hERG tail current

5 posts / 0 new
Last post
zhangxu04
zhangxu04's picture
hERG tail current

hi I have got some questions when recording hERG tail current.

protocol: -90mv for 300ms, +40mv for 2s , -10mv for 2s and -90mv for 300ms

1: I get largest tail current amplitude at -10mv, but it is reported that the largest tail curent amplitude should be induced at about -50 or -40mv

2: sometimes I can not record the tail current while the stable current (at +40mv) is good.

can soembody give me suggestions?

lazy
lazy's picture
(I am using the 1st voltage

(I am using the 1st voltage step to +20 mV instead of +40 mV, and the 2nd voltage step to -50 mV).

1: Did you try to create an I/V curve by changing the 2nd voltage steps? I did it from -120 mV to +20 mV, and I got a nice bell-shape I/V curve. The peak of the bell shape curve is around -50 mV. Try and see how it looks like. You can also check your reversal potential with this method. From that, you will see if your voltage protocol works correctly or not. In my case, the I/V curve crosses with voltage axis around -90 mV which is in good accrdance with the theoretical K-reversal potential.

2: I have sometimes the similar observation. In such case, I change the cell. But mostly, when I rupture the membrane, I know the cell has a good hERG current or not. When the capacity current in whole cell donfig is huge, I have a good hERG current, while the capacity current is small, then mostly I do not have a good hERG current. Probably it depends on the protein expression.

Hope this helpful.  

zhangxu04
zhangxu04's picture
thank you veru much. 

thank you veru much. 

zhangxu04
zhangxu04's picture
I used -40mv for second

I used -40mv for second voltage step previously. but I the tail current is very small. so I changed to -10mv.

now I even can not record the tail current at -10mv. and the tail current is  negative  at -40mv 

I used the EPC10 amplifier. I am not sure where the issue lie

lazy
lazy's picture
If the tail current is

If the tail current is negative (inward direction) at -40 mV, that means the reversal potential is higher than -40 mV (more positive than -40 mV). Just try several different values of  the 2nd voltage step and if you see a change of the polarity of the tail current, your experimental reversal potential is around that value.  You should have approximate value.

Try to calculate the K equilibrium potential using Nernst equation. Just enter the values of your external and internal K concentration into the equation. You will get your thoretical K reversal potential.

If your reversal potentials are significantly different between experimental and theoretical values, as my opinion, there must be a problem in your system (cell, solutions, machine, software, setting the voltage protocol,  etc.).

You are using EPC-10. So, probably you use patchmaster or pulse software for data acquisition. Using those softwares in correct way is important, but finding the correct way is complicate and not easy.