High-resolution microcomputed tomography technology has allowed researchers to use live mice to address questions that previously could be answered only at necropsy. Serial analyses of the same mouse allow tissue changes to be followed over time. The ability to follow a single mouse noninvasively can decrease the total number of mice required for the study. The magnitude of inter-mouse variation for matched mice undergoing microcomputed tomography has not been determined previously. We selected lung and contrast-enhanced stomach as tissues of standard size and anatomical structure that were hypothesized to vary minimally between mice. The analyses of the tissue volumes from matched mice showed considerable variation among mice, among multiple sequential scans of the same mouse, and even among multiple evaluations of the same scan. More variation occurred with repeated scans of the same mouse (intramouse variation) than between mice (intermouse variation). In addition, significant variation and obvious bias was detected between the 2 scan evaluators. These data suggest that to obtain the widest range of possible values, among which the true value would be found, multiple analyses of multiple scans of the same mouse must be performed by multiple scan evaluators.

Euthanasia is one of the most commonly performed procedures in laboratory animal settings. The method of euthanasia may affect experimental results in studies using animals and must be compatible with research objectives including subsequent tissue analyses. Our present study was performed to evaluate the effects of 7 euthanasia methods on sperm motility in mature rats. Rats were euthanized using CO₂, 2 commercially available euthanasia solutions (Beuthanasia-D and Sleepaway), and 4 volatile anesthetics (enflurane, halothane, isoflurane, and sevoflurane). Rats euthanized by rapid decapitation alone served as negative controls, and α-chlorohydrin-treated rats euthanized by rapid decapitation were positive controls for sperm impairment. For 5 of these methods, we also measured time to ataxia, recumbency, respiratory arrest, and no auscultable heartbeat. Immediately after euthanasia of each rat, distal caudal epididymides were removed; 1 was processed for automated sperm motility analysis, and the other was frozen for subsequent concentration analysis. Time to all measured parameters was less for volatile anesthetics than for Beuthanasia-D. Times to last respiration and no heartbeat were less for halothane and isoflurane than for enflurane and sevoflurane. Percentage motile sperm did not differ significantly between methods. Percentage progressively motile sperm did not vary significantly between methods except for Beuthanasia-D, for which it was significantly less than the negative control value. Specific sperm motion parameters for each euthanasia method except CO₂ and Sleepaway varied significantly from the negative control. Our results indicate that the method of euthanasia is an important consideration when rat sperm motility parameters must be evaluated.

We present a list of simple sequence length polymorphisms (SSLPs) that was used in a polymerase chain reaction (PCR)-based marker-assisted selection protocol to facilitate the generation of a C57BL/6J Rj congenic strain from a mixed 129S2/SvPasCrl × C57BL/6J Rj background. We chose informative SSLP markers that would permit evaluation of the PCR products on NuSieve agarose or nondenaturing polyacrylamide gels. This list of oligonucleotide pairs is useful for analyzing and backcrossing knockout or transgenic strains derived from 129S2/SvPasCrl or C57BL/6J Rj backgrounds.

Inbred strains of the laboratory rat (Rattus norvegicus) are commonly used models in biomedical and behavioral research. Genetic contamination caused by breeding errors, incomplete inbreeding with residual allogenicity, mutation, and genetic drift all are known to contribute to substrain divergence. Therefore, colonies of inbred strains from various suppliers likely contain differing amounts of genetic variation. We used microsatellite markers to scan the genomes and compare the genetic composition of 3 Lewis substrains and 2 Wistar-Furth substrains obtained from 3 different suppliers. LEW/SsNHsd rats showed approximately 37% genomic difference as compared with LEW/MolTac rats, and 8% difference as compared with LEW/Crl rats. WF/NHsd rats demonstrated a difference of approximately 8% as compared with WF/CrCrl rats. Investigators should consider the background genetics of the particular strains for their research projects and should use strains from a single source when feasible.

