Moving from the evidence that activation of type 4 metabotropic glutamate (mGlu4) receptors inhibits proliferation and promotes differentiation of cerebellar granule cell neuroprogenitors, we examined the expression and function of mGlu4 receptors in medulloblastoma cells. mGlu4 receptors were expressed in 46 of 60 human medulloblastoma samples. Expression varied in relation to the histotype (nodular desmoplastic>classic>>large-cell anaplastic) and was inversely related to tumor severity, spreading, and recurrence. mGlu4 receptors were also found in D283med, D341med, and DAOY medulloblastoma cell lines, where receptor activation with the selective enhancer PHCCC inhibited adenylyl cyclase and the phosphatidylinositol-3-kinase pathway without affecting the mitogen-activated protein kinase, Sonic Hedgehog, and Wnt pathways. Interestingly, mGlu4 receptor activation reduced DNA synthesis and cell proliferation in all three cell lines. This effect was abrogated by the phosphatidylinositol-3-kinase inhibitor LY294002 [2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one]. In in vivo experiments, repeated subcutaneous injections of N-phenyl-7-(hydroxyimino)cyclopropa[b]chromen-1a-carboxamide (PHCCC) reduced the growth of D283med and DAOY cell xenografts in nude mice. More remarkably, subcutaneous or intracranial injections of PHCCC during the first week of life prevented the development of medulloblastomas in mice lacking one Patched-1 allele and x-irradiated 1 d after birth. These data suggest that mGlu4 receptor enhancers are promising drugs for the treatment of medulloblastomas.