Showing papers in "Comparative Medicine in 2005"

Hearing in laboratory animals: strain differences and nonauditory effects of noise.

Abstract: Hearing in laboratory animals is a topic that traditionally has been the domain of the auditory researcher. However, hearing loss and exposure to various environmental sounds can lead to changes in multiple organ systems, making what laboratory animals hear of consequence for researchers beyond those solely interested in hearing. For example, several inbred mouse strains commonly used in biomedical research (e.g., C57BL/6, DBA/2, and BALB/c) experience a genetically determined, progressive hearing loss that can lead to secondary changes in systems ranging from brain neurochemistry to social behavior. Both researchers and laboratory animal facility personnel should be aware of both strain and species differences in hearing in order to minimize potentially confounding variables in their research and to aid in the interpretation of data. Independent of genetic differences, acoustic noise levels in laboratory animal facilities can have considerable effects on the inhabitants. A large body of literature describes the nonauditory impact of noise on the biology and behavior of various strains and species of laboratory animals. The broad systemic effects of noise exposure include changes in endocrine and cardiovascular function, sleep-wake cycle disturbances, seizure susceptibility, and an array of behavioral changes. These changes are determined partly by species and strain; partly by noise intensity level, duration, predictability, and other characteristics of the sound; and partly by animal history and exposure context. This article reviews some of the basic strain and species differences in hearing and outlines how the acoustic environment affects different mammals.

Reference cardiopulmonary values in normal dogs.

Abstract: The purpose of this project was to collate canine cardiopulmonary measurements from published and unpublished studies in our laboratory in 97 instrumented, unsedated, normovolemic dogs. Body weight; arterial and mixed-venous pH and blood gases; mean arterial, pulmonary arterial, pulmonary artery occlusion, and central venous blood pressures; cardiac output; heart rate; hemoglobin; and core temperature were measured. Body surface area; bicarbonate concentration; base deficit; cardiac index; stroke volume index, systemic and pulmonary vascular resistance indices; left and right cardiac work indices; alveolar partial pressure of oxygen (pO2) ; alveolar-arterial pO2 gradient (A-apO2); arterial, mixed-venous, and pulmonary capillary oxygen content; oxygen delivery; oxygen consumption; oxygen extraction; venous admixture; arterial and mixed-venous blood CO2 contents; and CO2 production were calculated. In the 97 normal, resting dogs, mean arterial and mixed-venous pH were 7.38 and 7.36, respectively; partial pressure of carbon dioxide (pCO2), 40.2 and 44.1 mm Hg, respectively; base-deficit, -2.1 and -1.9 mEq/liter, respectively; pO2, 99.5 and 49.3 mm Hg, respectively; oxygen content, 17.8 and 14.2 ml/dl, respectively; A-a pO2 was 6.3 mm Hg; and venous admixture was 3.6%. The mean arterial blood pressure (ABPm), mean pulmonary arterial blood pressure (PAPm), pulmonary artery occlusion pressure (PAOP) were 103, 14, and 5.5 mm Hg, respectively; heart rate was 87 beats/min; cardiac index (CI) was 4.42 liters/min/m2; systemic and pulmonary vascular resistances were 1931 and 194 dynes.sec.cm-5, respectively; oxygen delivery, consumption and extraction were 790 and 164 ml/min/m2 and 20.5%, respectively. This study represents a collation of cardiopulmonary values obtained from a large number of dogs (97) from a single laboratory using the same measurement techniques.

Behavioral differences among fourteen inbred mouse strains commonly used as disease models.

Abstract: We compared the behavior of 14 inbred mouse strains and an F1 hybrid commonly used in transgenic and knockout production. These strains were 129P3/J, 129S1/SvImJ, 129S6/SvEvTac, 129T2/SvEmsJ, 129X1/SvJ (formerly 129/J, 129/Sv-p+Tyr+Kitl+/J, 129/SvEvTac, 129SvEmsJ, and 129/SvJ, respectively), A/JCrTac, BALB/cAnNTac, C3H/HeNTac, C57BL/6J, C57BL/6NTac, DBA/2NTac, FVB/NTac, NOD/MrkTac, SJL/JCrNTac, and the hybrid B6129S6F1Tac. Performance in three behavioral tests (rotorod, open-field activity-habituation, and contextual and cued fear conditioning) was determined. On the rotorod assay, SJL/JCrNTac mice had the shortest latencies to fall on the first day of testing, and DBA/2NTac mice showed impaired motor learning. Open-field behavior was analyzed using the parameters total distance, center distance, velocity, and vertical activity. 129T2/EvEmsJ and A/JCrTac were least active in the open field, whereas NOD/MrkTac mice were most active. Contrary to earlier studies, we found that all strains habituated to the open field in at least one of these parameters. In contextual and cued fear conditioning, all strains displayed activity suppression. However, FVB/NTac mice reacted less strongly to both context and cue than did most of the other strains. There were no significant behavioral differences between C57BL/6J and C57BL/6NTac, except for higher open-field activity in C57BL/6J female mice. These findings illustrate the importance of the appropriate selection of background strain for transgenic, gene targeting, or drug research.

