Neurons in the paraventricular nucleus (PVN) are important in regulating autonomic function through projections to the brain stem and spinal cord. Although the vanilloid receptors (VR(1)) are present in the PVN, their physiological function is scarcely known. In this study, we determined the role of VR(1) receptors in the regulation of synaptic inputs and the excitability of spinally projecting PVN neurons. Whole cell patch-clamp recordings were performed on the PVN neurons labeled by a retrograde fluorescence tracer injected into the thoracic spinal cord of rats. Capsaicin significantly increased the frequency of glutamatergic miniature excitatory postsynaptic currents (mEPSCs) without changing the amplitude and decay time constant of mEPSCs. On the other hand, capsaicin had no effect on GABAergic miniature inhibitory postsynaptic currents (mIPSCs). The effect of capsaicin on mEPSCs was abolished by a specific VR(1) antagonist, iodo-resiniferatoxin (iodo-RTX), or ruthenium red. Importantly, iodo-RTX per se significantly reduced the amplitude of evoked EPSCs and the frequency of mEPSCs. Removal of extracellular Ca(2+), but not Cd(2+) treatment, also eliminated the effect of capsaicin on mEPSCs. Furthermore, capsaicin caused a large increase in the firing rate of PVN neurons, and such an effect was abolished in the presence of ionotropic glutamate receptor antagonists. Additionally, the double-immunofluorescence labeling revealed that all of the VR(1) immunoreactivity was colocalized with a presynaptic marker, synaptophysin, in the PVN. Thus this study provides the first evidence that activation of VR(1) receptors excites preautonomic PVN neurons through selective potentiation of glutamatergic synaptic inputs. Presynaptic VR(1) receptors and endogenous capsaicin-like substances in the PVN may represent a previously unidentified mechanism in hypothalamic regulation of the autonomic nervous system.