Glutamatergic transmission is mediated by ionotropic receptors that directly gate cationic channels and metabotropic receptors that are coupled to second messenger generating systems and to ionic channels via heterotrimeric guanine-nucleotide binding- (G) proteins. This distinction cannot be made for the ionotropic receptor subclass activated by alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA), which has been shown to be physically associated with the alpha-subunit of Gi1 protein and activates this G-protein. Here, we report that, in addition to a Ca2+ influx, AMPA induces the mobilization of Ca2+ from the mitochondrial pool by reversing the mitochondrial Na+/Ca2+ exchanger in mouse neurons in primary culture. Both processes required the activation of tetrodotoxin-sensitive Na+ channels. AMPA receptor activation modified the gating properties of the Na+ channel, independently of the AMPA current, suggesting a G-protein-mediated process. Indeed, co-immunoprecipitation experiments indicated that AMPA receptor activation induced the association of Gbeta with the alpha-subunit of the Na+ channel. These results suggest that, in addition to its ionic channel function, the AMPA receptor is coupled to Na+ channels through G-proteins and that this novel metabotropic function is involved in the control of neuronal excitability.