Administration of substances to laboratory animals requires careful consideration and planning to optimize delivery of the agent to the animal while minimizing potential adverse experiences from the procedure. For all species, many different routes are available for administration of substances. The research team and IACUC members should be aware of reasons for selecting specific routes and of training and competency necessary for personnel to use these routes effectively. Once a route is selected, issues such as volume of administration, site of delivery, pH of the substance, and other factors must be considered to refine the technique. Inadequate training or inattention to detail during this aspect of a study may result in unintentional adverse effects on experimental animals and confounded results.

Administration of substances to laboratory animals requires careful consideration and planning to optimize delivery of the agent to the animal while minimizing potential adverse experiences from the procedure. The equipment selected to deliver substances to animals depends on the length of the study and the nature of the material being administered. This selection provides a significant opportunity for refining animal treatment. Similarly, when substances are administered as solutions or suspensions, attention should be given to selection of vehicles and methods used for preparing the solutions and suspensions. The research team, veterinarian, technical personnel, and IACUC members should be aware of reasons underlying selection of equipment for substance delivery and should consider carefully how substances will be prepared and stored prior to administration to animals. Failure to consider these factors during experimental planning may result in unintentional adverse effects on experimental animals and confounded results.

Anatomic knowledge of the cardiovascular system, including arterial diameters and arterial segment lengths, is important for appropriate selection of animal size. Here we correlate the diameter and segment lengths of the femoroiliac artery of Yorkshire swine with animal weight. Angiographic images were acquired by using fluoroscopic equipment, and quantitative vascular analysis software was used to measure the internal iliac, external iliac, external femoral, internal femoral, and circumflex femoral arteries. Results indicated that swine weight appeared to have a positive but moderate statistical correlation with arterial diameter, with the exception of the correlation between femoroiliac arterial segment length and swine weight, which was low and sometimes negative. Morphometric data and correlation between animal weight and anatomic dimensional assessments can benefit preclinical researchers and the medical device industry, assisting them to determine their specific animal needs and allowing study design to be performed with a more realistic and educated approach.

The South African clawed frogs Xenopus laevis and X. tropicalis are fully aquatic amphibians and well-established animal models. Because genetically engineered laboratory Xenopus are now being produced, the establishment of normal reference ranges for serum biochemical and hematologic parameters is essential for phenotyping and as a diagnostic aide. We determined normal reference ranges for hematologic values from 3 populations of X. laevis: wild-caught frogs (n = 43) and frogs from 2 commercial sources (A, n = 166; B, n = 109). For serum biochemistry, we determined normal reference ranges for frogs from source A and wild-caught frogs divided by sex and season. Significant differences across populations were found in WBC and RBC counts, hemoglobin concentration, hematocrit, mean corpuscular hemoglobin concentration, and mean corpuscular volume. Among serum biochemical analytes, significant differences were found for albumin:globulin ratio, anion gap, and concentrations of albumin, globulin, total protein, lipase, alanine transaminase, γ-glutamyl transpeptidase; creatine phosphokinase; indirect, direct, and total bilirubin; cholesterol, low-density lipoprotein lipase, carbon dioxide, glucose, lactacte dehydrogenase, calcium, chloride, and sodium. We hypothesize that these differences can be attributed to differences in water quality, habitat, ambient temperature, diet, sex, recent transport or shipment, and genetic background. However, testing that hypothesis is beyond the scope of the current study. In addition, clinical chemistry and hematologic reference range values Xenopus laevis are quite distinct from those for other species and are most consistent with the only values published for another fully aquatic amphibian, the Eastern hellbender (Cryptobranchus alleganiensis).

Periparturient manipulation of mice is a valuable tool for modern research facilities. Although fostering and Caesarian section frequently are used to eradicate pathogens, an often overlooked use is to rescue poorly breeding strains of mice. Here we characterized the weaning success rates after fostering outbred pups of variable ages (younger than 24 h; 5 to 7 d; 10 to 12 d) with full or partial replacement of litters and multiparous dams. There were no significant differences between most groups when analyzed by full or partial replacement or age of donor pups as compared with control groups, in which pups were manipulated but returned to the birth dam or the birth dam was not disturbed. However, significant differences were associated with fostering of 10- to 12-d-old pups in combination with younger pups. Overall, these findings suggest that limiting fostering to pups that are within 48 h of age and age-matching litters when fostering are unnecessary.

