Evaluation of Rodent Diet Stability when Stored in Conditions that Diverge from Guide Parameters.

An essential aspect of animal resource programs is the storage and provision of food for a variety of species. Environmental parameters for feed storage conditions (temperature less than 70 °F; relative humidity less than 50%) are recommended in the Guide for the Care and Use of Laboratory Animals, along with aspects of nutrition such as palatability, vermin-control measures, diet quality, and integrity of feed bags. After receiving a suggestion for improvement for environmental conditions in 2 feed storage locations during an AAALAC accreditation visit, we hypothesized that the packaging of contemporary rodent feed could sustain wider environmental variations in temperature and humidity without adverse impact on integrity and palatability. This study evaluated representative feed storage sites across campus buildings to capture the variation in environmental conditions that are inherent to large and diverse animal care programs. Each test storage location held 2 identical bags of feed (same type, lot, and expiration date) that were stored from June to September of 2021; some aspects of the project were repeated during summer 2022 with a similar rodent feed. Baseline nutrients were analyzed from feed samples collected at time 0 (control) and again after 1 and 3mo of storage. The overall nutritional values measured in feed at the end of the study were not significantly different from control values, regardless of test site and variation in environmental parameters. Retinol (as a measure of Vitamin A) was the only component that decreased significantly; however, final retinol levels were consistently above those necessary for appropriate nutrition for mice. Our animal care program stakeholders were briefed on the outcomes of this study with the intent to verify at future AAALAC site visits that our storage conditions are adequate for maintaining the nutritional quality of packaged rodent feed.

Evaluation of Active Warming and Surgical Draping for Perioperative Thermal Support in Laboratory Mice.

Surgical procedures are commonly performed using mice but can have major effects on their core body temperature, including development of hypothermia. In this study, we evaluated active perioperative warming with and without surgical draping with adherent plastic wrap to refine practices, improve animal welfare, and optimize research experiments. Mice were randomized into treatment groups (n = 6; 8 CD1 mice per group). Treatments included placement within a small-animal forced-air incubator at 38 ° C for 30 min before surgery (Pre), after surgery (Post), or before and after surgery (Both). To explore the effect of surgical draping, one group received incubator warming before and after surgery in addition to surgical draping (Both/ Drape), whereas another group received surgical draping only without incubator warming (Control/Drape). The final group of mice received neither warming nor draping (Control). Subcutaneous temperature transponders were placed in all mice. Approximately 5 d after transponder placement, mice were anesthetized with ketamine-xylazine and underwent laparotomy. Subcutaneous body temperatures were collected perioperatively from transponders, and rectal temperatures were taken every minute during surgery. For recovery from anesthesia, mice were placed either in a standard cage on a warm water blanket set to 38 °C (100.4 °F) or in the incubator. Subcutaneous body temperatures were significantly higher in mice prewarmed for 30 min (Pre, Both, Both/Drape) as compared with mice that were not prewarmed. Anesthetic recovery times were significantly longer for mice placed in the incubator (Pre, Post, Both, Both/Drape) than for those that did not receive incubator warming (Control, Control/Drape). Mean intraoperative rectal temperatures of Both/Drape mice tended to be greater than those of mice in the Both group, suggesting a warming benefit of surgical draping. Using a forced air incubator and adherent plastic draping mitigated body temperature loss in mice during both surgery and postoperative recovery.

Using Waterless Alcohol-based Antiseptic for Skin Preparation and Active Thermal Support in Laboratory Rats.

