Cell surface stories: miniature electrodes probe diminutive domains

Science News,  Oct 7, 2006  by Aimee Cunningham

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The rat in the plastic box has a drug habit. Every 5 minutes or so, he presses a lever that sends a shot of cocaine through a catheter into his veins. Even more unusual is his "rat hat," the data-transmitting headgear that monitors the animal's brain activity without immobilizing the head. The hat positions several insulated wires within the rat's brain.

For more than 50 years, scientists have been inserting electrodes into tissue samples and animals' bodies to eavesdrop on electrical activity. But the latest-generation electrodes go further. They detect the ebbs and flows of chemicals at the surfaces of cells.

Researchers first developed electrodes that could measure chemical compounds in the early 1980s, but it took a couple of decades for scientists to figure out the best ways to fabricate and use them, says R. Mark Wightman, an analytical chemist at the University of North Carolina in Chapel Hill.

"Once that was done, people started in on applications," says Wightman, who is using microelectrodes in rats to study how cocaine influences the brain circuitry that rewards behaviors.

An electrical property of these microelectrodes "enables chemical events occurring on the sub-microsecond time scale to be monitored" Wightman noted in the March 17 Science.

Moreover, the tools' small size makes a large difference in their capabilities. The microelectrodes, also called ultra-microelectrodes, are typically 50 to 100 micrometers ([micro]m) long. The wire's uninsulated tip, which does the sensing, is 10 [micro]m or less in diameter.

With tips in the same size range as cells, microelectrodes are reaching into biological realms that can't be accessed by their bigger cousins. "If you want to look at chemistry next to a single cell or some other very small space, you would want a microelectrode," says Wightman.

AT THE PUMP Drugs intended to kill microbes and tumors are often foiled by busy cellular pumps. One research team is using microelectrodes to investigate how cells expel those drugs.

A microelectrode senses what's going on at the cell's surface by exchanging electrons with chemicals in solution surrounding the cell. By carefully moving a microelectrode across the surfaces of one or many cells, researchers can map a compound's concentration.

Allen J. Bard of the University of Texas at Austin applies this technique to cells as they rid themselves of toxic substances. He and his colleagues have used the information from concentration maps in a mathematical model that calculates how quickly a cell pumps out material across its entire surface.

The researchers scanned lab-cultured liver cells as they encountered menadione, an analog of a drug used against cancer. Menadione changes to a toxic form inside the cell. But the cell can tag menadione with another chemical group and pump out the resulting compound, called thiodione, through channels in the cell membrane.

"What we find is that the cell is very efficient at getting rid of the menadione" says Bard. His group determined that each cell exports 6 million molecules of thiodione per second. That's almost as fast as the rate at which menadione gets in, he says.

Bard's group is beginning to study whether cancer cells discharge chemotherapy drugs with a similar speed.

In the long term, Bard's goal is to shut down the export processes that cancer cells and infectious microbes use to resist drugs intended to kill them. If cells are "using that kind of pump mechanism, and if you could shut that off with another drug, you could get rid of that resistance," Bard says.

EYEING CELLULAR EXITS Detailed monitoring of cell surfaces can also provide insight into the release of chemical messengers, such as hormones, that carry signals to other cells. Scientists have developed a tool to examine the molecular anatomy of the opening where the cell discharges signal molecules.

During the secretion process, a sac, or vesicle, containing a cargo of molecules inside a cell moves to the cell's membrane and fuses with it. The cargo then leaves the cell through a channel called a fusion pore that opens between the membranes the vesicle and the cell.

Manfred Lindau, a biophysicist at Cornell University, and his colleagues are working to identify the proteins that form the fusion pore. Toward that goal, they've developed a microelectrode array to locate a single active channel. Each microelectrode reports hormone concentration in its vicinity.

The researchers patterned four platinum electrodes onto a glass coverslip, 'like a printed circuit board," Lindau says. In the space at the middle of the four electrodes, they placed a cell that in the adrenal gland releases the stress hormones adrenaline and noradrenaline. The cells, called chromaffin cells, have large vesicles, 200 nanometers (nm) across.

"When you use several electrodes, you can sort of triangulate the position where that release occurs," says Lindau. His team applied a computer analysis to locate the site secreting adrenaline or noradrenaline.