The opossum Monodelphis domestica is the most commonly used marsupial in biomedical research. At our institution, these opossums are housed in polycarbonate (35.6 cm × 25.4 cm × 17.8 cm) individually ventilated cages. Previous studies of the cage microenvironment of rodents housed in individually ventilated cages have demonstrated that the cage-change frequency could be extended from 7 to 14 d, without detriment to the animals' wellbeing. We sought to determine whether the cage change frequency for opossums housed in individually ventilated cages could be extended to 14 d. Opossums were placed into 3 experimental groups: singly housed males, singly housed females, and females housed with litters. The 14-d testing period was repeated twice, with temperature, relative humidity, and ammonia levels tested on days 0, 7, and 14. Acceptable ranges for the cage microenvironment were based on standards followed by our institution for housing rodents: temperature between 22 to 26 °C, relative humidity between 30% to 70%, and ammonia less than 25 ppm. Throughout both 14-d testing periods, temperature, relative humidity, and ammonia levels for singly housed male and singly housed female opossums were within acceptable ranges. However, ammonia levels exceeded the recommended 25 ppm on day 7 of both testing periods for female opossums housed with litters. In summary, the cage-change frequency for a singly housed opossum in an individually ventilated cage can be extended to 14 d.

The current study was performed to determine the vibration levels that were generated in cages on a ventilated rack by common construction equipment in frequency ranges likely to be perceived by humans, rats, and mice. Vibration generated by the ventilated rack blower caused small but significant increases in some of the abdominal, thoracic, and head resonance frequency ranges (RFR) and sensitivity frequency ranges (SFR) in which each species is most likely to be affected by and perceive vibration, respectively. Vibration caused by various items of construction equipment at 3 ftfrom the cage were evaluated relative to the RFR and SFR of humans, rats, and mice in 3 anatomic locations. In addition, the vibration levels in the RFR and SFR that resulted from the use of a large jackhammer and were measured at various locations and distances in the facility and evaluated in terms of humans, rats, and mice in 3 anatomic locations. Taken together, the data indicate that a given vibration source generates vibration in frequency ranges that are more likely to affect rats and mice as compared with humans.

The restrictive nature of an Animal Biosafety Level 4 (ABSL4) laboratory complicates even simple clinical evaluation including data capture. Typically, clinical data are recorded on paper during procedures, faxed out of the ABSL4, and subsequently manually entered into a computer. This system has many disadvantages including transcriptional errors. Here, we describe the development of a highly customizable, tablet-PC-based computerized data-capture system, allowing reliable collection of observational and clinical data from experimental animals in a restrictive biocontainment setting. A multidisciplinary team with skills in containment laboratory animal science, database design, and software engineering collaborated on the development of this system. The goals were to design an easy-to-use and flexible user interface on a touch-screen tablet PC with user-supportable processes for recovery, full auditing capabilities, and cost effectiveness. The system simplifies data capture, reduces the necessary time in an ABSL4 environment, offers timely reporting and review of data, facilitates statistical analysis, reduces potential of erroneous data entry, improves quality assurance of animal care, and advances the use and refinement of humane endpoints.

Regular work tasks in the washroom of laboratory animal facilities include cleaning of cages and bottles and handling of chow and bedding. These operations largely are carried out by hand. We quantitatively determined the musculoskeletal load on the trunk and upper limbs of washroom employees in an animal facility with a holding capacity of 35,000 rodent cages by using a computer-assisted, quantitative, recording, and long-term analysis (CUELA) system, which volunteers wore during routine work. Parallel video recording allowed exact assignment of each movement of body and limbs to the data recorded by the sensors. For the most part, trunk movements were unassociated with risk of injury. Evaluation of upper limb movements by CUELA indicated elevated burden on shoulder, elbows, and wrists due to the high repetitiveness and range of movements and postures. However, after additional work factors like low effort and the presence of micropauses were taken into account, workers were not at risk for the development of musculoskeletal disorders of the upper limbs. Handling bottles, chow, and bedding and maneuvering trolleys that entailed greater musculoskeletal loads did not yield evidence of overstraining, because the actions typically were executed alternately and were of short duration during daily shifts. The results represent quantitative information on the musculoskeletal load of regular washroom operations in a laboratory animal facility. These data provide the basis for ergonomic redesign of operations and implementation of automation for highly repetitive movements.

