Alexandra L. Whittaker

Effects of space allocation and housing density on measures of wellbeing in laboratory mice: a review:

In the majority of countries where there are legislative requirements pertaining to the use of animals in research, figures are quoted for minimum cage sizes or space allocation to be provided per animal. These figures are generally based on professional judgement and are in common usage. However, there is a growing trend and expectation that welfare science should inform regulatory decision-making. Given the importance of the potential welfare influences of cage size on the animals themselves, this paper presents the latest scientific knowledge on this topic in one of the most commonly used animals in research, the mouse. A comprehensive review of studies in laboratory mice was undertaken, examining the effects of space allocation per animal and animal density on established welfare indicators. To date, animal density studies have predominated, and the effects of space allocation per se are still relatively unclear. This information will guide those involved in facility management or legislative review, and provide a more solid foundation for further studies into the effects of routine husbandry practices on animal welfare.

Evaluation of Facebook© to create an online learning community in an undergraduate animal science class

There has been widespread comment on the use and impact of Web 2.0 technologies in education. Given the use of such technologies, particularly social networking sites such as Facebook amongst the student body, it would be remiss of educators to not consider their use as part of a pedagogical strategy. This paper provides a preliminary investigation into the success of Facebook as a tool for formation of an online learning community amongst a group of undergraduate science students. A quantitative examination of participant numbers and traffic on the site was undertaken. In order to determine functionality of the group, a semi-qualitative examination of category of response posting was also performed. It was concluded that Facebook is a promising tool to establish an online educational community under the conditions imposed in our study. The main benefits are in establishment of affective communication, social support, and a problem-solving strategy.

Effects of Streptococcus thermophilus TH-4 in a rat model of doxorubicin-induced mucositis

Abstract Background. Mucositis is a debilitating intestinal side effect of chemotherapeutic regimens. Probiotics have been considered a possible preventative treatment for mucositis. Streptococcus thermophilus TH-4 (TH-4), a newly identified probiotic, has been shown to partially alleviate mucositis induced by administration of the antimetabolite chemotherapy drug, methotrexate in rats; likely mediated through a mechanism of folate production. However, its effects against other classes of chemotherapy drug have yet to be determined. Aims. The authors investigated the effects of TH-4 in a rat model of mucositis induced by the anthracycline chemotherapy drug, doxorubicin. Methods. Gastrointestinal damage was induced in female Dark Agouti rats (148.3 ± 1.5 g) by intraperitoneal injection of doxorubicin (20 mg/kg). Animals recieved a daily oral gavage of TH-4 at 109 cfu/ml or skim milk (vehicle) from days 0 to 8. At day 6, rats were injected with either saline or doxorubicin. At kill, small intestinal tissues were collected for determination of sucrase and myeloperoxidase (MPO) activities and histological assessment. Results. Body weight was significantly decreased by doxorubicin compared with normal controls (p < 0.05). Histological parameters, such as crypt depth and villus height, were also significantly decreased by doxorubicin. TH-4 partially prevented the loss of body weight induced by doxorubicin (2.3% compared with 4%), but provided no further therapeutic benefit. Conclusions. The minimal amelioration of doxorubicin-induced mucositis by TH-4 further supports folate production as a likely mechanism of TH-4 action against methotrexate-induced mucositis. Further studies into TH-4 are required to confirm its applicability to other conventional chemotherapy regimens.

The assessment of general well-being using spontaneous burrowing behaviour in a short-term model of chemotherapy-induced mucositis in the rat

Mucositis is a common and serious side-effect experienced by cancer patients during treatment with chemotherapeutic agents. Consequently, programmes of research focus on the elucidation of novel therapeutics for alleviation of mucositis symptoms, and these frequently use animal models. However, although these models are assumed to be painful and distressing to the animal, endpoints are difficult to determine. The aim of this study was to evaluate whether a change in burrowing behaviour could provide an indication of disease onset and potentially be applied as a humane endpoint. Baseline burrowing behaviour was measured in healthy animals on three occasions by determining the weight of gravel displaced from a hollow tube. Mucositis was then induced in the same animals by intraperitoneal injection of 5-fluorouracil (150 mg/kg) and burrowing behaviour recorded over three consecutive days. Standard measures of disease progression, including body weight loss and clinical score, were also made. The presence of mucositis was confirmed at necropsy by findings of decreased duodenal and colon lengths, and reduced liver, spleen and thymus weights in comparison with non-treated control animals. Histological score of the jejunum and ileum was also significantly increased. Mucositis onset coincided with a decrease in mean burrowing behaviour which was progressive, however this result did not achieve statistical significance (P = 0.66).We conclude that burrowing may be a useful indicator of inflammation in the mucositis model, although this requires further characterization. Pre-selection of animals into treatment groups based on their prior burrowing performance should be pursued in further studies.

