ELM 6.1

Question 1

The fastest events that electrophysiology is capable of measuring occur on the following time scale

Answer 1

tens to hundreds of µs

Question 2

You wish to study an ion channel by measuring currents through single channels. You know that for the ion channel you are studying, it is important to keep the cell intact. A suitable mode of the patch-clamp would be:

Answer 2

cell-attached
In the cell-attached configuration, the electrode is sealed onto the outside of the cell and the membrane is left intact. In all other types of patch clamp, the membrane is disrupted in some way.

Question 3

When patch clamping in tissue slices, positive pressure is applied to the patch electrode in order to:

Answer 3

remove debris when “neuron hunting”
The jet of solution from the electrode abrades away the tissue debris to allow a pathway to be cleared to a neurone. A second function is to prevent the electrode from becoming blocked.

Question 4

The patch clamp techniques was invented by:

Answer 4

Sakmann and Neher

Question 5

1. Move electrode onto cell
2. Form a seal on the cell surface.
3. Disrupt area sealed under the electrode
4. Pull electrode away from cell
5. Move electrode out of the bath solution

Move above are manoeuvres that can be used in patch clamping. In order to record in inside-out mode, which of these manoeuvres would be needed?

Answer 5

1, 2, 4, 5
Steps 1 and 2 result in the formation of a “gigaseal” and entry into cell-attached mode. To progress to inside-out mode, the electrode is simply moved away from the cell and then moved out of the bath to ensure that the sticky ends of the membrane stick to the pipette, instead of each other (or disrupt the membrane if it has already re-sealed to form a vesicle). If instead, the area under the electrode is disrupted by further suction, use of a permeabilising agent or a strong electrical “buzz”, whole-cell mode would be achieved. To go into outside out patch mode, further steps are required after whole cell mode has been entered.