Characterization of cyclicity and hormonal profile with impending ovarian failure in a novel chemical-induced mouse model of perimenopause.

Abstract: 4-Vinylcyclohexene diepoxide (VCD) causes early, gradual ovarian failure in mice because it specifically targets small pre-antral ovarian follicles. The period between loss of these follicles and ovarian failure is analogous to perimenopause in women. We sought to characterize the period of onset of ovarian failure in VCD-treated mice in regard to estrous cycle length and hormonal changes. Female C57Bl/6 mice (age, 28 days) were dosed daily for 15 days with VCD (160 mg/kg intraperitoneally) to cause early ovarian failure or with vehicle only (control animals). Cycle length was monitored by vaginal cytology. Plasma levels of 17beta-estradiol (E2), progesterone (P4), and follicle-stimulating hormone (FSH) in control and VCD-treated animals were measured during proestrus of cycles 1 through 12. Cycle length (mean, 5.8 days) did not differ between groups for cycles 1 through 4. In contrast, cycle length during cycles 5 through 12 was increased (mean length, 10.9 days; P < 0.05 versus control) in VCD-treated animals, which also showed an apparent increase in plasma FSH levels. Plasma E2 and P4 at proestrus did not differ between groups during any cycle. Ovarian failure in VCD-treated mice was confirmed by histological evaluation on day 156 after onset of dosing, whereas control animals were still cycling. Therefore, despite compromised cycle length in VCD-treated mice, peak ovarian steroid production in preovulatory follicles at proestrus is adequate. These results demonstrate that the VCD-treated mouse can serve as an appropriate model to mimic hormonal changes during the perimenopausal transition in women.

Susceptibility of rats to corneal lesions after injectable anesthesia.

Abstract: Corneal injury is not a commonly reported side effect after injectable or inhalation anesthesia in rats, but a number of surgery studies at our facility resulted in a high incidence of these injuries To explore the potential association of various anesthetic protocols with the development of corneal lesions in rats, we retrospectively evaluated clinical records and sections of eyes from 215 male and 187 female Wistar rats used in eight intravenous infusion toxicology studies None of the studied compounds was associated with eye toxicity For placement of jugular vein vascular access ports, rats were anesthetized with enflurane, isoflurane, ketamine-xylazine, or Hypnorm-midazolam Histologically, corneal changes were scored from 0 to 4 in light of degree of mineralization, leukocytic infiltrates, neovascularization, fibrosis, and ulceration Prestudy (postsurgical) ophthalmic examination findings of corneal opacities were correlated with mineralization of the anterior limiting membrane and corneal ulceration Corneal lesions were more severe in animals anesthetized with ketamine-xylazine, and minimal changes occurred after anesthesia with either enflurane or isoflurane The results of further analysis suggest that corneal lesions can be observed within 24 h after injectable anesthetic administration and are not reversible The severity of corneal changes was reduced when ketamine-xylazine anesthesia was reversed with yohimbine Compared with Sprague-Dawley and Lewis rats, Wistar, Long-Evans, and Fischer 344 rats had increased incidence and severity of corneal lesions after anesthesia with ketamine-xylazine, suggesting that these three strains are at increased risk for developing postanesthetic corneal lesions with this regimen

Behavioral and clinical pathology changes in koi carp (Cyprinus carpio) subjected to anesthesia and surgery with and without intra-operative analgesics.

Abstract: Fish surgery is becoming increasingly common in laboratory and clinical settings. Behavioral and physiologic consequences of surgical procedures may affect experimental results, so these effects should be defined and, if possible, ameliorated. We document behavioral and clinical pathology changes in koi carp (Cyprinus carpio) undergoing surgery with tricaine methanesulphonate (MS-222) anesthesia, with and without intraoperative administration of the opiate butorphanol (0.4 mg/kg intramuscularly) or the nonsteroidal antiinflammatory analgesic ketoprofen (2 mg/kg intramuscularly). For all fish combined, surgery resulted in reduced activity, lower position in the water column, and decreased feeding intensity at multiple time points after surgery. The butorphanol-treated group was the only one not to experience significant (P < 0.05) alterations from presurgical behaviors. Clinical pathology changes at 48 h after anesthesia and surgery included decreased hematocrit, total solids, phosphorus, total protein, albumin, globulin, potassium, and chloride and increased plasma glucose, aspartate aminotransferase, creatine kinase, and bicarbonate. The only clinical pathology difference between treatment groups was a lower increase in creatine kinase in the ketoprofen-treated group. No adverse effects of butorphanol or ketoprofen at these doses were identified. These results suggest a mild behavioral sparing effect of butorphanol and reduced muscle damage from the antiinflammatory activity of ketoprofen.

