The maintenance of synaptic functions is essential for neuronal information processing, but cellular mechanisms that maintain synapses in the adult brain are not well understood. Here, we report an activity-dependent maintenance mechanism of parallel fiber (PF)-Purkinje cell (PC) synapses in the cerebellum. When postsynaptic metabotropic glutamate receptor (mGluR) or inositol 1,4,5-trisphosphate (IP(3)) signaling was chronically inhibited in vivo, PF-PC synaptic strength decreased because of a decreased transmitter release probability. The same effects were observed when PF activity was inhibited in vivo by the suppression of NMDA receptor-mediated inputs to granule cells. PF-PC synaptic strength similarly decreased after the in vivo application of an antibody against brain-derived neurotrophic factor (BDNF). Furthermore, the weakening of synaptic connection caused by the blockade of mGluR-IP(3) signaling was reversed by the in vivo application of BDNF. These results indicate that a signaling cascade comprising PF activity, postsynaptic mGluR-IP(3) signaling and subsequent BDNF signaling maintains presynaptic functions in the mature cerebellum.