Fish & Invertebrate Restraint & Anesthesia

Question 1

What are some of the anatomical and physiologic considerations that need to be planned for when anesthetizing fish?

What are some of the different respiratory strategies? How does that affect your anesthesia?

What are the two types of muscle? How does that affect absorption?

How do fish regulate temperature and metabolism?

Answer 1

Respiratory Systems

• · most fish-> gas exchange through forcing water over the gill surface.
• o tuna/pelagic sharks -> ram ventilation -> forward propulsion of the body forces water through gills.
• o other sp -> gas exchange through outpouchings of alimentary track or modified swim bladders.
• § some are obligate nasal breathers.
• § adapted to hypoxic environments 🡪 pacu, bettas, gouramis, snakeheads.
• § usually require higher immersion anesthetic concentrations.
• o skin -> up to 30% O2 uptake in some sp. -> higher in marine fish, young fish, scaleless/fine-scaled sp.
• § immersion anesthesia may be lower compared to freshwater sp.

Skin and Muscle.

• · scales can make IM/IV injections challenging without restraint/sedation/anesthesia in immersion.
• · 2 types of muscle.
• o red - aerobic, slow-oxidative 🡪 dorsal midline of body 🡪 anesthetic injections may have more rapid induction/clearance.
• o white - anaerobic, fast-twitch.

Temperature and Metabolism.

• · most are ectotherms - colder temp = colder metabolism of drugs.
• · tuna, some laminid sharks - endothermic = may need higher dosages.

Question 2

What considerations about water quality need to be made while anesthetizing fish?

How does DO change? What physiologic changes occur during induction of anesthesia that may make low DO worse?

How should pH be monitored throughout anesthesia?

How does ammonia accumulate throughout the procedure?

Answer 2

Water Quality Considerations.

Dissolved Oxygen (DO) and Temperature.

· induction - hypoventilation and reflex bradycardia.

o + low DO - hypoxia.

· higher water temp - tissue oxygen demand increases, DO decreases.

o hypoxia more likely to occur during anesthesia in higher temps - tropical marine fish most at risk due to few hypoxic adaptations.

pH and Nitrogenous Waste.

· marine environment – narrow pH 8-8.3 range.

· buffer all immersion anesthetics, check pH levels prior to immersion and throughout procedures.

· ammonia accumulates in all anesthesias – risk of toxicity is greater in marine/brackish systems 🡪 higher pH favors presence of more toxic unionized ammonia (NH3).

· visual indicators of deteriorating water quality – surface foam formation, mucus in the water.

· change 20-25% of the water.

Ionic Balance.

· ionic and organic compounds differ between water sources.

· use source water for induction, maintenance, and recovery.

Question 3

What preparations should be made prior to anesthetizing a fish?

What considerations for human safety should be made?

How long should fish be fasted for?

Answer 3

Anesthetic Techniques and Drugs.

Human Safety.

• · fish as hazard to humans – trauma (teeth, spines), electric shocks, venom, zoonotic disease.
• · gloves worn at all times.

Preanesthetic Preparation.

• · baseline behavior – ventilation. color, caudal fin stroke rate, position in water column, activity level.
• · consider 12-24 hour fast to reduce risk of regurgitation – dirties gill tissue, increases ammonia levels.
• o continue to feed fish that are young/small and fed frequently.
• · physical and chemical variables should match water used for all components of anesthesia.
• · provide supplemental ventilation, ensure adequate moisture maintenance when out of the water.

Question 4

Describe immersion anesthesia in fish.

What makes MS-222 so quick to be absorbed?

How does ventilation change over time? How should you adjust?

For long procedures, how should concentrations be adjusted?

What is an appropriate flow rate for recirculation of anesthetic water?

What potential for gill damage is present with anesthesia?

Answer 4

Immersion Anesthesia.

· most common method used.

· agents enter the bloodstream via the gills, accessory respiratory organs, ± skin.

o rapidly pass to CNS.

o efficiency of uptake related to lipid solubility – MS22 has high lipid solubility and rapid uptake.

