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1.
STAR Protoc ; 2(2): 100536, 2021 06 18.
Article in English | MEDLINE | ID: mdl-34027493

ABSTRACT

Gnotobiology has revolutionized the study of microbiota-host interactions. This protocol explains how to generate, maintain, and monitor gnotobiotic mice. Three monitoring methods are presented and compared: bacterial culture, microscopy to visualize the presence (or absence) of bacteria using Gram staining or DNA staining, and 16S rRNA gene amplification and sequencing. The generation and maintenance of gnotobiotic animals should be performed in a germ-free and gnotobiotic facility to guarantee sterility and precision of gnotobiotic conditions. For complete details on the use and execution of this protocol, please refer to McDonald et al., 2020.


Subject(s)
Bacteria , Germ-Free Life , Laboratory Animal Science , Animals , Bacteria/genetics , Bacteria/isolation & purification , Bacteriological Techniques , DNA, Bacterial/genetics , Laboratory Animal Science/methods , Laboratory Animal Science/standards , Mice , RNA, Ribosomal, 16S/genetics , Specific Pathogen-Free Organisms
2.
Exp Anim ; 70(3): 372-377, 2021 Aug 06.
Article in English | MEDLINE | ID: mdl-33840658

ABSTRACT

It is known that administering a gavage to rodents evokes a cardiac reflex, due to gastrointestinal stimulation. Consequently, it is difficult to evaluate changes in hemodynamics after a single oral dose of a pungent or astringent, which alters the circulation by increasing sympathetic activity. In the present study, we developed a method for administering a gavage without significantly affecting hemodynamics measurements. We marked a gastric tube at 10 cm from the tip, to mark the distance from the oral cavity to the stomach body of Wistar male rats. Rats were intubated under urethane anesthesia.After 10-15 min of stabilization, we measured the mean blood pressure (MBP), heart rate (HR), and blood flow (BF) in the cremaster arteriole under two different conditions; condition 1: a pointed gastric tube, room temperature distilled water, and injected at normal speed (approximately 3 ml/min); condition 2: a rounded gastric tube, 37°C distilled water, and injection at 1.0 ml/min. Under condition 1, we observed striking hemodynamic alterations, due to the somatic afferent reflex. In contrast, under condition 2, these hemodynamic changes were nearly eliminated. In addition, we could clearly detect hemodynamic changes in rats after a single gavage treatment of pungent (capsaicin) or astringent (cinnamtannin A2). We observed transient increases in the HR and MBP soon after treatment with capsaicin. Moreover, cremasteric BF was elevated with cinnamtannin A2. These results confirmed the utility of the gavage method developed in this study.


Subject(s)
Hemodynamics , Intubation, Gastrointestinal/methods , Laboratory Animal Science/methods , Rats/blood , Animals , Male , Rats, Wistar
3.
Lab Anim ; 55(1): 76-80, 2021 Feb.
Article in English | MEDLINE | ID: mdl-32883167

ABSTRACT

Drug administration to experimental rodents is often invasive and stressful, thus reducing animal welfare and potentially confounding experimental results. Methods of oral drug delivery in which rodents cooperate voluntarily minimize stress, pain and morbidity. We herein describe a method for oral administration through voluntary intake of strawberry jam, developed for C57BL/6J mice. During a 3-day habituation period, animals were placed in individual cages once daily and presented with a drop of jam. Five days later, the jam was again offered with admixed drug. Mice ingested it in less than 5 min, with latency times below 1 min, confirming the suitability of the administration method.


Subject(s)
Administration, Oral , Laboratory Animal Science/methods , Models, Animal , Animals , Female , Male , Mice
4.
Sci Rep ; 10(1): 20835, 2020 11 30.
Article in English | MEDLINE | ID: mdl-33257713

ABSTRACT

Experiments involving food restriction are common practice in metabolic research. Under fasted conditions, mice supplement their diet with cage bedding. We aimed at identifying metabolic and microbiota-related parameters affected by the bedding type. We exposed mice housed with wooden, cellulose, or corncob cage beddings to ad libitum feeding, caloric restriction (CR), or over-night (ON) fasting. Additionally, two subgroups of the ON fast group were kept without any bedding or on a metal grid preventing coprophagy. Mice under CR supplemented their diet substantially with bedding; however, the amount varied depending on the kind of bedding. Bedding-related changes in body weight loss, fat loss, cecum size, stomach weight, fecal output, blood ghrelin levels as well as a response to glucose oral tolerance test were recorded. As fiber is fermented by the gut bacteria, the type of bedding affects gut bacteria and fecal metabolites composition of CR mice. CR wood and cellulose groups showed distinct cecal metabolite and microbiome profiles when compared to the CR corncob group. While all ad libitum fed animal groups share similar profiles. We show that restriction-related additional intake of bedding-derived fiber modulates multiple physiological parameters. Therefore, the previous rodent studies on CR, report the combined effect of CR and increased fiber consumption.


