Transcript Cyclic Voltammetry 1..

Cyclic Voltammetry for the
Detection of Dopamine in vivo
Dopamine
• 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
Structure of Dopamine.
4-(2-aminoethyl)
benzene-1,2-diol
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
excytos dopamine vesicles into
the extracellular space where its
contents can be detected by
target neurons.
Detection of Dopamine
• Exocytosis of dopamine from vesicle occurs on
a milisecond time scale
• Sensor must be fast, sensitive, and selective
since dopamine concentrations are low
• Fast scan cyclic voltammetry 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 Scan voltammetry (Polarography)
– Differential pulse polarography
– Square-wave voltammetry
– Cyclic Voltametry
Three Electrode Set Up
1. Working Electrode: Redox
reaction
• limited reaction surface
area in order to limit
current flow
2. Auxillary Electrode: Consists of
an inert material (Hg, Pt)
• does not participate in
redox reaction
• completes electrical circuit
3. Reference Electrode: constant
potential reference (SCE)
• establishes electrical
potential
Modern Three Electrode Set Up
Cyclic Voltammetry
•A potential is applied between the reference
electrode and the working electrode.
•The electrode potential ramps linearly at the
experimental scan rate (V/s) and 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
analytes that have been reduced in scanned
potential range.
Cyclic Voltammetry
• The direction of the potential is reversed
at the end of the each scan in cyclic
voltammetry
• Current increases as the potential
reached the reduction potential of the
anlayte
• Current decreases as the analyte is
depleted in the redox reaction
Experimental Results
•In a recent study by Venton and Wightman (2003), rats were
implanted the carbon-fiber microelectrode electrode at thieir
nucleus accumbens, a part of the brain associated with reward
among other functions.
•A biopolar stimulating electrode was implanted in the substantia
nigraventra until its electrically evoked release was observable at the
carbon-fiber microelectrode.
•After collection, background subtraction and digital filtering were
performed.
•To remove the background and charging currents, the collected set
of cyclic voltammograms was subtracted from the average of the
first 10.
•The peak oxidation potential for dopamine is 0.65 V, which was the
condition use.
Results
Electrical Stimulation
•Subtracted voltamogram taken at
the end of stimulation (above)
resembles one recorded in a solution
containing dopamine.
•Subtracted voltamogram taken five sconds
after stimulation ended (below) is due to
changes in the residual current caused by a
basic pH change that originates from a change
in local blood.
Results
Cocaine Influsion
22s after infusion
52s after infusion
62s after infusion
•Subject rats were infused with saline (i.v., 20 s per infusion), followed by 0.3
mgkg, 1.0 mgkg, and 3.0 mgkg cocaine infusions at 15-min intervals.
•Subtracted cyclic voltammograms recorded at various times after the infusion
as seen above.
•The cyclic voltammograms represented with the dashed lines are those
obtained with electrical stimulation.
Results
• The similarities between subtracted cyclic
voltamograms of electrical stimulation and
cocaine infusion indicate a similar change in
electroactive species after both events.
• While this change in electroactive species is a
composite of multiple substances, the two
major contributors are dopamine and pH,
determined through regressional analysis.
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
Carbon Fiber Microelectrodes
• 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
voltammetry
Carbon Fiber Microelectrode