Contemporary Topics 42 (2)

Comparison of three commercially available activated charcoal canisters for passive scavenging of waste isoflurane during conventional rodent anesthesia. Contemporary Topics 42 (2): 10.
In humans, chronic exposure to low levels of waste anesthetic gases is linked to increased incidences of neurologic and reproductive dysfunction, renal and hepatic toxicity and neoplasia. Those at increased risk include not only health care workers such as veterinarians and veterinary technicians, but also those individuals that interact with exposed health care professionals due to the retention within the body of the older inhalation agents (e.g. halothane, methoxyflurane, nitrous oxide), resulting in exposure to appreciable amounts of exhaled gases. This study explored the efficacy of three different activated charcoal canister brands (Breath Fresh, EnviroPure, F/Air) used in conjuction with standard rodent anesthesia equipment, to remove waste isoflurane from the work environment. Administration of isoflurane was accomplished using a precision isoflurane vaporizer. Each vaporizer was then connected to two circuits, one being a modified Bain, non rebreathing circuit attached to a sealed rodent face mask, and the other a 2L acrylic induction box. Exhaust ports of both systems were then attached to passive gas scavenging canisters. All circuit components were pretested for leakage. Quantification of waste isoflurane into the work environment was achieved using an ambient air analyzer containing a single beam infrared spectrophotometer. Placed horizontally, air was sampled approximately 2 cm from each canister's exit hole. Values above 100 ppm were designated as ">100", with further breakdown into either high emission, indicating a rapid increase (less than 15 sec) to 100 ppm and designated as "fast", or low emission indicating a gradual increase to 100 ppm, designated as "slow". Canisters (6 from each brand) were from identical manufacturing lots. Canisters were weighed weekly and were not rotated between anesthetic units, however, they were alternated between the two circuits every hour to ensure adsorption of isoflurane by each canister. At the conclusion of each study, the canister was attached to the facemask circuit, with the induction box circuit closed. Isoflurane measurements were obtained using a uniform, continuous concentration of 2% and an oxygen flow rate of 1L/min until the canister neared or surpassed their recommended use life (a weight of 50g). Air turnover rate in the animal procedure rooms was 26 nonrecirculating changes/hour. Emissions of isoflurane varied among the different canister brands, with used canisters of all brands emitting between 5 and 100 ppm, specifically, 46% of Breath Fresh, 8% of EnviroPure and 27% of F/Air canisters. All Breath Fresh and F/Air canisters showed this finding after only 60% of their rated use life had been surpassed. Unused canisters of each brand were also tested. A significant percentage of unused Breath Fresh units, failed to scavenge isoflurane (emissions >100 ppm). Finally, emissions exuded from both Breath Fresh and F/Air units varied inconsistently with time, compared the low, consistent levels emitted by EnviroPure units. Explanations for differences in isoflurane emissions from different brands of canisters may be due to creation of low resistance channels as a result of redistribution of charcoal granules during shipping and handling. Another potential issue may be as a result of the charcoal particle size and degree of charcoal saturation specific to each manufacturer. In order to further minimize risk to exposure of workers to waste anesthetic gases, an active exhaust system in conjunction with passive scavenging may be a solution. Active waste gas scavenging include using facemasks equipped with evacuation lines, and increasing air exchanges in animal procedure rooms. Finally, consideration should be given to implementation of programs to diligently monitor anesthetic gas levels within animal facility procedure rooms.
1. What is the currently acceptable level of exposure to newer halogenated gases according to NIOSH ?
2. How else can an active scavenging system be readily implemented ?
1. 2 ppm
2. Place the entire gas scavenging canister or entire anesthetic unit in a nonrecirculating fume hood.

Successful management of long-term general anesthesia in rabbits used as an animal model of human disease. Contemporary Topics 42 (2): 16.