Subject(s)
Feeding Behavior/physiology , Gastrointestinal Microbiome/drug effects , Laboratory Animal Science/methods , Animals , Bacteria/genetics , Body Weight , Caloric Restriction/methods , Cellulose/adverse effects , Diet, High-Fat/methods , Dietary Fiber/metabolism , Dietary Supplements/adverse effects , Energy Intake/physiology , Feces/microbiology , Gastrointestinal Microbiome/genetics , Gastrointestinal Microbiome/physiology , Housing, Animal , Male , Mice , Mice, Inbred C57BL , Obesity/microbiology , Wood/adverse effects
5.
PLoS One ; 15(8): e0236290, 2020.
Article in English | MEDLINE | ID: mdl-32785214

ABSTRACT

High and variable pre-weaning mortality is a persistent problem in laboratory mouse breeding. Assuming a modest 15% mortality rate across mouse strains, means that approximately 1 million more pups are produced yearly in the EU to compensate for those which die. This paper presents the first large study under practical husbandry conditions to determine the risk factors associated with mouse pre-weaning mortality. We analysed historical records from 219,975 pups from two breeding facilities, collected as part of their management routine and including information on number of pups born and weaned per litter, parents' age and identification, and dates of birth and death of all animals. Pups were counted once in their first week of life and at weaning, and once every one or two weeks, depending on the need for cage cleaning. Dead pups were recorded as soon as these were found during the daily cage screening (without opening the cage). It was hypothesized that litter overlap (i.e. the presence of older siblings in the cage when new pups are born), a recurrent social configuration in trio-housed mice, is associated with increased newborn mortality, along with advanced dam age, large litter size, and a high number and age of older siblings in the cage. The estimated probability of pup death was two to seven percentage points higher in cages with litter overlap compared to those without. Litter overlap was associated with an increase in death of the entire litter of five and six percentage points, which represent an increase of 19% and 103% compared to non-overlapped litters in the two breeding facilities, respectively. Increased number and age of older siblings, advanced dam age, small litter size (less than four pups born) and large litter size (over 11 pups born) were associated with increased probability of pup death.


Subject(s)
Animals, Laboratory/physiology , Breeding/methods , Laboratory Animal Science/methods , Litter Size/physiology , Mice, Inbred C57BL/physiology , Age Factors , Animals , Female , Male , Maternal Age , Mice , Pregnancy , Reproduction/physiology , Weaning
6.
PLoS One ; 15(4): e0228059, 2020.
Article in English | MEDLINE | ID: mdl-32294094

ABSTRACT

Assessing the well-being of an animal is hindered by the limitations of efficient communication between humans and animals. Instead of direct communication, a variety of parameters are employed to evaluate the well-being of an animal. Especially in the field of biomedical research, scientifically sound tools to assess pain, suffering, and distress for experimental animals are highly demanded due to ethical and legal reasons. For mice, the most commonly used laboratory animals, a valuable tool is the Mouse Grimace Scale (MGS), a coding system for facial expressions of pain in mice. We aim to develop a fully automated system for the surveillance of post-surgical and post-anesthetic effects in mice. Our work introduces a semi-automated pipeline as a first step towards this goal. A new data set of images of black-furred laboratory mice that were moving freely is used and provided. Images were obtained after anesthesia (with isoflurane or ketamine/xylazine combination) and surgery (castration). We deploy two pre-trained state of the art deep convolutional neural network (CNN) architectures (ResNet50 and InceptionV3) and compare to a third CNN architecture without pre-training. Depending on the particular treatment, we achieve an accuracy of up to 99% for the recognition of the absence or presence of post-surgical and/or post-anesthetic effects on the facial expression.


Subject(s)
Animal Welfare , Animals, Laboratory/physiology , Deep Learning , Laboratory Animal Science/methods , Pain, Postoperative/diagnosis , Anesthetics/administration & dosage , Animals , Behavior, Animal/physiology , Castration/adverse effects , Datasets as Topic , Facial Expression , Female , Male , Mice/physiology , Pain, Postoperative/etiology
7.
ALTEX ; 37(2): 167-186, 2020.
Article in English | MEDLINE | ID: mdl-32242634

ABSTRACT

Seven years after the last release, the European Commission has again collated and released data on laboratory animal use. The new report is the first to correspond to the requirements of the new Directive 2010/63/EU. Beside minor problems in reporting, the new reporting format is a major step forward, with additional new categories like severity allowing insight into animal use related questions that goes far beyond the previous reports. An in-depth analysis confirms a slight decrease in animal use from 2015 to 2017, but also compared to the 2005, 2008 and 2011 reports, though the new reporting scheme makes this comparison difficult. Notable success is evident for replacing rabbit pyrogen testing but, in general, the implementation of accepted alternative methods lags behind expec-tations. Beside the roughly 10 million animals per year covered in the report, about 8 million animals were identified that fall under the Directive but are not included in this number. Their omission downplays the impact of REACH on animal use. The report, second to none in its detail internationally, represents an important instrument for benchmarking and strategi-cally focusing activities in the 3Rs.