Effects of Metabolic Cage Housing on Rat Behavior and Performance in the Social Interaction Test

Although the metabolic cage is commonly used for housing nonhuman animals in the laboratory, it has been recognized as constituting a unique stressor. Such an environment would be expected to affect behavioral change in animals housed therein. However, few studies have specifically addressed the nature or magnitude of this change. The current study sought to characterize the behavioral time budget of rats in metabolic cage housing in comparison to that of individually housed animals in standard open-top cages. Rats in metabolic cages spent less time moving, manipulating enrichment, and carrying out rearing behaviors, and there was a corresponding shift toward inactivity. In an applied Social Interaction Test, behavioral scoring implied that metabolic cage housing had an anxiogenic effect. In conclusion, metabolic cage housing produces measurable effects on spontaneous and evoked behavior in rats in the laboratory. These behavioral changes may lead to a negative emotional state in these animals, which could have negative welfare consequences. Further research is needed to quantify the existence and magnitude of such an effect on rat well being.

Effects of acute chemotherapy-induced mucositis on spontaneous behaviour and the grimace scale in laboratory rats

Intestinal mucositis is a frequent side-effect of chemotherapy treatment. Many oncological research programs aim to identify novel treatments for this distressing condition, and these programs frequently use rat models. Little is known about the presence and progression of pain in these models and how this can best be treated by analgesic therapy. We used a number of behaviour-based methods of pain assessment to determine which tools were best suited for pain identification. Baseline measures for behavioural assessment, rat grimace score and sociability were determined through analysis of continuously recorded video data and an applied social interaction test (n = 16). Mucositis was then induced by intraperitoneal injection of 5-fluorouracil (150 mg/kg) and further behavioural analyses undertaken. An assessment of enrichment interaction was also made by determining the mass of a plastic chew toy gnawed both pre- and post-chemotherapy injection. Behavioural scoring was performed 1, 6, 12, 24 and 48 h after injection, with facial expression being scored at the 12, 24 and 48 h time-points. Sociability testing was performed once during the post-injection period. No significant differences were found in grimace scores between baseline and later daily measures. Behaviours similar to those previously reported post-laparotomy were observed. Writhing, twitching and back-arching behaviours were most evident in rats affected by mucositis and were increased in frequency (respective P values: 0.002, 0.004 and 0.008) 48 h after chemotherapy injection compared with baseline, implying that pain onset occurred around this time-point. Social investigatory behaviour was also increased (P = 0.002) following disease onset. Each day, rats post-5FU injection gnawed a greater percentage of their nylabone enrichment by weight than the saline-injected control rats (P = 0.046). These data suggest that, of the tools tested, behavioural assessment scoring may find greatest utility in rodent models of intestinal mucositis and should be investigated further.

Assessment of housing density, space allocation and social hierarchy of laboratory rats on behavioural measures of welfare

Minimum space allowances for laboratory rats are legislated based on weight and stocking rates, with the understanding that increased housing density encourages crowding stress. However, there is little evidence for these recommendations, especially when considering positive welfare outcomes. This study consisted of two experiments which investigated the effects of housing density (rats per cage), space allocation (surface area per rat) and social rank (dominance hierarchy) on the ability to perform simple behavioural tests. Male Sprague Dawley (SD) rats (n = 64) were allocated to either high-density (n = 8) or low-density (n = 8) cages. The second experiment investigated the effects of surface area. SD rats (n = 40) were housed in dyads in either the large (n = 10) or small (n = 10) cage. In both experiments, animals were tested on a judgment bias paradigm, with their responses to an ambiguous stimulus being ascribed as optimistic or pessimistic. Animals were also tested on open-field, novel-object recognition and social-interaction tests. Recordings were taken from 1700-2100h daily for rat observation and social rank establishment. Dominant animals responded with significantly more optimistic decisions compared to subordinates for both the housing density (p<0.001) and space allocation (p = 0.0015) experiment. Dominant animals responded with increased social affiliative behaviours in the social-interaction test, and spent more time in the centre of the open-field test for both experiments. No significance was detected between housing density or space allocation treatments. These findings suggest that social rank is a significantly greater modifier of affective state than either housing density or space allocation. This finding has not yet been reported and suggests that future drafts of housing guidelines should consider animal social status in addition to floor space requirements.