Rodent models of rheumatoid arthritis.

Abstract: The study of rheumatoid arthritis is greatly facilitated by animal models that enable investigation of a complex system involving inflammation, immunological tolerance, and autoimmunity. Although the models cover several species and pathogenetic mechanisms and can be classified as induced or spontaneous, all converge on arthritis. However, because each model features a different mechanism driving disease expression, the merits of each should be evaluated carefully in making the appropriate choice for the scientific question to be addressed. In addition, because the incidence and kinetics of disease vary by model, careful thought should be given to protocol design to minimize animal use.

Relation between storage temperature and fertilizing ability of freeze-dried mouse spermatozoa

Abstract: The advantage of freeze-dried mouse spermatozoa is that samples can be stored in the refrigerator (+4 degrees C). Moreover, the storage of freeze-dried spermatozoa at ambient temperature would permit spermatozoa to be shipped easily and at low cost around the world. To examine the influence of the storage temperature on freeze-dried spermatozoa, we assessed the fertilizing ability of spermatozoa stored at different temperatures. Cauda epididymal spermatozoa were freeze-dried in buffer consisting of 50 mM ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid, 50 mM NaCl, and 10 mM Tris-HCl (pH 8.0). Samples of freeze-dried spermatozoa were stored at -70, -20, +4, or +24 degrees C for periods of 1 week and 1, 3, and 5 months. Sperm chromosomes were maintained well at -70, -20, and + 4 degrees C for 5 months, and oocytes fertilized with these spermatozoa developed to normal offspring. Moreover, the chromosomal integrity of spermatozoa stored at -20 or + 4 degrees C did not decrease even after 17 months. In contrast, the chromosomes of spermatozoa stored at +24 degrees C were maintained well for 1 month but became considerably degraded after 3 months. In addition, to investigate the cause of deterioration of sperm chromosomes during storage at +24 degrees C, spermatozoa were freeze-dried in buffer containing DNase I. The chromosomes of spermatozoa freeze-dried with 1 or 0.2 units/ml of DNase I, 100% or 72%, respectively, exhibited chromosomal abnormalities. Our findings suggest that freeze-dried spermatozoa can be stored long-term with stability at +4 degrees C, and the suppression of nucleases present in the buffer or spermatozoa during storage led to the achievement of long-term storage of freeze-dried spermatozoa.

Evaluating postoperative analgesics in mice using telemetry.

Abstract: The study examined the efficacy of preemptive or postoperative analgesia on surgical pain in the mouse. Radiotelemetry transmitters were surgically implanted in 28 female ICR mice. A mock ova implantation surgery was then performed. Mice were treated with a single dose of buprenorphine or flunixin meglumine prior to or after surgery, three doses of buprenorphine, or were untreated. Heart rate, blood pressure, home cage activity, food and water consumption, and body weight were measured. The no-analgesia group showed no significant differences between any parameters collected prior to surgery and those collected at similar times during the day of surgery. Significant increases in mouse activity on the day of surgery occurred with all analgesic treatments, compared with pre-surgical activity. There were no consistent significant changes in any other telemetry parameter after treatment with analgesics compared with no analgesia. Food consumption and body weight the day after surgery were reduced significantly in the animals treated with three doses of buprenorphine compared with untreated mice and mice given a single dose of buprenorphine. We conclude that the mock ova implant procedure does not induce sufficient pain to cause alterations in heart rate and blood pressure in the mouse. Activity was significantly reduced in the first 6 h after surgery in mice without analgesia, compared with activity prior to surgery. There were no significant differences between pre-emptive and postoperative analgesia. Body weight and food and water consumption were poor measures of pain because analgesia alone affected these parameters.

Overview movement as an index of anesthetic depth in humans and experimental animals.

Abstract: 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.

NIH Swiss and Black Swiss mice have retinal degeneration and performance deficits in cognitive tests.

