Vagus nerve stimulation (VNS) is an adjunctive treatment for resistant epilepsy and depression. Electrophysiological recordings in the rat brain have already shown that chronic VNS increases norepinephrine (NE) neuronal firing activity and, subsequently, that of serotonin (5-HT) neurons through an activation of their excitatory α1-adrenoceptors. Long-term VNS was shown to increase the tonic activation of post-synaptic 5-HT1A receptors in the hippocampus. This study was aimed at examining the effect of VNS on extracellular 5-HT, NE and dopamine (DA) levels in different brain areas using in vivo microdialysis, on NE transmission in the hippocampus, and DA neuronal firing activity using electrophysiology. Rats were implanted with a VNS device and stimulated for 14 d with standard parameters used in treatment-resistant depression (0.25 mA, 20 Hz, 500 μs, 30 s on-5 min off). The results of the present study revealed that 2-wk VNS significantly increased extracellular NE levels in the prefrontal cortex and the hippocampus and enhanced the tonic activation of post-synaptic α2-adrenoceptors on pyramidal neurons. The electrophysiological experiments revealed a significant decrease in ventral tegmental area DA neuronal firing rate after long-term VNS; extracellular DA levels were nevertheless increased in the prefrontal cortex and nucleus accumbens. Chronic VNS significantly increased extracellular 5-HT levels in the dorsal raphe but not in the hippocampus and prefrontal cortex. In conclusion, the effect of VNS in increasing the transmission of monoaminergic systems targeted in the treatment of resistant depression should be involved, at least in part, in its antidepressant properties observed in patients not responding to many antidepressant strategies.