The effect of light level, CO2 flow rate, and anesthesia on the stress response of mice during CO2 euthanasia

Abstract

Euthanasia protocols are designed to mitigate the stress experienced by animals, and an environment that induces minimal stress helps achieve that goal. A protocol that is efficient and practical in a typical animal research facility is also important. Light intensity, isoflurane, and CO2 flow rate were studied for their impact on the stress response of mice during CO2 euthanasia. Behavior was observed and scored during euthanasia and serum corticosterone was measured immediately after death. Unsurprisingly, animals euthanized with a high-flow rate of CO2 became unconscious in the least amount of time, while animals euthanized with a low-flow rate required the most time to reach unconsciousness. There was a significant increase in anxious behaviors in animals in the isoflurane group (F1,12 = 6.67, P = 0.024), the high-flow rate CO2 group (F1,12 = 10.24, P = 0.007), and bright chamber group (F1,12 = 7.27, P = 0.019). Serum corticosterone was highest in the isoflurane group (124.72 ± 83.98 ng/ml), however there was no significant difference in corticosterone levels observed for the other study variables of light and flow-rate. A darkened chamber and low CO2 flow rates help to decrease stress experienced during CO2 euthanasia, while the use of isoflurane was observed to increase the stress response during euthanasia.

Gene silencing of indoleamine 2,3‑dioxygenase 1 inhibits lung cancer growth by suppressing T‑cell exhaustion

“…Tumor onset was considered to have occurred when the tumor diameter reached 5 mm. On day 21, all mice were sacrificed with 30%/min carbon dioxide for ~180 sec prior to cervical dislocation (31,32) and were confirmed dead based on cardiac arrest and respiratory arrest 40-60 sec later. Tumors, lymph nodes and spleens were collected, tumors were weighed and tumor volumes were measured.…”

Welfare Impact of Carbon Dioxide Euthanasia on Laboratory Mice and Rats: A Systematic Review

“…Of the 108 full text articles reviewed, 71 articles were excluded as they did not meet eligibility criteria and 37 articles were included. In total, 15 papers were reviewed for mice (16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28)(29)(30), 21 papers for rats (16,22,23,(31)(32)(33)(34)(35)(36)(37)(38)(39)(40)(41)(42)(43)(44)(45)(46)(47)(48), and 5 papers were reviewed for neonatal rodents (21,(49)(50)(51)(52), with three studies reporting results for both adult mice and rats (16,22,23) and one study reporting results for both adult female mice and their pups (21,(49)(50)(51)…”

“…Some studies evaluated different VDRs or FRs of CO 2 , some compared one or more VDR/FR of CO 2 with one or more inert or inhalant anesthetic gas and some studies compared one or more VDR/FR/percentage of CO 2 with other physical or chemical methods of euthanasia, including pentobarbital sodium, potassium chloride, cervical dislocation, and decapitation. One study evaluated the effects of different VDRs of CO 2 with or without premedication with acepromazine or midazolam (30) and one study evaluated the effects of different flow rates of CO 2 when given with differing ambient light intensities (28). Flow or volume displacement rates of carbon dioxide evaluated ranged from 0.2 L/min to 15-100%.…”

“…In studies evaluating physiologic parameters in mice, time to LORR following CO 2 exposure were consistently in three circumstances: when compared to other tested agents, such as isoflurane in oxygen (18,27), for increasing VDRs or flow rates of CO 2 (17,28,30), and when CO 2 was combined with nitrous oxide (N 2 O) (29). Use of a sedative, such as acepromazine or midazolam, prior to administering CO 2 did not shorten time to LORR (30).…”

Euthanasia via CO2 inhalation causes premature cortical granule exocytosis in mouse oocytes and influences in vitro fertilization and embryo development

“…It was suggested that while fast displacement of 100% CO 2 decreases the duration of the stress to mice, slow displacement of CO 2 prolongs the time period of experiencing distress, ataxia, stress on breathing (Creamer-Hente et al, 2018), and decreases the duration of dyspnea (Moody, Chua, & Weary, 2014). To this end, a recent study found that a darkened chamber with slow displacement of CO 2 may alleviate some of the stress during CO 2 euthanasia (Powell, Ethun, & Taylor, 2016).…”