Endotracheal intubation in the rabbit is challenging due to the difficulty in accessing the animal's airway and sensitivity to common anesthetic agents. The authors evaluated a method modified from a retrograde orotracheal intubation technique using straight transoral intubation under direct visualization. They hypothesized that this technique with guide wire assistance would improve airway access success and reduce perioperative mortality in the rabbit. 39 NZW rabbits (Oryctolagus cuniculi) were used in a research project inducing acute hindlimb ischemia. Ketamine (30 mg/kg im), Xylazine (6 mg/kg im) and isoflurane (3-5%) with face mask were used for anesthesia induction. The rabbits were placed with its head in a tipped, upright, and stretched position to bring the oral cavity, larynx, and cervical tracheal in linear alignment. The larngoscope with a #1 Miller blade was advanced through the oropharynx. The guide wire was passed through the vocal folds, and a lubricated sterile 2.5 to 3.0 mm ID endotracheal tube was advanced over the guide into the trachea. The animal was connected to an anesthesia machine with a rebreathing circuit and ventilator, providing intermittent positive pressure ventilation. Of the 39 animals, one rabbit died as a result of airway injury during intubation, and one died due to respiratory failure after extubation. After the technique was modified from using a rigid to a flexible guide wire, there were no deaths and no airway injuries in the subsequent 33 animals. Two animals required orogastric tube placement for gastric distention, but there were no other complications resulting from intubation or anesthesia. The death of the rabbit seems to be attributed to the intubation technique using the inflexible guide. Although adequate ventilation appeared problematic in their study, there was no correlation observed between time elapsed between anesthesia administration and intubation and observed hypercarbia. The authors conclude that the wire-guide transoral intubation under direct visualization with passive inhaled anesthesia using 3 to 5 % isoflurane in oxygen represents an excellent technique for induction and securing airway access in rabbits. A flexible guide wire should be used rather than a rigid one, to avoid mortality.
1. List the unfavorable anatomical characteristics in the rabbit that make endotracheal intubation difficult.
2. T/F A rigid guide wire, rather than a flexible wire should be used for rabbit endotracheal intubation.
3. T/F There was no correlation between high perioperative ETCO2 and time to recovery from anesthesia.
1. large incisors, a large tongue, a restricted mobility of the maxillary joints, a narrow oropharynx
2. F
3. T

Acetaminophen as a postsurgical analgesic in rats: a practical solution to neophobia. Contemporary Topics 42 (2): 20.
Acetaminophen administration via drinking water or gelatin is a common method of providing post-operative analgesia in rodents, despite recent reports that animals may consume suboptimal doses as a result of taste neophobia (fear of the new). Neophobia is commonly observed in rodents. The purposes of this study were to determine the length of time necessary for the extinction of any neophobic response to acetaminophen in adult Long Evens Rats and to determine whether the patterns of neophobic acquisition and extinction differed according to the sex of the animal.
A 7-day baseline was established for 32 rats (20 female and 12 male) in which average water consumption, food consumption, and body weight were measured. Cherry-flavored acetaminophen was then administered at 6 mg/ml for 7 days in the drinking water. Fluid consumption, food consumption, and weight were monitored during this period of drug exposure.
Results: Male rats had a 1-day reduction in fluid consumption followed by elevated fluid consumption of subsequent days. Female rats displayed elevated fluid consumption on all days of drug administration. Both male and female rats decreased food intake after drug exposure and subsequently lost weight.
Discussion: Neophobic response to acetaminophen in drinking water was mild in male rats and absent in female rats. Within 3 days of drug exposure, fluid consumption had increased 13.87% from baseline. There was a significant decline in food consumption and body weight for both sexes during drug exposure. These results were in partial contrast to the recent study by Speth, et al (CT 40(6), 2001) who found that adult female Sprague-Dawley rats showed decreased water intake during the 24 hours after administration of cherry-flavored acetaminophen administered in the drinking water. The authors suggest this may be the result of the use of Sprague-Dawley rats in the 2001 study and the use of Long Evans rats in the present study. The authors also speculate that differences could result from the use of different brands of cherry-flavored acetaminophen.
The use of oral acetaminophen offers some practical advantages over opioids. Although not a potent as opioids, they offer continuous analgesia without disruptions for re-dosing. Opiates can suppress the respiratory, immune, and cardiovascular systems. Side effects of acetaminophen include nephrotoxicity and hepatotoxicity.
