A variety of extracellular signals are transduced across the cell membrane by the enzyme phosphoinositide-specific phospholipase C-β (PLC-β) coupled with guanine-nucleotide-binding G proteins. There are four isoenzymes of PLC-β, β1-β4, but their functions in vivo are not known. Here we investigate the role of PLC-β1 and PLC-β4 in the brain by generating null mutations in mice: we found that PLCβ1−/− mice developed epilepsy and PLCβ4−/− mice showed ataxia. We determined the molecular basis of these phenotypes and show that PLC-β1 is involved in signal transduction in the cerebral cortex and hippocampus by coupling predominantly to the muscarinic acetylcholine receptor, whereas PLC-β4 works through the metabotropic glutamate receptor in the cerebellum, illustrating how PLC-β isoenzymes are used to generate different functions in the brain.