Dopamine transport into a single cell in a picoliter vial.
The analysis of chemical events in small volumes requires careful manipulation of samples and sensitive detection methods. Here, we describe the measurement of the neurotransmitter dopamine in a picoliter vial with electrochemical techniques. The vials were fabricated from fused-silica capillaries that provided a transparent container suitable for the observation and manipulation of a biological cell, sample solutions, and electrodes. Evaporation of the sample was prevented with a mineral oil layer, allowing for experiments lasting for several minutes. The small volume of these vials (100-200 pL) allows rapid mixing of all of the solution reagents. Similarly, the small volume allows exhaustive electrolysis of the vial contents with a 3-microm radius, disk-shaped carbon fiber microelectrode within 60 s. Fast-scan cyclic voltammetry at carbon fiber microelectrodes was used to monitor the concentration of analyte in the vial without depleting its contents. The concentration of dopamine introduced by pneumatic injection remained stable when sampled by cyclic voltammetry, and no evidence for adsorption to the walls was observed. However, when the vial contained a single HEK-293 cell transfected to express the dopamine transporter, the dopamine concentration decreased with time at a rate consistent with the uptake kinetics mediated by the transporter located on the cell membrane.
Troyer
Kevin P
KP
University of North Carolina at Chapel Hill, Department of Chemistry, 27599, USA.
Wightman
R Mark
RM
D002460:Cell Line; D004298:Dopamine / Q000737:chemistry* / Q000378:metabolism; D004563:Electrochemistry; D006801:Humans; D007202:Indicators and Reagents; D008839:Microelectrodes; D036103:Nanotechnology
D007202:Indicators and Reagents; D004298:Dopamine