Abstract: Swiss mice are among the most commonly used outbred strains in biomedical research. Because prior knowledge of the baseline phenotypes of mouse strains will allow informed selection of strains for particular experiments, we sought to characterize the behavior of two previously untested outbred Swiss strains--NIH Swiss and Black Swiss--in the two most widely used paradigms for evaluating the cognitive abilities of mice. Unlike the C57BL/6J and C57BL/6J-Tyr(c-2J) controls, animals of both outbred Swiss strains were unable to demonstrate learning in the Morris water maze and contextual fear conditioning paradigms. A polymerase chain reaction assay revealed that all of the NIH Swiss and Black Swiss mice tested were homozygous for the recessive retinal degeneration 1 mutation of the Pde6b gene. Histological examination of NIH Swiss and Black Swiss mouse eyes confirmed the presence of retinal degeneration, which causes visual image blindness. These findings indicate that NIH Swiss and Black Swiss mice are visually im paired and thus may be unsuitable for use in some experiments.

The effects of fluoxetine and buspirone on self-injurious and stereotypic behavior in adult male rhesus macaques.

Abstract: The effects of two serotonergic agents--fluoxetine, a serotonin (5-HT) reuptake inhibitor, and buspirone, a 5-HT 1a agonist--on rates of self-injurious and stereotypic behavior were examined in 15 adult male Macaca mulatta. All animals received a placebo for 2 weeks followed by either buspirone or fluoxetine for 12 weeks. Behavior was monitored using a focal sampling technique throughout the study and for 2 weeks post-study. Cerebrospinal fluid (CSF) samples and body weights were obtained pre-study, at the ends of placebo and treatment phases, and post-study. Fluoxetine and buspirone were significantly effective in reducing rates of self-biting during treatment weeks 1 to 8 and self-directed stereotypic behavior during weeks 5 to 12 and post-treatment. No significant effect of either treatment on hair-plucking, stereotypic pacing, saluting, or head tossing was identified. The duration of neutral behavior increased, and rates of scratching and yawning decreased in the buspirone-treated condition. In the fluoxetine-treated condition, rates of yawning, scratching, and self-directed grooming were higher overall compared with those of buspirone-treated animals, and rates of scratching increased significantly (P < 0.05) in weeks 9 to 12; these findings suggest that animals in the fluoxetine-treated condition experienced higher levels of anxiety throughout the study. In both treatment conditions, concentrations of CSF 5-HIAA (5-HT metabolite) were significantly lower (P < 0.05) than placebo concentrations. Fluoxetine and buspirone may be efficacious for treatment of self-injurious and self-directed stereotypic behavior in macaques. Further studies are required to determine the optimal dosages and treatment length.

Effects of housing density and cage floor space on three strains of young adult inbred mice.

Abstract: Some recommendations in the Guide for the Care and Use of Laboratory Animals (the Guide) are based on best professional judgment. Our current efforts are directed toward replacement with data-driven standards. We demonstrated earlier that young adult C57BL/6J mice could be housed with half the floor space recommended in the Guide without discernable negative effects. This report extends that work by examining optimal housing densities for young adult male and female BALB/cJ, NOD/LtJ, and FVB/NJ mice. These 8-week studies were initiated with 3-week-old BALB/cJ and NOD/LtJ mice and 3- to 5-week-old FVB/NJ mice housed in three cage types. We adjusted the number of mice per cage to house them with the floor space recommended in the Guide (approximately 12 in2 [ca. 77 cm2] per mouse) down to 5.6 in2 [ca. 36 cm2] per mouse. Early-onset aggression occurred among FVB/NJ male mice housed at all densities in cages having 51.7 in2 (ca. 333 cm2) or 112.9 in2 (ca. 728 cm2) of space. FVB/NJ male mice housed in shoebox (67.6 in2 [ca. 436 cm2]) cages did not exhibit aggression until the fifth week. Urinary testosterone output was density-dependent only for BALB/cJ male mice in shoebox cages (output decreased with increasing density) and FVB/NJ male mice. We conclude that all but FVB/NJ male mice can be housed with half the floor space specified in the Guide. The aggression noted for male FVB/NJ mice may have been due to their age span, although this did not impact negatively on the female FVB/NJ mice.

Genetic characterization of Indian-origin and Chinese-origin rhesus macaques (Macaca mulatta).

Abstract: Genetic differences between Indian-origin and Chinese-origin rhesus macaques are as great as those between some primate species and can influence the results of experiments in which both are used as animal models for the study of the same human diseases. Unfortunately, many breeding facilities do not know with certainty the origin of the founders of their rhesus breeding colonies. Here I summarize the most definitive of the genetic traits among the microsatellite (STR) loci and mitochondrial DNA sequences that my laboratory previously reported to characterize Indian-origin and Chinese-origin rhesus macaques and then estimate the frequencies of these traits and their reliability as indicators of country of origin. The expression of diagnostic traits at two or more of four different unlinked loci provides a nearly 100% reliability in distinguishing rhesus macaques of Indian and Chinese origin.