The authors suggest that acetaminophen administered in the drinking water may serve as a reliable analgesic provided that drug exposure begins 1 to 2 days before surgery.
1. True/False: When acetaminophen is administered to rats via drinking water, significant reduction in food consumption and weight loss occurs.
2. Regarding the Long Evans rat's neophobic response to acetaminophen administered in the drinking water, which of the following in TRUE?
a. The neophobic response was transient
b. The neophobic response was absent in females
c. The neophobic response was absent in males
d. A and B
e. A and C
1. True
2. D

An outbreak of Pasteurella pneumotropic in genetically modified mice: treatment and elimination. Contemporary Topics 42 (2): 26.
Pasturella pneumotropica is an opportunistic gram-negative bacteria regarded as a secondary invader with low pathogenicity. Its primary site of colonization is the oropharynx, and immunodeficiency (specifically B cell immunity) has been reported to play a role in pathogenesis. Factors such as rodent genotype, coinfection with other pathogenic microbes, experimental manipulations, and stressful husbandry practices can predispose to disease. Infected immunocompetent animals are frequently asymptomatic.
Fluoroquinolone antibiotics possess a broad spectrum against gram-negative and gram-positive aerobes, absorption is rapid and complete after oral or parenteral administration, and widespread distribution with good tissue penetration is achieved. They also have a low potential for host toxicity and prolonged elimination half-lives.
The affected colony outlined here composed of knockout, transgenic, wild-type, and stock mice. Clinical symptoms (bilateral mucopurulent conjunctivitis and panophthalmitis) presented initially in a knockout strain. Clinical disease eventually disseminated throughout the entire colony, and clinical cases were manifested in all strains, including stock mice. The colony had been closed for several years, and other than husbandry maintenance of the breeding colony, no other experimental manipulations occurred. Quarterly and comprehensive annual sentinel tests were consistently negative in this colony. The colony was housed under barrier conditions.
Microbiologic testing via blood agar plates, colony morphology, gram-stain reaction, and the results from a panel of standard biochemical tests identified the organism as P. pneumotropica. Antibiotic sensitivity to fluoroquinolones was determined; enrofloxacin selected because of its availability in a convenient oral formulation and a previous report documenting the efficacy.
Triple antibiotic ophthalmic ointment was used in severely affected animals. All animals housed in the room were treated with 3.23% enrofloxacin in the drinking water with a final calculated dose of 85mg/kg per animal daily for 14 days. During the treatment period, there was no appreciable increase in morbidity, no alteration in breeding or water consumption.
One month post treatment completion, 22 mice with a history of clinical signs and 20 mice that had not been previously affected were selected with a variety of ages and strains represented. Multiple organ bacteriological testing was negative for P. pneumotropica. Thus, symptomatic animals cleared the infection and there were no asymptomatic carriers within the sampled population.
P. pneumotropica infection is usually subclinical in immunocompetent animals, and under the best conditions difficult to culture. While several reports clearly implicate immunosuppression as a predisposing factor for disease; some reports of clinical disease in wild-type animals indicate the role of the immune system in the disease pathogenesis is inconclusive at best. Nine months later, there has still been no cultured P. pneumotropica from any animal originating from this colony. The authors conclude that oral administration of enrofloxacin for P. pneumotropica was easy and inexpensive, had no detectable side effects, and resulted in both rapid and long-term eradication of clinical signs making it a feasible alternative to embryo transfer for an outbreak of P. pneumotropica in large mouse colonies.
1. The primary site of colonization of P. pneumotropica in the mouse is:
2. Frequently, immunocompetent mice affected with P. pneumotropica are:
3. True/False: P. pneumotropica is easy to grow in microbiologic culture.
4. True/False: It was the conclusion of these authors that enrofloxacin oral treatment is a feasible alternative to embryo transfer for an outbreak of P. pneumotropica in large mouse colonies.
1. the oropharynx
2. subclinical/asymptomatic
3. false
4. true

Validation study of social housing of canines in toxicology studies. Contemporary Topics 42 (2): 29.
The study was performed primarily to validate social housing for canines in toxicology studies. Studies have proven that socially housed canines display a closer human animal relationship compared to individually housed canines. However when changing from a system of individual housing of canines to social housing, it is prudent to ensure that the change would not effect the integrity of the test system and data collection.