A Policy at the University of Adelaide for Student Objections to the Use of Animals in Teaching

In veterinary medical education, the use of animals or cadaveric tissue as a component of teaching practice is common. Teachers are required, during the process of ethical review, to apply the 3 Rs principle (replacement, refinement, reduction) whenever they consider using animals during a teaching exercise. This often involves use of replacement strategies, such as utilization of video footage or simulation-based training. However, aside from legislative or ethical requirements imposed by a countrys regulatory framework on the institution, students are often the key advocates for using alternative teaching practices that do not make use of animals. This has prompted many institutions with veterinary and other life sciences teaching programs to develop student-conscientious objection policies to the use of animals in teaching. In this article, we discuss the procedures implemented to make provision for student-conscientious objectors at a new Australian Veterinary School, at the University of Adelaide. We also describe the processes to provide information to students and faculty on this issue and to facilitate information gathering on alternatives.

Female rats display fewer optimistic responses in a judgment bias test in the absence of a physiological stress response

Metabolic cages are a type of housing used in biomedical research. Metabolic cage housing has been demonstrated to elicit behavioural and physiological changes in rodents housed within them. The nature of this effect has been characterized as anxiogenic. However, few studies have evaluated positive affect in response to metabolic cage housing and the interaction between this, sex and traditional physiological measures of stress. Cognitive biasing, as measured through a judgment bias paradigm has proven a reliable measure of animal affective state, particularly through its ability to measure positive affect. The current study investigated differences in cognitive biasing between male and female rats when transferred from open-top, grouped housing to a metabolic cage. Rats (Rattus norvegicus) (n=60) were trained in a judgment bias paradigm previously validated for use in the rat model. Upon exposure to an intermediate, ambiguous probe rats responded with either an optimistic or pessimistic decision. The animals were also subjected to the sucrose preference test to identify the presence of anhedonia. Faecal corticosterone and changes in adrenal tyrosine hydroxylase were also measured to establish whether a stress-like state was experienced. There was a significant interaction between sex and metabolic cage housing on the number of optimistic decisions made F (1, 56)=7.461, p=0.008. Female rats that remained in control housing responded with a reduced number of days featuring an optimistic decision compared to males in control housing (p=0.036). However, both males and females responded with significantly fewer optimistic decisions in the metabolic cage compared to control (p<0.001). There was a significant negative correlation between treatment and sucrose consumption (rpb=-0.654, n=195, p<0.001). There was also a significant sex effect for faecal corticosterone concentrations F (1, 30)=6.305, p=0.018) with female rats (4.050±1.285), displaying greater corticosterone concentrations than males (2.291±0.495). No differences between treatment were observed for either corticosterone or tyrosine hydroxylase levels. This data demonstrates that movement into a metabolic cage resulted in rats displaying significantly greater pessimistic cognitive biases as determined through the judgment bias test. Interestingly, male rats that remained in control housing demonstrated cognitive biases that were not equivalent to female rats. Furthermore, despite a behavioural change being evident, a physiological change in corticosterone or tyrosine hydroxylase levels was not observed.

Oral Nucleotides only Minimally Improve 5-Fluorouracil-Induced Mucositis in Rats

Chemotherapy-induced mucositis is characterized by inflammation and ulceration of the intestinal mucosa, compromising intestinal function. Exogenous nucleotides have been reported to repair the mucosa. The nucleotide preparation, Nucleoforce F0328 (Nucleoforce), was investigated for its potential to ameliorate intestinal mucositis in rats. Female Dark Agouti rats (n = 8/group) were gavaged once daily with Nucleoforce (175 mg/kg) or water from Days 0 to 8 and injected (i.p.) with 5-fluorouracil (5-FU; 150 mg/kg) or saline on Day 5. Histological parameters (disease severity, crypt depth, and villus height measurements) and myeloperoxidase activity were quantified. P < 0.05 was considered significant. Jejunal and ileal histological disease severity scores were significantly increased by 5-FU, compared to normal controls (P < 0.05). Nucleoforce treatment in 5-FU-injected rats significantly reduced jejunal and ileal disease severity compared to 5-FU controls (P < 0.05). In 5-FU-injected rats, jejunal and ileal villus heights and crypt depths were significantly decreased compared to 5-FU controls, with no additional Nucleoforce effect (P > 0.05). Intestinal myeloperoxidase activity was significantly elevated by 5-FU (8.8-fold), compared to normal controls (P < 0.05), which was not normalized by Nucleoforce treatment (P > 0.05). Nucleoforce only partially improved parameters associated with experimentally-induced mucositis. Future studies could investigate increased concentrations, more frequent administration, or protective microencapsulation delivery methods, to increase bioavailability.