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Microelectrode arrays reveal spatial variations in cell signaling Nerve cells communicate with one another by releasing chemical messengers across cell–cell gaps called synapses. Electrochemical analysis has been particularly valuable in studying this communication process, called exocytosis, in which small quantities of these easily oxidized neurotransmitters are released from intracellular vesicles. But single microelectrodes miss spatial heterogeneity, which has emerged as an important factor in nerve cell communication, says Andrew Ewing of Pennsylvania State University and Göteborg University (Sweden). “We’ve been looking at exocytosis from [model] PC12 cells since 1994,” says Ewing. “We wanted to be able to record from more than one site, but we didn’t have the capability we needed.” As Ewing and colleagues report in their latest paper in AC (2008, 80, 1394– 1400), they found a solution in carbon microelectrode arrays (MEAs). “You can get a lot of information about single-cell secretion processes with a traditional, single carbon fiber electrode,” says Bo Zhang, first author on the paper. “But you can’t get spatial information from a single electrode, and exocytosis is actually a very dynamic and spatially heterogeneous process.” Multiple discrete carbon microelectrodes are very difficult to manipulate. Existing microchip-based MEAs are too large—each electrode tip is 10–20 µm in diameter, about the same size as a PC12 cell. The challenge, says Zhang, “was to find smaller electrodes—and a way to put them together.” Zhang turned to carbon fiber electrodes. Typically, a single carbon fiber would be threaded through a glass capillary, which is then pulled into a fine, 5–10 µm diam tip. Zhang says, “We thought we might be able to use a multibarreled capillary and make a carbon fiber array” by threading each of the conjoined barrels with a single, 5 µm diam carbon fiber before pulling. Zhang reasoned if he could produce 1356
5 µm seven 5 µm diam electrodes with (b) (a) 10–20 µm diam tips, isolated Carbon from one another by the intervenfiber ing glass, he would have the resolution needed for simultaneous recording of exocytosis at distinct 20 µm sites around a PC12 cell. The approach worked—eventually. Zhang quickly found that the fabrication wasn’t going to be trivial. “We had to do quite a bit of work to get it to function,” says Ewing. A major challenge, according to Zhang, was learning to handle the 5 µm diam carbon 20 µm fibers. It initially took him most of an afternoon to thread a sevenbarrel capillary, but Zhang says he can now produce a dozen or more seven-electrode arrays in a day. The team worked to establish and model electrode–cell connec20 µm tions and successfully monitored changes in the rate and location of exocytosis from a stimulated PC12 (a) Schematic diagram and (b) SEM images of cell. “There was quite a bit of colcarbon microelectrode arrays composed of lective-brain collaboration to decide two, three, and seven individually addressable how to use it,” Ewing recalls. “But electrodes. Each electrode tip measures ~5 µm in it was Bo’s spark that got it started, diameter. and his effort that got it to work.” The persistence paid off. Using of probing within active synaptic gaps. MEAs with two, three, and seven Zhang is continuing to develop the discrete, individually addressable eleccarbon MEAs, including modified trodes, the group was able to follow arrays that would allow simultanecell secretions over time and to collect ous recording from two or more cells detailed records of exocytosis “hot” and in a network. He plans to develop “cold” spots—areas of heightened or multifunctional arrays, in which each decreased activity. “Bo’s data shows that electrode would be capable of sensing hot and cold spots can move around,” a distinct analyte, such as glucose or says Ewing, “and I don’t think that’s oxygen, or pH. In the meantime, says been shown clearly before.” Ewing, plenty can be learned with the The biological significance of the new MEAs by recording exocytosis observation is still unknown, but Ewing dynamics under treatment with estrowould like to find out whether there’s a gen or other hormones, for example. link to neuronal plasticity. “My feeling, “We’re just trying to see where these though there isn’t strong evidence yet, is events are happening, and when, so that that cells can control the rate and place we can figure out how the cell works,” of individual exocytosis events, and that says Ewing. “It’s very basic cell biology, may be part of memory and learning,” but we had to develop a new analytical he says. That intriguing idea, however, technology to be able to do it.” a awaits even smaller electrodes, capable —Thomas Hayden
A n a l y t i c a l C h e m i s t r y / Ma r c h 1 , 2 0 0 8
