How to isolate GABAa currents?

Thomas Gabriel its_me72 at hotmail.com
Mon Apr 4 08:51:02 EST 2005


Hi!
Does anybody know an easy way to isolate GABAa currents? I want to do
a test if my bicuculine works properly. But to my surprise i found
this test to be kinda tricky. First i tried to block all glutamatergic
currents with DNQX/APV and fixed the Em at ~ -85 mV via injetion of
negative DC current (since I am using a Axoclamp 2B amplifier i prefer
current-clamp mode). I expected the remaining PSPs to be gabaergic and
thought that i should be able to block them with bicuculine. But with
application of DNQX/APV all synaptic activity died and my signal
became a flatline. Neither at normal Em nor at hyperpolarised Em,
there is nothing that may be interpreted as gabaergic potential.
I left out the APV and added a little NMDA (5 µM, for a slightly
compensation of the blocked synaptic drive via AMPA) and selective
GABAa-agonist isoguvacine (10 µM). But still I can't manage to
identify gabaergic events. The Em-signal is stil a flat line with no
sign of activity.

Does anybody know a simple experimental procedure to isolate GABAa
currents?

Every comment is appreciated!
Thanks,
Thomas



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