Pavel Takmakov

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.

Higher sensitivity dopamine measurements with faster-scan cyclic voltammetry.

Fast-scan cyclic voltammetry (FSCV) with carbon-fiber microelectrodes has been successfully used to detect catecholamine release in vivo. Generally, waveforms with anodic voltage limits of 1.0 or 1.3 V (vs Ag/AgCl) are used for detection. The 1.0 V excursion provides good temporal resolution but suffers from a lack of sensitivity. The 1.3 V excursion increases sensitivity but also increases response time, which can blur the detection of neurochemical events. Here, the scan rate was increased to improve the sensitivity of the 1.0 V excursion while maintaining the rapid temporal response. However, increasing scan rate increases both the desired faradaic current response and the already large charging current associated with the voltage sweep. Analog background subtraction was used to prevent the analog-to-digital converter from saturating from the high currents generated with increasing scan rate by neutralizing some of the charging current. In vitro results with the 1.0 V waveform showed approximately a 4-fold increase in signal-to-noise ratio with maintenance of the desired faster response time by increasing scan rate up to 2400 V/s. In vivo, stable stimulated release was detected with an approximate 4-fold increase in peak current. The scan rate of the 1.3 V waveform was also increased, but the signal was unstable with time in vitro and in vivo. Adapting the 1.3 V triangular wave into a sawhorse design prevented signal decay and increased the faradaic response. The use of the 1.3 V sawhorse waveform decreased the detection limit of dopamine with FSCV to 0.96 ± 0.08 nM in vitro and showed improved performance in vivo without affecting the neuronal environment. Electron microscopy showed dopamine sensitivity is in a quasi-steady state with carbon-fiber microelectrodes scanned to potentials above 1.0 V.

In Vivo Comparison of Norepinephrine and Dopamine Release in Rat Brain by Simultaneous Measurements with Fast-Scan Cyclic Voltammetry

J. Neurochem. (2011) 119, 932–944.

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.

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.

Brain dopamine and serotonin differ in regulation and its consequences

Dopamine and serotonin (5-hydroxytryptamine or 5-HT) are neurotransmitters that are implicated in many psychological disorders. Although dopamine transmission in the brain has been studied extensively in vivo with fast scan cyclic voltammetry, detection of 5-HT using in vivo voltammetric methods has only recently been established. In this work we use two carbon-fiber microelectrodes to simultaneously measure dopamine release in the nucleus accumbens and 5-HT release in the substantia nigra pars reticulata, using a common stimulation in a single rat. We find that 5-HT release is profoundly restricted in comparison with dopamine release despite comparable tissue content levels. Using physiological and pharmacological analysis, we find that 5-HT transmission is mostly sensitive to uptake and metabolic degradation mechanisms. In contrast, dopamine transmission is constrained by synthesis and repackaging. Finally, we show that disruption of serotonergic regulatory mechanisms by simultaneous inhibition of uptake and metabolic degradation can have severe physiological consequences that mimic serotonin syndrome.

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.

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.

Instrumentation for fast-scan cyclic voltammetry combined with electrophysiology for behavioral experiments in freely moving animals.

Fast-scan cyclic voltammetry is a unique technique for sampling dopamine concentration in the brain of rodents in vivo in real time. The combination of in vivo voltammetry with single-unit electrophysiological recording from the same microelectrode has proved to be useful in studying the relationship between animal behavior, dopamine release and unit activity. The instrumentation for these experiments described here has two unique features. First, a 2-electrode arrangement implemented for voltammetric measurements with the grounded reference electrode allows compatibility with electrophysiological measurements, iontophoresis, and multielectrode measurements. Second, we use miniaturized electronic components in the design of a small headstage that can be fixed on the rats head and used in freely moving animals.

Water Confinement in Hydrophobic Nanopores. Pressure-Induced Wetting and Drying

Wetting and drying of hydrophobic pores with diameters lower than 0.2 μm by aqueous solutions at different hydrostatic pressures is investigated by measuring the ionic conductance variation through the nanopores. The critical pressure for water intrusion into the nanopores increases with lowering the pore diameter and the surface tension of the hydrophobic modification, in agreement with the Laplace equation. Nevertheless, restoring the pressure to the atmospheric one does not result in spontaneous pore dewetting unless bubbles are left inside the pores. Such bubbles can appear at the regions of narrowing cross section and/or varying quality of the hydrophobic modification and thus can be engineered to control water expulsion.