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Johns Hopkins Bloomberg School of Public HealthCAAT

7th Annual 3Rs Symposium: Practical Solutions and Success Stories

rat image © Cathy Schuppli

7th Annual 3Rs Symposium: Practical Solutions and Success Stories

June 4-5, 2020

The 7th Annual 3Rs symposium, co-hosted by the USDA Animal Welfare Information Center (AWIC), NIH Office of Laboratory Animal Welfare (OLAW), the Johns Hopkins Department of Molecular and Comparative Pathobiology, and the Johns Hopkins Center for Alternatives to Animal Testing (CAAT), will be held online on June 4-5, 2020. The goal of this year’s symposium is to bring together experts in replacement, reduction, and refinement of animal experimentation to exchange information with scientists, IACUC members, veterinarians, and animal care technicians about practical solutions and recent success stories to reduce the use of animals in research and improve their welfare.
The format includes 1.5 days of lectures and panel discussions with interactive breakout sessions in the afternoon on day two. These lectures give participants a strong foundation in the relevant research underlying breakthroughs in the 3Rs, while the breakout sessions allow participants to receive feedback specific to their own facilities from experts and colleagues. We will still be able to offer RACE credits for online participation. Information on new pricing and registration will follow soon. For questions, please contact Camila Januario at


Day 1 - 6/4/2020 

8:30 - 8:45
Organizer’s Welcome

8:45 - 9:45
The Zoobiquity Partnership and What Animal Adolescents Can Teach Us About Growing Up as Human Adults
Barbara Natterson-Horowitz, University of California Los Angeles and Harvard University

Recent Success Stories

9:45 - 10:15
Minibrain Organoids
Helena Hogberg, Center for Alternatives to Animal Testing, Johns Hopkins

Helena Hogberg is the Deputy Director of the Center for Alternatives to Animal Testing (CAAT) at the Johns Hopkins Bloomberg School of Public Health, Baltimore, MD. She received her PhD in Toxicology from the Physiology Department at Stockholm University, Sweden in 2009. The scientific work during her PhD was performed at the European Centre for the Validation of Alternative Methods (ECVAM), European Commission, Ispra, Italy and aimed to develop new in vitro approaches to detect chemicals with developmental neurotoxicity (DNT) potential, with a focus on gene expression and electrical activity recordings.

Together with CAAT Director Thomas Hartung she started the current laboratory at CAAT as a Postdoctoral Fellow in 2010. She was later promoted to a faculty position and was appointed Deputy Director of the center in the beginning of 2016. Her current research activity is still in the field of DNT with the use of emerging tools, such as 3D organotypic cell models, induced pluripotent stem cells, and omics (transcriptomics and metabolomics) approaches.

10:15 - 10:45
A Mouse is not a Rat, and Neither is a Human: A Conundrum for Translational Research in Neurodegenerative Diseases
Barry Greenberg, Alzheimer's Disease Research Center, Johns Hopkins University
Bio (MS Word doc)

The use of rodent models in translational research has provided invaluable information in understanding selected aspects of human neurodegenerative disease biology. However, because there are no valid animal models of human neurodegenerative disease, many nuances of such studies are often underappreciated and misinterpreted, obfuscating rather than elucidating the biology of the human disease that investigators are attempting to address. It is critical to understand the limitations of cross-species translation, particularly in diseases that are inherently heterogeneous in which impacts of aging, environment, co-morbidities, behavior and cognition are uniquely human attributes that cannot be modeled in animals. Moreover, co-occurring mechanisms of resistance and resilience that impact on disease pathogenesis and progression are critical to understand in developing therapeutic interventions in humans, aspects of which are difficult or potentially impossible to study in non-human systems without an adequate understanding of interspecific comparative biology. Human-based model systems that are integrated with studies employing corresponding biological samples from well-characterized longitudinal human cohorts are sorely needed to overcome issues related to reliance on animal models, and to serve as a prospective guide for when research in non-human species may be rational, warranted and informative.

