UF Vision Researchers Use Transgenic Tadpoles To Prove Crucial Protein Moves Inside Eye Cells
GAINESVILLE, Fla. — University of Florida researchers have obtained the first photographic proof that a protein crucial to vision moves inside eye cells in response to light, which may help explain how people and animals can see in a wide range of conditions.
Scientists say insight about the protein’s movement could one day lead to better understanding diseases such as night blindness or macular degeneration, the leading cause of vision loss in Americans older than 65. The UF findings were published online this month in the journal Experimental Eye Research.
The protein, visual arrestin, regulates a chemical reaction responsible for vision that begins in the retina, the delicate layer of light-sensitive cells at the back of the eyeball where visual information is converted into nerve impulses, said W. Clay Smith, an assistant professor of ophthalmology at UF’s College of Medicine.
“The movement of arrestins probably impacts how we’re able to regulate the light sensitivity of our eyes,” Smith said. “If you go from a darkened theater to bright sunlight, the light intensity can increase by a factor of 10 billion. Not many receptors are capable of dealing with that kind of range, but our eyes can.”
UF researchers examined retinal cells called rods, which operate in low-light conditions but do not perceive color. Other photoreceptor cells called cones, concentrated near the center of the retina, are color-sensitive but function only in bright light.
To trace arrestin’s movements in rod cells, UF researchers introduced a gene derived from luminous jellyfish into African clawed frog tadpoles, Smith said. The tadpoles’ eyes produced arrestin that glowed bright green when exposed to blue light, making the protein easy to observe and – for the first time – photograph.
Smith and UF ophthalmology postdoctoral student James Peterson showed that in low-light conditions arrestin is stored at the back of rod cells, an area called the inner segment. But in the presence of bright light, arrestin moves to the opposite end of the cell – the outer segment – where incoming light is captured.
Arrestin binds to another protein that starts the chemical reaction responsible for vision once light strikes the retina, Smith said, stopping the conversion of visual information into electrical impulses that travel to the brain. Once stopped, the entire process can begin again.
Because of arrestin, light-sensitive cells in the retina can process multiple new images. As a result, vision is akin to a seamless video instead of a series of still photos.
A human retina contains about 100 million rod and cone cells combined, each type of cell with its own form of arrestin. Arrestin found in human cone cells also can move, Smith said.
“Arrestins are present in every eye that’s been studied, from nematodes (a type of worm) to humans,” Smith said. “If you have a visual organ, you have a visual pigment and you have arrestins.”
In collaboration with colleagues at the University of Connecticut, UF researchers gathered thousands of eggs from female African clawed frogs and fertilized them with sperm modified to carry a jellyfish gene. The procedure yielded several dozen healthy, transgenic tadpoles.
In the study, 21 groups of four or five transgenic tadpoles were kept in darkness overnight, then exposed to specific lighting conditions for periods ranging from 15 minutes to four hours. Another five groups went through the same procedure, then were exposed to darkness again for five to 45 minutes.
When scientists examined retinal cells under a special microscope that emitted a blue laser light, the arrestin glowed green, revealing its exact location.
Arrestin began migrating from the inner part of retinal cells to the outer segment after 15 minutes of exposure to light, a move it had almost completed after an hour.
UF researchers were surprised to find arrestin began returning to the inner segment after two-and-a-half hours of light exposure and returned almost completely after four hours.
“We’re not sure why arrestin moves back to the inner segment after prolonged exposure to light,” Smith said. “We hypothesize that arrestin stays in the outer segment until its job of quenching the visual pigment is done. Then some other mechanism must take over, releasing the arrestin.”
The research was funded by grants from the National Eye Institute, the Research to Prevent Blindness Foundation and the Foundation Fighting Blindness. Smith and Peterson are applying for an additional $1.8 million grant from the National Institutes of Health for further research.
Smith is particularly interested in targeting proteins involved in directing arrestin to the correct destinations in retinal cells. Defects in those proteins could cause problems in the movement, or translocation, of arrestin and may play a role in some retinal disorders, said Smith, who will discuss his findings May 9 at the Association for Research in Vision and Ophthalmology annual meeting in Fort Lauderdale.
Smith’s hypothesis is supported by recent findings at the University of Southern California and the University of Utah showing that arrestin also moves in cone cells, said James F. McGinnis, a professor of ophthalmology and cell biology at the Dean A. McGee Eye Institute at the University of Oklahoma Health Sciences Center in Oklahoma City.
“This suggests the possibility that some forms of macular degeneration, a disease primarily affecting cones, may be due to a malfunction of this translocation mechanism,” said McGinnis, who researches arrestin.