· spontaneous ventilation decreases esp at longer events or greater depth.

o bulb/other syringe to flush water over the gills -> non-recirculating anesthetic water -> suitable for short procedures.

o flow should follow the normal direction of water movement over the gills to maximize gas/anesthetic exchange.

o recirculating systems for larger fish, longer procedures, anesthetic drug cost/wastewater concerns exist.

§ gravity-driven or pump-driven delivery system that feeds water over the gills via tubing.

§ close monitoring of DO and ammonia.

· maintaining a fish for longer than 15 minutes at induction dose can lead to excessive anesthetic depth.

o prepare multiple anesthetic solutions at different concentrations.

o pre-measures volumes of water and anesthetic/buffer to titrate the dose.

o deliver anesthetic-free water directly to the gills via bulb or syringe.

§ can directly adjust the dose through here as well.

· flow rate to regulate anesthetic depth -> 1-3 L/min/kg.

o too high -> gastric dilatation.

o too slow -> lack of fluid exiting both opercula.

· gills are very fragile -> trauma, inadequate perfusion, prolonged exposure to inadequate buffered immersion agents.

· recovery -> accessory respiratory organs, skin, kidneys also assist w. elimination of immersion agents through gills.

o consider changing out some of the water -> eliminated agents can be reabsorbed.

Question 5

What immersion agents are used in the anesthesia of fish?

What is the mechanism of MS-222? How safe is this drug?

What are the safety concerns with eugenol?

Describe the use of alfaxalone, propofol, and metomidate as immersion agents.

Answer 5

• Tricaine Methanesulfonate (MS-222).
  
  • most common agent.
  • benzocaine derivative.
  • reduces water pH -> use buffer like sodium bicarbonate (baking soda) or sodium carbonate (soda ash).
    
    • sodium carbonate is much more potent buffer.
  • wide safety index.
  • higher concentrations needed -> freshwater, obligate/facultative air-breathing species, individuals undergoing regular anesthesias.
  • lower concentrations -> brackish/SW species, young fish, soft/warm water.
  • FDA approved in teleosts.
  • reversible retinal deficits in humans w. long-term exposure.
• Clove Oil, Eugenol, Isoeugenol, and Aqui-S.
  
  • clove oil = 84% eugenol + isoeugenol + methyleugenol.
  • available over-the-counter, much less expensive than MS-222.
  • incompletely water-soluble especially in cold temperatures.
    
    • common to mix a small amount of 95% ethanol as a solvent.
  • Aqui-S -> synthetic isoeugenol, more expensive water-soluble alternative.
  • consistent anesthesia, more rapid induction, longer recoveries.
    
    • less excitation during induction in elasmobranchs.
  • safety margin depends on taxa.
    
    • overall safety vs narrow safety -> ventilatory failure, medullary collapse.
    • possible direct neurotoxic/hepatotoxic effects, oil interference w. gill epithelia/gas exchange.
    • mild gill necrosis shown w. repeated exposure to eugenol.
  • NOT FDA APPROVED.
• Alfaxalone.
  
  • synthetic steroid -> binds to GABA receptors in CNS -> muscle relaxation, sedation, anesthesia.
  • water-soluble, neutral pH.
  • achieves surgical anesthesia in several FW species -> goldfish, koi, Oscars.
    
    • dose-dependent ocular rate depression.
  • no literature readily available on alfaxalone in marine species.
• Propofol.
  
  • ultra-short-acting alkylphenol derivative hypnotic -> GABA receptors in CNS (true MOA unknown).
  • studies mostly in FW fish -> whitefish, catfish, tilapia, sturgeon, tetra, trout, koi, goldfish.
  • prolonged recoveries and mortalities w. long procedures/high dosages.
  • opaque nature of solution makes good observation of fish challenging.
• Metomidate Hydrochloride.
  
  • nonbarbiturate imidazole -> hypnotic agent -> sleep rather than general anesthesia.
    