Subject(s)
Animal Testing Alternatives/statistics & numerical data , European Union , Laboratory Animal Science/methods , Laboratory Animal Science/statistics & numerical data , Animals , Benchmarking , Data Interpretation, Statistical
9.
Adv Physiol Educ ; 44(2): 163-168, 2020 Jun 01.
Article in English | MEDLINE | ID: mdl-32167832

ABSTRACT

Blood flow through the cardiovascular system is governed by the same physical rules that govern the flow of water through domestic plumbing. Using this analogy in a teaching laboratory, a model of the cardiovascular system constructed of pumps and pipes was used to demonstrate the basic interactions of pressure, flow, and resistance in a regulated system, with student volunteers providing the operational actions and regulatory components. The model was used to validate predictions and explore solutions prompted by student discussion. This interactive teaching laboratory provides an engaging experiential exercise that demonstrates regulation of flow and pressure in an intact cardiovascular system with apposite changes in heart rate and resistance. In addition, the system provides strong clinical correlates and illustrates how that regulated system responds to challenges such as heart failure, inappropriate vasodilation, and hemorrhage. The results demonstrate that, with limited practice, the instructor can effectively guide the students to reliably reproduce physiologically appropriate results.


Subject(s)
Cardiovascular Physiological Phenomena , Laboratory Animal Science/education , Laboratory Animal Science/methods , Physiology/education , Problem-Based Learning/methods , Schools, Veterinary , Cardiovascular System , Exercise/physiology , Heart Rate/physiology , Hemodynamics/physiology , Humans
10.
Elife ; 82019 12 11.
Article in English | MEDLINE | ID: mdl-31825307

ABSTRACT

Open Science has changed research by making data accessible and shareable, contributing to replicability to accelerate and disseminate knowledge. However, for rodent cognitive studies the availability of tools to share and disseminate data is scarce. Automated touchscreen-based tests enable systematic cognitive assessment with easily standardised outputs that can facilitate data dissemination. Here we present an integration of touchscreen cognitive testing with an open-access database public repository (mousebytes.ca), as well as a Web platform for knowledge dissemination (https://touchscreencognition.org). We complement these resources with the largest dataset of age-dependent high-level cognitive assessment of mouse models of Alzheimer's disease, expanding knowledge of affected cognitive domains from male and female mice of three strains. We envision that these new platforms will enhance sharing of protocols, data availability and transparency, allowing meta-analysis and reuse of mouse cognitive data to increase the replicability/reproducibility of datasets.


Subject(s)
Cognition/physiology , Laboratory Animal Science/instrumentation , Laboratory Animal Science/methods , Rodentia , Alzheimer Disease , Animals , Behavior, Animal , Choice Behavior , Databases, Factual , Disease Models, Animal , Female , Learning/physiology , Male , Memory/physiology , Mice , Neuropsychological Tests , Reproducibility of Results , Rodentia/genetics , Software
11.
ILAR J ; 60(1): 9-16, 2019 12 31.
Article in English | MEDLINE | ID: mdl-31711238

ABSTRACT

Biomedical research has made great strides in the past century leading to rapid advances in human life expectancy, all derived from improved understanding, prevention, and treatment of many diseases and conditions. Research involving laboratory animals has played a significant role in this medical progress. However, there continues to be controversy surrounding the use of animals in research, and animal models have been questioned regarding their relevance to human conditions. While research fraud and questionable research practices could potentially contribute to this problem, we argue that a relative ignorance of laboratory animal science has contributed to the "uncontrolled vivarium experiment" that runs parallel to the more controlled scientific experiment. Several variables are discussed, including husbandry, animal environment, social housing, and more, that can contribute to this uncontrolled experiment, and that can simultaneously decrease quality of life for rodent test subjects when ignored. An argument is put forward that laboratory animal veterinarians and scientists can and should play an important role in better controlling such variables. Similarly, the laboratory animal veterinarian and scientist should play an important role in responsible science by addressing complex interdisciplinary challenges.