Noninvasive measures of body fat percentage in male Yucatan swine.

Abstract: 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.

A national survey of laboratory animal workers concerning occupational risks for zoonotic diseases.

Abstract: In this cross-sectional survey of laboratory animal workers in the United States, 23 of 1367 persons reported 28 cases of infection with zoonotic agents from research animals at their workplace during the past 5 years, with six persons indicating that their infections were medically confirmed Based on these data, the annualized incidence rate for work-related transmission of zoonotic agents from laboratory animals was 45 cases per 10,000 worker-years at risk (95% confidence interval, 30 to 65 cases), approximating the rate for nonfatal occupational illnesses in the agricultural production-livestock industry and for those employed in the health services during 2002 Logistic regression analysis found various characteristics of persons and their employers that were significantly associated with the likelihood of having been medically evaluated for exposure to a zoonotic agent from laboratory animals Most (95595% +/- 11%) persons working with laboratory animals or their tissues indicated that they knew whom to talk to at their institution for medical evaluation and care should they be concerned about the possibility of an occupationally acquired zoonotic disease in future However, occupational illnesses and exposures among laboratory animal workers was underreported, as 10 of the 28 (36%) alleged zoonotic disease cases were not communicated to the employee's supervisor Lack of concern about the potential significance to their health and the perception of punitive consequences to the employee were some of the reasons cited for underreporting, an issue which must be vigorously addressed in the interests of continuing progress toward zoonotic disease prevention in this field

The efficacy of diazepam treatment for the management of acute wounding episodes in captive rhesus macaques.

Abstract: The spontaneous development of self-injurious behavior (SIB) in singly housed monkeys poses a challenge for their management and well-being in captivity. Relatively little information is available on effective treatments for SIB. This study examined the effects of diazepam (Valium) on self-wounding and other abnormal behaviors in eight individually housed male rhesus monkeys (Macaca mulatta). Each monkey's response to an anxiolytic dose of diazepam (1 mg/kg or greater orally) was compared with the animal's behavior during drug-free periods. When examined across all animals, treatment with diazepam did not significantly alter wounding frequency or rates of self-directed biting without wounding. However, closer examination of the data revealed that four of the animals showed significant decreases in self-biting and wounding frequency (positive responders, PR group), whereas the remaining monkeys showed a trend towards increased wounding frequency (negative responders, NR group). Subsequent examination of colony and veterinary records demonstrated that compared with NR monkeys, PR monkeys had spent significantly more years in individual cage housing and had experienced a greater number of minor veterinary procedures. PR animals also were significantly less likely to have a documented history of self-biting behavior. Our findings suggest that SIB is not a homogeneous disorder in rhesus monkeys; rather, distinct subtypes exist that require different treatment approaches.

Duration of effects on clinical parameters and referred hyperalgesia in rats after abdominal surgery and multiple doses of analgesic.

Abstract: This study evaluated the duration of clinical effects and referred hyperalgesia in rats (n = 10 per group) undergo ing abdominal surgery with analgesics (ketoprofen at 3 mg/kg and buprenorphine at 0.01 or 0.1 mg/kg) administered intramuscularly twice daily for 72 h beginning prior to surgery; no-surgery and no-analgesia control groups were included. Food and water consumption and body weight were monitored daily. As a measure of referred hyperalgesia, tail-flick latency was measured daily, before and 4 h after analgesia administration. Compared with those of the no-surgery controls, significant decreases in food consumption and body weight occurred 24 h after surgery without analgesics. There were nonsignificant reductions in these effects by analgesics, but the benefits were not significantly different than those of saline. These parameters continued to be decreased with variable significance in the buprenorphine groups at 48 and 72 h after surgery. In both buprenorphine-treated groups, water consumption was significantly increased at 24 h after surgery but not at 48 or 72 h. Tail-flick latency was not significantly different between the no-surgery and no-analgesia groups but was significantly increased 4 h after high-dose buprenorphine administration and declined nonsignificantly over time in the other groups. We conclude that painful effects from surgery are present primarily during the first 24 h after surgery. The analgesic regimens tested did not completely reduce these effects. Buprenorphine was associated with adverse effects for as long as 72 h after surgery. Referred hyperalgesia from this abdominal surgery could not be measured using the tail-flick assay.

Characterization of immune responses during regression of rabbit oral papillomavirus infections.

Abstract: 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.

Canine transmissible venereal tumor: a large-animal transplantable tumor model.

