RESEARCH OF A DIFFERENT STRIPE

WHEN YOU IMAGINE an animal research facility, the first thing that comes to mind is likely a room with racks of small plastic boxes with scurrying white mice, or rows of metal cages housing monkeys. However, over the past few decades, another sight has become increasingly common: dozens, sometimes hundreds, of racks of small, plastic aquaria containing thousands of minnowsized fish.

The zebrafish (Danio rerio) is a small, freshwater fish that—like its namesake— has narrow dark stripes along its body. While fish might not be the first lab animal you think of, zebrafish have become a popular model organism for biomedical and toxicological research, and their use has expanded significantly since the first studies using zebrafish were published in the 1980s.

WHY ZEBRAFISH?

Researchers cite several reasons for the increasing popularity of zebrafish. They are relatively small and easy to keep (com- pared to mice and other mammals), and they reproduce and grow quickly. The zebrafish genome was sequenced in 2013, and researchers have created many wellcharacterized genetic mutations for study. All these factors make zebrafish a convenient model organism. That said, most zebrafish research is not actually focused on adult fish. Instead, the life stage of greatest research interest is the zebrafish embryo. In most research labs, adult fish are primarily used for breeding purposes to obtain embryos.

Zebrafish embryos are widely used in biomedical research because they are transparent and develop outside the mother’s body. This makes it easy for researchers to watch embryos grow in real time under a microscope, something that is very difficult for a mouse embryo. Zebrafish embryos are also used in toxicity testing and to help determine if or how a particular chemical affects embryonic development. Ironically, the use of zebrafish embryos in toxicity testing is sometimes considered an “alternative” to animal use, since they are used in place of other “higher” animals.

Welfare Concerns And Legal Protections

The rapid increase in zebrafish use raises numerous welfare concerns, since researchers acknowledge there is not enough information about welfare to develop best practices. However, even in cases where good evidence currently exists, uptake of best practices and the implementation of additional protections have been disappointingly slow.

For decades, many scientists insisted that fish do not feel pain, and this was used as justification for not providing welfare protections on par with other research animals. However, it is now well established that fish can feel pain, to the extent that researchers are now working on methods to use zebrafish in studies about pain (Ohnesorge, 2021). As evidence accumulates regarding the ability of fish to feel pain and experience other environmental harms, there is an increased urgency for welfare protections.

Fish are not protected by the Animal Welfare Act (AWA), an unfortunate similarity they share with reptiles and cephalopods used in research in the United States. They are also not covered by the AWA’s reporting requirements. Around 5 million zebrafish are estimated to be used in over 3,000 research facilities worldwide, but the absence of reporting requirements makes it difficult to know how many are used in the United States (Lidster, 2017). The European Union animal use reporting database (ALURES) indicates that 5 percent (about half a million) of the approximately 10.4 million animals used in EU research in 2019 were zebrafish. The National Institutes of Health RePORTER database lists more than 900 projects with “zebrafish” in the title or keywords for fiscal year 2021, with federal funding over $460 million.

Although fish are not covered by the AWA, they are afforded consideration under the Health Research Extension Act, which applies to vertebrate animals used in federally funded research. This means their use is subject to review by an Institutional Animal Care and Use Committee and some minimum standards of care. However, the significant knowledge gaps regarding welfare mean that even the most well-intentioned caretakers in labs lack sufficient guidance.

Urgent Need For Better Practices

A 2016 survey of 98 laboratories worldwide noted several welfare concerns regarding zebrafish care, from housing and handling to euthanasia, and provides a possible roadmap for future welfare studies (Lidster, 2017). One of the most disconcerting findings involves practices for euthanasia and anesthesia. The most common method for anesthesia, which is often also the first step of euthanasia, uses a chemical called tricaine (MS-222). In some countries, tricaine is the only approved anesthetic for zebrafish in research. However, its use raises serious welfare concerns as researchers have reported adverse effects; in fact, over half of respondents to the survey reported observing behaviors such as rapid swimming, gasping, and