    • blocks cortisol synthesis -> increase in melanocyte stimulating hormone production -> fish appear darker in color.
  • Aquacalm -> indexed drug for ornamental fish.
  • water-soluble; immersion or oral.
  • opercular ventilation maintained for longer than other immersions
  • low dosages -> muscle fasciculations, incomplete relaxation.
  • sedation/anesthesia for minor procedures, sedation during transport to limit trauma.
  • likely contraindicated in gouramis, cichlids when pH < 5.
  • recovery times increase w. increasing body weight.
• Other Immersion Agents.
  
  • benzocaine, 2-phenoxyethanol, quinaldine ± sulfate, azaperone.
  • isoflurane.
    
    • poor solubility -> high local concentrations can lead to anesthetic overdose -> control of anesthetic depth can be difficult.
    • drug volatilization, difficulty in scavenging waste gas -> hazard to personnel and environment.

Question 6

Describe the use of parenteral anesthesia in fish.

What route is least reliable?

what is the most common route?

What drugs are commonly used?

What complications have been observed?

Answer 6

Parenteral Anesthesia.

• oral -> least reliable.
• intracoelomic, IV -> manual ± chemical restraint for administration.
  
  • IV > ICe -> rapid induction, short predictable duration of effect, no risk to visceral damage.
• IM.
  
  • most common parenteral route.
  • manual restraint for hand injection, pole syringe, darting system.
  • dorsal epaxials -> expect drug leakage.
  • sedentary sp may have longer induction -> movement helps vascular circulation in fish.
  • large volumes may be required -> risk of sterile abscess formation.
• compared to immersion -> less predictable, longer recoveries requiring ventilatory support.
• Ketamine Hydrochloride.
  
  • short-acting cyclohexamine dissociative.
  • alone -> apnea, prolonged recoveries w. excitement, seizure-like muscle spasms in elasmos.
  • respiratory depression, bradycardia, incomplete immobilization.
  • combination protocols always recommended -> eg w. alpha-2s.
  • capture + initial restraint -> move to MS222/other for maintenance.
• Alpha-2-Adrenergic Agonists (Xylazine, Medetomidine, Dexmedetomidine).
  
  • xylazine-ketamine in sharks -> safe.
    
    • teleosts (esp salmonids) -> apnea, ECG abnormalities, seizure activity.
  • instead use medetomidine/dexmed + ketamine.
  • yohimbine for xylazine, atipamezole for medetomidine/dexmed.
• Benzodiazepines (Midazolam, Diazepam).
  
  • midaz/ket/dexmed -> safe in black seabass.
  • other sp -> fatal lactic acidosis.
  • beneficial in elasmos.
• Propofol.
  
  • IV in elasmos/sturgeon.
  • significant resp depression, prolonged recoveries in some sharks.
  • not practical for ram-ventilating sp.
• Alfaxalone.
  
  • prolonged excitement in higher doses in 2 marine sp.
  • FW koi -> unpredictable duration of anesthesia/opercular rate, high mortalities at higher doses.

Question 7

What are the stages of anesthesia in fish?

What cardiopulmonary monitoring can be performed during anesthesia?

What water quality monitoring should be performed?

How should fish be recovered?

How are fish resuscitated if issues arise?

Answer 7

Anesthetic Depth

• activity compared to normal, reaction to stimuli, righting reflex, jaw/muscle tone, ventilation, heart rate.
• stages.
  
  • sedation -> decrease in caudal fin stroke activity may be first sign of effect.
  • loss of equilibrium -> decreased swimming behavior, RR, reaction to stimuli.
  • short excitement phase on induction -> trauma (jumping from container), rapid/repeated flaring of opercular (“coughing” reflex).
  • anesthesia -> further decrease in RR/RE, total loss of muscle tone, no rxn to firm squeeze at base of tail, ventrodorsal recumbency (may also be stress response in some sp).
• immersion induction within 5-10 minutes.

Cardiopulmonary Activity.