The Zebrafish International Resource Center (ZIRC) supplies wildtype, mutant, and transgenic zebrafish (Danio rerio) to the international research community. In 2005, the ZIRC halted shipment of adult Tübingen (TU) zebrafish, a popular wildtype line, after diagnosis of asymptomatic Mycobacterium chelonae infections in a high proportion of the TU stock. Mycobacterium presents a zoonotic risk to fish handlers. In addition, the presence of underlying chronic disease in a model organism is unacceptable. The TU stock was depopulated and replaced by a new import of TU with the intent of reducing disease prevalence. In the current study, we sampled the new population of TU and fish of the AB, Tupfel long-fin (TL), TAB5 and TAB14 (2 AB × TU hybrid lines), and wildtype-in-Kalkutta (WIK) lines for histologic evaluation and acid-fast staining and compared the prevalence of subclinical mycobacteriosis between these lines. Although prevalence in the new TU stock was lower than that of the original TU stock, asymptomatic infections with Mycobacterium remained high (10%) in the new TU stock held in 20-gal tanks. The prevalence was similar (10%) in the TAB5 line compared with other wildtype lines held in similar conditions. Prevalence of infections in TU can be minimized by husbandry adjustments, including tank size, population density, and cleaning method. Application of these findings has allowed us to decrease mycobacteriosis in TU zebrafish and resume shipment of TU adults to the research community.

We assessed hematologic recovery, body weight, and behavior after serial blood collection in 10- to 14-wk-old C57BL/6 mice. Male and female mice (5 to 11 mice for pilot groups, 23 to 35 mice for full study groups) had either 15%, 20%, or 25% of their estimated total blood volume (TBV) collected once weekly for 6 wk. Except for those of the 25% TBV male pilot group, the weights of all mice recovered or increased from one collection to the next. The behavior of all mice, with the exception of the 25% TBV male pilot group, appeared normal throughout the study. Erythrogram value changes from baseline were analyzed at each weekly blood collection. Recovery was defined as the return of mean hemoglobin values to within 2 SD of mean baseline values. According to this definition, mice in the 15% TBV male group and 15%, 20%, and 25% TBV female groups recovered hematologically. To support the statistical definition of recovery, we compared our data with human anemia categories to assess the clinical relevance of the mouse hemoglobin values. On the basis of these data, we conclude that as much as 25% TBV can be collected once weekly from female mice for 6 wk and as much as 15% TBV can be collected once weekly from male mice for 6 wk without producing weight loss, behavioral changes, or clinically significant anemia.

Contemporary laboratory animal guidance suggests that tail biopsy of laboratory mice can be performed before 21 d of age without anesthesia, whereas older mice must receive anesthesia before biopsy. Our objective was to determine whether administration of isoflurane anesthesia before tail biopsy produced a measurable effect on the behavior of mice (n = 196). We evaluated C57BL/6 and BALB/c mice at 21 to 24 (weaning), 28 to 31 (delayed weaning), and 42 to 45 (adult) d of age. Mice were observed at the time of biopsy and then twice within the first hour after a sham or tail biopsy. Anxiety-like responses were assessed by using an elevated plus-maze. Activity was evaluated remotely for 120 min. Isoflurane did not diminish acute responses to tail biopsy in mice 31 d or younger compared with sham-biopsied animals but had a significant effect in C57BL/6 biopsied adult mice. In addition, mice of all ages and strains that received anesthesia, regardless of biopsy, spent more time in the enclosed maze arms and had decreased activity up to 5 h after isoflurane exposure. Although tail biopsy should be performed in young mice to avoid transection of distal mature vertebrae, our experimental paradigm indicates that isoflurane anesthesia does not appreciably enhance wellbeing over that of mice biopsied without anesthesia at weaning ages. The influence of inhaled isoflurane was demonstrable and indicated that acute and prolonged alterations in anxiety and activity must be considered when interpreting the impact of anesthesia on tail biopsy across various ages and strains of laboratory mice.

