The physical dimensions of an ion channel pore can be estimated in patch-clamp experiments using various charged and noncharged molecular probes. Measuring the permeability to organic ions of different size and shape yields an estimate of the size of the narrowest part of the selectivity fi ...
When a patch pipette approaches a cellular surface at distances comparable to the size of the tip opening, the monitored pipette current drops before the pipette touches the membrane. Fine nano-tipped glass electrodes can effectively function as “spherical sensors” that can “roll” over s ...
If gene expression could be analyzed after electrophysiological recording of a single cell, it would enhance our molecular understanding of neuronal physiology and help to interpret the results obtained from patch-clamp recordings. For instance, after completing a recording from a ...
This chapter describes practical techniques for electrophysiological experiments in the heterologous expression systems using mammalian cell cultures and Xenopus laevis oocytes. These two experimental systems are the most commonly used for expression of ion channels and t ...
Since the cloning of the first thermosensitive TRP channel, a capsaicin receptor TRPV1, in 1997, nine thermosensitive TRP channels have been identified. Thermosensitive TRP channels can be used to detect local temperatures by expression in HEK293 cells as a kind of biosensor, as described ...
Cells or membranes with ligand-gated ion channels endogenously expressed in cultured cells or heterologously expressed in HEK293 cells can be used as biosensors with a patch-clamp technique to detect small amounts of locally released bioactive substances that are known to activate i ...
Patch-clamping is an essential technique in the field of ion channel drug discovery and safety pharmacology. Conventional patch-clamp methods provide high data quality and can be applied to a wide variety of cells, but the methods are laborious and time-consuming. Automated patch-clamp ...
A historical overview describes how classic electrophysiological techniques, such as membrane potential measurement, seal formation, and the voltage clamp, have evolved into modern patch-clamp techniques. We show that old ideas from seminal papers on ion channels (i.e., gating, in ...
When the second patch-clamp book of Sakmann and Neher appeared in 1995 (Sakmann and Neher, Single-channel recording, 2nd edn. Plenum Press, New York, 1995), the molecular nature of plant ion channels was still in its infancy. Since 1995, various members of the Shaker-, Two-Pore-, and KCO-type potass ...
The standard giga-seal patch-clamp techniques opened the way to monitor ion-channel activity in the native organelle membrane in situ. One has to have an organelle preparation, covered with the intact membrane carrying rightly oriented channel proteins and suitable for proper patch- ...
Not only electrical conductance but also electrical capacitance of the cell membrane can be measured by patch-clamp techniques. Exocytotic events can be detected by recording changes in membrane capacitance. The membrane capacitance, which reflects the surface area of the plasma me ...
The planar lipid bilayer method is another way to examine channel molecules functionally at the single-molecule level. In contrast to patch-clamping, channel molecules are isolated from various biological resources and are reconstituted into an artificial membrane that has a defi ...
The piette perfusion technique is a version of patch-clamp techniques and provides a greater intracellular access during electrophysiological recordings. This internal perfusion technique offers the intracellular change of not only ions but also substances with higher mole ...
The giant patch method was first developed in 1989. The major characteristic is the use of a pipette with a large tip diameter, which dramatically broadened the applications of the excised patch. The giant patch method enables (1) the current recording of transporters/channels with slow turn ...
Na+/K+ pump, Na+/Ca2+ exchanger, glutamate transporter and other transporters for dopamine, norepinephrine, GABA, and others, are all electrogenic. These transporters or exchanger transport Na+ ions, with or without transmitters, in or out of the cells, while K+ or Ca2+, sometimes with anion ...
The advent of two-photon microscopy has enabled us to visualize individual neurons in the intact brain. This technique, used in combination with whole-cell patch-clamp recordings, has facilitated targeted intracellular recording from particular neurons of interest. This chap ...
The methods for in vivo blind patch-clamp recording from the spinal cord, brain stem, and cortex neurons are described herein, including the (1) technique for in vivo preparations, (2) recordings of action potential and synaptic potentials under current-clamp conditions and excitatory ...
Dendritic patch-clamp recordings have recently become popular as an electrophysiological technique to investigate the functional properties of ion channels in neuronal dendrites. During the past decade, experimentalists have pioneered recording from dendrites of neur ...
Over a period spanning some 15 yr, we have exploited microinjected oocytes and embryos of Xenopus laevis to clone and characterize the human genes encoding AChE and BuChE and their protein products. Subsequently, we tapped into the enormous versatility of Xenopus to continue into the realm of t ...
A DNA sequence encoding the brain and muscle form of human AChE (AChE-T) was constructed in our laboratory from cloned cDNA and genomic sequences, and tentatively identified by its homology to known ChEs (Soreq et al., 1990). This putative AChE-coding sequence, bearing the 3′ alternative exon E6, w ...
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