Imaging modalities such as micro-computed tomography (micro-CT), micro-positron emission tomography (micro-PET), high-resolution magnetic resonance imaging (MRI), optical imaging, and high-resolution ultrasound are rapidly becoming invaluable research tools. These advanced imaging technologies are now commonly used to investigate rodent biology, metabolism, pharmacokinetics, and disease in vivo. Choosing an appropriate anesthetic regimen as well as monitoring and supporting the animal's physiologic balance is key to obtaining images that truly represent the biologic process or disease state of interest. However, there are many challenges in rodent bioimaging such as limited animal access, small sample volumes, anesthetic complications, strain and gender variability, and the introduction of image artifacts. Because each imaging study presents unique challenges, a thorough understanding of the imaging modality used, the animal's health status, and the research data desired is required. This article addresses these issues along with other common laboratory animal clinical considerations such as biosecurity and radiation safety in in vivo rodent bioimaging.

In vivo bioluminescent imaging (BLI) is a versatile and sensitive tool that is based on detection of light emission from cells or tissues. Bioluminescence, the biochemical generation of light by a living organism, is a naturally occurring phenomenon. Luciferase enzymes, such as that from the North American firefly (Photinus pyralis), catalyze the oxidation of a substrate (luciferin), and photons of light are a product of the reaction. Optical imaging by bioluminescence allows a low-cost, noninvasive, and real-time analysis of disease processes at the molecular level in living organisms. Bioluminescence has been used to track tumor cells, bacterial and viral infections, gene expression, and treatment response. Bioluminescence in vivo imaging allows longitudinal monitoring of a disease course in the same animal, a desirable alternative to analyzing a number of animals at many time points during the course of the disease. We provide a brief introduction to BLI technology, specific examples of in vivo BLI studies investigating bacterial/viral pathogenesis and tumor growth in animal models, and highlight some future perspectives of BLI as a molecular imaging tool.

Optical imaging can advance knowledge of cellular biology and disease at the molecular level in vitro and, more recently, in vivo. In vivo optical imaging has enabled real-time study to track cell movement, cell growth, and even some cell functions. Thus, it can be used in intact animals for disease detection, screening, diagnosis, drug development, and treatment evaluation. This review includes a brief introduction to fluorescence imaging, fluorescent probes, imaging devices, and in vivo applications in animal models. It also describes a quantitative fluorescence detection method with a reconstruction algorithm for determining the location of fluorophores in tissue and addresses future applications of in vivo fluorescence imaging.

Ovarian epithelial carcinoma is the most common form of ovarian cancer, causing more deaths than any other cancer in women with gynecologic malignancies in the United States. The poor outcome for women with late-stage ovarian cancer underscores the need for early detection strategies and for new treatment options to be developed. High-intensity focused ultrasound (HIFU) is a therapeutic modality that is safe and effective against various types of solid tumors, and has the potential to serve as a treatment for ovarian cancer. Use of an appropriate animal model is important in obtaining relevant data for translational research. The purpose of the study reported here was to modify and create an immunocompetent orthotopic tumor isograft model for evaluation of intra-operative HIFU application. This model would resemble the clinical presentation of human patients with late-stage ovarian cancer. We were able to consistently produce a surgical model that presented with a single large, intra-abdominal tumor nodule within the left ovarian bursa, as well as multiple small nodules on the surface of other organs and tissues. This technique may also be used to refine other tumor models, using the ovarian bursa as an implant site for heterotopic tumor isografts.

