Abstract5-HT is an important molecule in the brain that is implicated in mood and emotional processes. In vivo, its dynamic release and uptake kinetics are poorly understood due to a lack of analytical techniques for its rapid measurement. Whereas fast-scan cyclic voltammetry with carbon fiber microelectrodes is used frequently to monitor sub-second dopamine release in freely-moving and anesthetized rats, the electrooxidation of 5-HT forms products that quickly polymerize and irreversibly coat the carbon electrode surface. Previously described modifications of the electrochemical waveform allow stable and sensitive 5-HT measurements in mammalian tissue slice preparations and in the brain of fruit fly larvae. For in vivo applications in mammals, however, the problem of electrode deterioration persists. We identify the root of this problem to be fouling by extracellular metabolites such as 5-HIAA, which is present in 200-1000 times the concentration of 5-HT and displays similar electrochemical properties, including filming of the electrode surface. To impede access of the 5-HIAA to the electrode surface, a thin layer of Nafion®, a cation exchange polymer, has been electrodeposited onto cylindrical carbon-fiber microelectrodes. The presence of the Nafion® film was confirmed with environmental scanning electron microscopy and was demonstrated by the diminution of the voltammetric signals for 5-HIAA as well as other common anionic species. The modified microelectrodes also display increased sensitivity to 5-HT, yielding a characteristic cyclic voltammogram that is easily distinguishable from other common electroactive brain species. The thickness of the Nafion® coating and a diffusion coefficient (D) in the film for 5-HT were evaluated by measuring permeation through Nafion®. In vivo, we used physiological, anatomical and pharmacological evidence to validate the signal as 5-HT. Using Nafion®-modified microelectrodes, we present the first endogenous recording of 5-HT in the mammalian brain.

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 Vs −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 towards 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.

Data analysis is an essential tenet of analytical chemistry, extending the possible information obtained from the measurement of chemical phenomena. Chemometric methods have grown considerably in recent years, but their wide use is hindered because some still consider them too complicated. The purpose of this review is to describe a multivariate chemometric method, principal component regression, in a simple manner from the point of view of an analytical chemist, to demonstrate the need for proper quality-control (QC) measures in multivariate analysis and to advocate the use of residuals as a proper QC method.

Fast-scan cyclic voltammetry has been used in a variety of applications and has been shown to be especially useful to monitor chemical fluctuations of neurotransmitters such as dopamine within the mammalian brain. A major limitation of this procedure, however, is the large amplitude of the background current relative to the currents for the solution species of interest. Furthermore, the background tends to drift, and this drift limits the use of digital background subtraction techniques to intervals less than 90 s before distortion of dopamine signals occurs. To minimize the impact of the background, a procedure termed analog background subtraction is reported here. The background is recorded, and its inverse is played back to the current transducer during data acquisition so that it cancels the background in subsequent scans. Background drift still occurs and is recorded, but its magnitude is small compared to the original background. This approach has two advantages. First it allows the use of higher gains in the current transducer, minimizing quantization noise. Second, because the background amplitude is greatly reduced, principal component regression could be used to separate the contributions from drift, dopamine, and pH when appropriate calibrations were performed. We demonstrate the use of this approach with several applications. First, transient dopamine fluctuations were monitored for 15 min in a flowing injection apparatus. Second, evoked release of dopamine was monitored for a similar period in the brain of an anesthetized rat. Third, dopamine was monitored in the brain of freely moving rats over a 30 min interval. By analyzing the fluctuations in each resolved component, we were able to show that cocaine causes significant fluctuations in dopamine concentration in the brain while those for the background and pH remain unchanged from their predrug value.