Twenty Beagles, 10 males and 10 females, were enrolled in a study, which consisted of a 15-day pretest and a 28-day dosing period. Dogs 1 to 2 years of age were selected since sexually mature males have a higher potential to demonstrate incompatibility. The dogs were equally distributed into individually housed and socially housed groups and males and females were housed in separate rooms. Clinical signs, food consumption, body weight, hematology, clinical chemistry, urinalysis and EKG were monitored at different points during the study. Social housing occurred overnight and the dogs were separated for approximately 7 hours during data collection. During the late pretest period, one male socially housed Beagle was removed and replaced with a "spare male" to demonstrate the feasibility of replacement of rejected canines within a social setting.
Results:
There were no incidences of trauma due to fighting among the socially housed dogs. There were no significant changes in any of the parameters evaluated. There was no effect on the ability to dose the animals however the ability to evaluate fecal consistency on a single animal was affected.
Conclusions:
Social housing of canines in toxicology studies can be performed if planned appropriately. Other than collection of fecal data, there is no effect on the collection of clinical sign data. A simple modification of the method of fecal collection proved successful. Social housing according to the described study design was validated for the facility. Some questions from the research staff, which could not be addressed during this validation study, remain unanswered. The unanswered questions do not prohibit the use of social housing for canines.
1. Toxicology studies typically are conducted using dogs 7 to 9 months of age. What was the rational for selecting dogs one to two years of age for this study?
2. T/F Dogs should not be socially housed during studies due to the risk of fighting and competition for food that could result in a decrease in body weight of some dogs.
1. Typically signs of incompatibility will occur in older males.
2. F In the present study there were no significant changes in body weight in social housed dogs.

A combined and comparative study of physiologic and behavioral parameters in a systemic toxicity test. Contemporary Topics 42 (2): 31.
Acute systemic toxicity testing is used to assess safety and adverse effects of test substances. More refined tests than the traditional LD50 test have been proposed and in used in some EU countries. In the search for a more refined method, the present investigation used radiotelemetry and behavior to identify mild toxicity to lipopolysaccharide (LPS) and nickel injections in male Sprague Dawley rats. Body temperature and body weight gains were compared to behavioral indices such as activity, food and water intake. Both diurnal and nocturnal data were collected and compared.
LPS injection: body temperature, and heart rate were significant elevated during the daylight phase but there was no detectable effect on body temperature during the night. activity and weight gains were significantly reduced during the nocturnal phase when compared with controls (injected with saline).
Ni injection: body temperature, and body weight gain were significantly reduced during the day light phase but there were no effects on heart rate and activity. During the dark phase, there were significant hypothermia, significant reduction in body weight gain, minor reduction in water intake and no effect on heart rate and activity.
Significant increase in heart rate was detected with the higher dose of LPS (25 ug/kg) but not the lower dose of 5 ug/kg. Hence, radiotelemetry is not as accurate to detect mild toxicity. There were no changes in heart rate detected with both doses of Ni that were injected, but both showed hypothermia. Since reduced body temperature is a good sign of stress, body temperature seems to be more accurate than heart rate changes, according to the results. Still, further development of complimentary noninvasive techniques are necessary.
According to this study, which of the following is more accurate in detecting mild toxicity in rats both during daytime and nighttime?
A. body temperature
B. body weight gain
C. heart rate
D. activity
A. Body temperature. "Body temperature is the most sensitive physiological parameter that can be measrued diurnally or nocturnally, depending on whether a hyper- or hypothermic response, resppectively, to the test solution is anticipated. Overall, weight gain is the most sensitive index of all, responding to the lowest dose of NiCl2, but the response is dependednt on nocturnal administration of the test substance." p.37, last paragraph.

Quantification of TNF-alpha and IL-6 bioactivity in response to lipopolysaccharide in the degu (Octodon degus). Contemporary Topics 42 (2): 39.
