Characterization of local pH changes in brain using fast-scan cyclic voltammetry with carbon microelectrodes.

Transient local pH changes in the brain are important markers of neural activity that can be used to follow metabolic processes that underlie the biological basis of behavior, learning and memory. There are few methods that can measure pH fluctuations with sufficient time resolution in freely moving animals. Previously, fast-scan cyclic voltammetry at carbon-fiber microelectrodes was used for the measurement of such pH transients. However, the origin of the potential dependent current in the cyclic voltammograms for pH changes recorded in vivo was unclear. The current work explored the nature of these peaks and established the origin for some of them. A peak relating to the capacitive nature of the pH CV was identified. Adsorption of electrochemically inert species, such as aromatic amines and calcium could suppress this peak, and is the origin for inconsistencies regarding in vivo and in vitro data. Also, we identified an extra peak in the in vivo pH CV relating to the presence of 3,4-dihydroxyacetic acid (DOPAC) in the brain extracellular fluid. To evaluate the in vivo performance of the carbon-fiber sensor, carbon dioxide inhalation by an anesthetized rat was used to induce brain acidosis induced by hypercapnia. Hypercapnia is demonstrated to be a useful tool to induce robust in vivo pH changes, allowing confirmation of the pH signal observed with FSCV.

Microfabricated FSCV-compatible microelectrode array for real-time monitoring of heterogeneous dopamine release.

Fast scan cyclic voltammetry (FSCV) has been used previously to detect neurotransmitter release and reuptake in vivo. An advantage that FSCV has over other electrochemical techniques is its ability to distinguish neurotransmitters of interest (i.e. monoamines) from their metabolites using their respective characteristic cyclic voltammograms. While much has been learned with this technique, it has generally only been used in a single working electrode arrangement. Additionally, traditional electrode fabrication techniques tend to be difficult and somewhat irreproducible. Described in this report is a fabrication method for a FSCV compatible microelectrode array (FSCV-MEA) that is capable of functioning in vivo. The microfabrication techniques employed here allow for better reproducibility than traditional fabrication methods of carbon fiber microelectrodes, and enable batch fabrication of electrode arrays. The reproducibility and electrochemical qualities of the probes were assessed along with crosstalk in vitro. Heterogeneous release of electrically evoked dopamine was observed in real-time in the striatum of an anesthetized rat using the FSCV-MEA. The heterogeneous effects of pharmacology on the striatum were also observed and shown to be consistent across multiple animals.

Enhancing Electrochemical Detection by Scaling Solid State Nanogaps.

The ability to quickly and inexpensively fabricate planar solid state nanogaps has enabled research to be effectively performed on devices down to just a few nanometers. Here, nanofabricated electrode pairs with electrode-to-electrode spacings of <4, 6 and 20 nm are utilized for monitoring an electroactive molecules, dopamine, in ionic solution. The results show a several order of magnitude enhancement of the electrochemical signal, collected current, for the solid state nanogaps with 6 nm electrode-electrode spacings as compared to traditional microelectrodes. The data from the <4 nm and 20 nm solid state nanogaps verify that this enhancement is due to cycling of the redox molecules in the confined geometry of the nanogap. In addition the data collected for the <4 nm nanogap emphasizes and reinforces that scaling does have limits and that as device sizes move to the few nanometer scale, the influence of a molecule's size and other physical properties becomes increasingly important and can eventually dominate the generated signals.

Carbon microelectrodes with a renewable surface.