Subject(s)
Biomedical Research/methods , Laboratory Animal Science/methods , Animal Husbandry/methods , Animals , Gastrointestinal Microbiome , Mice , Rats
16.
Sci Rep ; 9(1): 11403, 2019 08 06.
Article in English | MEDLINE | ID: mdl-31388041

ABSTRACT

The black soldier fly, yellow mealworm and house fly are known for their wide distribution, ease of breeding, and environmental and nutritional attributes. Diets based on these fly proteins for the rearing of mosquito larvae are more accessible and affordable when compared to the reference IAEA diet which consists largely of costly livestock products such as bovine liver powder. Following a step-by-step assessment, we developed diet mixtures based on insect meal for the optimal mass production of Aedes albopictus and Ae. aegypti. Based on the assessed parameters including mosquito egg hatch, body size, flight ability, longevity and diet cost reduction, two mixtures are recommended: 1/2 tuna meal (TM) + 7/20 black soldier fly (BSF) + 3/20 brewer's yeast and 1/2 TM + 1/2 BSF. These findings, which could be adapted to other mosquito species, provide alternative protein sources for mass rearing insects for genetic control strategies.


Subject(s)
Aedes/physiology , Animal Feed , Breeding/methods , Laboratory Animal Science/methods , Simuliidae , Animals , Female , Larva/physiology , Male , Mosquito Control/methods , Pest Control, Biological/methods
18.
Lab Anim (NY) ; 48(7): 207-216, 2019 Jul.
Article in English | MEDLINE | ID: mdl-31217565

ABSTRACT

Deep phenotyping is an emerging conceptual paradigm and experimental approach aimed at measuring and linking many aspects of a phenotype to understand its underlying biology. To date, deep phenotyping has been applied mostly in cultured cells and used less in multicellular organisms. However, in the past decade, it has increasingly been recognized that deep phenotyping could lead to a better understanding of how genetics, environment and stochasticity affect the development, physiology and behavior of an organism. The nematode Caenorhabditis elegans is an invaluable model system for studying how genes affect a phenotypic trait, and new technologies have taken advantage of the worm's physical attributes to increase the throughput and informational content of experiments. Coupling of these technical advancements with computational and analytical tools has enabled a boom in deep-phenotyping studies of C. elegans. In this Review, we highlight how these new technologies and tools are digging into the biological origins of complex, multidimensional phenotypes.


Subject(s)
Caenorhabditis elegans/genetics , Genetic Techniques , Laboratory Animal Science/methods , Phenotype , Animals , Genetic Techniques/instrumentation , Laboratory Animal Science/instrumentation
19.
Lab Anim ; 53(3): 232-243, 2019 Jun.
Article in English | MEDLINE | ID: mdl-31096878

ABSTRACT

Gnotobiotics or gnotobiology is a research field exploring organisms with a known microbiological state. In animal research, the development of gnotobiotics started in the late 19th century with the rederivation of germ-free guinea pigs. Cutting-edge achievements were accomplished by scientists in the Laboratories of Bacteriology at the University of Notre Dame (LOBUND). The primary goals of gnotobiotics were not only the development of the equipment required for long-term husbandry but also phenotypic characterization of germ-free animals. The first isolators were designed by Reynolds and Gustafsson as rigid-wall stainless steel autoclave-like chambers, which were subsequently replaced by Trexler's flexible-film polyvinyl plastic isolators. Flexible-film or semi-rigid isolators are commonly used today. The long-term maintenance of gnotobiotic rodents is performed in positive-pressure isolators. However, to facilitate gnotobiotic experimental procedures, short-term husbandry systems have been developed. Gnotobiotic animal husbandry is laborious and requires experienced staff. Germ-free animals can be rederived from existing rodent colonies by hysterectomy or embryo transfer. The physiology and anatomy of germ-free rodents are different from those of specified pathogen-free (SPF) rodents. Furthermore, to guarantee gnotobiotic status, the colonies need to be regularly microbiologically monitored. Today, gnotobiotics provides a powerful tool to analyse functional effects of host-microbe interactions, especially in complex disease models. Gnotobiotic models combined with 'omics' approaches will be indispensable for future advances in microbiome research. Furthermore, these approaches will contribute to the development of novel therapeutic targets. In addition, regional or national gnotobiotic core facilities should be established in the future to support further applications of gnotobiotic models.


Subject(s)
Animal Husbandry/methods , Germ-Free Life , Laboratory Animal Science/methods , Microbiota , Models, Animal , Animals
20.
Lab Anim ; 53(5): 508-510, 2019 Oct.
Article in English | MEDLINE | ID: mdl-31042100

ABSTRACT

There are different husbandry situations that lead to social isolation of male mice. Besides legal considerations, single housing has a negative impact on behavioural and molecular studies. We have implemented two strategies, the 'companion mouse' and just-weaned male grouping, to reduce the number of male mice housed singly in our facility. We have achieved a progressive reduction (42% in three years) without an increase in aggression.


Subject(s)
Animal Husbandry/methods , Housing, Animal , Laboratory Animal Science/methods , Mice , Social Isolation , Animals , Male
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