How small zebrafish are contributing to science

They number 15,000, the colony of zebrafish in the Core Zebrafish Facility Ghent at the site of UZ Gent. They are nurtured and cared for under optimum conditions as laboratory animals by the researchers of the Centre for Medical Genetics. “But research into genetic disorders is not all we do here,” professor Paul Coucke emphasises.

The Core Zebrafish Facility offers comprehensive support to both medical and non-medical researchers. External research groups can also use this facility. It started in 2011 with two racks of fish. Initially they were used as a model organism for the connective tissue lab. Together with Core Facility Manager Andy Willaert, Paul Coucke threw his weight behind the expansion of this core facility (see box). A visit to the US where zebrafish were used as a model organism convinced them of the many assets of this small tropical fish.

Today the lab has 17 racks with some 15,000 zebrafish and they work with research groups from different faculties at Ghent University. Amongst others, the zebrafish are used for toxicological studies, the screening of active substances, research into genetic disorders, behavioural studies...

Significant genetic similarities to humans

“But they are still relatively unknown, many researchers still prefer to use mice as a model organism. Until we tell them about the versatility of our zebrafish and the medical breakthroughs they have led to,” Paul argues his case. Research with zebrafish has also proven very valuable for patients with osteogenesis imperfecta, a rare genetic disorder that causes very brittle bones.

Paul: “Our Centre for Medical Genetics specialises amongst others in connective tissue disorders. There are plenty of disorders for which we can conduct studies with humans but with rare diseases this is not an option because the groups are not large enough to allow research. That is why we introduced mutations in fish, in the same gene as in human patients. A key asset of zebrafish is their genetic similarity to humans: we share up to 70% of our genes.”

For instance, there is hot debate in the medical world whether swimming helps improve bone quality in patients with brittle bones (osteogenesis imperfecta). “That is why we designed a special swimming tunnel where fish with brittle bones have to swim upstream to train their muscles and skeleton. Several experiments are ongoing to determine whether such training actually helps,” says Andy.

The most recent study could even result in a treatment for connective tissue disorders. Paul: “We know that the genetic mutation of this disorder is in the collagen. At the same time we see that in a single family one brother has a mild form whereas another brother is wheelchair-bound. So there has to be more going on than just the one mutation. To find the gene that explains that difference, we need a host of test subjects.”

“Therefore it is a major advantage that we have large numbers of zebrafish. We just published a paper in which we found a modifier (a genetic factor that can influence the severity of a disorder) that the fish with a mild form have but not the fish with a more serious disorder,” Andy continues.

The many assets of the zebrafish

The strong genetic match between humans and zebrafish is not the only advantage. Another are the numerous embryos in a single spawning. Andy: “A single couple of zebrafish easily produce 100 to 200 eggs, which is sufficient to set up an experiment.”

Their fast development is also an asset. Andy: “Especially the first few days are extremely fast. Within three days you have a complete fish with functioning organs. At three months they are fully grown and we can crossbreed them.” Within an hour of the egg being laid, the mutations are introduced in the embryos with a very fine injection needle using the CRISPR-Cas method. The zebrafish larvae can be studied after just 48 hours. Andy: “Since they are so small there is no need to add a lot of compound - the substance we wish to test - for drug screens or toxicity studies. And that’s a good thing because those compounds are not exactly cheap. Consequently zebrafish are also highly cost-effective for research purposes.”

The zebrafish as an avatar

More and more cancer researchers are also using zebrafish as a model organism. Breast cancer is just one example, since many of the genes that mutate in patients are also found in zebrafish. But even more miraculous is the so-called xenografting: “With xenografting the zebrafish becomes a patient avatar because a small amount of tumour tissue can be implanted in the larvae. We then test the various medicines to see which of them shrinks the tumour. After those tests we can recommend the optimum treatment for the patient.”

Behavioural research

The larvae and the adult fish are also used for behavioural experiments. “This is especially the case for research into neurogenetic disorders such as epilepsy,” Andy clarifies. “We record the movements of the larvae and how much they deviate from their natural behaviour. Do they swim faster or slower, are they more frightened, do they swim in groups, ...” Paul: “A colleague is currently conducting a pain study among fish that have a certain connective tissue disease. If they swim to the right in the tank, they are fed. If they swim to the left, they get a painkiller. The more pain they have, the faster they swim to the left. Then we add a substance that can alleviate the pain. Depending on their choice - food or painkiller - we can instantly see whether the substance has an effect on the pain.”

The services of the Core Zebrafish Facility

Researchers who are interested in working with zebrafish but don’t know how to go about it can rely on support from Andy and lab technician Hanna De Saffel. Hanna provides technical support in conducting experiments in the core facility. Hanna: “We can train the researchers so they can run their own experiments and breed fish. In addition we offer a range of services where we set up and follow up an experiment on behalf of the researchers. We offer tailor-made projects for users who lack sufficient knowledge.”