Anesthesia is used widely in animal research, but there are diverse opinions regarding acceptable anesthetic depth. Excessive anesthesia is associated with increased morbidity and mortality. Traditionally, researchers have been taught that animal movement during surgical and experimental procedures indicates that the animal is 'underanesthetized.' Complex movement, however, can be initiated and propagated within the spinal cord, with little input from supraspinal structures. For example, frogs with high spinal-cord transections still maintain the wiping reflex, whereby the hindlimb can move to the forelimb to wipe away a noxious stimulus. Rats that have been decerebrated can perform complex tasks, such as grooming. Brain-dead humans can have spontaneous movement of the arms, legs, and head. Consistent with these phenomena, emerging evidence suggests that, in anesthetized animals, movement in response to noxious stimulation is abolished primarily via anesthetic action in the spinal cord. When isoflurane, halothane, or thiopental is delivered selectively to the brain circulation in goats, substantially greater anesthetic concentrations in brain are needed to ablate movement, as compared with those required upon delivery of anesthetic to the entire body. Rats that have had a precollicular decerebration require the same isoflurane concentrations to prevent movement as compared to intact rats. Furthermore, data from both humans and animals indicate that memory and awareness are ablated at anesthetic concentrations that are <50% of those needed to abolish movement. Collectively, these data indicate that animals can be anesthetized at depths that, although they do not abolish movement, still produce unconsciousness and amnesia.

Mycoplasma pulmonis induces persistent infections in laboratory mice and rats and can contaminate biological materials. We developed a fluorogenic nuclease polymerase chain reaction (fnPCR) assay to detect M. pulmonis specifically. Primer and probe sequences for the assay were targeted to 16S rRNA sequences specific to M. pulmonis. The assay consistently detected the equivalent of fewer than 10 copies of template DNA. When evaluated against a panel of 24 species of bacteria, the M. pulmonis assay detected only M. pulmonis isolates. Evaluation of 10-fold serial dilutions of cultured M. pulmonis showed that the M. pulmonis fnPCR assay and culture on Dutch agar had comparable sensitivity in detecting viable M. pulmonis organisms, whereas the mouse antibody production test displayed positive serologic results at dilutions higher than those in which viable organisms could be detected. Finally, the M. pulmonis fnPCR assay was able to detect M. pulmonis DNA in nasopharyngeal wash fluid and trachea, lung, and uterus tissue collected from mice naturally infected with M. pulmonis but did not detect the organism in similar samples collected from uninfected, negative control mice. The M. pulmonis fnPCR assay provides a high-throughput, PCR-based method to detect M. pulmonis in infected rodents and contaminated biological materials.

Cercopithecine herpesvirus 16 (Herpesvirus papio 2; HVP2) is an α-herpesvirus of baboons (Papio spp.) that generally causes minimal to inapparent disease in the natural host species. HVP2 is very closely related genetically and antigenically to Cercopithecine herpesvirus 1 (monkey B virus; BV) of macaques, which is well known for its extreme lethality in nonmacaque species including humans. Preliminary evidence suggests that a mouse model of HVP2 would be an excellent tool for studying zoonotic BV infections. Although the pathogenicity of different BV isolates in mice spans the full range of severity from apathogenic to extremely neurovirulent, testing of multiple HVP2 isolates revealed only two distinct phenotypes in mice regardless of route of inoculation: apathogenic (HVP2ap) and highly neurovirulent (HVP2nv). For the HVP2nv mouse model to truly reflect BV infection in both its natural host and the differential pathogenicity of BV in aberrant host species, HVP2nv should not produce severe disease in its natural host. To test this, juvenile baboons were inoculated with doses of 10⁶ or 10⁴ plaque-forming units of HVP2ap or HVP2nv by using an oral subdermal inoculation route. Parameters followed included the appearance of lesions, shedding of infectious virus, general health, and the immune response to the infection. Regardless of the inoculum dose used, no differences were noted between the two HVP2 subtypes in baboons in any of the parameters measured. These findings further support the use of the HVP2nv mouse system as a model to elucidate and study the viral determinants associated with cross-species BV neurovirulence.

