Transplanting the light fantastic; cells from eye donors may someday restore vision in some blind individuals.

Transplanting the Light Fantastic

Reading these words requires the cooperation of more than a quarter of a billion specialized, light-sensitive nerve cells neatly arranged along the backs of your eyes. There, upon an exquisitely organized neuronal array called the retina, these alphabet-shaped patters of darkness and light get translated into patterns of electrical potential. Thus encoded, the written words rush to the brain for processing and, ultimately, for decisions -- such as "keep reading" or "I'm bored, turn the page."

But what about the estimated 5 million people in the United States who, through disease or injury, have lost some of their light-sensitive retinal cells? For these individuals, this eyeball-initiated bio-chemical and electrical cascade proceeds sloppily at best, leaving them visually compromised or completely blind. And because mammalian photoreceptor cells so far appear incapable of significant regeneration, visual recovery for these people remains impossible today.

Researchers experimenting with photoreceptor transplants hope to change that prognosis. Recent progress in the ability to graft healthy, light-sensitive neurons inside the eyes of blind animals suggests partial restoration of vision for people with photoreceptor damage may someday become feasible.

"The mindset was that it would be impossible," says Martin S. Silverman, a neurobiologist affiliated with Washington Univeristy and the Central Institute for the Deaf, both in St. Louis. Now, he says, that view has begun to change. In the August INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE, Silverman and Stephen E. Hughes describe the first successful translplants of gelatin patches containing organized arrays of photoreceptor cells. The grafts thrive in the eyes of blinded rats, and tests suggest the transplanted receptors respond properly to light.

Photoreceptor transplants performed in test animals have yet to yield evidence of visual improvement. Indeed, investigators remain uncertain whether the transplanted cells have made the neuronal connections necessary to send messages to the brain. But researchers experimenting with various transplant techniques express increasing optimism that grafted photoreceptors can perform the basic functions needed to tell other cells what they "see."

"It's not something you would necessarily expect to work," says University of Pittsburgh neuroscientist Raymond D. Lund, commenting on Silverman's unique method for splicing photoreceptor patches into damaged retinas. "But it looks like a very promising technique."

Indeed, says James E. Turner, a retinal transplant researcher at Wake Forest University's Bowman Gray School of Medicine in Winston-Salem, N.C., "if such a technique becomes practical, it certainly would be more than helpful for some major classes of retinal dystrophies." Retinal dystrophies refer to a class of diseases, including retinitis pigmentosa and macular degeneration, that result in the dradual deterioration of photoreceptor cells. "There's no known cure for any of these diseases," Turner says.

Photoreceptor cells--known as rods and cones -- form one of 10 cell layers in the human retina. Each photoreceptor cell contains a light-reactive protein called an opsin, which converts light into an electrical potential. These minute power surges must leap across tiny gaps, or synapses, to trigger similar potentials in bipolar cells and ganglion cells that reside in neighboring retinal layers. Ganglion cells then conduct the current via the optic nerve to the brain, which reconstructs a "map" of the visualized image in much the same way a television produces pictures from rows of colored dots.

Sight restoration in people with disrupted ganglion cells seems impossible for now because of difficulties in getting nerve cells to regenerate the long distances traversed by these cells. But many retinal diseases result in a loss of photoreceptors alone, while other parts of the retina remain healthy and intact. These are the disease that researchers hope to treat with photoreceptor transplants.

Some researchers have attempted to restore light responsiveness in rats afflicted with retinal dystrophies by injecting solutions of fresh photoreceptors into the rats' eyes. Results have been largely disappointing -- perhaps because the transplanted cells in these experiments become so disrupted in the process.

In an attempt to leave undisturbed the intrinsic, orderly arrangement of donor photoreceptors, Silverman takes a different approach. First, he immerses donor-rat retains in gelatin. After chilling this biological gel, he uses a micromilling machine to shave consecutive horizontal slices until he gets to the retinal layer containing photoreceptors. Then, performing eye surgery on rats lacking photoreceptors, he slips this cell-laden gelatin slab between the appropriate layers of retinal cells. Hours later the gelatin dissolves and disappears, leaving the transplanted cells in position.

Tests indicated the cells remain alive for at least six weeks and produce large amounts of opsin. Moreover, the transplanted cells utilize increased quantities of glucose after exposure to light -- providing indirect evidence that they are performing their intended electrical functions in response to light.

Those results look promising, but they fall short of demonstrating restoration of true neural function. Glucose uptake tests are fairly messy as a means of confirming specific neuronal activity, notes Lund. Light may simply accelerate the transplanted cells' metabolic rates--and hence their glucose consumption -- without actually inducing within the cells an electrical potential. "We need very careful electrophysiological studies [in these cells] to show that electrical responses can be generated in response to light," Lund says.

While silverman has not yet provided that proof, other researchers using a different transplant technique recently reported evidence of electrical activity in their grafts. Robert J. Collier of the University of Rochester (N.Y.) and his colleagues looked at electroretinograms -- tracings of retinal electrical activity that resemble the electrocardiograms used to measure heart functions -- recorded from retinal tissue they had transplanted into rats.

Unlike Silverman, Collier's group grafted retinal cells into the front of rats' eyes -- far from the retina where any functional transplant must ultimately reside but in a location that allowed the researchers to observe the transplanted cells more easily. At the annual meeting of the Association for Research in Vision and Ophthalmology in May, Collier showed electroretinograms indicating that the transplanted cells, like normal photoreceptors, conduct waves of electrical potentials in response to light.

Those results still don't show that transplanted cells, when placed in the rear of the eye, can grow the micron or so necessary to come within shouting distance of ganglion cells. "The retinogram tells you that these retinal cells are talking, but it does not yet tell you that they are talking to the brain," says Collier's co-worker Manuel del Cerro. Proof of that ultimate electrophysiological link, he says, will require measuring evoked potentials in the brain's visual cortex in response to illumination or to patterns shown on a screen.

No researcher has shown such responses in the brains of photoreceptor recipients, says Silverman, in part because rats have very poor visual acuity in the first place. Silverman says he's preparing to perform transplants in rabbits, cats and primates, which are easier to test for specific brain responses to visual stimuli.

Even if photoreceptor connections prove neurologically sound, researchers foresee other potential complications. For example, transplanted photoreceptors tend to grow abnormally into so-called rosette formations that create bumps on the normally smooth retinal layer. Rosettes are definitely a concern, says Silverman. "We'd expect any disruption to degrade the visual image."

Unfortunately, says del Cerro, "nobody is sure how photoreceptors orient themselves during development. We know the rosettes are linked to abnormal development, but since we do not know what normal is, it's very difficult to know what we should be doing differently in transplantation to avoid having rosettes."

Graft rejection remains another potential problem. Nervous tissue rarely triggers immune responses, making photoreceptors an ideal material for transplantation. But the presence of contaminating, non-neural retinal cells within a graft could trigger an immune response, warns del Cerro. In theory, that could lead not only to graft rejection but also to a sight-threatening antibody attack against the recipient's other eye.

So far, Silverman notes, no such complications have arisen. And even if immunosuppresent drugs become necessary in some cases--as they were in his recent successful transplants of human retinal cells into rats -- they can be applied directly to the eye to prevent the side effects that go along with systemic use of such drugs.

Nobody knows to what extent photoreceptor transplants may restore vision in people with retinal diseases. However, del Cerro emphasizes, patients regard even poor vision as a vast improvement over no vision at all.

Before transplantation, "the rats we are working with cannot see anything at all; the transduction of light is totally demolishedM" he says. "So anything that can say to the retina, and eventually to the brain, 'Here there is light,' would be a considerable jump."