10:45 -11:05

11:05 - 12:05
Grimace Scale Panel
Dale Langford (Mice), Univ. of Washington, Anne Burrows (Nonhuman primates), Duquesne University

12:05 - 1:00

Hot Topics: Best Data from Design to Publish

1:00 - 2:00
Improving the design and reporting of animal experiments: the Experimental Design Assistant and the ARRIVE guidelines 2019
Nathalie Percie du Sert, National Centre for Replacement, Refinement and Reduction of Animals in Research

Nathalie Percie du Sert is Head of Experimental Design and Reporting at the National Centre for the Replacement, Refinement and Reduction of Animals in Research (NC3Rs), which she joined in 2010. Her programme of work includes the development of the Experimental Design Assistant, an online tool to guide researchers through the design of in vivo experiments and dissemination of the ARRIVE guidelines to improve the design and reporting of animal research.

She holds a PhD from St George's University of London and worked as a post-doctoral researcher in the field of nausea and emesis at the University of California, San Francisco and at the Chinese University of Hong Kong, where she developed expertise in in vivo research and systematic reviews and meta-analysis of animal models.


Many factors influence the reproducibility of preclinical experiments, with issues around experimental design and reporting estimated to account for half of irreproducible research. Researchers often have limited knowledge and understanding of experimental design and statistics, and may not appreciate the importance of rigorous methodology. This has an impact on the way we design and conduct experiments and it also affects the way we assess manuscripts and grant proposals in our capacity as journal editors, reviewers and funding panel members. The NC3Rs has developed a suite of tools to support researchers. This includes the recently revised ARRIVE guidelines and accompanying Explanation and Elaboration document, and the online Experimental Design Assistant which provides bespoke feedback on individual experimental plans. These resources provide extensive guidance on how to design, conduct and analyse animal experiments, and what crucial information to report in scientific publications. This presentation will include a live demonstration of the Experimental Design Assistant.

2:00 – 2:30
The Center for Open Science: openness, integrity, and reproducibility of research
Cynthia (Cyndy) Parr, National Agricultural Library

2:30 - 3:00
The PRIMatE Data Exchange: an Open Science Resource for Non-human Primate Imaging
Michael Milham, Center for the Developing Brain, Child Mind Institute

3:00 - 3:15

3:15 – 3:45
PREPARE Guidelines
Adrian Smith, NORECOPA
Bio (MS Word doc)

Efforts to improve the scientific rigour and reproducibility of animal experiments have tended to focus on improved reporting and "experimental design"—where the focus is often on the more mathematical elements of design such as randomization, blinding and statistical analysis.

The PREPARE guidelines were written by Norwegian and British scientists as the result of dialogue over a 30-year period, from courses in Laboratory Animal Science for researchers and technicians, supervision of animal research and through management of laboratory animal facilities, including AAALAC accreditation. The name PREPARE implies that we encourage scientists to seek collaboration from day 1 of planning with the animal facility where they intend to work. This ensures that all aspects of the planned experiment, including the practicalities, timeframe, distribution of labour and costs, are discussed with all those who will be involved.

PREPARE consists of 15 main topics, organised in the form of a checklist, which has been translated into 20 languages. The checklist is not meant to add to the administrative burden, and should be used as a basis for dialogue rather than as a mandatory requirement.

Importantly, the checklist is supported by a website where each of the 15 topics is described in more detail. The website is updated continuously with links to new resources within each field. We have made a 3-minute cartoon film illustrating the main principles in PREPARE, making comparisons with the aviation industry with their excellent record of safety and reliability. PREPARE is available at


Smith AJ, Clutton RE, Lilley E, Hansen KEA & Brattelid T (2017): PREPARE: Guidelines for Planning Animal Research and Testing. Laboratory Animals, 52(2): 135-141.