• opercular rate (OR) typically easy to observe.
  
  • EXCEPT -> syngnathids (seahorses, pipefish), ram-ventilation sp (tuna, pelagic sharks).
• HR -> US, ECG, Doppler.
  
  • electrodes to pectoral and anal fins.
• pulse ox not effective.
• respiratory arrest occurs long before cardiovascular collapse.
  
  • decreased gill blood flow may lead to hypoxemia -> gill/fin pallor.
• periodic blood gas analyses.
  
  • pH and lactate -> higher accuracy than other blood gas analytes in fish esp elasmos.
  • trends over time still useful.

Water Quality Monitoring.

• temperature, DO, ammonia, pH at a minimum.
  
  • DO/pH tends to decrease, ammonia tends to increase.
• problems -> prolonged anesthesias, fish sequentially anesthetized in the same container.
• water movement and presence of surface foam (protein buildup).

Recovery.

• immersion (place in anesthetic-free water), injectables (metabolism/excretion of drug, reversals).
• all fish should be placed in fresh water regardless of anesthetic protocol.
• adequately aerated water at all times.
• ventilatory support PRN.
• immersion -> full recovery within 5 minutes -> if more than 10 minutes then excessive anesthetic dose or compromised animal.
• increased RR/RE, return of muscle tone/fin movement, progressively less ataxia until full equilibrium returned.
  
  • excitement phase may occur -> limit risk of trauma.

Resuscitation.

• bradypnea -> pass water over gills, decrease [anesthetic], increase water flow.
• apnea -> remove from anesthesia, pass water over gills.
  
  • buccal flow/HR reflex -> HR increases in response to increase in water flow.
  • consider doxapram? marked stimulant in elasmos (dangerous).
• physiologic collapse/shock -> epi, corticosteroids?

Question 8

Describe analgesia in fish.

What oopioid receptors are present in fish brains?

What opioids have been used?

What local anesthetics have been used?

What NSAIDs have been used?

Answer 8

Analgesia.

• limited information.
• mu and kappa opiate receptors present in fish brains -> antinociceptive effects have been shown.
  
  • morphine, tramadol, buprenorphine, dermorphine -> trout, cod, koi, goldfish, zebrafish, flounder.
  • naloxone negates/decreases effectiveness of morphine/buprenorphine.
• butorphanol -> mu antagonist + kappa agonist.
  
  • koi -> postsurgical analgesia, respiratory depression.
  • catsharks, dogfish -> no effect.
• lidocaine -> SQ, immersion -> analgesic in trout/zebrafish (respectively).
• NSAIDS -> variable effects.

Question 9

Describe the euthanasia methods of fish.

What is the primary method?

Why are secondary methods recommended and what methods are they?

What are some methods that are NOT approved?

Answer 9

Euthanasia.

• immersion alone may be insufficient -> two-step euthanasia is preferred.
• primary method.
  
  • buffered MS-222 at 5-10x anesthetic dose for that sp for at least 15-30 min after reflexes/OR cease.
    
    • recovery is possible in some sp.
    • Doppler for asystole, check gills for pallor.
• secondary methods.
  
  • pithing, decapitation + pithing, macerating, exsanguination, rapid freezing, pentobarbital.
  • pentobarbital -> administer and then leave in anesthetic solution for at least 15-30 min after reflexes/OR cease.
• other options.
  
  • large fish -> sedation then pentobarbital IV.
  • air-breathing sp -> sedation then pentobarbital IV or physical methods.
    
    • never rely on immersion euthanasia alone.
  • small tropical/subtropical sternotropic sp -> immediate immersion in cold water at least 10-20 min after reflexes/OR cease.
  • other methods described by AVMA.
• UNAPPROVED METHODS.
  
  • slow chilling/freezing of unanesthetized animals.
  • death by anoxia and desiccation.
• AVMA Guidelines “euthanasia methods should be tested in one animal or a small group of animals prior to use in a large population for an unfamiliar species.”