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AIM: To determine the possible role of hyperpolarization-activated cyclic nucleotide-gated cation channels in the regulation of the mechanosensitivity of vagal and spinal afferents innervating the small intestine.METHODS: The mechanosensory responses of mesenteric afferent activity was measured in an ex vivo murine jejunum preparation.HCN channel activity was recorded through voltage and current clamp in acutely dissociated dorsal root ganglia (DRG) and nodose ganglia (NG) neurons retrogradely labeled from the small intestine through injection of a fluorescent marker (DiI).The isoforms of HCN channels expressed in DRG and NG neurons were examined by immunohistochemistry.RESULTS: Ramp distension of the small intestine evoked biphasic increases in the afferent nerve activity, reflecting the activation of low and high threshold fibers.HCNs blocker CsC1 (5 mmol/L) preferentially inhibited the responses of low threshold fibers to distension and was without significant effect on the high threshold responses.The effect of CsCl was mimicked by the more selective HCN blocker ZD7288 (10 μmo1/L).In 71.4% of DiI labeled DRG neurons (n=20) and 90.9% of DiI labeled NG neurons (n=10), an inward current (Ih current) was evoked by hyperpolarization pulses which was fully eliminated by extracellular CsCl.In neurons expressing Ih current, a typical "sag" was observed upon injection of hyperpolarizing current pulses in current-clamp recordings.CsCl abolished the sag entirely.In some DiI labeled DRG neurons, the Ih current was potentiated by 8-Br-cAMP, which had no effect on the Ih current of DiI labeled NG neurons.Immunohistochemistry revealed differential expression of HCN isoforms in vagal and spinal afferents, with HCN2 and HCN3 seemingly to be the dominant isoform in DRG and NG, respectively.CONCLUSION: HCNs differentially regulate the excitability ofvagal and spinal afferent of murine small intestine.