Cyclic Voltammetry for the Detection of Dopamine in vivo

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Dopamine

Structure of Dopamine. 4-(2-aminoethyl) benzene-1,2-diol

Neurotransmitter small molecule chemical messenger Important for motor and cognitive functions Deficts in dopamine levels cause Parkinson Disease Regulates reward Dopamine increases after drugs of abuse like cocaine

Dopaminergic Neurons

Dopamine is synthesized in dopaminergic neurons and packaged into membrane bound vesicles. Electrical action potential initiates the release of dopamine. Dopamine vesicles undergo exocytosis. Spills out into the extracellular space Can be detected

The dompaminergic neuron can release dopamine into the extracellular space where its contents can be detected by target neurons. (Venton & Wightman 2003)

Detection of Dopamine

• Exocytosis of dopamine from vesicle occurs on a millisecond time scale
• Sensor must be fast, sensitive, and selective since dopamine concentrations are low
• Fast scan cyclic voltametry is the dominant electrochemical technique used

Volametry

• Electrochemical technique in which the current (I) is measured as a function of voltage (Eapp)

There are several kinds:

• Linear ccan voltametry (Polarography)
• Differential pulse polarography
• Square-wave voltametry
• Cyclic voltametry

Cyclic Voltametry

• A potential is applied between the reference electrode and the working electrode.
• The electrode potential ramps linearly at the experimental scan rate (V/s)
• The current is measured between the working electrode and auxillary electrode
• Data is plotted as Current (I) vs. Potential(E). 

• Forward scan produces a current peak for the analystes that have been reduced in scanned potential range.
• The direction of the potential is reversed at the end of the each scan in cyclic voltametry
• Current increases as the potential reached the reduction potential of the anlayte
• Current decreases as the analyte is depleted in the redox reaction

Carbon Fiber Microelectrodes

http://www.clinchem.org/cgi/content/full/49/10/1763 Carbon Fiber Microelectrode

Carbon Fiber Microelectrodes (CFME) are electrodes in which a carbon fibers serve as the electroactive area Advantages: Carbon fibers are biological compatible to cells Small size (less than 10um in diameters) allows for implantation in vivo Commonly used with cyclic voltametry

Fast Scan Cyclic Voltametry



Scan electrode from a holding potential to a switching potential and back at a
high scan rate.  Repeat these scans every 100 ms.  



As potential is ramped up, dopamine is oxidized to dopamine-o-quinone.
As potential is ramped down, dopamine-o-quinone reduced back to dopamine.

Cyclic Voltammogram Fast scan rate cause a large background charging current due to charging the double layer. When dopamine is added (red line), the differences are small.

Background-subtracted cyclic voltammogram of dopamine The background current is stable and can be subtracted out to obtain a background- subtracted cyclic voltammogram for dopamine. The position of the peaks help identify the molecule being detected.

Data can also be depicted as a color plot to show many CVs over time.
Current is in color and shows when dopamine is present. 

Experimental Results

• Carbon-fiber microelectrode implanted in nucleus accumbens
• Nucleus accumbens regulates reward
• Electrically stimulate cell bodies in ventral tegmental area

Electrically-Stimulated release in a rat

(Addy, et al.  2010)

• Size of dopamine evoked depends on frequency of stimulation
•  Color plots look like dopamine

Spontaneous dopamine transients in a rat


Spontaneous dopamine transients in a rat after cocaine


Cocaine increases the concentration and the length of time for dopamine signaling (Aragona et. al. 2008).

Advantages

• Allows for the product of the electron transfer reaction that occurred in the forward scan to be probed again in the reverse scan

• Can be used to determine of formal redox potentials, detect of chemical reactions that occur before or after a electrochemical reaction, and evaluate electron transfer kinetics

References

Huffman, M.L. and Venton, B.J.  "Carbonfiber Microelectrodes for in vivo applications." Analyst (2009): 18-24. 

Venton, B. J. and Wightman, M. R. "Psychoanalytical Chemistry: Dopamine and Behavior."  Analytical Chemistry (2003): 414-421.