Rabbit oral papillomavirus (ROPV) is a mucosatropic papillomavirus that causes small benign discrete papillomas within the oral cavity of domestic rabbits. The goal of this study was to characterize the immune cell infiltrate over the course of regression of oral papillomas. ROPV-infected oral tissues were harvested at various time points after infection and analyzed by immunohistochemistry for papilloma morphology, viral capsid proteins, and associated immune infiltrates. The results of this study indicated that the L1 and L2 viral capsid proteins were lost rapidly at a time that coincided with an inflammatory response from the rabbit. This inflammatory response began with a rapid rise in numbers of CD11c⁺ cells at early regression. CD11c⁺ cells continued to increase in frequency through mid-regression and remained the most-represented cell through late regression. The initial rise in CD11c⁺ cells was followed by an infiltrate containing increased numbers of activated T cells, including CD4⁺ and CD25⁺ cells, during mid-regression. Mid-regression coincided spatially with a loss of viral capsid stain, suggesting that immune cells or cytokines or both were playing a key role in clearance of the papillomas. CD8⁺ cells increased at the lowest rate and were at low levels in the papilloma epidermis even at mid-regression. All cell types decreased by late regression. CD11c⁺ and major histocompatibility class II⁺ cells were the last populations of cells to decrease in number.

The goal of this study was to assess the duration of pain-related clinical effects and referred hyperalgesia after surgery in rats. Isoflurane anesthesia with or without femoral vein cannulation was performed (n = 6 per group). Body weight and food and water consumption were monitored daily for 48 h, and tail-flick latency was measured twice daily for 24 h after surgery. Water consumption at 24 h after surgery was significantly decreased in the surgical group compared with baseline values and those of the anesthesia group. Body weight change and food consumption showed nonsignificant decreases compared with baseline in both groups 24 h after the procedure. There was a trend toward decreased food consumption after surgery compared with that for the anesthesia-alone group. Tail-flick latency was nonsignificantly decreased the afternoon after surgery compared with baseline values or that after anesthesia alone. Tail-flick latency was similar to baseline and between groups 24 h after surgery. All parameters were similar between groups and compared with baseline by 48 h after surgery. Our results show some changes in postsurgical pain-related parameters only during the initial 24-h period after femoral cannulation surgery, but only the change in water consumption was significant. Although this study involved only a small number of animals, our findings suggest that femoral vein cannulation produces a less painful stimulus than that seen in studies assessing these parameters after abdominal surgery. Hyperalgesia from a distant painful stimulus could not be measured in this model by using the tail-flick assay.

The purpose of this study was to assess the use of body circumference, ultrasonography, and serum leptin levels as noninvasive measures to estimate body fat percentage in adult, male, Yucatan swine, which are widely used in biomedical research models. Swine (ages, 8 to 15 months) were maintained for 20 weeks: control (n = 7); high-fat, high-cholesterol diet (hyperlipidemic; n = 8); alloxan-induced diabetes with high-fat, high-cholesterol diet (diabetic dyslipidemic; n = 7); and diabetic dyslipidemic plus exercise-trained (n = 6). Anesthetized swine were positioned on their dorsum for the following measurements: 1) neck, mid-abdomen, and widest abdominal girth circumferences; and 2) neck and mid-abdomen ultrasound measurements. Blood samples were obtained for quantification of serum leptin levels. After euthanasia, the carcass and viscera were separated for chemical composition analysis, which demonstrated a significant increase in carcass and visceral fat in the diabetic dyslipidemic swine compared to controls. Serum leptin levels were also increased in the hyperlipidemic and diabetic dyslipidemic swine. Regression analyses demonstrated a significant correlation between carcass fat, visceral fat, and all of the circumference, ultrasound, and serum leptin measures. In conclusion, the widest abdominal girth circumference was the noninvasive measure with the highest predictive value for estimating carcass and visceral fat in adult, male Yucatan miniature swine.

The purpose of this study was to develop a model of vascular injury in 8-week-old C3H/HeJ mice (weight, 25 to 30 g) by using air desiccation. The carotid arteries were excised 1 to 8 weeks postinjury and evaluated by Verhoeff's stain and immunocytochemistry. In the first group of mice studied (n = 107), neointimal formation occurred and peaked at Day 14. In addition, medial cell division (measured by bromodeoxyuridine labeling) peaked at Day 3, whereas intimal cell proliferation increased gradually throughout the experimental period of 21 days. In addition, extensive thrombus formation occurred within 3 days after injury. The next experiment involved 124 mice and evaluated the effect of anticoagulants on the neointimal and thrombotic response. Mice received aspirin, heparin, or vehicle-only time-release pellets. Both anticoagulants significantly decreased the neointimal and thrombotic responses. The results of this study validate our animal model as being consistent with the Response to Injury Hypothesis of atherogenesis.