In vivo imaging of rats represents an important tool for outcome evaluation in research on stroke, brain trauma, and other neurologic diseases. Since sedation of animals is necessary to avoid artifacts, a mixture of ketamine and xylazine is frequently used for anesthesia. We assessed the suitable dosage of narcotics and its correlation to severe respiratory adverse events in 269 cases of ketamine/xylazine anesthesia in male Wistar rats for performance of magnetic resonance imaging after middle cerebral artery occlusion (MCAO) or sham surgery. Anesthesia depth was not measured. Anesthesia was efficacious in avoiding movement artifacts during imaging. Necessary dosage was lower if rodents were subjected to MCAO instead of sham surgery, if body weight was below baseline, and if time since surgery was short. If anesthesia was induced during the first 2 days after surgery in animals with body weight loss, necessary dose rates were 27% below doses required for rats more than 10 days post-surgery with body weight above baseline (91.4/8.3 versus 125.1/11.3 mg of ketamine/xylazine/kg). A dose adaptation scale for the prediction of necessary dose rates was developed. Apnea developed in 3.3% of all animals.Use of ketamine/xylazine anesthesia for imaging procedures is feasible and safe, though it is associated with a small risk of respiratory arrest. In case of apnea, inspiration can be provoked by a puff of air into the rat's pelt. If unsuccessful, endotracheal intubation and mechanical ventilation are needed until spontaneous breathing is restored or xylazine effects are antagonized.

The Guide for the Care and Use of Laboratory Animals (the Guide) is widely accepted as the housing standard by most Institutional Animal Care and Use Committees. The recommendations are based on best professional judgment rather than experimental data. Current efforts are directed toward replacing these guidelines with data-driven, species-appropriate standards. Our studies were undertaken to determine the optimum housing density for C57BL/6J mice, the most commonly used inbred mouse strain. Four-week-old mice were housed for 8 weeks at four densities (the recommended ∼12 in² [ca. 77.4 cm²]/mouse down to 5.6 in²[ca. 36.1 cm²]/mouse) in three cage types with various amounts of floor space. Housing density did not affect a variety of physiologic parameters but did affect certain micro-environmental parameters, although these remained within accepted ranges. A second study was undertaken housing C57BL/6J mice with as little as 3.2 in²/mouse (ca. 20.6 cm²). The major effect was elevated ammonia concentrations that exceeded limits acceptable in the workplace at increased housing densities; however, the nasal passages and eyeballs of the mice remained microscopically normal. On the basis of these results, we conclude that C57BL/6J mice as large as 29 g may be housed with 5.6 in² of floor space per mouse. This area is approximately half the floor space recommended in the Guide. The role of the Guide is to ensure that laboratory animals are well treated and housed in a species-appropriate manner. Our data suggest that current policies could be altered in order to provide the optimal habitation conditions matched to this species' social needs.

Rodents and dogs are frequently used for preclinical toxicologic assessment of candidate iron chelators. Although the iron-clearing profile of a ligand often is known in rodents, and sometimes in primates, such information in dogs is rarely, if ever, available. Because of this, toxicity studies in dogs could be misleading; chelators that may otherwise be suitable for human clinical studies may be abandoned as being unacceptably toxic, simply because, unknown to the investigator, these drugs remove more iron in this species than would have been expected on the basis of iron clearance results in other species. This is a scenario that we encountered during toxicity trials of (S)-β,β-dimethyl-4′-hydroxydesazadesmethyldesferrithiocin in dogs. Thus, we developed an iron-overloaded dog model in which it is possible to evaluate iron-clearing efficiencies of potential therapeutic ligands. Seven deferration agents have been screened in this model, and the results were compared with the iron-clearing efficiency of the same ligands in an iron-loaded Cebus apella monkey model. The data suggest that while the iron-clearing efficiencies of most of the drugs were similar between the two species, there can be profound differences. This is consistent with the idea that caution needs to be exercised when carrying out preclinical toxicity evaluations of a chelator in dogs without first measuring the drug's iron-clearing efficiency in this species.