Electrode fouling decreases sensitivity and can be a substantial limitation in electrochemical experiments. In this work we describe an electrochemical procedure that constantly renews the surface of a carbon microelectrode using periodic triangle voltage excursions to an extended anodic potential at a scan rate of 400 V s(-1). This methodology allows for the regeneration of an electrochemically active surface and restores electrode sensitivity degraded by irreversible adsorption of chemical species. We show that repeated voltammetric sweeps to moderate potentials in aqueous solution causes oxidative etching of carbon thereby constantly renewing the electrochemically active surface. Oxidative etching was established by tracking surface-localized fluorine atoms with XPS, by monitoring changes in carbon surface morphology with AFM on pyrolyzed photoresist films, and also by optical and electron microscopy. The use of waveforms with extended anodic potentials showed substantial increases in sensitivity toward the detection of catechols. This enhancement arose from the adsorption of the catechol moiety that could be maintained with a constant regeneration of the electrode surface. We also demonstrate that application of the extended waveform could restore the sensitivity of carbon microelectrodes diminished by irreversible adsorption (electrode fouling) of byproducts resulting from the electrooxidation and polymerization of tyramine. Overall, this work brings new insight into the factors that affect electrochemical processes at carbon electrodes and provides a simple method to remove or reduce fouling problems associated with many electrochemical experiments.

Simultaneous monitoring of dopamine concentration at spatially different brain locations in vivo.

When coupled with a microelectrode, background-subtracted fast scan cyclic voltammetry (FSCV) allows fast, sensitive and selective determination of analytes within a small spatial location. For the past 30 years experiments using this technique have been largely confined to recordings at a single microelectrode. Arrays with closely separated microelectrodes would allow researchers to gain more informative data as well as probe regions in close spatial proximity. This work presents one of the first FSCV microelectrode arrays (MEA) implemented in vivo with the ability to sample from different regions in close spatial proximity (equidistant within 1mm). The array is manufactured from fused silica capillaries and a microfabricated electrode spacer. The functionality of the array is assessed by simultaneously monitoring electrically stimulated dopamine (DA) release in the striatum of anesthetized rat. As expected, heterogeneous dopamine release was simultaneously observed. Additionally, the pharmacological effect of raclopride (D(2) receptor antagonist) and cocaine (monoamine uptake blocker) on the heterogeneity of DA release, in spatially different brain regions was shown to alter neurotransmitter release at all four electrode sites.

Simultaneous decoupled detection of dopamine and oxygen using pyrolyzed carbon microarrays and fast-scan cyclic voltammetry.

Microfabricated structures utilizing pyrolyzed photoresist have been shown to be useful for monitoring electrochemical processes. These previous studies, however, were limited to constant-potential measurements and slow-scan voltammetry. The work described in this paper utilizes microfabrication processes to produce devices that enable multiple fast-scan cyclic voltammetry (FSCV) waveforms to be applied to different electrodes on a single substrate. This enabled the simultaneous, decoupled detection of dopamine and oxygen. In this paper we describe the fabrication process of these arrays and show that pyrolyzed photoresist electrodes possess surface chemistry and electrochemical properties comparable to PAN-type, T-650, carbon fiber microelectrodes using background-subtracted FSCV. The functionality of the array is discussed in terms of the degree of cross talk in response to flow injections of physiologically relevant concentrations of dopamine and oxygen. Finally, other applications of pyrolyzed photoresist microelectrode arrays are shown, including spatially resolved detection of analytes and combining FSCV with amperometry for the detection of dopamine.

Electrochemical Dopamine Detection: Comparing Gold and Carbon Fiber Microelectrodes using Background Subtracted Fast Scan Cyclic Voltammetry.

Electrochemical detection is becoming increasingly important for the detection of biological species. Most current biological research with electrochemical detection is done with carbon fiber electrodes due to their many beneficial properties. The ability to build electrochemical sensor from noble metals instead of carbon fibers may be beneficial in developing inexpensive multiplexed electrochemical detection schemes. To advance understanding and to test the feasibility of using noble metal electrochemical sensors the detection of dopamine, a biologically important small molecule was studied here. Specifically, dopamine detection on gold microelectrodes was characterized and compared to P-55 carbon fiber microelectrodes of the same geometry, using background subtracted fast scan cyclic voltammetry. While not as sensitive to dopamine as carbon fibers, it was observed that gold microelectrodes have six times the saturation coverage per area and 40 times the linear working range. Selectivity to dopamine, in comparison to several other neurotransmitters and their derivatives, is also quantitatively described.