Theiler's murine encephalomyelitis virus (TMEV), a member of the genus Cardiovirus, is an enteric pathogen of mice that causes acute encephalomyelitis followed by persistent central nervous system infection with chronic inflammation and demyelination after intracerebral inoculation. Although TMEV is a mouse pathogen, antibodies against TMEV strain GDVII have been detected in conventional rat colonies. Natural infection of rats by Cardiovirus has not yet been described. The purpose of this study was to demonstrate TMEV infection of rat colonies by using serologic assays, reverse transcription–polymerase chain reaction (RT-PCR) analysis, and clinical characterization. Indirect immunofluorescence assay of rat serum samples demonstrated antibodies against TMEV-GDVII in 86.3% of samples analyzed, and 77.2% of the antibody-positive samples had neutralizing antibodies. To determine whether rats can be infected experimentally with TMEV-GDVII, specific pathogen-free newborn mice and rats were inoculated intracerebrally with intestinal suspensions from seropositive rats. Both species showed the typical clinical signs of TMEV infection in mice, which is characterized by flaccid hindlimb paralysis and tremor. RT-PCR in brain tissue of experimentally infected animals detected RNA sequences corresponding to the 5′ noncoding region of Cardiovirus known as the 'internal ribosome entry site.' These results suggest that rats can be naturally infected with TMEV and related Cardiovirus. Therefore, continued health monitoring for TMEV infection should be included in rat colonies mainly because these animals are used for various experimental purposes.

There is an ongoing need to eradicate intercurrent disease from research mouse colonies. Commonly used surgical methods, however, are expensive and time-consuming. The purpose of this study was to determine the percentage of litters that could be rederived from infected mouse colonies by neonatal transfer. We immersed neonatal mice in a dilute iodine solution and transferred them to disease-free foster mothers within 48 h of birth. Donor and foster mothers were evaluated for pathogens by serology and fecal polymerase chain reaction (PCR) assay. Of 55 donor mothers, 100% were positive serologically and 59% were positive by fecal PCR for one or more tested organisms, including mouse hepatitis virus, Theiler's murine encephalomyelitis virus, mouse rotavirus, and Helicobacter hepaticus. At 4 to 6 weeks after neonatal transfer, 95% of foster mothers (which served as sentinels for the transferred pups) tested free of pathogens, the exceptions being one case of mouse parvovirus 1 and two of Helicobacter spp. We suggest that cross-fostering is a viable low-cost method for rederivation of mouse colonies contaminated with pathogens such as mouse hepatitis virus, Theiler's murine encephalomyelitis virus, mouse rotavirus, and H. hepaticus.

Male SS/Jr rats were placed on a specially formulated, high-cholesterol, low-sodium diet at 3 weeks of age. Of the 50 animals on the diet, 40 developed skin lesions ranging from focal areas of alopecia to diffuse areas of moist dermatitis on the head, face, ear pinnae, and neck. Similar lesions were noted later in 17 of 36 SS/Jr rats in a second study group. Histopathologic findings from two affected animals revealed diffuse, hyperplastic, ulcerative dermatitis, with bacterial colonies of cocci in superficial crusts, as well as chronic hepatic inflammation with hepatocellular glycogen and sinusoidal macrophage aggregates suggestive of lipidosis. Results of a fatty-acid profile of the affected rats showed serum linoleic acid levels of 931 to 1566 μmol/liter, whereas those for control (SS/Jr) samples ranged from 2711 to 3145 μmol/liter. Dietary analysis of the specially formulated diet showed that it contained only 0.225% linoleic acid, which is below the recommended level of 0.3 to 0.6%. In light of the clinical and dietary findings, a diagnosis of linoleic acid deficiency was made. The food manufacturer revised its dietary formulation to increase the linoleic acid content to 1.05%, and no further cases of dermatitis developed in any subsequent groups of rats maintained under the same study protocol.