switch from I-clamp to V-clamp

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EGC
EGC's picture
switch from I-clamp to V-clamp

Recently I'm looking at neuron action potentials (AP). I generally get tight seal under V-clamp and switch to I-clamp. When I switch back to V-clamp, however, I sometimes lost the seal. To switch, I turn off ex command knob for I-clamp, then switch mode to I = 0, then switch the mode to V-clamp, turn command for V-clamp on and go to V-clamp protocol.  Is it correct? I also noticed that sometimes I run seal test under I-clamp and the seal seemed lost. What could be the problem? Thanks a lot.

Fraser Moss
Fraser Moss's picture
what amplifier are you using?

what amplifier are you using?
 
The Axopatch 200B can perform seal tests in curent clamp mode by applying current steps, however if you do not configure the software correctly in pClamp then it will think that you are still in voltage clamp mode and you'll just get a flat line.
in the pClamp menus look at the configuration settings and check that the amplifier is telegraphed correctly through the digidata. Also check the "lab bench" to make sure that all your input and output siganls are scaled and set up correctly and that all your protocols are using the correct input and output signals, including those for the seal and capacitance tests.
 
When you are switiching between I-clamp and V-clamp, just make sure that the amplifier and the protocol on your computer are set up correctly.  You could be unintentionally jumping the cells to massively depolarized potentials as you swtich if you dont have the holding potntial set properly before you switch.  Ideally you want the current clamp to be eliciting a given membrane potential and when you switch to voltage clamp it should be help at that same potential when you perform the switch.