Rodents are frequently used for models that require surgical procedures. At our institution, laboratory rats are increasingly preferred for investigations of neurologic disorders, cardiovascular interventions, and assessment and treatment of addictive and depressive behaviors. For these types of studies, surgical preparations of the head and neck areas are necessary for catheterization and instrumentation. Based upon our former work in laboratory mice, we sought to improve rat surgery outcomes and confirm the efficacy of a waterless alcohol-based (WAB) antiseptic for skin disinfection prior to incision. In addition, we wanted to investigate whether active warming efforts improved perioperative body temperatures for rats to aid in return to consciousness. Prior to cranial surgical incision and placement in stereotactic equipment, rats were assessed after skin preparation with WAB and after thermal interventions, including prewarming cages for 30 min before anesthesia and delivery of warmed fluid (NaCl) supplementation. Core temperatures were recorded and aerobic culture swabs collected from surgical sites at multiple time points. As previously shown in mice, bacterial counts in rats were effectively diminished by WAB agents. Assessment of intraoperative body temperature trajectories did not identify appreciable differences between control rats and rats that were exposed to prewarming or warmed fluid supplementation or both. However, heavier male rats recovered more rapidly from isoflurane anesthesia than did lighter male and female rats. Although these thermal support measures did not significantly improve anesthetic recovery times in rats, animals warmed for 30 min trended toward a faster return to righting reflex after exposure to isoflurane. These findings confirm that WAB antiseptic is an acceptable option for skin preparation in rats and suggest that continued evaluation of thermal interventions remains of interest for improved outcomes in rat surgery.

Nonexperimental Xenobiotics: Unintended Consequences of Intentionally Administered Substances in Terrestrial Animal Models.

Review of the use of nonexperimental xenobiotics in terrestrial animal models and the potential unintended consequences of these compounds, including drug-related side effects and adverse reactions.

Vaporized Hydrogen Peroxide Decontamination of N95 Respirators in a Dedicated Animal Research Facility for Reuse During a Novel Coronavirus Pandemic.

Introduction: During the COVID-19 pandemic, health care systems and safety providers have faced an unprecedented challenge of limited access to personal protective equipment (PPE) to conduct patient and public care. In federal emergencies, reuse of PPE after disinfection can occur by processes, like vaporized hydrogen peroxide (VHP), recommended by the Centers for Disease and Control and Prevention. We identified a vacant animal holding facility at our institution to repurpose into a regional VHP decontamination center. Methods: The facility is a multiroom, 20 000 ft2 building with control of HVAC to adjust to VHP conditional requirements. H2O2 was delivered to rooms using robotic HaloFoggers, dispersing H2O2 vapor and increasingly concentrated microdroplets as a fog for a timed period based on cubic footage of rooms. Results: Fogging cycles eliminated 6-log Geobacillus stearothermophilus up to 7 days postcycle. Functional efficacy of treated N95s was confirmed by fit tests of institutional personnel. Signage, process flow mapping, and training materials facilitated ease of workflow and adherence to safety expectations within the building. Discussion and Conclusion: Our study determined that a variety of N95 respirator types and sizes were able to be cleared of potential bacterial and viral agents using VHP in a controlled fog/dwell/exhaust cycle. This repurposed animal facility has the capacity to decontaminate up to 6700 respirators daily, which will address the predicted surge of COVID-19 cases in the state, and ultimately allow each respirator to be reused multiple times. There is no other public site in the region with our capacity to offset the continued supply chain issues for PPE needs.

Investigator Engagement: Somewhat Radical Considerations on Practices to Improve Animal Care Program Compliance.

The authors discuss approaches to bolster investigator engagement, inviting investigators to be partners within the Animal Care Program. Regulatory burden in animal research endeavors continues to be reviewed and critiqued; therefore, this article intends to encourage Animal Care Programs to promote transparency and incorporation of unique educational training initiatives to tailor and focus compliance efforts across research programs. Borrowing from concepts of patient engagement, adherence, and enrollment efforts within the medical profession, it is likely that gains in trust, understanding, and communication between stakeholders within animal programs can be achieved without excessive efforts to alter existing approaches. Institutions will continue to be challenged to balance animal welfare expectations with promotion of research missions. This article provides a framework for somewhat radical ideas, including the use of collaborative orientations, assistance with self-evaluations, timely self-reporting, and meaningful and directed trainings, that are all aimed to resonate in contemporary animal care programs and foster investigator engagement in ongoing compliance efforts.

Effects of Rodent Thermoregulation on Animal Models in the Research Environment.

