Abstract
Transparent and Flexible Low Noise Graphene Electrodes for Simultaneous Electrophysiology and Neuroimaging
Kuzum D, Takano H, Shim E, Reed JC, Juul H, Richardson AG, de Vries J, Bink H, Dichter MA, Lucas TH, Coulter DA, Cubukcu E, Litt B. Nat Commun 2014;20(5):5259. doi: 10.1038/ncomms6259.
Calcium imaging is a versatile experimental approach capable of resolving single neurons with single-cell spatial resolution in the brain. Electrophysiological recordings provide high temporal, but limited spatial resolution, because of the geometrical inaccessibility of the brain. An approach that integrates the advantages of both techniques could provide new insights into functions of neural circuits. Here, we report a transparent, flexible neural electrode technology based on graphene, which enables simultaneous optical imaging and electrophysiological recording. We demonstrate that hippocampal slices can be imaged through transparent graphene electrodes by both confocal and two-photon microscopy without causing any light-induced artefacts in the electrical recordings. Graphene electrodes record high-frequency bursting activity and slow synaptic potentials that are hard to resolve by multicellular calcium imaging. This transparent electrode technology may pave the way for high spatio-temporal resolution electro-optic mapping of the dynamic neuronal activity.
Commentary
Electrophysiology technology has seen very few changes since its inception. Over the past 30 years, new developments in digital amplifiers, microfabricated microelectrodes, and signal processing have greatly improved the resolution and capabilities of this technique in both research and clinical venues. However, at its core (or rather, at its tip), electrophysiology has always required metal. Whether it be silver-chloride electrodes in brain slices, tungsten sharp electrodes, platinum-iridium disks in human intracranial studies, gold tabs on flexible ribbons, or platinum tips on microarrays—the business end of nearly all electrophysiological electrodes is metal. Metal is a good conductor, flexible, easily acquired, and durable. But choosing the right metal is complicated. The underlying need for all of these devices is to create an electrode–tissue interface, a transition zone in which electrical current in the wires is transformed into physiological (ionic) current in the tissue. This interface is very complex, and choosing materials that can successfully transmit the current while not harming the tissue or degrading the electrode is an ongoing challenge (1). The majority of in vivo electrodes use noble metals—gold, platinum, or platinum-iridium—not because they are effective, but because they are safe. In fact, their electrical properties are often quite poor, and at microelectrode sizes, they filter out lower frequencies and provide limited fidelity of recorded or stimulated signals (2). The filtering removes direct current (DC) from clinical EEG signals, although DC may be helpful in localizing seizures (3). Other limitations of metal are more subtle: nonreversible reactions can cause electrode corrosion over time, and metal always visually blocks the tissue it is recording. Thus, although there is a rich history with metal electrodes, there are many areas for potential improvement.
The information age has provided us with a wealth of new materials with many useful and unique properties. Many of these have led to next-generation electrodes, such as carbon fibers (4), carbon nanotubes (5), and poly(3,4-ethylenedioxy-thiophene) (PEDOT)-coated metal (6), all of which represent improvements over traditional metal microelectrodes. One new material that has been at the center of many new applications is graphene. Graphene is a two-dimensional lattice of carbon atoms, connected in a honeycomb, hexagonal structure. It forms into a single atomic layer with remarkable properties: graphene acts like a semiconductor or a conductor; and it is flexible, clear, and stronger than diamond or steel. Originally discovered in 2005 by Geim and Novoselev (7), it very quickly led them to the Nobel Prize in Physics (2010), and it has become one of the most intriguing materials for electronics.
In this study, Kuzum and colleagues developed a method of producing graphene electrodes on flexible cables and demonstrated the utility of this new material in electrophysiology. They first developed a polyimide ribbon cable with gold wires but did not extend the gold into the electrode sites. They then deposited a layer of graphene in the electrode locations and connected it to the gold wires. The result was a thin, translucent cable with invisible electrodes (see Figure 1b in Kuzum et al. 2014). The authors demonstrate several advantages of this new electrode type. Graphene can be used either in its basic form or “doped” to conduct electricity more freely. The doped version has very favorable electrical properties that are clearly superior to gold: lower impedance, more uniform response across frequencies, and fewer chemical reactions with tissue. Graphene also generates considerably less noise than gold electrodes. But the most unique property of the graphene is its transparency, which has never been available in prior electrode technology. Kuzum and colleagues exploited this feature by performing a series of optical imaging experiments using calcium-indicator dyes. For the first time, they were able to record local field potentials from the same cells that were being recorded or illuminated optically, allowing them to combine the spatial resolution of two-photon imaging with the temporal resolution of microelectrodes. The optical imaging demonstrated that most pyramidal cells in the field fired simultaneously, while the electrode recorded neural potentials that were either too fast or too slow to resolve with the calcium dyes. Finally, they demonstrated the protective properties of graphene. They fabricated gold electrodes, coating half of them with graphene, then monitored the electrical properties and Raman spectroscopy over 6 months. The graphene maintained its integrity over the entire period, improving the efficacy of the gold electrode. They repeated the experiment with silver electrodes and found that graphene prevented degradation of the metal, which is a known limitation of bare silver electrodes. Together, these experiments demonstrated that graphene can be used either as a protective coating for traditional metal contacts or as an effective, transparent electrode by itself.
Graphene is already recognized for its tremendous versatility in next-generation electronics, but it now appears to have several unique characteristics that make it especially suitable for epilepsy and neuroscience research as an electrode. There are certainly some limitations: being just one-atomic-layer thick, its durability is concerning; the electrochemistry is unknown; its biocompatibility has not been rigorously tested; and it is unclear whether it can produce DC signals. But the benefits are significant. It is flexible, protects underlying electronics, and is extremely strong. It maintains its integrity for months and improves the signal-to-noise ratio of microelectrodes. But perhaps the most ideal characteristic is its transparency. Simultaneously recording the local field potential while performing optical experiments such as calcium imaging or optogenetics thru the electrode is unprecedented. This technology opens the door to many experiments that have previously been impossible, such as combining high spatial and temporal resolution to investigate how how various local field potential phenomena are produced. Graphene provides an alternative to the inherent limitations we have come to accept with metal electrodes and allows more sophisticated and versatile types of recording and stimulation in neuroscience research.