Presentation on theme: "Cyclic Voltametry for the Detection of Dopamine in vivo"— Presentation transcript:
1 Cyclic Voltametry for the Detection of Dopamine in vivo
2 Dopamine Neurotransmitter Important for motor and cognitive functions small molecule chemical messengerImportant for motor and cognitive functionsDeficts in dopamine levels cause Parkinson DiseaseRegulates rewardDopamine increases after drugs of abuse like cocaineVenton, B. J. and Wightman, M. R. “Psychoanalytical Chemistry: Dopamine and Behavior.” Analytical Chemistry (2003):Structure of Dopamine.4-(2-aminoethyl)benzene-1,2-diol
3 Dopaminergic NeuronsDopamine is synthesized in dopaminergic neurons and packaged into membrane bound vesiclesElectrical action potential initiates the release of dopamineDopamine vesicles undergo exocytosisSpills out into the extracellular spaceCan be detectedVenton, B. J. and Wightman, M. R. “Psychoanalytical Chemistry: Dopamine and Behavior.” Analytical Chemistry (2003):The dompaminergic neuron can release dopamine into the extracellular space where its contents can be detected by target neurons.
4 Detection of DopamineExocytosis of dopamine from vesicle occurs on a millisecond time scaleSensor must be fast, sensitive, and selective since dopamine concentrations are lowFast scan cyclic voltametry is the dominant electrochemical technique usedVenton, B. J. and Wightman, M. R. “Psychoanalytical Chemistry: Dopamine and Behavior.” Analytical Chemistry (2003):DopamineDopamine-o-quinone+ 2 H+
5 Carbon Fiber Microelectrodes Carbon Fiber Microelectrodes (CFME) are electrodes in which a carbon fibers serve as the electroactive areaAdvantages:Carbon fibers are biological compatible to cellsSmall size (less than 10um in diameters) allows for implantation in vivoCommonly used with cyclic voltametryHuffman, M.L. and Venton, B.J. “Carbon fiber Microelectrodes for in vivo applications.” Analyst (2009):Image:Carbon Fiber Microelectrode
6 VoltammetryElectrochemical technique in which the current (I) is measured as a function of voltage (Eapp).There are several kinds:Linear Scan voltametry (Polarography)Differential pulse polarographySquare-wave voltametryCyclic Voltametry
7 Fast-scan Cyclic Voltametry 400 V/s-0.4 V100 ms8.5 msScan electrode from a holding potential to a switching potential and back at ahigh scan rate. Repeat these scans every 100 ms.DopamineDopamine-o-quinone+ 2 H+As potential is ramped up, dopamine is oxidized to dopamine-o-quinone.As potential is ramped down, dopamine-o-quinone reduced back to dopamine.
8 Fast-Scan Cyclic Voltametry Cyclic voltammogramsBackground-subtractedcyclic voltammogram of dopamineNo dopamineDopamine presentoxidationreductionFast scan rate cause a large backgroundcharging current due to chargingthe double layer.When dopamine is added (red line),the differences are small.The background current is stable and canbe subtracted out to obtain a background-subtracted cyclic voltammogram for dopamine.The position of the peaks help identify themolecule being detected.
9 Fast-scan cyclic voltametry -53i, nAE vs Ag/AgCldopaminereductiondopamineoxidationdopamine presentData can also be depicted as a color plot to show many CVs over time.Current is in color and shows when dopamine is present.
10 Results Electrically-Stimulated release in a rat Addy, N. A. et. al. "Sensitization of Rapid Dopamine Signaling in the Nucleus Accumbens Core and Shell After Repeated Cocaine in Rats." Journal of Neurophysiology (2010). 104:Carbon-fiber microelectrode implanted in nucleus accumbensNucleus accumbens regulates rewardElectrically stimulate cell bodies in ventral tegmental areaSize of dopamine evoked depends on frequency of stimulationColor plots look like dopamine
11 Results Spontaneous dopamine transients in a rat Spontaneous dopamine transients in a rat after cocaineAragona, J.B, et. al. “Preferential Enhancement of Dopamine Transmission within the Nucleus Accumbens Shell by Cocaine is Attributed to a Direct Increase in Phasic Dopamine Release Events.” Journal of Neuroscience (2008). 28:Cocaine increases the concentration and the length of time for dopamine signaling.
12 Advantages Allows real-time detection of dopamine in behaving animals Can determine during what behaviors dopamine is releasedCan determine how drugs affect dopamine signaling in the brain