We have investigated the presence of histamine H(3) receptors (H(3)Rs) on rat thalamic isolated nerve terminals (synaptosomes) and the effect of their activation on glutamate and GABA release. N-alpha-[methyl-(3)H]histamine ([(3)H]-NMHA) bound specifically to synaptosomal membranes with dissociation constant (K(d)) 0.78+/-0.20 nM and maximum binding (B(max)) 141+/-12fmol/mg protein. Inhibition of [(3)H]-NMHA binding by histamine and the H(3)R agonist immepip fit better to a two-site model, whereas for the H(3)R antagonist clobenpropit the best fit was to the one-site model. GTPgammaS (30 microM) decreased [(3)H]-NMHA binding by 55+/-4% and made the histamine inhibition fit better to the one-site model. Immepip (30 nM) induced a modest, but significant increase (113+/-2% of basal) in [(35)S]-GTPgammaS binding to synaptosomal membranes, an effect prevented by clobenpropit (1 microM) and by pre-treatment with pertussis toxin. In thalamus synaptosomes depolarisation-induced, Ca(2+)-dependent glutamate release was inhibited by histamine (1 microM, 25+/-4% inhibition) and immepip (30 nM, 38+/-5% reduction). These effects were reversed by clobenpropit (1microM). Conversely, immepip (up to 1 microM) had no effect on depolarisation-evoked [(3)H]-GABA release. Extracellular synaptic responses were recorded in the thalamus ventrobasal complex by stimulating corticothalamic afferents. H(3)R activation reduced by 38+/-7% the glutamate receptor-mediated field potentials (FPs), but increased the FP2/FP1 ratio (from 0.86+/-0.03 to 1.38+/-0.05) in a paired-pulse paradigm. Taken together, our results confirm the presence of H(3)Rs on thalamic nerve terminals and show that their activation modulates pre-synaptically glutamatergic, but not GABAergic neurotransmission.