3:45 - 4:15
NINDS/NIH Approach to Rigor and Transparency
Shai Silberberg, National Institute of Neurological Disorders and Stroke, NIH

4:15 – 5:00
Refinement WIKI and International Culture of Care Network
Adrian Smith, NORECOPA


This presentation will highlight two recent collaborative projects designed to refine animal studies and to improve the atmosphere in which we work—for the benefit of both the animals and personnel.

All those who have participated in congresses or email discussion forums know that there are many people developing or using refinements to animal care and use which have not been published in the scientific literature. This is often because of financial or time constraints, or because the refinement was not considered worthy of a publication in its own right.

To bridge the gap between scientific papers and discussion groups, we have constructed a Refinement Wiki (, where improvements can be posted. We hope that it will also function as a hub where those investigating the effects of a potential refinement can identify collaborators, and that it will encourage colleagues to share experiences or develop new strategies to solve a current problem.

The contents of the Wiki are not curated. The quality of the Wiki is determined by registered bona fidemembers of the research animal community. No one else can add, delete or comment upon material. Wiki content is retrievable from Norecopa's main search engine, and the Wiki has also its own search engine. Those interested in participating are encouraged to take contact.

There is increasing focus internationally on fostering a climate of care around animal research - also known as a "culture of care". In Europe this is mentioned specifically in the text accompanying the EU Directive on the protection of animals used for scientific purposes. Culture of care means a commitment to improving animal welfare, scientific quality, care of the staff and transparency for the stakeholders. The EU Commission has produced guidance documents on how to implement such a culture, and these have been endorsed by all Member States.

An international Culture of Care Network was established following the FELASA congress in Brussels in 2016. Norecopa acts as the host website for this network: Members of the network include animal technicians, veterinarians, members of animal welfare bodies, representatives of competent authorities, communications experts and members of animal welfare organisations. The primary aim of the network is to share and publish examples of activities fostering a culture of care. Emphasis is also placed on methods for improving work conditions for all those who care for or use animals in research and testing. The website is used to post practical examples of how to implement a culture of care.

Closely related to a culture of care is the concept of a Culture of Challenge: looking for the acceptable, rather than choosing the accepted. This is much easier to achieve if a caring atmosphere has already been created at the workplace.

5:30 pm
Speaker dinner

Day 2 6/5/2020 

Practical Solutions

8:30 - 9:00
How Dogs are Helping Us Understand Cancer
Amy LeBlanc, Comparative Oncology Program, National Cancer Institute

9:00 - 9:30
Image Guided Animal Therapy
Dara Kraitchman, Johns Hopkins Center for Image Guided Animal Therapy

9:30 - 10:20
Pig handling
Derrick Brocksmith, Sinclair Biosciences

10:20 - 10:40

10:40 - 11:30
Behavioral Training as Part of the Health Care Program
To be Invited: Steven Schapiro, Department of Comparative Medicine, MD Anderson Cancer Center

11:30 - 12:30
Lunch (Chat room planned with attendees and available speakers)

12:30 - 1:00
Hydrophobic Sand as an Alternative to Metabolic Cages
Jessica Hoffman, Uniformed Services University of the Health Sciences

Jessica Hoffman’s greatest passion is in integrative, translational research in the field of mental health, with a strong interest in policy, ethics, and advocacy for better research and treatment. She received a BS in Biochemistry and Molecular Biology and a BA in Psychology from UMBC in 2008, and her PhD in Neuroscience from UMB-SOM in 2014, where her thesis work used cellular and behavioral techniques to discover a sensitive period in which inflammation affected cerebellar development and led to an increased risk of autism-like behavior in the rat. Her postdoctoral work with the National Institutes of Health (NIH, NIMH and NIAAA used cell lines from human clinical trials to integrate physiological and biochemical/genetic research to achieve a more complete understanding of hormone-related mood disorders such as perimenopausal depression (PMD) and premenstrual dysphoric disorder (PMDD). As a Research Biochemist at AFRRI she uses a rat model of embedded metals (i.e. shrapnel wounds) to examine the health effects of military-relevant metals across a multitude of tissues, collaborating with the Maryland VA program and the FDA to identify health risks and early warning biomarkers to improve treatment policy. In addition to her research, she enjoys serving on the AFRRI/USU IACUC.


