There is now substantial evidence, from single-cell imaging, that complex patterns of release from Ca(2+) stores play an important role in regulating synaptic efficacy and plasticity. Moreover, the major mechanism of store release depends on the generation of inositol 1,4,5-trisphosphate [Ins(1,4,5)P(3)] through the action of phospholipase(s) C on phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P(2)], and several neurotransmitters can enhance receptor-mediated activation of this enzyme. The recent development of techniques to image real-time changes in PtdIns(4,5)P(2) hydrolysis according to generation of Ins(1,4,5)P(3) and diacylglycerol in single cells has significantly advanced our ability to investigate these signalling pathways, particularly in relation to single-cell Ca(2+) signals. This article reviews these new approaches and how they have provided novel insights into mechanisms underlying spatio-temporal Ca(2+) signals and phospholipase C activation in neurons.