I am going to start doing perforated patch in mouse brain slices to record action potential activity. I am a bit confused as to what the best perforating agent is. I am not specifically recording any chloride-dependent processes, although the action pottentials themselves could be affacted by GABA input. Do I need to use gramicidin? I am put off by the long wait for stable perforation.
My second issue is how you actually do this in a brain slice. It seems you can not use positive pressure on the pipette (because of the need for tip-filling of agent-freesolution) and I currently rely on this to get through the slice without debris buildup on the pipette and for cleaning the cell surface for seal formation. How can I get around this issue?
Any info would be greatly appreciated!