Endocrine-disrupting chemicals (EDCs) are giving rise to serious concerns for humans and wildlife. Phytoestrogens, such as daidzein and genistein in plants, and organochlorine pesticides are suspected EDCs, because their chemical structure is similar to that of natural or synthetic estrogens and they have estrogenic activity in vitro and in vivo. We assessed estrogenic activity and dietary phytoestrogen and organochlorine pesticide contents of various fish diets made in the United Kingdom, and compared them with those features of diets made in Japan that were tested in a previous study. Genistein and daidzein were detected in all of the diets. Using an in vitro bioassay, many of these diets had higher activation of estrogen β-receptors than estrogen α-receptors. Organochlorine pesticides such as hexachlorobenzene, β-benzene hexachloride (BHC), and γ-BHC were detected in all fish diets. On the basis of these data, we investigated the effect of differing dietary phytoestrogen content in Japanese fish diets on hepatic vitellogenin production and reproduction (fecundity and fertility) in medaka (Oryzias latipes). Assessment of the effects of a 28day feeding period on reproduction of paired medaka did not indicate significant differences in the number of eggs produced and fertility among all feeding groups. However, hepatic vitellogenin values were significantly higher for male medaka fed diet C (genistein, 58.5 ± 0.6 μg/g; daidzein, 37.3 ± 0.2 μg/g) for 28 days compared with those fed diet A (genistein, < 0.8 μg/g; daidzein, < 0.8 μg/g) or diet B (genistein, 1.4 ± 0.1 μg/g; daidzein, 2.0 ± 0.1 μg/g). Our findings indicate that fish diets containing high amounts of phytoestrogens, such as diet C, have the potential to induce hepatic vitellogenin production in male medaka, even if reproductive parameters are unaffected. Therefore, some diets, by affecting vitellogenin production in males, may alter estrogenic activity of in vivo tests designed to determine activity of test compounds added to the diet.

Mouse hepatitis virus (MHV) is one of the most prevalent viruses infecting laboratory mice. Most natural infections are caused by enterotropic strains. Experiments were done to compare the pathogenesis of enterotropic strain MHV-Y in immunocompetent BALB/c and C57BL/6 mice with that in B and T cell-deficient mice. In situ hybridization was used to identify sites of virus replication, and reverse transcriptase-polymerase chain reaction analysis was used to detect viral RNA in feces and blood. MHV-Y caused acute subclinical infections restricted to the gastrointestinal tract in BALB/c and C57BL/6 mice. Viral RNA was detected in small intestine and associated lymphoid tissues of immunocompetent mice for 1 week and in cecum and colon for 2 weeks. Infected B cell-deficient mice developed chronic subclinical infection also restricted to the gastrointestinal tract. Viral RNA was detected in the small intestine, cecum, colon, and feces for 7 to 8 weeks. In contrast, infected T cell-deficient mice developed multisystemic lethal infection. During the first week, viral RNA was restricted to the gastrointestinal tract. However, by 2 weeks, mice developed peritonitis, and viral RNA was detected in mesentery and visceral peritoneum. Three to four weeks after virus inoculation, T cell-deficient mice became moribund and viral RNA was detected in multiple organ systems. These results suggest that B cells promote clearance of MHV-Y from intestinal mucosa and that T cells are required to prevent dissemination of MHV-Y from the gastrointestinal tract and associated lymphoid tissues.

Although the N-type Ca²⁺ channel plays a role in a variety of neuronal functions, N-type Ca²⁺ channel α_1B-deficient mice exhibit normal life span without apparent behavioral or histologic abnormalities. To examine whether the reason for their normal behavior is compensation by other Ca_v2 channel α₁ or β subunit genes and to analyze whether genetic background influences the subunit expression pattern, we studied the α_1A, α_1E, β_1b, β₂, β₃ and β₄ subunit mRNA levels in cerebellum of α_1B-deficient mice with CBA × C57BL/6 or CBA/JN background. In cerebellum of the mice with a CBA × C57BL/6 background, α_1A mRNA was expressed at a higher level than that in wild-type or heterozygous mice, but difference in the expression levels of α_1E, β_1b, β₂, β₃, and β₄ subunits was not found among wild-type, heterozygous, and homozygous mice. In cerebellum of α_1B-deficient mice with CBA/JN background, β₄ mRNA was expressed at a higher level than that in wild-type or heterozygous mice, but α_1A, α_1E, β_1b, α₂, and β₃ transcripts were expressed at similar levels in all genotypes. Therefore, a possible explanation of the normal behavior of α_1B-deficient mice is that Ca_v2 channel family members compensate for the deficiency, and that the change of functional subunit expression pattern for compensation differs in animals with different genetic backgrounds.