The degu is the only diurnal rodent. The circadian rhythms of pro inflammatory cytokines (i.e. TNF-alpha and IL-6) and the fact that humans are diurnal suggests the degu may be a useful rodent model. The study was designed to use standard techniques to create an inflammatory response in the degu and then attempt to measure pro inflammatory cytokines.
The inflammatory response was created by the addition of lipopolysaccharides (LPS) into the blood for in vitro studies and intraperitoneally (IP) for in vivo studies. In the in vitro studies plasma was used for cytokine measurements. For in vivo studies broncheoalveolar lavage (BAL) and peritoneal lavage fluid samples were used for cytokine measurement. The cytokine measurements were taken using standard bioassays.
Results;
In Vitro - the addition of LPS lead to an increase in TNF-alpha but not IL-6. Controls showed no detectable pro inflammatory cytokines.
In Vivo - peritoneal fluid showed a significant increase in neutrophils post LPS injection confirming a strong inflammatory response compared to controls which lacked the neutrophil response. The BAL samples revealed no such inflammation in either controls or treated animals. TNF-alpha was found in high amounts in plasma, and peritoneal lavage but not in the BAL fluid. IL-6 was not detected in any of the samples.
Discussion
The inflammatory response seen in degu's post exposure to LPS is similar to that seen in other rodents. the TNF-alpha increase seen in the degu as part of the inflammatory response is similar to that seen in other species. However, unlike other species, IL-6 was not detectable in the presence of a marked inflammatory response. Suggested reasons for not finding IL-6 were; degu's do not have the IL-6 gene, the IL-6 levels were too low to detect, or most likely the bioassay used was incapable of detecting IL-6 in the degu.
In summary TNF-apha can be used as a marker for acute inflammatory responses in the degu.
1. T or F degu's are the only diurnal rodent?
2. T or F TNF-alpha and IL-6 are anti inflammatory cytokines?
3. T or F when LPS is injected IP an acute inflammatory response is detectable in BAL fluid?
4. T or F post exposure to LPS in the degu, the authors were able to demonstrate a rise in both TNF-alpha and IL-6.
5. List 3 reasons why IL-6 may not have been detected in the degu?
1. T
2. F
3. F
4. F
5. degu's may not have an IL-6 gene, IL-6 levels were too low to detect, problems with the bioassay used.

Polycystic and chronic kidney disease in a young degu (Octodon degus). Contemporary Topics 42 (2): 43.
The degu is a nontraditional laboratory animal and member of the order Rodentia, suborder Histricomorpha and family Octodontidae. It is used as an animal model for studies on sleep biology, circadian rhythm, jet lag, neurobiological development, drug tolerance, diabetes and cataract development. This report details to findings of polycystic and chronic didney disease in a young (20 month old) female degu who had been a member of a breeding pair. The animal was reported with bloody vaginal discharge, cahexia, molting, slight matting of the hair, a hunched, thin appearance and moderate dehydration. Hematology results included animia and leukopenia with lymphocytopenia. Biochemical abnormalities included severe azotemia and phosphatemia. Urine specific gravity was 1.016. Necropsy results included a small cystic right kidney and pale, slightly enlarged left kidney. Histologically, the left kidney had multiple regions of chronic diffuse interstitial nephritis. A diagnosis of renal failure secondary to chronic nephritis and polycystic kidney disease was made. Chronic renal failure characterized by irreversible loss of renal function may be congenital, familial or acquired. The anemia seen in this animal was most likely secondary to chronic renal failure due to inadequate formation of erythropoietin. Polycystic kidney disease is characterized by replacement of normal renal parenchyma by variably sized cysts (nephrons or collecting ducts). Congenitally acquired polycystic kidney disease has been reported in the Cairn terrier, long haired cats, woodchuck, striped skunk and Mongolian gerbil. In man, it has been demonstrated to be a result of mutation in 3 separate genes and can be either autosomal dominantly or recessively inherited.
1. What is the genus and species of degus?
2. Anemia secondary to renal failure is due to:
a. Lack of erythropoietin production
b. Rennin:angiotensin imbalance
c. Lymphocytic infiltration
d. Destruction of the primary site of erythropoiesis in the degu.
3. The normal life span of the Degu is:
a. 1 year
b. 18 months
c. 3-5 years
d. 7-10 years
1. Octodon degus
2. a.
