Here, we uncover a mechanism for regulating the number of active presynaptic GABA_B receptors (GABA_BRs) at nerve terminals, an important determinant of neurotransmitter release. We find that GABA_BRs gain access to axon terminals by lateral diffusion in the membrane. Their relative accumulation is dependent upon agonist activation and the presence of the two distinct sushi domains that are found only in alternatively spliced GABA_BR1a subunits. Following brief activation of NMDA receptors (NMDARs) using glutamate, GABA_BR diffusion is reduced, causing accumulation at presynaptic terminals in a Ca²⁺-dependent manner that involves phosphorylation of GABA_BR2 subunits at Ser783. This signaling cascade indicates how synaptically released glutamate can initiate, via a feedback mechanism, increased levels of presynaptic GABA_BRs that limit further glutamate release and excitotoxicity.