Little is known about the natural history of herpesviruses indigenous in baboons. Here, we describe the development of ELISAs for five herpesviruses. These assays were used to test more than 950 serum samples collected from approximately 210 infant/juvenile and 130 adult baboons in a captive breeding colony over a period of seven years. Results indicated that baboon cytomegalovirus, lymphocryptovirus, and rhadinovirus are transmitted efficiently within the colony and are acquired at an early age. Baboon α-herpesvirus HVP2 and polyomavirus simian virus 40 (SV40) were acquired later and by fewer juveniles than were the other three herpesviruses. More than 60% of baboons acquired HVP2 before reaching sexual maturity, indicating that oral infection of infants and juveniles, rather than sexual transmission between adults, is the predominant mode of transmission for this virus. Antibody to simian varicella virus (SVV) was found in about 40% of baboons. SVV was acquired principally by infants and juveniles; few adults seroconverted despite seronegative adults being in constant contact with infants and juveniles undergoing primary infection. Time of seroconversion was not statistically correlated to specific individual herpesviruses, suggesting that each virus is acquired as an independent infection event rather than multiple viruses being acquired at the same time. Several baboons that were delivered by cesarean section and were housed separate from, but in close proximity to, other baboons remained free of many or all viruses for several years, suggesting that, similar to human herpesviruses, baboon herpesviruses and SV40 are transmitted principally by direct contact.

Lactobacilli are the predominant microorganisms in the vaginal flora of human beings, and are known to play an important role in protecting them from genital infections. On the other hand, the composition of the vaginal flora differs among laboratory animal species, and lactobacilli are not the predominant vaginal microorganism in many laboratory animals. We speculated that the vaginal flora of chimpanzees would be more similar to those of human beings than to those of other animal species, because chimpanzees are phylogenetically close to human beings, and their reproductive physiology is similar to that of human beings. To clarify our speculation, we examined the development of the vaginal flora in chimpanzees (Pan troglodytes). Streptococci, lactobacilli, and members of the family Bacteroidaceae were the most predominant bacteria in the vagina of mature chimpanzees (9 to 22 years old). During development of the vaginal flora of chimpanzees, the total number of bacteria increased with age and reached a plateau just before sexual maturity (5 to 7 years of age; juvenile period). Lactobacilli were already one of the predominant bacteria before sexual maturity. In mature chimpanzees, the total number of bacteria (aerobes and anaerobes) in the vagina was highest during the swelling phase of the menstrual cycle. During the swelling phase in mature chimpanzees, streptococci, lactobacilli, and Bacteroidaceae were the most frequently isolated (100%) organisms, and the total number of organisms recovered from vaginal specimens from these three groups was the highest. In mature chimpanzees in which the number of bacteria was the highest, lactobacilli were the predominant bacteria. Taken together, these results suggest that these three bacterial groups (streptococci, lactobacilli, and Bacteroidaceae) are indigenous to the vagina of chimpanzees, and chimpanzees would be the most suitable laboratory animals for studying the role of lactobacilli in the vagina of human beings.

Increased mortality was observed in a single colony of 50 Xenopus laevis. The frogs were used as oocyte donors in developmental biology studies. Necropsy findings included dermal erythema and petechiation consistent with red leg syndrome; dermal ulcerations and white, filamentous growths on the skin were consistent with Saprolegnia sp. Microscopic evaluation of the skin and fungus revealed an astigmatid mite similar to those of the genus Rhizoglyphus. The mite was also found in the water and the biological filter of the tanks housing the frogs. This mite is considered not to be a parasite of X. laevis; instead, it feeds off moss, fungi, and detritus. Subsequent evaluation of the sphagnum moss used for shipping the frogs from the supplier revealed the same mite in the moss. Our hypothesis is that the mite was introduced into the tank with the shipment of new frogs in sphagnum moss. The mites lived within the biological filter, and were only found after the growth of Saprolegnia sp. attracted the mites to the frogs. Laboratory animal care and veterinary personnel should consider non-pathogenic species of mites in the differential diagnosis of acariasis in Xenopus frogs.