I am doing whole cell LTP in rat hippocampal slices. I am stimulating for 10 minutes prior to patching my cell. I use two monopolar tungsten stimulators, one on the CA3 side of the cell that I patch and the other offset and on the Entorhinal cortex side of the cell. I record a baseline for 5 minutes after patching the cell and then I use pairing to induce LTP. When I get LTP in one pathway I almost always get a run up in the other pathway. Why is this? How can I prevent it from running up?
You need to be sure the pathways are not overlapping. A simple test for this is to set the amplitude of the two pathways to be the same and compare the paired-pulse ratio in pathway 1 and pathway 2 to stimuli in each pathway (1 in pathway 1 and the other in pathway 2) separated by the same interval.
Thank you for your response. I will try that...so I hold the cell in current clamp mode to test the pathways?
Any suggestions on how to set up the stimulators so that the pathways do not overlap? I have tried several configurations, and have taken pictures of what seemed to be good set up that did not run up in the second pathway, and one that caused run up...when I put the pictures side by side they are the exact same configurations.
How do you confirm that your pathways are actually independent? Do you check to make sure there is no short term plasticity between the pathways? It can be quite time consuming, as you need to get a good bit of statistical power.
There are plenty of options here. You could be initiating some form of intrinsic plasticity in the cell. Is the cell's input resistance change? Or other I/V behaviour? You could also be getting heterosynaptic plasticity. There are numerous recorded examples of this, e.g. Lee et al., (2012) J Physiol. If you want to avoid it, I would suggest a less aggressive induction protocol. Depending on your goals, you might want to try and get inputs that are more spatially separated, so try to place one stim electrode towards the apical dendrites, and one near the basals (and of course keep your stim current low). None of the are guarantees of course.