Large animal models are still required for many experimental purposes. The aim of the current study was to define a viable narcotic procedure for experimental cardiovascular interventions and imaging in pigs. A total of 32 domestic pigs were used. Animals received propofol, midazolam, and fentanyl as continuous intravenous infusion anesthesia for complex vascular interventions, angiographic X-ray imaging, and magnetic resonance imaging (MRI). Anesthesia was maintained for 6 to 10 h. The initial hourly doses were 2.29 mg/kg of propofol, 1.14 mg/kg of midazolam, and 0.009 mg/kg of fentanyl, with controlled ventilation. Anesthesia, interventions, imaging, periods of apnea of as long as 2 min, and transportation were well-tolerated. Stress-induced arrhythmias were not noted, and artifact-free imaging was achieved. The combination of propofol, midazolam, and fentanyl is well-suited for experimental angiographic interventional studies, experimental cardiovascular MRI, and MR-guided interventions in pigs.

A variety of rehabilitation methods that increase social interaction and locomotor activity are reported to yield positive benefits in humans and animals with spinal cord injury (SCI). Environmental enrichment often incorporates group housing, increased cage size, and objects to increase social interaction and stimulate locomotor activity of animals. Others have reported that adult rats housed in enriched environments immediately after moderate contusion thoracic SCI show improvements in locomotion, but not in neurotransmission through or anatomy at the SCI site. In the present study, in contrast to previous reports, environmental enrichment did not improve the locomotion of rats with contusion thoracic SCI. Furthermore, as in previous reports, improvements were not observed for either electrophysiologic measures of neurotransmission through (transcranial magnetic motor-evoked potentials) and caudal to (magnetic-evoked interlimb reflex) the injury site or the amount of spared white matter at the epicenter. Determining the effectiveness of environmental enrichment to improve locomotor recovery in the SCI model requires standardization of housing procedures, outcome measures, and analyses.

We evaluated the efficacy of fenbendazole (FBZ) and milbemycin oxime (MO) in the treatment of baboons (Papio cynocephalus anubis) with naturally acquired Trichuris trichiura infection by comparing fecal egg count reduction (FECR) tests. We assigned 7 baboons, each singly housed and confirmed infected with T. trichiura, to treatment groups of FBZ (n = 3) or MO (n = 3), or as a control (n = 1). All (100%) baboons that received FBZ stopped shedding T. trichiura eggs within 6 d of treatment, and fecal egg counts remained negative at 65 d after treatment. Although the number of T. trichiura eggs shed per gram of feces from 2 (67%) baboons decreased significantly after the second treatment with MO, this regimen never totally eliminated eggs of T. trichiura. The results of our study indicate that FBZ was more effective for treating baboons with T. trichiura than was MO.

A widely used in vivo technique in mice and other species is the surgical implantation of transmitters for telemetric monitoring of core body temperature, locomotor activity, and other variables. However, these devices are quite large relative to the size of the mouse abdomen. We report here on the results of several related studies that we conducted to evaluate refinement strategies relevant to implantation of abdominal devices in mice. First, we evaluated survival from surgery as a function of strain and body weight and found that both parameters influence the proportion of mice that survive. Second, we assessed the effect of several interventions on postsurgical recovery of food and water intakes, core temperature, and locomotor activity. Some of the interventions were associated with increased mortality (atipamezole) or were otherwise detrimental (the abdominal lubricant carboxymethylcellulose), whereas others had little or no effect on recovery (thermal support). These findings indicate that interventions presumed to promote recovery from surgery that are based on data from other species may not always have the anticipated positive effect in mice. This study therefore underscores the need to carefully assess the effect of modifications in experimental procedures to avoid causing unexpected complications in mice.

For a molecular identification and typing tool, we developed a specific polymerase chain reaction (PCR) based on the gyrB gene sequence and a subsequent restriction fragment length polymorphism (RFLP) analysis using the products amplified from the specific PCR to facilitate discrimination of biotypes of Pasteurella pneumotropica from laboratory mice. Appropriate PCR products, a 1039-basepair fragment for biotype Jawetz and a 1033-basepair fragment for biotype Heyl, were amplified by use of the primers CZO-1 and NJO-2 from all 105 P. pneumotropica isolates tested and the 2 reference strains but not from other bacterial species tested. MspI digestion of PCR-generated products showed 3 RFLP patterns among the 105 isolates, and these patterns correlated with the biotype of the isolate (RFLP pattern 1, biotype Jawetz; RFLP pattern 2, biotype Heyl; and RFLP pattern 3, biotype Jawetz with β-hemolytic activity). Our procedure identifies and biotypes isolates of P. pneumotropica from laboratory mice, using simple PCR and enzymatic restriction techniques. Therefore, this procedure is useful as a rapid identification and biotyping tool for isolates of P. pneumotropica from laboratory mice.

