14.11:
Patch Clamp
The patch-clamp technique is an electrophysiological tool to study the ion channel behavior of excitable cells like neurons and muscle fibers.
The patch-clamp setup includes a glass micropipette placed against a small area of cell membrane containing one or more ion channels. It consists of an electrolyte solution and a chlorinated silver electrode to measure voltages and currents.
When a small suction is applied, the ‘membrane patch' gets tightly sealed against the pipette tip.
This omega-shaped area of the membrane, called the gigaohm seal, is electrically isolated, ensuring the ions flowing through the channel pass only into the pipette.
The recording electrode within the micropipette is connected to a highly sensitive amplifier that amplifies current or voltage fluctuations resulting from the movement of ions through the ion channels, allowing for the measurement of generated electrical signals.
In the ‘voltage clamp', the voltage is set at a particular value, and the resulting changes in current flow are measured. Alternatively, the current can be clamped to a set value, and the voltage variation across the membrane can be measured.
14.11:
Patch Clamp
Many fundamental cell functions such as muscle contraction and nerve transmission rely on the electrical signals produced by the movement of positively and negatively charged ions across the cell membrane. One competent method to record current flowing across the whole cell or single ion channel is the patch-clamp technique.
In this method, a glass micropipette containing electrolyte solution is tightly sealed against a small portion of the cell membrane. As a result, a patch of the cell membrane is electrically isolated, ensuring that the ions moving across the channels flow only into the micropipette. Further, this tight sealing of the cell membrane prevents ions from escaping into the bath solution in which the cell is suspended.
Different Patch-clamp Methods:
Depending upon the research interest, several variations of the patch-clamp method can be used for measuring the cell's biophysical properties. For example, researchers can clamp or control the membrane voltage and measure the current passing through it. Alternatively, they can clamp the current and measure any variation in the membrane voltage.
In cell-attached mode, the membrane patch containing a single or a few ion channels remains intact. As a result, the current flowing through the membrane patch alone can be measured. In contrast, the whole-cell method involves disruption of the membrane patch by briefly applying strong suction. Consequently, the interior of the pipette becomes continuous with the cytoplasm. This mode enables the measurement of electrical current and voltage from the entire cell.
Another patch-clamp method requires gentle retraction of the attached pipette. As a result, the membrane patch is excised without affecting the tight seal. In this inside-out configuration, the intracellular portion of the membrane is exposed to the bath solution, allowing the study of intracellular factors affecting the channel functions.
Suggested Reading
- Obergrussberger, Alison, Tom A. Goetze, Nina Brinkwirth, Nadine Becker, Søren Friis, Markus Rapedius, Claudia Haarmann et al. "An update on the advancing high-throughput screening techniques for patch clamp-based ion channel screens: implications for drug discovery." Expert opinion on drug discovery 13, no. 3 (2018): 269-277
- Stoelzle, Sonja, Alison Obergrussberger, Andrea Bruggemann, Claudia Haarmann, Michael George, Ralf Franz Kettenhofen, and Niels Fertig. "State-of-the-art automated patch clamp devices: heat activation, action potentials, and high throughput in ion-channel screening." Frontiers in pharmacology 2 (2011): 76