of small animals under general anesthesia (1). Unpublished experience by the authors with birds administered doses of 1 to 2 mEq/kg BW was that additional drug was sometimes necessary.

Electrocardiographic changes observed after administration of KCl to the birds of this study included an initial elevation of the S-T segment followed by its depression, prolongation of the P-R intervals, widening of the QRS complexes, progressive bradycardia, atrioventricular dissociation and finally ventricular electrical asystole. Electrocardiographic changes reported in mammals with hyperkalemia include prolongation of the P-R intervals, peaking of T waves, widening of QRS complexes, absence of a P wave and finally sine wave appearance at extremely high potassium levels (7,8,15,16). Peaking of T waves, one of the classic ECG changes in mammals with hyperkalemia, was not observed in our study.

Involuntary skeletal muscle contractions were seen after cessation of the heartbeats in birds administered the high dose of KCl. Potassium is an important intracellular cation that maintains cell membrane potential and participates actively in depolarization and contraction of muscle fibers. Hyperkalemia initially affects the more sensitive cardiac muscle fibers leading the heart to arrest in systole. Higher systemic concentrations likely had a similar effect on the skeletal muscle fibers causing depolarization and sustained muscle contractions leading to limb movement, most noticeably of the legs. The only high dose bird in which limb movement was not noted was Bird 11, whose legs were restrained at the time of KCl administration. These muscle contractions might be unacceptable in clinical practice when the owner of a bird is witnessing the procedure. In this situation, the lower dose of potassium chloride may be more appropriate; however, as 1 bird that received the lower dose of the drug showed mild vocalization, some involuntary muscle activity may still occur. Deepening the anesthetic plane with a higher concentration of isoflurane might negate this effect. In our study, birds were at a moderate anesthetic plane with the isoflurane vaporizer set at 2.5%.

None of the histopathological sections examined, including lung and heart, had artefacts that could be attributable to the euthanasia agent (Figure 1). This is likely because the KCl solution does not contain any directly cytotoxic chemicals, solvents, or other agents that can damage tissues or cells, unlike barbiturates and T-61.

Figure 1.

Microscopic images of the lungs of birds euthanized with 3 euthanasia agents. Note excellent anatomic detail seen in the lung of a bird euthanized with (a) potassium chloride compared to (b) T-61, and (c) pentobarbital sodium. The latter 2 show erythrolysis with many erythrocyte bare nuclei and nuclear remnants, and accumulation of eosinophilic granular material and fluid in airways. PB — parabronchus, A — atria, PP — pulmonary parenchyma. Bar = 50 μm. Hematoxylin and eosin.

In conclusion, our recommended method of euthanasia for psittacine birds whose tissues will be submitted for histopathologic assessment is the intravenous injection of KCl, at 3 mEq/kg BW. For humane reasons, birds must be under at least a moderate plane of anesthesia prior to injection. While the lack of histopathologic artefacts would apply to any species of animal euthanized with KCl, the minimal effective dose of the agent may vary.