Clinical pathology is a valuable means for assessing specific organ pathology and a screening tool for general animal health. Routine clinical pathology evaluation in mice usually includes whole blood for a complete blood count (CBC) and a clinical biochemistry analysis. Acquisition and analysis of these samples can be problematic due to the small volumes of blood that can be obtained from a mouse. Typically, a complete blood count requires blood from a tube containing an anticoagulant, whereas a clinical biochemistry profile needs blood from a serum clot tube. Because of the small volume that can be obtained, splitting the blood from a single mouse into 2 different tubes may result in inadequate samples to perform the desired tests or introduce inaccuracies. We explored the feasibility of using a single lithium heparin tube for generation of a CBC, biochemistry profile, and serology profile. We also evaluated the consistency of CBC data, including the quality of a peripheral blood smear taken from a lithium heparin or EDTA tube after various storage times. We found that CBC, biochemistry, and serology profiles could be obtained more readily when blood samples were placed in a single lithium heparin tube than in 2 separate tubes. In addition, the quality of blood smears and CBC results from the lithium heparin tube were comparable (with few exceptions) to those from an EDTA tube after prolonged storage.

We developed a CO₂ euthanasia system that functions as an individually ventilated caging system and that accommodates the simultaneous euthanasia of as many as 70 cages of mice. The automated, logic-controlled system allows euthanasia of mice in their home cage, provides consistent and reproducible delivery of CO₂, permits visualization of animals during euthanasia, and integrates various safety features. Requirements for the safe use of this system are that all cage locations are to be filled and that engineering controls (that is, a thimble connection) be used to minimize CO₂ contamination of the immediate environment. The system was evaluated using mice that were nongravid and greater than 6 d of age. CO₂ measurements were made over time to assess the reproducibility of intracage CO₂ levels and the effect of 3 supply plenum pressures (0.35, 0.25, and 0.15 in. H₂O) on maximal intracage CO₂ concentration, CO₂ fill slope, time until CO₂ detection, and time until maximal CO₂ concentration. Results indicate that both supply plenum pressure and cage position on the rack affect intracage CO₂ concentrations. We also conducted behavioral assessments of mice undergoing euthanasia to evaluate distress during euthanasia at 2 plenum pressures (0.15 and 0.35 in. H₂O). Personnel experienced with laboratory mice did not discern differences in mouse distress associated with either cage location or plenum pressure. This system was safe, effective, and labor-saving for euthanizing large numbers of mice in an aesthetically acceptable and humane manner compatible with recommendations provided in the ACLAM 2005 Report on Euthanasia.

We allocated 35 male Sprague-Dawley rats into 7 groups and anesthetized each by using one of the following regimens: ketamine 50 mg + xylaxine 5 mg; ketamine 75 mg + xylazine 5 mg; pentobarbital 45 mg; and Telazol 30, 40, 50, and 60 mg/kg; supplemental doses were used as required. Respiratory rate, heart rate, mean arterial pressure, cardiac index, and stroke index were measured every 30 min for 4 h. The Telazol groups showed a dose-dependent increase in duration of anesthesia. Duration of anesthesia was significantly shorter for the ketamine and pentobarbital groups than for any of the Telazol doses. Heart rate showed a dose-dependent decrease among the Telazol groups, but overall heart rate in these groups was higher than in the ketamine and pentobarbital groups. Mean arterial pressure in the Telazol 40 and 50 groups was significantly higher than the pentobarbital and higher ketamine groups yet lower than that of the Telazol 60 group. Overall animals anesthetized with Telazol showed the highest cardiac index, ketamine intermediate, and pentobarbital the lowest; cardiac index was higher in the Telazol 50 group than in either the Telazol 30 or pentobarbital groups. The pentobarbital group exhibited the lowest stroke index, whereas ketamine-treated animals had an intermediate stroke index. These differing effects of anesthetics on cardiovascular parameters must be considered when choosing an anesthesia regimen or comparing data from different studies. In our model, the Telazol 40 and 50 groups appeared to exhibit the fewest adverse cardiovascular effects.

