Story posted July 01, 2004
It takes a certain appreciation for the bizarre to be a DNA sharpshooter.
Take Hadley Wilson Horch, an assistant professor of biology and neuroscience at Bowdoin. Her daily rounds include a repugnant swipe into a bin filled with crickets, the chief organism of her genetic research. ("They bite sometimes," she says.)
Under a dissecting microscope, she extracts from them nerve-cell clusters, or ganglia, that contain auditory neurons, which she breaks up into individual cell bodies. Horch then mixes trace amounts of gold dust with DNA-encoding fluorescent jellyfish protein. She loads this micromix into tiny tubing and cuts it into half-inch "bullets," which she loads into a handheld plastic biolistic gun.
Aiming her gun at the Petri dish of auditory neurons, Horch takes aim and "blasts, blasts, blasts," she says.
Hopefully, the gilded blast will be sufficient to transfer some of the DNA into the cricket neurons, through a process known as transfection. If successful, this transfection will cause the cells to become fluorescent, making them wonderfully visible under Bowdoin's powerful, confocal microscope -- one of only a handful in use at a teaching institution in the Northeast.
Why go to this trouble?
Crickets have the ability to regenerate auditory neurons after their ears have been amputated. While the gene responsible for this plasticity has not been identified, Horch is attempting to identify candidate genes in the cricket that might be involved.
If scientists can identify these genes, they can study their role in regeneration - adding a piece of knowledge to the vast network of current research in this field.
"I believe in the power of basic research," says Horch. "You can never know or predict what you'll learn when you ask a basic question. You can have an interesting phenomenon or molecule in a fruit fly. If you figure out how it works there, it may turn out to be relevant to larger fields of inquiry, or even clinical work, such as cancer research. You often find very surprising things. If you are doing clinical work you will also find wonderful things, but you have to ask your question in a very narrow way, going towards a cure."
The goals of Horch's research are hardly basic. Before she can begin studying and developing 3-D models of the dendrites of auditory neurons, she must adapt biolistic technology backwards across the evolutionary chain. Translation: She has to get the gun to work on crickets.
"This blasting technique was developed 15 or 20 years ago for work on plant cells," says Horch. "Then a few people worked on adapting the technique to animal cells. While there are a few labs using biolistics for invertebrates, as far as I know, there isn't anyone focusing on getting the genes introduced specifically into invertebrate neurons."
If Horch's Petri-dish transfection is successful, she will be able to apply the technology to work with living insects and thus begin a deepened study of neuronal regeneration. "We may be able to shoot a live cricket with the GFP, survive it for a few days, then look at the glowing cell. Our goal is to actually watch regeneration happening in a living animal.
"We suspect there are guidance cues, or molecules, that are normally used during development that may be recycled during regeneration. If you can understand their role, you might be able to facilitate it to make regeneration happen, possibly even in human spinal cord situations. There are key differences between human and cricket neuronal regeneration, but we also think there may be some key similarities."
Thus far, says Horch, the transfection process hasn't been fully successful. "We've just gotten started and there is a huge number of parameters to work out. It may be a while before this happens, but I'm confident we'll work the technology out."
Horch's unspoken partner in her enterprise is Bowdoin's confocal microscope, a highly sophisticated piece of equipment funded by grants from the National Science Foundation and Maine Biomedical Research Fund. It uses a laser to "excite" fluorescent molecules, allowing scientists to make a computerized image of the sample. That image can then be made into a 3-D model on a computer.
"Look at this," calls out Horch. On a computer screen in front of her, the lurid, branching form of an auditory neuron hangs suspended like an exotic deep-sea creature. The image was made from samples from the confocal microscope.
"The fact that we have one of these microscopes at Bowdoin is incredible," she says. "Many universities in the Northeast that don't have access to one. It allows me and others to perform all sorts of wonderful experiments."
Horch's research is drawing interest and support from several organizations, among them, the National Institutes of Health, The Grass Foundation, and the Maine Biomedical Research Infrastructure Network (BRIN). Horch hopes her work in neuronal transfection may be used by other researchers involved in invertebrate research, including the study of marine organisms, lobsters, skates and sharks.
"Once someone works on a technique, a number of people can use it. Someone talked to me about using transfection to study regeneration in lobsters. Last year, I had an honors student who looked at neuronal regeneration in crayfish. A technical breakthrough would be significant for me personally, as it would make it much easier for me to answer the actual research questions I'm interested in. But I think there could be a number of other labs that would be interested in something like this as well."
If nothing else, Horch can honestly say: Her work is a blast.