Anesthetic MS-222 eliminates nerve and muscle activity in frogs used for physiology teaching laboratories

Abstract

Frogs are routinely used in physiology teaching laboratories to demonstrate important physiological processes. There have been recent directives that promote the use of the anesthetic MS-222 (tricaine methanesulfonate), rather than lowering body temperature with a cold water bath to prepare reptiles and amphibians for physiological experiments or euthanasia. Indeed, the most recent edition of the American Veterinary Medical Association (AVMA) Guidelines for the Euthanasia of Animals proclaims that chilling in water is not an appropriate method and advocates for the usage of MS-222 or other anesthetics. However, prominent researchers have responded to this position by highlighting evidence that cooling ectothermic vertebrates is, in fact, an effective and appropriate method. Furthermore, MS-222 is a known voltage-gated Na+ channel blocker, and this anesthetic’s impact on the physiology of excitable tissues suggests that its use might be incompatible with experiments on nerve and muscle tissues. In the present study, I examined the effects of MS-222 at a concentration of 1.5 g/l on nerve, skeletal muscle, and cardiac muscle physiology of frogs. I found that immersion of frogs in this anesthetic blocked basic nerve and muscle physiology, making the frogs unsuitable for laboratory experiments. Applying MS-222 directly to the sciatic nerve dramatically blocked normal excitation-contraction coupling in skeletal muscle preparations, and direct application to the heart caused the organs to stop contracting. Based on these results, I conclude that MS-222 at the concentration studied may be incompatible with physiological preparations that rely on electrically excitable tissues for their normal function. Physiology educators who must use MS-222 with frogs should empirically determine an appropriate dosage and recovery time before using the anesthetic in the teaching laboratory.

Medium-throughput zebrafish optogenetic platform identifies deficits in subsequent neural activity following brief early exposure to cannabidiol and Δ9-tetrahydrocannabinol

“…We cannot exclude the possibility that due to longer photoconversion times some CaMPARI was converted into the red fluorescence version even when anesthetized. It is well established that when anesthetized with MS-222, neurons are barely active 63 , 64 . Indeed, the behavioural assay shows no locomotion in the presence of MS-222 and we used this assay as a supplement in this study.…”

Reference Intervals in Combined Veterinary Clinical Examinations of Male Black-Spotted Pond Frogs (Pelophylax nigromaculatus)

“…Amphibians and rep- For amphibians, the American Veterinary Medical Association (AVMA) recommends using anesthesia dissolved in water, such as MS-222 (tricaine methanesulfonate), instead of freezing anesthesia if their size is very small (<4 g) or if there is no scientific feasibility [53]. However, we determined that this method was not suitable for our study, as we measured the muscle ratio and mass, and MS-222 has been observed to affect muscle physiology [33]. Additionally, an expert group in the field of amphibians and reptiles has a positive opinion of freezing anesthesia in ectotherm animals [34].…”

“…As chemical anesthesia physiologically affects the muscle tissues of frogs [ 33 ], and supplementing anesthesia with ice-cold water helps to stop ectothermic vertebrates, such as amphibians and reptiles, from experiencing pain [ 34 ], for the immune assay, frogs were anesthetized in ice-cold water and blood was extracted by cardiac venipuncture. The extracted blood was transferred to a serum separator tube (SST) and centrifuged at 3000× g for 10 min.…”

Medium-throughput zebrafish optogenetic platform identifies deficits in subsequent neural activity following brief early exposure to cannabidiol and Δ-9-tetrahydrocannabinol

“…We cannot exclude the possibility that due to longer photoconversion some CaMPARI has converted into the red fluorescence version even when anesthetized. It is well established that when anesthetized with MS-222, neurons are barely active 63,64 . Indeed, our behavioural assay shows no locomotion in the presence of MS-222 and we used this assay as a supplement in this study.…”