Abstract: The canine transplantable venereal tumor is a naturally occurring transplantable round-cell tumor in dogs. Although experimental transplantable tumor models in rodents and rabbits are readily available, a reliable transplantable tumor model in a large animal that more closely resembles the physical dimensions of humans has not been available. A tumor model in a large animal would have a wide range of biomedical research applications, including the study of various interventional imaging techniques. In this report, we characterize the experimental transplantation of the canine transmissible venereal tumor in the brain, skin, muscle, prostate, lung, liver, and bone of dogs and provide X-ray computed tomographic and magnetic resonance imaging characteristics of the tumors in the brain, muscle, lung, and prostate.

Use of frozen-thawed oocytes for efficient production of normal offspring from cryopreserved mouse spermatozoa showing low fertility.

Abstract: Freezing of spermatozoa and unfertilized oocytes is a useful tool for the conservation of mouse genetic resources. However, the proportion of frozen-thawed oocytes fertilized with spermatozoa in vitro is low because spermatozoa, especially those frozen-thawed, can not penetrate into oocytes because of hardening of the zona pellucida following premature release of cortical granules. To produce offspring efficiently from cryopreserved transgenic mouse gametes, we fertilized frozen-thawed gametes by using intracytoplasmic sperm injection (ICSI) and assessed pre- and postimplantation development of embryos. Compared with fresh unfertilized oocytes, frozen-thawed unfertilized oocytes were highly tolerant to damage by injection, as the survival rates after injection of frozen spermatozoa were 51 and 78%, respectively. Frozen-thawed oocytes that survived after sperm injection developed normally to the blastocyst stage and gave rise to offspring. Moreover, offspring with transgenes also were obtained from frozen gametes fertilized by ICSI. These results demonstrate that ICSI is an efficient technique for producing offspring from transgenic spermatozoa showing low fertility and that use of frozen-thawed oocytes leads to conservation of genetic resources because suboptimally preserved gametes are not wasted.

Successful treatment of idiopathic dilated cardiomyopathy in an adult chimpanzee (Pan troglodytes).

Abstract: Various congenital and acquired forms of heart disease have been reported in captive lowland gorillas, and heart disease is a major cause of morbidity and mortality in geriatric humans. However, the prevalence of heart disease is unknown in nonhuman great apes species. Indeed, little is known about heart disease in chimpanzees, although the species has been used in research for decades. This report details the clinical presentation and diagnostics (thoracic radiography, electrocardiography, and echocardiography) utilized to diagnose idiopathic dilated cardiomyopathy in a 27-year-old male chimpanzee. Treatment decisions--indicated by follow-up diagnostics including repeat electrocardiography, echocardiography, and clinical laboratory data--over the 22-month period during which he continues to be treated are described. In addition, electrocardiographic and echocardiographic findings obtained from 20 clinically normal adult (11 female and 9 male) chimpanzees are presented for comparison.

Relationship of feline bispectral index to multiples of isoflurane minimum alveolar concentration.

Abstract: The study reported here was done to determine the relationship between bispectral index (BIS) values and minimum alveolar concentration (MAC) multiples of isoflurane in cats. Isoflurane MAC was determined using the tail-clamp method in eight domestic cats. Ten days later, the cats were anesthetized a second time with isoflurane at each of five MAC multiples administered in random order. Ventilation was controlled and, after a 20-min equilibration period at each MAC multiple of isoflurane, BIS data were collected for 5 min and the median BIS value calculated. Data from each isoflurane MAC multiple were compared using analysis of variance for repeated measures, and statistical significance was set at P < 0.05. The MAC of isoflurane (mean +/- 1 standard deviation) was 1.8% +/- 0.2%. BIS values at 0.5 MAC could not be recorded due to spontaneous movement in all eight cats. BIS values at 2.0 MAC were confounded by burst suppression in seven of the eight cats. Over the range of 0.8 to 1.5 MAC, BIS values decreased significantly with increasing end-tidal isoflurane concentrations. Mean (+/- 1 standard deviation) BIS measurements were 32 +/- 3 at 0.8 MAC, 20 +/- 4 at 1.0 MAC, and 5 +/- 3 at 1.5 MAC. Therefore, BIS values are inversely and linearly related to end-tidal isoflurane concentrations in anesthetized cats. However, the consistently low BIS values recorded in this study suggest that clinical BIS endpoints used to titrate anesthetic agents in humans may not be applicable to cats.

Evaluation of two combinations of Domitor, Zoletil 100, and Euthatal to obtain long-term nonrecovery anesthesia in Sprague-Dawley rats.