A common method for rodent urine collection requires a metabolic cage, exposing animals to extended periods of isolation in an unfamiliar cage with a wire-mesh floor. A new method involving hydrophobic sand, a material similar to bedding, has become available but has not been extensively compared with metabolic cages in regard to collection efficiency or stress. Using a within-subjects crossover design, we examined differences in stress markers, urinary markers, and urine volume of healthy male Sprague–Dawley rats during multiple collection sessions in hydrophobic sand and metabolic cages. Stress response markers of weight loss, fecal pellet output, or corticosterone did not differ between hydrophobic sand and metabolic cages, and observed behavior suggested that sand may be less stressful than metabolic cages. All clinically relevant urinary markers examined were normal, with no differences between collection methods. Total urine volume collected was greater from the metabolic cage than sand in most sessions, but the volume collected during the shortest session accounted for 62% of the total volume collected during the longest session. We found no contamination of the sand with several common metals of interest. We also isolated extracellular vesicles (EVs) containing miRNAs from the urine and found no significant differences in particle size, particle concentration, total RNA, or the type and abundance of miRNAs due to urine collection method. Our results suggest that hydrophobic sand is a refinement of urine collection methods for rats that decreases isolation time, risk of injury, and stress and maintains the integrity of urine samples.

Hoffman, J.F., Vechetti Jr., I.J., Alimov, A.P., Kalinich, J.F., McCarthy, J.J., Peterson, C.A. (2019) Hydrophobic sand is a viable method of urine collection from the rat for extracellular vesicle biomarker analysis. Molecular Genetics and Metabolism Reports. (e-pub, vol 21, article 100505, DOI 10.1016/j.ymgmr.2019.100505)

Hoffman, J.F., Fan, A.X., Neuendorf, E.H., Vergara, V.B., & Kalinich, J.F. (2018) Hydrophobic sand versus metabolic cages: a comparison of urine collection methods for the rat (Rattus norvegicus). Journal of the American Association for Laboratory Animal Science. 57(1):1-7

Hoffman, J.F., Vergara, V.B., Mog, S.R., & Kalinich, J.F. (2017) Hydrophobic sand is a non-toxic method of urine collection, appropriate for urinary metal analysis in the rat. Toxics 5(4), 25

Disclaimer: The views expressed in this presentation are my own and do not reflect the official policy or position of the Armed Forces Radiobiology Research Institute, Uniformed Services University for Health Sciences, Department of Defense, or US Government. I have no conflicts of interest to report. The use of specific products in this presentation does not constitute an endorsement by the US Government.

1:00 - 1:30
Mouse handling limits the impact of stress on metabolic endpoints
Parinaz Mahbod, University of Cinncinnati

1:30 - 2:20
State of the Art Rodent Handling: Can a Little Time Go a Long Way for Welfare?
Brianna Gaskill, Purdue University


Handling laboratory rodents and other common husbandry and experimental procedures can be a source of stress leading to the development of aversive responses to handling or human presence that are suggestive of emotional distress. Aversive handling of animals can lead to a fearful relationship with humans, making handling difficult, and increasing the risk of injury for both handler and animal, thus affecting human and animal welfare. Recent research has provided evidence that new handling techniques such as tickling, tube handling or, gentling with associative conditioning are effective at reducing fear of humans when rodents are handled for common husbandry activities and medical and research procedures such as injection. Some techniques such as tickling and associative conditioning techniques are even perceived as positive by rats, which improves the human-rodent relationship and rat welfare. The goal of the seminar is to review new state of the art rodent handling techniques and provide scientific data to support their effectiveness in improving rodent welfare as well as data quality.

2:20 - 2:35

Breakout Sessions (30 minites each) in the Zoom breakout room:

2:35 - 3:05 pm

3:10 - 3:40 pm

3:45 - 4:15 pm