Catherine J.A. Williams

Anaesthetic induction with alfaxalone in the ball python (Python regius): dose response and effect of injection site

OBJECTIVE To characterise the minimum dose of intramuscular alfaxalone required to facilitate intubation for mechanical ventilation, and to investigate the impact of cranial versus caudal injection on anaesthetic depth. STUDY DESIGN Randomised crossover study. ANIMALS Six healthy juvenile ball pythons (Python regius). METHODS Three dosages (10, 20 and 30 mg kg-1) of alfaxalone were administered to each python in a caudal location with a minimum 2 weeks washout. Induction and recovery were monitored by assessing muscle tone, righting reflex, response to a noxious stimulus and the ability to intubate. A subsequent experiment assessed the influence of injection site by comparing administration of 20 mg kg-1 alfaxalone in a cranial location (1 cm cranial to the heart) with the caudal site. Respiration rate was monitored throughout, and when intubation was possible, snakes were mechanically ventilated. RESULTS Regardless of dose and injection site, maximum effect was reached within 10.0 ± 2.7 minutes. When administered at the caudal injection site, intubation was only successful after a dosage of 30 mg kg-1, which is higher than in previous reports for other reptiles. However, intubation was possible in all cases after 7.2 ± 1.6 minutes upon cranial administration of 20 mg kg-1, and anaesthetic duration was significantly lengthened (p < 0.001). Both 30 mg kg-1 at the caudal site and 20 mg kg-1 at the cranial site led to apnoea approximately 10 minutes post-injection, at which time the snakes were intubated and mechanically ventilated. CONCLUSIONS AND CLINICAL RELEVANCE Alfaxalone provided rapid, smooth induction when administered intramuscularly to pythons, and may serve as a useful induction agent prior to provision of volatile anaesthetics. The same dosage injected in the cranial site led to deeper anaesthesia than when injected caudally, suggesting that shunting to the liver and first-pass metabolism of alfaxalone occur when injected caudally, via the renal portal system.

A critical test of Drosophila anaesthetics: Isoflurane and sevoflurane are benign alternatives to cold and CO2

Anaesthesia is often a necessary step when studying insects like the model organism Drosophila melanogaster. Most studies of Drosophila and other insects that require anaesthesia use either cold exposure or carbon dioxide exposure to induce a narcotic state. These anaesthetic methods are known to disrupt physiology and behavior with increasing exposure, and thus ample recovery time is required prior to experimentation. Here, we examine whether two halogenated ethers commonly used in vertebrate anaesthesia, isoflurane and sevoflurane, may serve as alternative means of insect anaesthesia. Using D. melanogaster, we generated dose-response curves to identify exposure times for each anaesthetic (cold, CO2, isoflurane and sevoflurane) that allow for five-minutes of experimental manipulation of the animals after the anaesthetic was removed (i.e. 5min recovery doses). We then compared the effects of this practical dose on high temperature, low temperature, starvation, and desiccation tolerance, as well as locomotor activity and fecundity of female flies following recovery from anaesthesia. Cold, CO2 and isoflurane each had significant or near significant effects on the traits measured, but the specific effects of each anaesthetic differed, and effects on stress tolerance generally did not persist if the flies were given 48h to recover from anaesthesia. Sevoflurane had no measureable effect on any of the traits examined. Care must be taken when choosing an anaesthetic in Drosophila research, as the impacts of specific anaesthetics on stress tolerance, behavior and reproduction can widely differ. Sevoflurane may be a practical alternative to cold and CO2 anaesthesia in insects - particularly if flies are to be used for experiments shortly after anesthesia.

