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

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

Content Provider	PubMed Central
Author	Ghosh, Jaya Liu, Xin Gillis, Kevin D.
Abstract	An electrochemical microelectrode located immediately adjacent to a single neuroendocrine cell can record spikes of amperometric current that result from exocytosis of oxidizable transmitter from individual vesicles, i.e., quantal exocytosis. Here, we report the development of an efficient method where the same electrochemical microelectrode is used to electropermeabilize an adjacent chromaffin cell and then measure the consequent quantal catecholamine release using amperometry. Trains of voltage pulses, 5–7 V in amplitude and 0.1–0.2 ms in duration, were used to reliably trigger release from cells using gold electrodes. Amperometric spikes induced by electropermeabilization had similar areas, peak heights and durations as amperometric spikes elicited by depolarizing high K+ solutions, therefore release occurs from individual secretory granules. Uptake of trypan blue stain into cells demonstrated that the plasma membrane is permeabilized by the voltage stimulus. Voltage pulses did not degrade the electrochemical sensitivity of the electrodes assayed using a test analyte. Surprisingly, robust quantal release was elicited upon electroporation in the absence of Ca2+ in the bath solution (0 Ca2+/5 mM EGTA). In contrast, electropermeabilization-induced transmitter release required Cl− in the bath solution in that bracketed experiments demonstrated a steep dependence of the rate of electropermeabilization-induced transmitter release on [Cl−] between 2 and 32 mM. Using the same electrochemical electrode to electroporate and record quantal release of catecholamines from an individual chromaffin cell allows precise timing of the stimulus, stimulation of a single cell at a time, and can be used to load membrane-impermeant substances into a cell.
Related Links	http://dx.doi.org/10.1039/c3lc41324a
Ending Page	2090
Page Count	8
Starting Page	2083
File Format	PDF
ISSN	14730189
e-ISSN	14730189
Journal	Lab on a chip
Issue Number	11
Volume Number	13
Language	English
Publisher Date	2013-06-07
Access Restriction	Open
Subject Keyword	Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Chemistry Nanoscience and Nanotechnology Biochemistry Bioengineering Biomedical Engineering

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