Several laboratories have provided immunohistochemical, molecular biological and electrophysiological evidence that the glutamatergic granule cells of the dentate gyrus can transiently express a GABAergic phenotype during development. Electrophysiological recordings on hippocampal slices obtained during this period have shown that stimulation of the mossy fibres (MFs) provokes simultaneous monosynaptic GABA(A) and glutamate receptor-mediated responses in their target cells,which have the pharmacological and physiological characteristics of MF neurotransmission. This evidence, although strongly supporting the hypothesis that MFs co-release glutamate and GABA, is indirect, as the extracellular stimulation used in slice experiments could activate fibres other than MFs. In this study, we show that selective stimulation of single, identified MF boutons (MFBs) attached to the apical dendrites of dissociated pyramidal cells of developing rats produced synaptic currents mediated by either glutamate receptors only or by both glutamate and GABA(A) receptors. By contrast, stimulation of MFBs of adult rats produced exclusively glutamate receptor-mediated responses. All responses evoked by stimulation of MFBs underwent strong frequency-dependent potentiation and were depressed by the activation of presynaptic metabotropic glutamate receptors. On the other hand, synaptic responses evoked by stimulation of interneuronal boutons located on the soma or on the basal dendrites of the same pyramidal cells were exclusively mediated by GABA(A) receptors, underwent frequency-dependent depression and were unaffected by mGluR agonists.We here demonstrate that the simultaneous glutamatergic and GABAergic responses evoked by MF stimulation in pyramidal cells of CA3 during development have a common origin in the giant MFBs.