We analyzed the cisplatin-induced emetic responses of Suncus murinus by observation of both videorecorded behavior and changes in peak expiratory flow (PEF) as monitored by whole-body plethysmography. Analysis of the PEF data by use of a macro program revealed emesis-related changes in PEF. Cisplatin dose-response curves obtained by both methods showed similar emesis profiles. In addition, our PEF-based analytical method could be used to assess emesis changes induced by various other emetogens (veratrine hydrochloride, nicotine bitartrate, and copper sulfate) and confirmed the efficacy of the antiemetic drugs ondansetron, metoclopramide, and GR205171 in the cisplatin-induced emesis model. These results demonstrate that the use of whole-body plethysmography to follow changes in PEF was an effective semiautomated method of assessing emetic responses in Suncus murinus. This novel semiautomated method may provide an objective, sensitive, and efficient way to study emesis in animal models.

This report describes the discovery and treatment of a multiagent infection in a captive colony of adult, female Xenopus laevis. Animals were determined to be infected with Saprolegnia sp, a relatively common fungal parasite in laboratory-housed frogs, and a less common ectoparasite, Epistylis sp, that had been described only once before in frogs. We discuss the diagnosis, pathology, and treatment of Epistylis and the importance of water-quality monitoring and husbandry in the care of these research animals.

A uterine mass was detected on physical exam in a multiparous African green monkey as an incidental finding, and the well-circumscribed mass was removed via hysterectomy. Histologically, the mass consisted of sheets, nests, and cords of uniform intermediate trophoblastic cells with eosinophilic or clear cytoplasm. These neoplastic cells aggregated around blood vessels, forming islands of viable tumor cells amid extensive areas of coagulative necrosis with calcification in a 'geographic' pattern of necrosis. Immunohistochemistry of the trophoblastic cells revealed strong and diffuse staining for pancytokeratin AE1/3 and p63, with weak and moderate staining for human placental lactogen and placental alkaline phosphatase, respectively. Immunohistochemical staining for smooth muscle actin, epithelial membrane antigen, and human chorionic gonadotropin was negative. Overall, the histologic and immunohistochemical features of this tumor were consistent with those of epithelioid trophoblastic tumor. This rare tumor type has not been reported previously to occur in African green monkeys.

The ACLAM Analgesia Task Force was appointed by ACLAM President Diane Gaertner in 2003. The charge to the Task Force was to develop guidelines that could be used by veterinarians, scientists and IACUCs in helping to provide appropriate assessment and management of pain in rodents, with the understanding that ultimately the clinical veterinarian on site at the institution must make decisions relevant to a specific animal or animals and/or protocol. The guidelines were not to be developed as, and should not be used as, requirements. To complete its charge the Task Force reviewed and cited, in a comprehensive manner, available data-based literature in writing the paper.In the course of completion of the document, ACLAM Board of Directors (BOD) reviewed an early draft and at that time also appointed 3 ACLAM diplomates with particular expertise in assessment and management of pain in rodents to act as reviewers of the draft. The Task Force members responded to the critiques and comments submitted by both the BOD and the 3 reviewers. This revised draft was then placed on the ACLAM website for comments from the entire College. The draft was well received by responding diplomates. Comments from the membership were reviewed and discussed by the Task Force, and most were incorporated into the final draft manuscript. The draft then received final review and editing by the ACLAM Publications Committee Chairman, and was accepted in the format presented here. Despite this extensive vetting process through ACLAM, readers should nonetheless be aware that because this document represents the approved statement of an AALAS affiliate organization, it has not undergone the usual JAALAS peer review process.I would like to acknowledge the efforts of the Task Force and the leadership of ACLAM for supporting this comprehensive and informative synthesis. The document should serve as a resource to the research community for years to come.