Phosphoinositides act as signaling molecules in the regulation of many cellular processes. We are interested in the regulation of ion channels such as KCNQ K⁺ channels by phosphatidylinositol 4,5bisphosphate (PIP₂) and related phosphoinositide species in their neuronal environment in order to understand the impact of such phospholipids on neuronal excitability. This requires examination of the dynamics of lipid concentrations in time and space. Recently, genetically encoded fluorescent probes for membrane lipids have become available. For example, GFP fusion-proteins of the PH domain of PLC-[delta]1 and of the C1a-domain of PKC bind to PIP₂ and diacylglycerol, respectively. We show that these probes are particularly potent tools in the investigation of lipid dynamics when combined with TIRF-microscopy, because TIRF selectively excites the plasma membrane-bound lipid probes that report the concentration of their specific lipid ligand. We expressed CFP/YFP-labeled PLC[delta]1-PH and PKC-C1a domains in CHO cells or primary neuronal cultures and induced activation of phospholipase C (PLC) via Gq-coupled receptors. Using matched excitation and emission wavelengths, the time course of concentration changes of PIP₂ and DAG upon PLC activation could be recorded directly and spatially resolved as a change in the TIRF fluorescence emission signals. Supported by DFG through grant OL240/2-1.