Category Archives: 3Rs

Research Roundup: Malaria vaccine, mouse sperm in space, animal welfare prizes, and more!

Welcome to this week’s Research Roundup. These Friday posts aim to inform our readers about the many stories that relate to animal research each week. Do you have an animal research story we should include in next week’s Research Roundup? You can send it to us via our Facebook page or through the contact form on the website.

  • New study finds that mouse sperm stored in space still functions on Earth. Increasingly in the news we read about the upcoming reality of commercial space travel (for example, here and here). Of course, with such advances there is caution with respect to feasibility — and of course imagination with respect to possibilities (e.g., colonizing Mars). With such goals on the horizons, these researchers investigated whether sperm that had been freeze dried, and transported to the International Space Station (ISS) and then back to Earth would be able to produce viable offspring. To accomplish this they used freeze dried mouse spermatozoa — which provided a unique advantage, as the addition of water — maintains the viability of the sperm to fertilize an egg and allows for the sperm to be stored at room temperature. Other sperm when freeze dried do not survive. Microinjection  of these “space” sperm into an egg on Earth — produced healthy viable  “space offspring”. Moreover, these offspring all grew to healthy adults and were able to produce offspring of their own. This study was published in the Proceedings of the National Academy of Sciences of the USA.

Space mouse and pups. Source: PNAS

Laboratory frogs. Source: University of Portsmouth

  • Modified experimental vaccine protects monkeys from deadly malaria. Researchers at the National Institute of Allergy and Infectious Diseases (NIAID), part of the National Institutes of Health, discovered that a modified version of an experimental malaria vaccine completely protected 4 of 8 monkeys from a malaria parasite, and delayed the first appearance of the parasites in 3 more monkeys. Scientists modified an existing malaria vaccine by including a particular protein, RON2L, so that it closely mimicked the protein complex used by the parasite to infect blood cells. Vaccination with the modified vaccine resulted in more neutralizing antibody, indicating a better quality response to parasitic infection. Additionally, the modified vaccine seemed to protect against other parasite strains that differed from those used to create the vaccine, suggesting that this new modified vaccine may protect against multiple parasite strains. This research will pave the way toward eventual human trials. The study was published in NPJ Vaccines.

A female Aedes mosquito. Source: NIAID.

 

 

 

 

 

 

 

 

 

 

 

  • Researchers at the University of Helsinki has found the lymphatic vessels extend into the brain – overturning 300 years of accepted wisdom. By genetically altering mice using the luminescent GFP gene, so that lymphatic vessels glowed under light, Aleksanteri Aspelund found that there were lymphatic vessels in the brain. The research was repeated by Karl Alitalo with the same results.  Other researchers have found evidence linking problems with the lymphatic and glymphatic systems to Alzheimer’s; one study in mice showed it could lead to the buildup of amyloid beta in the brain – a key sign of the Alzheimer’s. The study was published in the Journal of Experimental Medicine.

    Red fluorescence of the membrane protein aquaporin-4 in an individual with Alzheimer’s (left) and a healthy individual (right). Source: OHSU

  • Mice help researchers identify genes responsible for a severe congenital heart defect.  Congenital heart disease affect up to 1 percent of all live births. Hypoplastic Left Heart Syndrome (HLHS) is a rare congenital heart disease resulting in an inability to effectively pump blood  throughout the body.  Current treatment involves multiple complex surgeries during the first few years of a child’s life. For some, the surgical interventions improve heart function.  For others, it does not,  leading to heart failure and the need for heart transplants. It has been known that genetic risk factors play a role in HLHS but specific genes have been hard to identify.  Researchers at the University of Pittsburgh Schools of the Health Sciences used fetal ultrasound imaging to look for structural heart defects in genetically modified mice to identify HLHS.  Then by comparing the genomes of affected and non affected mice, and confirming using CRISPR technology they found that mutations in two specific genes that interact were required for HLHS.,   Dr. Cecilia Lo, a professor and the F. Sargent Cheever Chair in Developmental Biology at Pitt says, “Studying diseases with complex genetics is extremely challenging…By understanding the genetics and biology of HLHS, this can facilitate development of new therapies to improve the prognosis for these patients.” This study was published in the journal Nature Genetics.
  • The University of Bristol has awarded prizes in its first Animal welfare and 3Rs Symposium. The 3Rs, developed by Russel and Burch in 1954, have advanced the humane treatment of animals used in research by advocating for replacement (aiming to replace animals where possible, with alternatives), to reduce the number of animals used to the minimum required to answer and experimental question and and to refine their experiments to minimise any adverse effects experienced by the animals.These awards went to three research projects that have advanced the 3Rs in their various lines of research.