Dissection of living brain tissue for in vitro experiments requires the use of a rapid euthanasia method. However, the method must not subject animals to unnecessary pain and must be aesthetically acceptable to experimenters. The purposes of the current study were to assess the aesthetics of 6 euthanasia methods, measure the procedure duration, and evaluate brain for pathology after each procedure. We digitally recorded euthanasia of isoflurane-anesthetized rats by 6 physical methods: anesthetic overdose, cardiac exsanguination, decapitation, closed intrathoracic transection of the great vessels and heart, thoracic percussion, and thoracotomy with rupture of great vessels. Volunteer researchers and animal caretakers watched the video and completed an associated questionnaire. Anesthetic overdose and cardiac exsanguinations were rated most aesthetically pleasing, although these procedures took the longest to complete. In contrast, decapitation and thoracic percussion were the least aesthetically pleasing, but these methods were the quickest. No demographic factor was identified that could predict whether a given euthanasia procedure would be favored for aesthetic reasons, and participants provided a wide variety of rationales for the aesthetic ratings they assigned. Although all of these euthanasia methods meet the criteria of approved methods of euthanasia of anesthetized rats as defined by the AVMA, aesthetic features and the scientific need for rapid euthanasia are both considerations in selecting a method.

To establish a simple procedure for estimating the glomerular filtration rate (GFR) in conscious rabbits, we used the conventional multisample approach to develop a single-blood-sample method. A bolus injection of inulin was administered intravenously at a dose of 40 mg/kg to male New Zealand White rabbits, and blood was collected 30, 60, 90, and 120 min later. Serum inulin, urea nitrogen, and creatinine concentrations were determined. Using this multi-sample method, the reference GFR in clinically healthy rabbits was 4.01 ± 0.17 mL/min/kg (n = 17). In rabbits given an intravenous injection of the antitumor agent cisplatin, GFR fell before serum urea nitrogen and creatinine concentrations increased. Based on cumulative GFR data from healthy and nephropathy rabbits, the GFR obtained from the 3-sample method (30-, 60-, and 90-min samples) was closely correlated (r = 0.99) with that calculated from the estimated distribution volume and serum inulin concentration at 90 min after inulin injection in the single-blood-sample method. These results demonstrate that the single-blood-sample method supports sequential GFR measurements in rabbits and is a versatile procedure not only for research purposes but also in clinical settings.

Fluid therapy is an integral component of standard supportive care in veterinary medicine and is often given subcutaneously for ease of administration. Multiple animals may be treated by using the same bag. These bags often are used repeatedly until they are empty or until a time specified by an IACUC or similar advisory group. This practice introduces the risk of contamination. Here we assessed the sterility of multiple-use fluid bags in the laboratory setting for a maximum of 60 d. Bags were manipulated to mimic infrequent and frequent use. Bacterial cultures of fluid and bag wall and assays for endotoxin and ATP activity were negative at all time points through 30 d. Two fluid samples yielded bacterial growth at 60 d, although all other tests were negative. These results suggest that fluid bags used chronically can be maintained in a sterile condition for a maximum of 30 d.

Automated watering systems provide a reliable source of ad libitum water to animal cages. Our facility uses an automated water delivery system to support approximately 95% of the housed population (approximately 14,000 mouse cages). Drinking valve failure rates from 2002 through 2006 never exceeded the manufacturer standard of 0.1% total failure, based on monthly cage census and the number of floods. In 2007, we noted an increase in both flooding and cases of clinical dehydration in our mouse population. Using manufacturer's specifications for a water flow rate of 25 to 50 mL/min, we initiated a wide-scale screening of all valves used. During a 4-mo period, approximately 17,000 valves were assessed, of which 2200 failed according to scoring criteria (12.9% overall; 7.2% low flow; 1.6% no flow; 4.1% leaky). Factors leading to valve failures included residual metal shavings, silicone flash, introduced debris or bedding, and (most common) distortion of the autoclave-rated internal diaphragm and O-ring. Further evaluation revealed that despite normal autoclave conditions of heat, pressure, and steam, an extreme negative vacuum pull caused the valves' internal silicone components (diaphragm and O-ring) to become distorted and water-permeable. Normal flow rate often returned after a 'drying out' period, but components then reabsorbed water while on the animal rack or during subsequent autoclave cycles to revert to a variable flow condition. On the basis of our findings, we recalibrated autoclaves and initiated a preventative maintenance program to mitigate the risk of future valve failure.