To best promote animal wellbeing and the efficacy of biomedical models, scientific, husbandry, and veterinary professionals must consider the mechanisms, influences, and outcomes of rodent thermoregulation in contemporary research environments. Over the last 2 decades, numerous studies have shown that laboratory mice and rats prefer temperatures that are several degrees warmer than the environments in which they typically are housed within biomedical facilities. Physiologic changes to rodents that are cage-housed under standard temperatures (20 to 26 °C) are attributed to 'cold stress' and include alterations in metabolism, cardiovascular parameters, respiration, and immunologic function. This review article describes common behavioral and physiologic adaptations of laboratory mice and rats to cold stress within modern vivaria, with emphasis on environmental enrichment and effects of anesthesia and procedural support efforts. In addition, potential interventions and outcomes for rodents are presented, relative to the importance of repeating and reproducing experiments involving laboratory rodent research models of human disease.

Comparison of Aqueous and Alcohol-based Agents for Presurgical Skin Preparation Methods in Mice.

Preparing the skin of rodents for surgery often involves multiple applications of antiseptic agents. However, fewer applications may achieve the same antiseptic outcome. We evaluated the antimicrobial efficacy and effects on intraoperative body temperature of various surgical scrub agents, including novel waterless alcohol-based (WAB) options. Prior to ventral laparotomy, female C57BL/6 mice were treated with 0.9% saline (control); 70% ethanol; 10% povidone-iodine alternated with saline or 70% ethanol; 2% chlorhexidine digluconate alternated with saline or 70% ethanol; or 1 of 3 WAB products-commercial surgical scrub A, commercial surgical scrub B, or a common commercial hand sanitizer. Core temperatures were recorded, and aerobic culture swabs were collected from the surgical site at multiple time points. Intraoperative temperature trajectories for animals treated with scrub B, 10% povidone-iodine with saline, or hand sanitizer did not differ from saline (control). Temperature trajectories of mice treated with other scrub agents did differ significantly from saline. Bacteria were not detected at the operative site after 3 scrubs of 70% ethanol or 10% povidone-iodine alternated with ethanol, 2 scrubs of scrub A or B, 1 scrub of hand sanitizer, and both 1 and 3 scrubs of 2% chlorhexidine alternated with ethanol. Scrub B and 2% chlorhexidine-ethanol demonstrated prolonged antibacterial efficacy. Histology of corresponding haired skin sections revealed no differences in postoperative healing between groups, and no postoperative infections occurred. These results indicate that various novel WAB disinfectants, particularly scrub B (61% ethanol and 1% chlorhexidine gluconate), mitigate intraoperative temperature effects associated with several traditional agents and combinations. Furthermore, reduction of skin bacterial load without adverse effects on healing was seen with fewer than triplicate applications of most tested agents. Ultimately effective skin preparation can be achieved by using only 1 or 2 applications of scrub, thus rendering the triplicate skin-prep method unnecessary in laboratory mice.

Quantification of Induced Hypothermia from Aseptic Scrub Applications during Rodent Surgery Preparation.

Laboratory mice (Mus musculus) are prone to develop hypothermia during anesthesia for surgery, thus potentially impeding anesthetic recovery, wound healing, and future health. The core body temperatures of isoflurane-anesthetized mice are influenced by the choice of supplemental heat sources; however, the contribution of various surgical scrubs on the body temperatures of mice under gas anesthesia has not been assessed. We sought to quantify the effect of using alcohol (70% isopropyl alcohol [IPA]) compared with saline to rinse away surgical scrub on the progression of hypothermia in anesthetized mice (n = 47). IPA, room-temperature saline, or warmed saline (37 °C) was combined with povidone-iodine and then assessed for effects on core (rectal) and surface (infrared) temperatures. Agents were applied to a 2×2-cm shaved abdominal area of mice maintained on a water-recirculating blanket (at 38 °C) under isoflurane anesthesia (1.5% to 2.0% at 0.6 L/min) for 30 min. Although all scrub regimens significantly decreased body temperature at the time of application, treatments that included povidone-iodine led to the coldest core temperatures, which persisted while mice were anesthetized. Compared with room-temperature saline and when combined with povidone-iodine, warming of saline did not ameliorate heat loss. IPA alone demonstrated the most dramatic cooling of both surface and core readings at application but generated an unanticipated warming (rebound) phase during which body temperatures equilibrated with those of controls within minutes of application. Although alcohol is inappropriate as a stand-alone agent for surgical skin preparation, IPA is a viable alternative to saline-based rinses in this context, and its use should be encouraged within institutional guidance for rodent surgical procedures without concern for prolonged hypothermia in mice.