“The research project that won first prize has developed a refined method for producing aortic aneurysms in mice.  An aortic aneurysm is a bulge in a section of the aorta, which is the body’s main artery, and if the bulge ruptures it can cause sudden death. The research team has also developed a new human aortic aneurysm model in the laboratory, potentially replacing the need for animal models, using arteries taken from the discarded umbilical cord of newly born babies.

The second prize was awarded to a research team who has developed a method for giving oral drugs using solutions that mice and rats both like and which avoids the need for restraint and reduces stress in the animals. The research team found that liquid foods such as condensed milk, milkshake and fruit puree baby food are good solutions to use for giving a wide range of drugs.
The final prize was awarded to a research team who has developed photographic techniques that can be used in conscious animals.  This new technique has revolutionised preclinical eye research and has markedly reduced the number of animals needed for research studies.”

The 3Rs. Source: Bayer

How animal enclosures are designed to meet the needs of laboratory animals

Having worked in animal research for over 14 years now I have not only gained a comprehensive knowledge of the requirements for animals used in research but have also seen significant improvements in this field. Currently, I work at King’s College London as a Site Manager where I oversee three animal units.

The role of an animal technologist varies dependent on experience but all are there to provide the best possible life to animals in research. Trainee animal technologists will often perform general husbandry duties such as cleaning cages, feeding, and watering, whereas senior technologists may be involved in colony management, scientific procedures etc.

During my career, and the many tours of research labs I’ve given, one of the common discussion is the type of cages used and how they vary so much between species.

Requirements for housing research animals in the UK are stipulated by Home Office and Animals (Scientific Procedures) Act, 1986 as well as any additional institutional requirements beyond this law. Providing the correct type of environment is essential for species to exhibit their natural behaviour.

Example of UK minimum cage sizing for M. mulatta

Housing requirements vary between species but here are some examples of why cages are designed in such a way:

Primates

Primate caging is typically tall as this enables the animals to feel more secure, as in the wild they would use the trees to climb high and get away from prey. Providing higher cages also allows for a more complex environment. Bars are often horizontal to allow the animal to climb the cage and maximise this as much as possible. Cages are normally made out of steel to ensure the animals are safely contained and also withstand potential damage in what are often a strong and intelligent species.

Cages are often multi-tiered to allow primates better utilisation of cage height and enable primates to get away from each other when necessary.  Environmental enrichment such as mirrors and perches provide further security to primates.

Primates are socially housed in multi-tier caging. The perches allow the primates to watch what is going on around the room.

Rodents

Rodents have much smaller cages which are normally made up of a plastic, such as polysulfone. These plastics can withstand high temperatures during cleaning and have been shown to last a long time. Traditionally, animals were kept in open top caging but in recent years there has been a movement towards individually ventilated cages (IVCs). IVCs provide a more stable environment by having sealed caging and using air handling units for filtration; this has, in turn, provided a better environment for animal welfare and research. Controlling for the environment can both help control experimental variables, and prevent risks to animal health from external pathogens.

While the caging appears to be relatively small for rodents it is designed around the need of the animals. Rodents are often social species and in some cases larger spaces can cause anxiety due predator/prey relationships.

Environmental enrichment is used to encourage natural nesting behaviours which can be seen in the wild. In recent years red boxes have been implemented in some cages, humans can see through these but animals don’t see through this colour in the same way, therefore this allows better monitoring while making animals feel safe and secure.

Individually Ventilated Cages

Rabbits

Rabbits are often housed in floor pens as this provides space to exercise and express their social behaviour. Rabbits which are kept grouped housed tend to show less stereotypic behaviour and greater activity. Previously, rabbits were predominantly housed in single cages which caused more stress to the animals.  Enclosures are normally made up of wood frame with metal bars or completely metal frame with very small holes to prevent animals escaping.

