What finding out worms, flies and fish says about autism | Spectrum
S.Scientists have also studied the nocturnal rituals of another model of autism: zebrafish (Danio rerio). Gene editing techniques allow scientists to create zebrafish with autism-related mutations, and then they can easily assess how the mutations are affecting behavior. “We can take our fish larvae – at 5 days of age they have this complex spectrum of behavior – we can easily pipette them into the wells of a 96-well plate and then monitor various aspects of their locomotor activity,” says Hoffman. The design offers the high throughput and easy replicability of a cell culture study with the ability to assess the effects on animal behavior.
As a postdoctoral fellow, Hoffman followed the behavior of 5-day-old zebrafish larvae that lack the autism-related gene CNTNAP2 and found that they are hyperactive at night. The fish also have fewer inhibitory neurons in the forebrain than usual, which dampen neural activity, which replicates the results of mice lacking the same gene and increases the fish’s importance as a model of autism.
Zebrafish are a useful tool for screening potential drugs for autism because chemical compounds can be added directly to the water the fish swim in. In their CNTNAP2 study, published in 2016, Hoffman and her colleagues tested the effects of 14 drugs on the larvae and showed that certain forms of the hormone estrogen can reverse the hyperactive behavior of the larvae.
The animals are small and relatively inexpensive, so the team can use them to study the effects of many genes linked to autism in parallel. Hoffman’s team studied brain activity, movement, and sleep-wake cycles in multiple zebrafish lines with mutations in the fish equivalents of 10 genes associated with autism, including CHD8, CNTNAP2, DYRK1A, GRIN2B, and SCN2A. The researchers aim to identify common traits between strains and identify drugs that could reverse changes in their behavior.
The CNTNAP2 zebrafish developed in Hoffman’s laboratory is set to appear on a list of validated zebrafish models curated by the Simons Foundation Autism Research Initiative (SFARI). (Spectrum is an editorially independent publication funded by SFARI.) The aim of this list is to make zebrafish research more reliable by directing researchers to models that pass a test of genetic quality, says Brigitta Gundersen, chief scientist at SFARI.
“It is important to match your question to the advantages and disadvantages of your model system.” Ethan Scott
Zebrafish larvae have another advantage: They are transparent in the first days of life. As a result, the researchers can see the internal organs of the larvae, including the intestines, and visualize the effects of autism mutations on bowel function, which is often disturbed in autism. In the larvae, researchers can observe the rhythmic movements of the intestinal muscles and the movement of food through the digestive system. “Things are playing out right before your eyes,” said Julia Dallman, associate professor of biology at the University of Miami in Coral Gables, Florida.
In studies published in 2019 and 2020, Dallman’s team showed that the intestinal muscles contract and food moves unusually slowly through the intestines in zebrafish with mutations in SYNGAP1 or SHANK3. In humans, mutations in these genes have been linked to both autism and gastrointestinal disorders, including constipation and acid reflux. “When we started studying bowel function in these models, my expectation was: [alterations] would be subtle, ”says Dallman. “It’s not subtle at all.” Her team’s studies suggest that the two fish strains have slightly different mechanisms underlying bowel problems, so treating constipation in autism may not be uniform. Dallman plans to test the effects of drugs on bowel function and fish behavior to help find autism drugs that won’t make constipation worse, she says.
The transparency of the zebrafish larvae also shows early brain development. Using special microscopes, researchers can visualize the activity of individual neurons and, because the fish are small, simultaneously track the activity of each neuron in the brain. “You are only observing the brain of an intact, alert, behavioral, perceptive animal,” says Ethan Scott, who studies sensory processing in zebrafish at the University of Queensland in Brisbane, Australia.
Zebrafish have a more similar brain structure to humans than invertebrate models, says Scott. And although fish lack a cerebral cortex, the structure on the surface of the human brain, they are useful for studying circuits in other parts of the brain. “The important thing is to match your question to the advantages and disadvantages of your model system,” says Scott.
Scott uses zebrafish larvae with mutations in autism-related genes to study changes in sensory processing in autism. He and his colleagues monitor their brain activity in response to images displayed on a computer screen and to sounds of varying volume. In a 2020 study, his team showed that zebrafish without FMR1, the gene mutated in Fragile X syndrome, are hypersensitive to sound. And in four regions of the brain, their neurons respond to sounds with more frequent or stronger bursts of activity than typical fish. The work could help explain sensory hypersensitivity in autism, says Scott.