3. c.

Shelter enrichment for rats. Contemporary Topics 42 (2): 46.
Nest boxes are a simple and effective form of environmental enrichment. The rats particularly like enclosed, opaque, thermoplastic boxes. Environmental enrichment is an increase in the physical complexity or naturalness of an enclosure with the goal of improving animal welfare. Many species have a basic need for shelter and security that is not satisfied with barren commercial caging. For both wild and domestic rats, shelter behaviors such as digging and nest building are important parts of the animal's repertoire of activities. Only social contact may be more important as a source of enrichment. Rats avoid open spaces and prefer cages with platforms and barriers. Studies have shown that rats prefer a cage containing a nest box. Nesting paper may substitute for nest boxes to some extent but rats prefer nest boxes when given the choice of the two. The aim of this study was to evaluate which nest box design was preferred by the rats by use of a T-maze. The rats were run individually in the T-maze. Each rat was allowed to make 10 choices per day for 6 days for a total of 60 choices. Each choice consisted of the rat being placed in front of the start tunnel. The rat then ran down the tunnel and then turned right or left into a cage. The rats chose from entering a cage with or without a nest box. Different types of nest boxes were utilized. Then the rats chose between two cages, both with nest boxes, but the nest boxes being of different configurations or material. The results demonstrated the rats to have a strong preference for cages with nest boxes vs. no nest boxes. Of the nest boxes, the rats preferred the enclosed, opaque, thermoplastic next boxes. The opaque thermoplastic nest boxes were preferred over cardboard or tin. As rat nest boxes are not commercially available at present, laboratories should consider constructing their own, using the simple designs (and pictures) presented in this study.
1. Although this study underscores the importance of providing shelter for caged rats, it also makes the point that social contact may be even more important as a source of enrichment. T or F
2. Define environmental enrichment as given in the article.
1. T
2. Environmental enrichment is an increase in the physical complexity or naturalness of an enclosure with the goal of improving animal welfare.

Morris Water Maze. Contemporary Topics 42 (2): 49.
Purpose To adapt to changing environments animals must demonstrate the ability to learn and subsequently remember what they have learned. As these processes are difficult to observe directly they can be assessed indirectly with the use of a Morris Water Maze (MWM). The MWM is based on the premise that rodents find swimming aversive and will work to avoid this task. Thus, the task is self-rewarded when the animal finds a platform to avoid further swimming. Environmental cues around the tank allow the animal to first learn then remember where the platform is. This type of testing is ideal for rodent models of neurodegenerative diseases like Alzheimer's and Parkinson's disease. The MWM is useful to study the effects of pharmacologic compounds on learning and memory. In addition the MWM is a productive adjunct to functional observation of genetically altered mice.
Method The tank is typically circular. The animal must not be able to see through the water nor have any part of its body be able to touch the bottom of the tank. High contrast geometric cues are made visible to assist the animal in spatial learning. Animals are pretrained and screened for their ability to perform the desired task. The two prominent trials involve either a visible or a hidden platform. After pretraining successful learning is demonstrated with a probe trial. In this trial the platform is removed. When introduced to the MWM the rodent will swim directly to where the platform used to be.
Animal Welfare Concerns,
Animals must be observed closely to ensure they are not overly stressed by the water. The swim time must be carefully monitored to prevent overtiring and hypothermia. The water should be kept fresh to avoid pathogen transmission and water should be warmed to at least room temperature. Rodents, especially mice, should be carefully dried prior to returning to their home cage.
1. T or F The MWM is a complicated system that is difficult to set up?
2. This test was first developed in which species?
3. The MWM is useful for
a) the study of neurodegenerative diseases
b) studying the effects of pharmacologic compounds on learning and memory
c) to assess genetically altered mice
d) all of the above
4. T or F Animals are pretrained to decrease confounding effects of stress on the animal.
5. Animal Welfare issues include
a) sinking when introduced the MWM
b) excessive stress in the maze
c) overtiring and hypothermia
d) infectious pathogens in the water
e) all of the above
1. F
2. rats
3. d
4. T
5. e