Environmental enrichment such as cardboard boxes, hay/straw and raised areas can also provide more security and natural behaviours therefore reducing any abnormal behaviour which may be seen otherwise.  As albino rabbits are often used in research, boxes also provide a darker place to prevent damage to the retina of the eye.

Final thoughts

As humans we often believe that larger housing is better, just look at people who often want a huge home, but this doesn’t mean that an animal will be comfortable with this. The key is to tailor this to each species/individual’s needs for the highest welfare standards. Animals which naturally live in holes, or nests, often feel comfortable with less space compared with other animals. Other additions to accommodation such as environmental enrichment can enable expression of natural behaviour further and have significantly increased in recent years, no more barren cages!

In my 14+ years working with research animals, I have seen a huge amount of change. Improvements in caging and enrichment benefit not only the animals, but the pursuit of good science as well, and we should welcome it. I am also a strong believer that this has also improved the morale of staff, after all we all want the best for animal welfare which in turn will lead to good science.

Stephen Woodley

Tail or Tunnel: Handling Methods Influence Mouse Behavior in Cognitive Tasks

  • A study funded by the NC3Rs explored how handling methods influenced mice’s behavior during cognitive tasks
  • Mice were either picked up by the tail or guided into a tunnel, then transferred to the testing arena
  • Mice that were transferred in the tunnel were far more exploratory during the cognitive task
  • Acclimation to handling procedures is important

A new study published today in Scientific Reports shows that the way experimenters pick up mice can affect their behavior during cognitive tasks. The study was funded by the NC3Rs, which is dedicated to replacing, refining, and reducing the use of animals in research and testing. This particular study focused on refinement: identifying optimal handling methods for mice has important implications for both the welfare of the animals and the validity and usability of data that are collected, which could potentially lead to a reduction in the need for animals in future studies.

Image Credit: Jane Hurst, University of Liverpool.

Drs. Kelly Gouveia and Jane Hurst first placed laboratory mice near a new, attractive stimulus – urine from a novel mouse of the opposite sex – that is known to stimulate approach and investigation. The mice were allowed three sessions to grow accustomed to the new scent. Throughout all three sessions, mice were either picked up by the tail (standard laboratory practice, though there is no obvious scientific reasoning for this method) or were guided into a clear tunnel that is both affordable and easily sterilized. (The method is also easy to learn.) Mice were then carried to the test arena either by the tail or in the tunnel and allowed to explore. Gouveia and Hurst report that mice picked up by the tail showed very little willingness to explore the test arena, and therefore investigate the new stimulus, whereas those transported in the tunnel showed much higher exploration and a strong interest in the new scent.

Importantly, Gouveia and Hurst then tested the mice’s ability to discriminate between the (formerly) new scent and a second, different urine stimulus. They report that since the mice picked up by the tail performed so poorly from the start, they did not discriminate between the two scents. However, those transported in the tunnel showed robust and reliable discrimination. These findings are noteworthy not only with respect to the psychological welfare of the animals, but also for the important effects that handling and habituation have on yielding usable, reliable data. With the potential to reduce the stress associated with handling, the tunnel method could reduce the anxiety that mice display upon tail handling – thereby resulting in more species-typical behaviors, such as exploration of a novel, conspecific scent. It could also reduce the uncontrolled variation that exists in animal studies and could ultimately produce more reliable data. Thus, identifying optimal handling techniques has the potential to reduce the number of animals needed in laboratory studies in addition to refining the techniques used to study them and enhance their welfare.

Image Credit: Jane Hurst, University of Liverpool.

It is worth noting that a study by Novak and colleagues (2015) found no difference in cognitive performance between mice that were handled by the tail or by a less invasive method (“cupping” in the experimenter’s hands). Why might no difference have been found in this study? One possibility is that the cognitive task the researchers used was different from the present study (a radial arm maze vs. novel scent), and the arm maze may probe for different behaviors than a novel odor task. Another possibility is that perhaps mice “prefer” the tunnel to both tail handling and cupping, but neither the 2015 nor the present study compared all three methods. Hurst and her colleague Rebecca West did compare all three methods in a 2010 study, however, and found that mice preferred both the tunnel and cupping method to tail handling (as assessed by voluntary interaction time with the experimenter); although the cupping method produced more variable results depending on strain and sex. However, in the Novak study, mice were handled daily for many weeks, whereas in the Hurst & West study they were handled only for nine days. Of course, the most parsimonious explanation is that in every handling study, experimenter interaction is confounded with handling. That is, are the mice acclimating to the experimenters, to the handling procedures, or both?

