Small Fish for a Large Task

David Hinton Takes His Medaka West to Monitor California's Drinking Water p.3

  Hinton’s own lab has shown how changes in medaka livers can signal the presence of cancer causing agents in water. He says their livers can be “primed” for detection duty by exposing the animals to doses too low to actually form tumors. That makes liver cells undergo changes that leave them more susceptible if exposed again later.

   Toxicologists usually have to “sacrifice” fish and extract their livers for microscopic examination before they can screen for tumors. But the Japanese have recently bred a strain that is transparent throughout its entire lifespan.

  Called SY II, the see-through form lacks the pigmentation of standard “orange-red” medakas. In the SY II type, “You can see the liver surface, the heart, the kidney, the spleen, and the developing eggs through the body wall in the living fish,” Hinton marvels. “We think we’re going to be able to use this mutant to ask questions about these fish without sacrificing them.” Researchers also have introduced a fluorescent “reporter” gene into the fish that lights up to signal whether eggs are developing normally.

  In 2002 Hinton spent three months at the Bioscience Center at Japan’s University of Nagoya evaluating the transparent fish’s potential and co-wrote a joint paper with research leaders there. The co-authors described their through-the-microscope observations of embryonic livers assembling themselves within living SY II medaka. Hinton’s team has since secured a new $713,000 National Institutes of Health grant to further study liver development in the see-through fish.

  Over the years, he and his co-workers have become adept at successfully raising medaka in the confines of aquarium tanks. “We’ve shown that they are very hardy, and there are a lot of diets we can feed them on,” he says. His lab uses a standard commercial food prepared by the Japanese, embellishing that with daily rations of brine shrimp.

  The researchers prompt the fish to reproduce by exposing them to daily schedules of 16 hours of light and eight hours of darkness. Females respond by depositing 20 to 30 eggs on their lower abdominal walls and males fertilize the emerging eggs. The scientists then remove individual fertilized eggs from their egg clusters, a problem complicated in medaka by the fact that each egg is attached to a filament.

  Hinton says the need to separate those filaments is one reason some researchers at Duke and elsewhere prefer using another aquatic species—the zebra fish—as animal models. Zebra fish, natives of India and parts of Southeast Asia, deposit eggs individually rather than in clusters.

  Japanese medaka breeders have responded with techniques to more easily isolate individual fertilized medaka embryos by using finger pressure on the egg masses. Once separated, it takes about nine days for each 1-millimeter in diameter medaka egg to hatch at prescribed water temperatures of 25-27 degrees Fahrenheit. During this time development of the embryo occurs while the organism is surrounded by the egg shell membrane. Nourishment is by the yolk sac, a part of the developing embryo. In fact, these developing organisms have their complete nutritional requirements to sustain them until hatching after which medaka swim and take on external food.

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