Automated Targeting of Cells to Electrochemical Electrodes Using a Surface Chemistry Approach for the Measurement of Quantal Exocytosis

Abstract

Here we describe a method to fabricate a multi-channel high-throughput microchip device for measurement of quantal transmitter release from individual cells. Instead of bringing carbon-fiber electrodes to cells, the device uses a surface chemistry approach to bring cells to an array of electrochemical microelectrodes. The microelectrodes are small and “cytophilic” in order to promote adhesion of a single cell whereas all other areas of the chip are covered with a thin “cytophobic” film to block cell attachement and facilitate movement of cells to electrodes. This cytophobic film also insulates unused areas of the conductive film, thus the alignment of cell docking sites to working electrodes is automatic. Amperometric spikes resulting from single-granule fusion events were recorded on the device and had amplitudes and kinetics similar to those measured using carbon-fiber microelectrodes. Use of this device will increase the pace of basic neuroscience research and may also find applications in drug discovery or validation.

Parallel recording of neurotransmitters release from chromaffin cells using a 10×10 CMOS IC potentiostat array with on-chip working electrodes

“…These include platinum (Dias et al 2002; Berberian et al 2008), pyrolyzed photoresist (Zachek et al 2009), carbon (Zhang et al 2008; Lin et al 2012), ITO (Kisler et al 2012), nitrogen-doped diamond-like carbon (DLC:N) (Barizuddin et al 2010), boron-doped nanocrystalline diamond (Carabelli et al 2010) and organic polymers (Yang et al 2011). The limitation of such microelectrode arrays is that for parallel recordings each electrode needs to be connected to an external amplifier, and such devices are therefore not scalable.…”

Quantification of noise sources for amperometric measurement of quantal exocytosis using microelectrodes

“…Therefore carbon-fiber electrodes are pressed gently to the surface of the cell whereas it is desirable for cells to tightly adhere to planar electrodes so they do not wash away upon solution exchange 13–14 . Gleixner and Fromherz 31 measure the sheet resistance (R sheet ) between adherent cells and a fibronectin-coated silica substrate to be ~10 MΩ in physiological bath solution.…”

“…These two materials also offer optical transparency to easily image cells over the electrodes 9–10, 21 and DLC:N has the further advantage that it promotes cell adhesion to electrodes 13–14 .…”

Electroporation followed by electrochemical measurement of quantal transmitter release from single cells using a patterned microelectrode

“…With existing systems the most common way of stimulating exocytosis is by perfusion of cells with a secratogogue 7, 11 leading to depolarization of the cell membrane, opening of voltage-gated Ca 2+ -channels, Ca 2+ influx and Ca 2+ -triggered exocytosis. Perfusion systems, however, often stimulate multiple cells and do not allow precise timing on the stimulus.…”