Abstract: We sought to evaluate a new protocol designed to maintain long-term, nonrecovery, surgical anesthesia in Sprague-Dawley rats. In the first phase, two groups of rats were anesthetized with two different dose combinations of Domitor (medetomidine) and Zoletil 100 (tiletamine-zolazepam) to investigate their efficacy in induction of anesthesia. One combination comprised Domitor at 35 microg/kg and Zoletil 100 at 40 mg/kg, whereas the other comprised Domitor at 50 microg/kg and Zoletil 100 at 20 mg/kg. Both combinations effectively induced deep anesthesia and caused no mortality, but the duration of anesthesia differed statistically. In the second phase, we induced anesthesia with both Domitor-Zoletil 100 dose combinations then investigated the possibility of maintaining anesthesia for 5 h by administering Euthatal (pentobarbitone) intra-arterially at 10 mg/kg hourly. Depth of anesthesia, mortality, physiological parameters, blood gas analysis, hematology, clinical chemistry, and postmortem histopathology were recorded. Euthatal provided stable long-term anesthesia with both dose combinations of Domitor-Zoletil 100. Seven of 8 (88%) animals anesthetized with Domitor at 50 microg/kg and Zoletil 100 at 20 mg/kg successfully were maintained under deep anesthesia for 5 h. Higher mortality (36% versus 12%) occurred in group of animals treated with Domitor at 35 microg/kg and Zoletil 100 at 40 mg/kg. This difference may be linked to dose-related respiratory depression, as alterations of arterial gas levels were noted. Our findings suggest that, when long-term nonrecovery anesthesia is required, doses of 50 microg/kg Domitor and 20 mg/kg Zoletil 100 are preferable when given with Euthatal to maintain physiological conditions in animals.

Successful cyclosporine treatment for atopic dermatitis in a rhesus macaque (Macaca mulatta).

Abstract: A juvenile (1 year old ) female rhesus macaque (Macaca mulatta) developed a chronic active skin condition characterized by pruritus, erythema, alopecia, scaling, exfoliation, and lichenification. Lesions were limited to the ventrum, specifically rostral mandible and neck, axilla and inguinal regions, distal extremities, and interdigital regions. Differential diagnoses included infection, dietary deficiency, metabolic abnormality, endocrinopathy, and immunological injury. Diagnostic tests included complete hemogram, serum chemistry, skin scrapes for ectoparasite detection, hair plucks for dermatophyte culture, and a serum-based hypersensitivity panel. All results were within normal limits. Dermal biopsies revealed lesions consistent with active allergic dermatitis, and a diagnosis of atopic dermatitis was made. Oral cyclosporine (5 mg/kg daily) rapidly eliminated clinical evidence of dermatitis. Histologically, lesions resolved after 12 months of treatment. Atopic dermatitis is an inflammatory skin condition for which there are neither pathognomonic clinical or diagnostic features nor a single successful therapy. Basic criteria such as pruritus, lichenification, a chronic course, and history of allergies strongly support the diagnosis. One successful therapeutic agent is a macrolide calcineurin inhibitor, cyclosporine. It represents a safer class of immunomodulatory drugs than corticosteroids and provides targeted alteration of lymphocyte function. To our knowledge this case represents the first reported successful treatment of atopic dermatitis in a nonhuman primate utilizing cyclosporine.

Toxic epidermal necrolysis in two rhesus macaques (Macaca mulatta) after administration of rituximab.

Abstract: A 2- and a 7-year-old rhesus macaque developed toxic epidermal necrolysis (TEN) after administration of ritux imab. Rituximab, a chimeric monoclonal antibody (mAb) directed against the CD20 antigen on B lymphocytes, is used to treat certain B cell neoplasias. The macaques were part of a gene therapy study that involved administering an adeno-associated viral vector encoding human factor IX (hFIX) to the animals. Both animals developed antibody against hFIX, which eliminated expression of the protein. Rituximab was administered to deplete the population of B cells producing antibodies against the protein. Two days after treatment, the 7-year-old animal developed erythemic skin lesions that rapidly progressed in severity, resulting in epidermal sloughing and ulceration. Despite aggressive treatment with analgesics, antibiotics, and corticosteroids, the animal had to be euthanized 5 days later. The 2-year-old macaque had no reaction to the initial dose of rituximab and received a second infusion 2 weeks later. Two days after drug administration, skin lesions developed; aggressive analgesic, antibiotic, and corticosteroid treatment was initiated, and the lesions resolved. A third rituximab dose was given approximately 2 months after the second. Skin lesions developed and were treated. The animal made a full recovery. In both cases, skin biopsies were taken and histopathologic findings were consistent with TEN. A severe, life-threatening condition, TEN manifests as an intolerance reaction in the skin. The most common cause of TEN is a response to previous drug administration. To our knowledge, this condition has not been reported in association with rituximab administration in macaques.