The influence of mechanical ventilation on physiological parameters in ball pythons (Python regius)

Mechanical ventilation is widely recommended for reptiles during anesthesia, and while it is well-known that their low ectothermic metabolism requires much lower ventilation than in mammals, very little is known about the influence of ventilation protocol on the recovery from anesthesia. Here, 15 ball pythons (Python regius) were induced and maintained with isoflurane for 60min at one of three ventilation protocols (30, 125, or 250mlmin-1kg-1 body mass) while an arterial catheter was inserted, and ventilation was then continued on 100% oxygen at the specified rate until voluntary extubation. Mean arterial blood pressure and heart rate (HR) were measured, and arterial blood samples collected at 60, 80, 180min and 12 and 24h after intubation. In all three groups, there was evidence of a metabolic acidosis, and snakes maintained at 30mlmin-1kg-1 experienced an additional respiratory acidosis, while the two other ventilation protocols resulted in normal or low arterial PCO2. In general, normal acid-base status was restored within 12h in all three protocols. HR increased by 143±64% during anesthesia with high mechanical ventilation (250mlmin-1kg-1) in comparison with recovered values. Recovery times after mechanical ventilation at 30, 125, or 250mlmin-1kg-1 were 289±70, 126±16, and 68±7min, respectively. Mild overventilation may result in a faster recovery, and the associated lowering of arterial PCO2 normalised arterial pH in the face of metabolic acidosis.

CARDIOVASCULAR EFFECTS OF ALFAXALONE AND PROPOFOL IN THE BULLFROG, LITHOBATES CATESBEIANUS

Abstract Alfaxalone is becoming a popular anesthetic for nonmammalian vertebrates, but the physiological effects of its administration remain largely unknown in these taxa. Therefore, the cardiovascular responses to a clinically relevant dose of alfaxalone (10 mg/kg) are reported in the bullfrog (Lithobates catesbeianus), following intramuscular (IM) and intravascular (IV) administration (via a femoral artery catheter) and compared with an IV dose of propofol, another parenteral GABA (γ-aminobutyric acid) agonist in common veterinary use as an induction agent. Heart rate (HR) and mean arterial blood pressure (MAP) (assessed by direct measurement from the catheter) are reported from under undisturbed conditions to assess both the direct effects of the drugs and the interaction with the stress of handling associated with IM injection of alfaxalone where IM administration is possible. Alfaxalone caused HR to increase significantly for over 45 min in both groups from a baseline of approximately 30 beats/min. This was significantly different from the lack of significant HR response on the IV administration of propofol. MAP increased in the peri-injection period with both routes of administration for alfaxalone but after IV use decreased significantly from 10 min following administration. Propofol did not affect blood pressure after 5 min from injection. Assessment of immobilization following intramuscular injection of alfaxalone in a pilot study was in accordance with the literature, as it provided no antinociception as a sole agent but did produce sedation and loss of righting reflex.

The effects of endogenous and exogenous catecholamines on hypoxic cardiac performance in red-bellied piranhas: JOYCE et al.

Catecholamines protect the fish heart during hypoxia. However, the humoral adrenergic stress response may only be invoked in extremis. We investigated the hypothesis that endogenous (e.g., neuronal) myocardial catecholamines may also impact cardiac performance during hypoxia in a hypoxia-tolerant tropical fish, the red-bellied piranha (Pygocentrus nattereri). First, we measured endogenous tissue catecholamines and in vitro catecholamine release from piranha myocardium using ultraperformance liquid chromatography. Ventricle homogenates contained detectable levels of both adrenaline (7.27 ng/g) and noradrenaline (14.48 ng/g), but only noradrenaline was released from ventricular tissue incubated in Ringers solution. Noradrenaline released in this assay was not affected by hypoxia but was promoted by the catecholamine releasing agent tyramine. Our second series of experiments explored cardiac contractile performance in vitro using tyramine, exogenous noradrenaline or adrenaline, and propranolol (a β-adrenoceptor antagonist). In ventricular strip preparations, β-adrenergic blockade with propranolol had no effects on twitch force or contraction kinetics in either normoxia or hypoxia, confirming that spontaneous endogenous catecholamine release did not impact cardiac performance. However, in the absence of propranolol, tyramine mimicked the positive inotropic effect of noradrenaline (10 µM) during hypoxia, although adrenaline was capable of generating larger effects. Our results suggest that, although it is not spontaneously released, inducible endogenous noradrenaline release may have a significant β-adrenoceptor-dependent impact on hypoxic performance in the fish heart.