Quantification of Induced Hypothermia from Aseptic Scrub Applications during Rodent Surgery Preparation.

Laboratory mice (Mus musculus) are prone to develop hypothermia during anesthesia for surgery, thus potentially impedinganesthetic recovery, wound healing, and future health. The core body temperatures of isoflurane-anesthetized mice areinfluenced by the choice of supplemental heat sources; however, the contribution of various surgical scrubs on the bodytemperatures of mice under gas anesthesia has not been assessed. We sought to quantify the effect of using alcohol (70%isopropyl alcohol [IPA]) compared with saline to rinse away surgical scrub on the progression of hypothermia in anesthetizedmice (n = 47). IPA, room-temperature saline, or warmed saline (37 °C) was combined with povidone-iodine and thenassessed for effects on core (rectal) and surface (infrared) temperatures. Agents were applied to a 2×2-cm shaved abdominalarea of mice maintained on a water-recirculating blanket (at 38 °C) under isoflurane anesthesia (1.5% to 2.0% at 0.6 L/min)for 30 min. Although all scrub regimens significantly decreased body temperature at the time of application, treatments thatincluded povidone-iodine led to the coldest core temperatures, which persisted while mice were anesthetized. Comparedwith room-temperature saline and when combined with povidone-iodine, warming of saline did not ameliorate heat loss.IPA alone demonstrated the most dramatic cooling of both surface and core readings at application but generated an unanticipatedwarming (rebound) phase during which body temperatures equilibrated with those of controls within minutes ofapplication. Although alcohol is inappropriate as a stand-alone agent for surgical skin preparation, IPA is a viable alternativeto saline-based rinses in this context, and its use should be encouraged within institutional guidance for rodent surgicalprocedures without concern for prolonged hypothermia in mice.

Intraperitoneal Continuous-Rate Infusion for the Maintenance of Anesthesia in Laboratory Mice (Mus musculus).

Intraperitoneal injectable anesthetics are often used to achieve surgical anesthesia in laboratory mice. Because bolus redosing of injectable anesthetics can cause unacceptably high mortality, we evaluated intraperitoneal continuous-rate infusion (CRI) of ketamine with or without xylazine for maintaining surgical anesthesia for an extended period of time. Anesthesia was induced in male C57BL/6J mice by using ketamine (80 mg/kg) and xylazine (8 mg/kg) without or with acepromazine at 0.1 mg/kg or 0.5 mg/kg. At 10 min after induction, CRI for 90 min was initiated and comprised 25%, 50%, or 100% of the initial ketamine dose per hour or 50% of the initial doses of both ketamine and xylazine. Anesthetic regimens were compared on the basis of animal immobility, continuous surgical depth of anesthesia as determined by the absence of a pedal withdrawal reflex, and mortality. Consistent with previous studies, the response to anesthetics was highly variable. Regimens that provided the longest continuous surgical plane of anesthesia with minimal mortality were ketamine-xylazine-acepromazine (0.1 mg/kg) with CRI of 100% of the initial ketamine dose and ketamine-xylazine-acepromazine (0.5 mg/kg) with CRI of 50% of the initial ketamine and xylazine doses. In addition, heart rate and respiratory rate did not increase consistently in response to a noxious stimulus during CRI anesthesia, even when mice exhibited a positive pedal withdrawal reflex, suggesting that these parameters are unreliable indicators of anesthetic depth during ketamine-xylazine anesthesia in mice. We conclude that intraperitoneal CRI anesthesia in mice prolongs injectable anesthesia more consistently and with lower mortality than does bolus redosing.

Intraperitoneal Injection of Ethanol for the Euthanasia of Laboratory Mice (Mus musculus) and Rats (Rattus norvegicus).