Age-related diffuse chronic telogen effluvium- type alopecia in female squirrel monkeys (Saimiri boliviensis boliviensis).

Abstract: We investigated the diffuse alopecia affecting some female squirrel monkeys (Saimiri boliviensis boliviensis) housed in a breeding facility. We randomly selected 100 female and 10 male animals and performed a complete physical exam and a hair assessment on all animals; blood tests, trichograms, hair density; and skin biopsies in representative cases; and a dominance behavioral assessment of 50 animals. Hair coat was normal in 35 female monkeys and all 10 male animals. Of the 65 females with diffuse alopecia, 17 had mild, 22 moderate, and 26 severe hair loss. The alopecia group had a mean age of 9.6 +/- 0.6 years, whereas that of the normal group was 4.7 +/- 0.6 years (P < 0.05). The parity in the alopecia group was 4.2 +/- 0.6 but 2.0 +/- 0.6 (P < 0.05) in the normal group. There were no statistically significant differences in body weight, hemoglobin, blood urea nitrogen, serum glucose, liver aspartate aminotransaminase, or free thyroxine. The trichogram demonstrated 20.8% +/- 1.6% (mean +/- standard error) of telogen hairs in the alopecia group compared with 9.5% +/- 2.8% of the control group (P < 0.05). The hair density in the alopecia group was 52.8 +/- 4.1/cm2 and 79.6 +/- 14.3/cm2 in the control group. Skin biopsies in affected monkeys demonstrated increased telogen follicles, with no fibrosis or inflammation. There were no statistically significant differences in the dominance behavioral analysis. The findings are consistent with chronic telogen effluvium (CTE). A number of organic, behavioral, and dominance-related stress causes of CTE were excluded. CTE appears to be predominantly age-related in this population. CTE in female squirrel monkeys may serve as an animal model for human diffuse alopecia.

Low-dose infectivity of Staphylococcus aureus (SMH strain) in traumatized rat tibiae provides a model for studying early events in contaminated bone injuries.

Abstract: Animal models of post-traumatic acute osteomyelitis (OM) that closely mimic human scenarios, including infection prophylactic procedures such as debridement and lavage, may provide a better understanding of OM. We contaminated mechanically traumatized rat tibiae (n = 69) with various doses of a Staphylococcus aureus strain (SMH) known to cause human OM and then performed curettage and lavage. Tibiae were harvested 24 h after lavage for assessment of bacterial load and determination of minimal infective doses for 50% (ID50) and 95% (ID95) of rats. Some experiments varied tibial harvest time after lavage (n = 10); for progressive infection, tibiae were evaluated at 7 and 15 days after contamination (n = 17 for each time point). At 24 h after contamination, the ID50 was 1.8 x 10(3) CFU, and the ID95 was 9.2 x 10(3) CFU. Tibial bacterial loads did not increase with inocula greater than the ID95. Lavage removed many bacteria from bone, but it did not prevent subsequent infection or disease. At 15 days after contamination, most tibiae (14 of 17) were infected, with macroscopic and radiological signs of established OM. This newly described rat OM model, with a low ID95 despite prophylactic curettage and lavage, closely mimics events in contaminated human bone injuries. This situation will allow study of early factors in contaminated bone injuries, including clinical interventions that may reduce infection and prevent disease.

Squamous cell carcinomas of the skin at ear tag sites in aged FVB/N mice.

Abstract: We report the development of squamous cell carcinomas (SCCs) of the skin at or near the site of ear tags composed of a nickel-copper alloy and used for identification during the course of a long-term study of incipient congenic FVB/N mice containing the human BCL6 transgene (FVB.Cg-Tg[tetO-BCL6]Bbn Tg[EmicroSR-tTa]83Bop), their littermate controls, and wild-type FVB/N. Of a total population of 160 mice, 14 (8.8%) developed SCCs in the tagged (right) ear after a median observation period of 25 months, but none of the animals developed tumors in the opposite ear (P = 0.0001). Nine of the fourteen mice with SCCs had to be euthanized because they were thought to be in distress from the ear condition, but the remaining five died or were euthanized for other reasons related to the research study. These animals ranged in age from 331 to 921 days at the time of death. Five of the tumors were well-differentiated (grade 1) SCCs; the remainder were grade 3 and tended to be deeply invasive neoplasms with undifferentiated areas containing a spindle cell component. One of these metastasized to kidney. When using the FVB/N mouse strain for long-term studies, it is necessary to consider that nearly 9% of the population may develop SCCs at or near ear-tag sites that may necessitate early removal of the animal.

Successful rederivation of contaminated immunocompetent mice using neonatal transfer with iodine immersion.

Abstract: 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.