EVALUATION OF FEEDING BEHAVIOR AS AN INDICATOR OF PAIN IN SNAKES

Abstract The necessity to prevent and manage pain in reptiles is becoming increasingly important, as their use in scientific research and popularity as exotic pets continues to rise. It was hypothesized that feeding behavior would provide an adequate indicator of pain perception in the ball python (Python regius). Normal feeding was defined the previous week, where a dead rodent was struck within 12 sec (n = 10). Eighteen pythons were randomly assigned to one of three treatments: anesthesia only (AO), chemical noxious stimulus (CS; capsaicin injection), or surgical noxious stimulus (SS; surgical incision). The time to strike was recorded 4 hr after the procedure and weekly during the subsequent 3 wk. Delayed feeding was observed in animals in the CS and SS groups, and normal feeding resumed after 1 and 3 wk, respectively. Spontaneous feeding remained uninterrupted for the AO group. These findings demonstrate feeding behavior as a potential model to assess pain in snakes.

Long-term surgical anaesthesia with isoflurane in human habituated Nile Crocodiles

A suitable long-term anaesthetic technique was required for implantation of physiological sensors and telemetric devices in sub-adult Nile crocodiles (Crocodylus niloticus) to allow the collection of physiological data. Five Nile crocodiles with a median body mass of 24 kg were used. After manual capture, they were blindfolded and 0.2 mL (1 mg/mL) medetomidine was administered intramuscularly in four of the animals which had an estimated body mass between 20 kg and 30 kg. One crocodile with an estimated body mass of 50 kg received 0.5 mL. For induction, 5 mL propofol (10 mg/mL) was injected intravenously into the occipital sinus. Additional doses were given when required to ensure adequate anaesthesia. Anaesthesia was maintained with 1.5% isoflurane. Ventilation was controlled. Local anaesthesia was administered for surgical incision and external placement of the radio transmitter. Medetomidine was antagonised with atipamezole at the end of surgery. Median heart rate during surgery was 22 beats/min, at extubation 32 beats per min and 30 beats per min the following day at the same body temperature as under anaesthesia. Median body temperature of the animals increased from 27.3 °C to 27.9 °C during anaesthesia, as room temperature increased from 24.5 °C to 29.0 °C during surgery. Anaesthesia was successfully induced with intramuscular medetomidine and intravenous propofol and was maintained with isoflurane for the placement of telemetric implants. Intraoperative analgesia was supplemented with lidocaine infiltration. Perioperative physiological parameters remained stable and within acceptable clinical limits. Multiple factors appear to influence these variables during the recovery period, including residual anaesthetic effects, environmental temperature and physical activity.

When local anesthesia becomes universal: Pronounced systemic effects of subcutaneous lidocaine in bullfrogs (Lithobates catesbeianus)

Sodium channel blockers are commonly injected local anesthetics but are also routinely used for general immersion anesthesia in fish and amphibians. Here we report the effects of subcutaneous injection of lidocaine (5 or 50mgkg-1) in the hind limb of bullfrogs (Lithobates catesbeianus) on reflexes, gular respiration and heart rate (handled group, n=10) or blood pressure and heart rate via an arterial catheter (catheterized group n=6). 5mgkg-1 lidocaine did not cause loss of reflexes or change in heart rate in the handled group, but was associated with a reduction in gular respiratory rate (from 99±7 to 81±17breathsmin-1). 50mgkg-1 lidocaine caused a further reduction in respiratory rate to 59±15breathsmin-1, and led to a progressive loss of righting reflex (10/10 loss by 40min), palpebral reflex (9/10 loss at 70min), and contralateral toe pinch withdrawal (9/10 loss at 70min). Reflexes were regained over 4h. Systemic sedative effects were not coupled to local anti-nociception, as a forceps pinch test at the site of injection provoked movement at the height of the systemic effect (tested at 81±4min). Amphibians are routinely subject to general anesthesia via exposure to sodium channel blockers such as MS222 or benzocaine, however caution should be exercised when using local injectable lidocaine in amphibians, as it appears to dose-dependently cause sedation, without necessarily preventing local nociception for the duration of systemic effects.