Compassion, professional ethics, and public sensitivity require that animals are euthanized humanely and appropriately under both planned and emergent situations. According to the 2013 AVMA Guidelines for the Euthanasia of Animals, intraperitoneal injection of ethanol is "acceptable with conditions" for use in mice. Because only limited information regarding this technique is available, we sought to evaluate ethanol by using ECG and high-definition video recording. Mice (n = 85) and rats (n = 16) were treated with intraperitoneal ethanol (70% or 100%), a positive-control agent (pentobarbital-phenytoin combination [Pe/Ph]), or a negative-control agent (saline solution). After injection, animals were assessed for behavioral and physiologic responses. Pain-assessment techniques in mice demonstrated that intraperitoneal injection of ethanol was not more painful than was intraperitoneal Pe/Ph. Median time to loss of consciousness for all mice that received ethanol or Pe/Ph was 45 s. Median time to respiratory arrest was 2.75, 2.25, and 2.63 min, and time (mean ± SE) to cardiac arrest was 6.04 ± 1.3, 2.96 ± 0.6, and 4.03 ± 0.5 min for 70% ethanol, 100% ethanol, and Pe/Ph, respectively. No mouse that received ethanol or Pe/Ph regained consciousness. Although successful in mice, intraperitoneal ethanol at the doses tested (9.2 to 20.1 g/kg) was unsuitable for euthanasia of rats (age, 7 to 8 wk) because of the volume needed and prolonged time to respiratory effects. For mice, intraperitoneal injection of 70% or 100% ethanol induced rapid and irreversible loss of consciousness, followed by death, and should be considered as "acceptable with conditions."

Evaluation of Presurgical Skin Preparation Agents in African Clawed Frogs (Xenopus laevis).

Despite the routine collection of oocytes from African clawed frogs (Xenopus laevis) for use in research, few studies have evaluated methods for preparing their skin for surgery. We evaluated 3 skin preparatory agents by examining their antibacterial efficacy and the gross and microscopic appearance of Xenopus skin after exposure. Frogs (n = 14) were sedated and treated (contact time, 10 min) with 0.9% sterile NaCl on one-half of the ventrum and with 0.5% povidone-iodine or 0.75% chlorhexidine on the other half. Bacterial cultures were obtained before and after skin treatment; bacteria were identified by mass spectrometry. To assess inflammation and degenerative changes, the incision sites were photographed and biopsied at 0, 1, and 7 d after surgery. We isolated at least 22 genera of bacteria from the skin of our frog population (mean ± SE, 5.21 ± 0.82 genera per frog). Iodine (2.00 ± 0.44 genera) and chlorhexidine (0.29 ± 0.76 genera) both had greater antimicrobial activity than did saline. Skin erythema did not correlate with treatment group. Histologic evidence of epidermal degeneration and necrosis was greater on days 1 and 7 after chlorhexidine treatment than after iodine or saline. In addition, frogs treated with chlorhexidine had a higher incidence of clinical illness associated with the exposure site. In summary, although chlorhexidine has adequate antimicrobial activity against organisms on X. laevis skin, it leads to skin damage and subsequent clinical complications. We therefore do not recommend chlorhexidine as a preoperative preparation agent in Xenopus.

Comparison of Heart Rate and Blood Pressure with Toe Pinch and Bispectral Index for Monitoring the Depth of Anesthesia in Piglets.

Determining depth of anesthesia (DOA) is a clinical challenge in veterinary medicine, yet it is critical for the appropriate oversight of animals involved in potentially painful experimental procedures. Here, we investigated various parameters used to monitor conscious awareness during surgical procedures and refined the application of noxious stimuli to anesthetized animals. Specifically we used a common stimulus, a compressive toe pinch (TP), to determine physiologic changes that accompanied a positive or negative motion response in isoflurane-anesthetized piglets. A positive response was defined as any reflexive withdrawal, whereas a negative response was defined as the absence of motion after stimulation. We also assessed the utility of the bispectral index (BIS) for its ability to predict a motion response to TP. The average of BIS values over 1 min (BISmean) was recorded before and after TP. In piglets with a positive response to TP, heart rate (HR), but not blood pressure (BP), increased significantly, but receiver operating characteristic (ROC) analysis revealed that HR was not a sensitive, specific predictor of TP motion response. Both before and after TP, BISmean was a strong predictor of a positive motion response. We conclude that HR and noninvasive BP changes are not clinically reliable indicators of anesthetic depth when assessed immediately after a peripherally applied compressive force as an indicator stimulus; however, BISmean and response TP are acceptable for assessing DOA in piglets maintained under isoflurane anesthesia.

