I've recently ran into trouble recording mIPSCs in a synaptically active in vitro neuron population. In all cases, I've been using whole-cell configuration and performing -70 VC recordings. For mEPSCs (in the presence of TTX and bicuculline), I use a 140 mM K-Gluconate based buffer and get beautiful recordings with mEPSC rates around 6-10 Hz. However, when recording mIPSCs (in the presence of TTX, CNQX, AP5) using either a 140 mM Cs-Cl based buffer or 140 mM KCl based buffer, I've recently started running into issues. Immediately after break-in, my holding current drops precipitously to around -500 pA. This leads to a horribly messy recording in which I can't even pick out events amongst the noise. Previously (as of a few weeks ago), I was reliably able to record mIPSCs with a frequency of around 1-3 Hz. Now, even if I am able to occasionally get a somewhat stable seal, the noise is terrible. I've tried making up new intracellular and extracellular buffers 3 times now. Sterile filtered just prior to use as always. The osmolarity seems right, 290. The pH seems right. I've tried new electrodes. Same result. Tried a whole other rig. Same result. :( The neurons look horrible after the patch is on. Has anyone ever ran into this problem? Any tips from any expert patch-clampers out there on getting good mIPSC recordings? I haven't tried using Cs-MeSO4 and recording at 0 mV yet, only because I previously have gotten such nice recordings with Cs-Cl at -70 mV. At this point I'm stuck, and seriously hating mIPSCs!