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Blitzkrieg 6
Patch Clamp and Single Channel Analysis
15
Biochemistry
10/16/2010

Term
 What is patch clamp used for?
Definition
 Patch clamp is used to measure the voltage and current of single channels.
Term
 How does patch clamp work?
Definition
 You put a pipette/electrode at the edge of a cell. You either leave it there and measure the activity of the channels under it, or you pull it away and excise a piece of cell membrane to record channel activity, or you poke a hole through the patch that is being sucked up a bit by the pipette so that you can change the intracellular contents and measure the macroscopic change of voltage that the rest of the channels will undertake.
Term
 Which types of patch clamp configurations can detect single channel activity?
Definition
 Only whole cell and excised membrane.
Term
 What is the unitary current done by single channels?
Definition
 This means that the currents are of the same size.
Term
 What version of Ohm's law can be used to calculate conductance of a single membrane channel?
Definition
 γ = i / V (where “V” = driving force)Gamma signifies G, but for single channel. i signifies current, but is written as i instead of I because it is a single channel.
Term
 What does it mean that single membrane channels are stochastic? How can you calculate this?
Definition
 They exhibit probabilistic behavior. Probability of being open = total time open/total time recorded
Term
 How can you calculate Veq from single channel data?
Definition
 Vary the Vm experimentally, and measure the Im (when the channel is open), and make a I-V plot
Term
 What would be a characteristic of leak channels?
Definition
 They pass ions equally well in terms of ingress and egress
Term
 What about the slope of leak channels' I-V plot is interesting?
Definition
 g is constant because leak channels don't change conductance, so the slope would be constant
Term
 What would you have to do to Vm to open a voltage gated K+ channel?
Definition
 You'd have to depolarize (less negative)
Term
 Do voltage gated K+ channels let stuff inwards, outwards, or both?
Definition
 Outwards only
Term
 Considering that voltage gated channels only go in one direction, what can you say about their current?
Definition
 It is related heavily to the driving force (proportional to it)
Term
 How could you test for inward rectification with the patch clamp method?
Definition
 Attach the patch clamp. See the beautiful inward rectification. Then excise the patch and put it into solution w/ low Mg, and you lose the rectification (the channels just have current directly proportional to driving force). If you add Mg+, you restore inward rectification activity--meaning that you only get channel activity when the solution is very hyperpolarized (when it is negative).
Term
 How can you test for membrane delimited drugs? Can you add then to the solution?
Definition
 Membrane delimited drugs will only open channels if you put them through the pipette.
Term
 Talk to me more about ruptured pipette method
Definition
 After you rupture, most of the cell's 2nd messengers will go away. Then you can add your own experimentally and test them out. However, the stuff that is attached to the membrane ("membrane delimited shit") will still usually be functional. yay, this is so relevant. maybe I should jerk it now out of pure excitement for this beautiful science because med school is so fun and its saturday night and I'm "learning" about this shit
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