The Effects of Acute Blood Loss for Diagnostic Bloodwork and Fluid Replacement in Clinically Ill Mice.

Despite the great value of diagnostic bloodwork for identifying disease in animals, the volume of blood required for these analyses limits its use in laboratory mice, particularly when they are clinically ill. We sought to determine the effects of acute blood loss (ABL) following blood collection for diagnostic bloodwork in healthy mice compared with streptozotocin-induced diabetic and dextran sulfate sodium (DSS)-treated dehydrated mice. ABL caused several mild changes in the control mice, with significant decreases in body weight, temperature, and activity in both experimental groups; increased dehydration and azotemia in the DSS-treated mice; and a significant drop in the blood pressure of the diabetic mice. To determine whether these negative outcomes could be ameliorated, we treated mice with intraperitoneal lactated Ringers solution either immediately after or 30 min before ABL. Notably, preABL administration of fluids helped prevent the worsening of the dehydration and azotemia in the DSS-treated mice and the changes in blood pressure in the diabetic mice. However, fluid administration provided no benefit in control of blood pressure when administered after ABL in the diabetic mice. Furthermore, fluid therapy did not prevent ABL-induced drops in body weight and activity. Although one mouse not receiving fluid therapy became moribund at the 24-h time point, no animals died during the 24-h study. This investigation demonstrates that blood for diagnostic bloodwork can be collected safely from clinically ill mice and that preemptive fluid therapy mitigates some of the negative changes associated with this blood loss.

Adverse effects of vapocoolant and topical anesthesia for tail biopsy of preweanling mice.

Tail biopsy of laboratory mice for genotyping purposes has been studied extensively to develop refinements for this common procedure. Our prior work assessed tail vertebral development in different mouse strains (age, 3 to 42 d) and analyzed behavior and activity in mice (age, 21 to 45 d) biopsied under isoflurane anesthesia. To assess the effects of biopsy on preweanling mice, we here evaluated BALB/cAnNCrl mice (n = 80; age, 18 to 21 d) that received topical vapocoolant (ethyl chloride), topical anesthetic (Cetacaine), or isoflurane anesthesia before undergoing a 5-mm or sham biopsy. Control mice did not receive any anesthetic intervention. Regardless of the anesthetic used, acute observation scores indicative of distress were increased at 10 min after biopsy, and locomotor activity was decreased, in biopsied compared with control mice. Acute observation scores at 10 min after biopsy were higher in mice that received ethyl chloride compared with isoflurane or no anesthesia. Microscopic analysis revealed that inflammatory changes in the distal tail remained elevated until 7 d after biopsy and were higher in tails exposed to ethyl chloride. Our findings indicate that vapocoolant, topical anesthesia, and inhaled isoflurane do not enhance the wellbeing of preweanling mice undergoing tail biopsy. Due to the lack of appreciable benefits and the presence of notable adverse effects, using vapocoolants or Cetacaine for this tail biopsy procedure in laboratory mice is unadvisable and we encourage the removal of these agents from institutional tail biopsy guidelines.

Type, duration, and incidence of pathologic findings after retroorbital bleeding of mice by experienced and novice personnel.