These questions underscore the need for replication before firm conclusions about optimal handling techniques can be drawn. Nevertheless, the findings published today in Scientific Reports are an important addition to the field of animal welfare, and they emphasize the importance of constant, rigorous studies surrounding welfare issues.

Amanda Dettmer

References:

Gouveia K, Hurst JL (2017) Optimising reliability of mouse performance in behavioural testing: the major role of non-aversive handling. Scientific Reports 7: 44999. doi: 10.1038/srep44999

Hurst JL, West RS (2010) Taming anxiety in laboratory mice. Nature Methods. Oct;7(10): 825-6

Novak J, Bailoo JD, Melotti L, Rommen J, Würbel H (2015) An Exploration Based Cognitive Bias Test for Mice: Effects of Handling Method and Stereotypic Behaviour. PLoS ONE 10(7): e0130718. doi:10.1371/journal.pone.0130718

University of Stirling improving animal welfare for dogs

A study, conducted by the University of Stirling, in collaboration with AstraZeneca and Charles River Laboratories, aimed to look at the impact of modern, purpose-built dog facilities, on the animals’ welfare. Dr Laura Scullion Hall and Professor Hannah Buchanan-Smith, from the Behaviour and Evolution Group (BERG) at the University of Stirling, published a paper (1) that aimed to validate the welfare benefits of the modern home design pens for dogs. The research was funded by the Biotechnology and Biological Sciences Research Council in the UK, and the National Centre for the Replacement, Refinement & Reduction of Animals in Research (NC3Rs).

There is a clear body of evidence showing the positive impacts of housing refinement on numerous species (2)(3)(4), however, according to Hall, the design of the home pens for dogs “has received little scientific attention since the 1990s, since when legislative minimum standards have improved”. Dogs spend most of their time in home pens, usually interspersed with occasional use of playrooms. The study compared animal welfare using the modern and traditional home pens.

different-pens-for-dogs-in-research

From left to right: modern home pen; traditional home pen; indoor play area. Image Credit: Behaviour and Evolution Research Group, University of Stirling.

These newer home pens are larger (around 4.8m2/animal compared with the EU minimum of 2.25m2/animal), provide good visibility for the dogs and staff, choice of resting places, noise reducing materials, horizontal rather than vertical bars and enrichment toys inside. The researchers concluded that “the Refinements described here are implemented consistently across industry and suggest that factors such as home pen design should be included in the design of experimental studies.”

Laboratory housed dogs in home pens, AstraZeneca facility. Credit: Laura Hall / Refining Dog Care

Laboratory housed dogs in modern home pens, AstraZeneca facility. Credit: Laura Hall / Refining Dog Care

Dr Hall had previously won an award from NC3Rs for her paper on improving techniques for oral dosing in dogs.  She also developed the “Refining Dog Care” website, to:

[I]mprove the welfare of dogs used in scientific research and testing worldwide, and to improve the quality of data which is obtained from their use. We do this by collaborating with our partners in industry, drawing on expertise and empirical data, to provide guidance on best practice for housing and husbandry, and provide online resources and hands-on training to staff to implement positive reinforcement training protocols for regulated procedures.

Around 4,000 procedures on dogs are carried out in the UK each year (around 0.1% of the total), these are mainly for safety testing, conducted at pharmaceutical or contract research organisations. The fact this research was conducted in collaboration with such organisations will hopefully speed its implementation.

Speaking of Research

References:

  1. Hall et al, 2016, “The influence of facility and home pen design on the welfare of the laboratory-housed dog” in Journal of Pharmacological and Toxicological Methods,
  2. Everds et al, 2013, “Interpreting stress responses during routine toxicity studies a review of the biology, impact, and assessment” in Toxicologic Pathology, 41 (2013)
  3. Hall, 2014, A practical framework for harmonising welfare and quality of data output in the laboratory-housed dog,D. thesis
  4. Tasker, 2012, Linking welfare and quality of scientific output in cynomolgus macaques (Macaca fascicularis) used for regulatory toxicology,D. thesis