Retroorbital blood collection is a common technique in laboratory rodents due to the ease with which it can be performed and the sample volumes obtained for subsequent blood analyses. However, its use has been discouraged recently due to aesthetic discomfort and anecdotal reports of potential for ocular injury during blood collection. We hypothesized that a single standardized session of in-person training would be sufficient to learn the appropriate technique and minimize the likelihood for adverse outcomes. Experienced instructors (n = 2) conducted hands-on training classes to teach novice personnel (n = 40) to perform this procedure. Blood was collected from anesthetized mice (n = 40) via a capillary tube first placed at the medial canthus of the right eye and then advanced into the retroorbital space; the left retroorbital spaces served as unmanipulated controls. For comparison, the experienced instructors similarly collected blood from 40 additional mice. The tube could be inserted only once in each mouse, with the goal of obtaining 50 to 100 µL blood. Overall, 79 of 80 mice (98.8%) showed normal body condition, posture, and behavior throughout the 14-d study. Thus, any clinical observation scores pertained specifically to ocular lesions, which occurred at least once after sampling in 43 (53.8%) of the mice. Clinical and histopathologic scores of mice after bleeding did not differ between experienced and novice personnel. We conclude that a coordinated hands-on training program can provide consistent and sufficient instruction for research personnel to conduct retroorbital blood collection with competence in anesthetized laboratory mice.

Antibiotic administration in the drinking water of mice.

Although antibiotics frequently are added to the drinking water of mice, this practice has not been tested to confirm that antibiotics reach therapeutic concentrations in the plasma of treated mice. In the current investigation, we 1) tested the stability of enrofloxacin and doxycycline in the drinking water of adult, female C57BL/6 mice; 2) measured the mice's consumption of water treated with enrofloxacin, doxycycline, amoxicillin, or trimethoprim-sulfamethoxazole; and 3) used HPLC to measure plasma antibiotic concentrations in mice that had ingested treated water for 1 wk. Plasma concentrations of antibiotic were measured 1 h after the start of both the light and dark cycle. The main findings of the study were that both enrofloxacin and nonpharmaceutical, chemical-grade doxycycline remained relatively stable in water for 1 wk. In addition, mice consumed similar volumes of antibiotic-treated and untreated water. The highest plasma antibiotic concentrations measured were: enrofloxacin, 140.1 ± 10.4 ng/mL; doxycycline, 56.6 ± 12.5 ng/mL; amoxicillin, 299.2 ± 64.1 ng/mL; and trimethoprim-sulfamethoxazole, 5.9 ± 1.2 ng/mL. Despite the stability of the antibiotics in the water and predictable water consumption by mice, the plasma antibiotic concentrations were well below the concentrations required for efficacy against bacterial pathogens, except for those pathogens that are exquisitely sensitive to the antibiotic. The findings of this investigation prompt questions regarding the rationale of the contemporary practice of adding antibiotics to the drinking water of mice for systemic antibacterial treatments.

Dose regimens, variability, and complications associated with using repeat-bolus dosing to extend a surgical plane of anesthesia in laboratory mice.

Extending a surgical plane of anesthesia in mice by using injectable anesthetics typically is accomplished by repeat-bolus dosing. We compared the safety and efficacy of redosing protocols administered either during an anesthetic surgical plane (maintaining a continuous surgical plane, CSP), or immediately after leaving this plane (interrupted surgical plane, ISP) in C57BL/6J mice. Anesthesia was induced with ketamine, xylazine, and acepromazine (80, 8, and 1 mg/kg IP, respectively), and redosing protocols included 25% (0.25K), 50% (0.5K), or 100% (1.0K) of the initial ketamine dose or 25% (0.25KX) or 50% (0.5KX) of the initial ketamine-xylazine dose. In the ISP group, the surgical plane was extended by 13.8 ± 2.1 min (mean ± SEM) after redosing for the 0.25K redose with 50% returning to a surgical plane, 42.7 ± 4.5 min for the 0.5K redose with 88% returning to a surgical plane, and 44.3 ± 15.4 min for the 1.0K redose, 52.8 ± 7.2 min for the 0.25KX redose, and 45.9 ± 2.9 min for the 0.5KX redose, with 100% of mice returning to a surgical plane of anesthesia in these 3 groups. Mortality rates for ISP groups were 0%, 12%, 33%, 12%, and 18%, respectively. Mice in CSP groups had 50% mortality, independent of the repeat-dosing protocol. We recommend redosing mice with either 50% of the initial ketamine dose or 25% of the initial ketamine-xylazine dose immediately upon return of the pedal withdrawal reflex to extend the surgical plane of anesthesia in mice, optimize the extension of the surgical plane, and minimize mortality.